Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia and Bernard-Soulier Syndrome

*Muhammet Mesut Nezir Engin*

## **Abstract**

Platelets, the smallest cells in the blood, are associated with hemostasis, bowel formation, tissue remodeling, and wound healing. Although the prevalence of inherited platelet disorders is not fully known, it is a rare disease group and is encountered in approximately between 10000 and 1000000. Glanzmann thrombasthenia (GT) and Bernard-Soulier syndrome (BSS) are more frequently observed in inherited platelet disorders. In GT, the platelet aggregation stage due to deficiency or dysfunction of the platelet GPIIb/IIIa complex cannot take place. BSS is a platelet adhesion disorder due to the absence or abnormality of GPIb/IX complex on the platelet surface. If there is bleeding after easy bruising, mucous and oral cavities, menorrhagia, tooth extraction, tonsillectomy, or other surgical interventions, inherited platelet dysfunction should be considered if the platelet count is normal while the bleeding time is long. Firstly, other causes should be investigated by making differential diagnosis of GT and BSS. In this chapter, the definition, etiology, historical process, epidemiology, genetic basis, pathophysiology, clinical findings, diagnosis, differential diagnosis, and the follow-up and treatment approach of GT and BSS will be reviewed according to the current medical literature.

**Keywords:** Glanzmann thrombasthenia, Bernard-Soulier syndrome, thrombocyte function disorder, thrombocyte transfusion, rFVIIa

## **1. Introduction**

Platelets, the smallest cells in the blood, are associated with hemostasis, bowel formation, tissue remodeling, and wound healing. Platelets perform their tasks in ensuring hemostasis in four stages: platelet adhesion, activation of platelet, platelet aggregation, and platelet procoagulant activity. When a damage occurs on the vascular endothelial surface, platelets bind to the collagen, fibronectin, von Willebrand factor, thrombospondin, and fibrinogen in the endothelial substrate with the glycoprotein receptors they carry on their surface. In this way, platelet adhesion takes place. Binding of platelet receptors to their respective ligands causes activation of the platelet. This activation occurs as a result of the change in the cytoskeleton system due to intracellular calcium. By importing the impulse from outside the cell, platelet α-granules secrete their contents. The released ADP causes structural


#### **Table 1.**

*Comparison of genetic mutation, pathophysiology, and affected platelet function status of Glanzmann thrombasthenia and Bernard-Soulier syndrome.*

change in GPIIb/IIIa on the platelet surface. Fibrinogen binds two or more platelets via GPIIb/IIIa receptors that are structurally altered, resulting in platelet aggregation. After aggregation of platelets, platelet plugs are formed at the damage site. Activation of platelets leads to changes in phospholipids on their surface. These phospholipids enable the activation of some clotting factors and perform platelet procoagulant activity [1–6].

The problem in any of the functions of platelets creates a tendency for the primary hemostatic plug not to form and therefore to bleed. Platelet dysfunctions can be hereditary or acquired. Although the prevalence of inherited platelet disorders is not fully known, it is a rare disease group and is frequently encountered in approximately between 10000 and 1000000. Glanzmann thrombasthenia (GT) and Bernard-Soulier syndrome (BSS) are more frequently observed in inherited platelet disorders.

In GT, the platelet aggregation stage due to deficiency or dysfunction of the platelet GPIIb/IIIa complex cannot take place. BSS is a platelet adhesion disorder due to the absence or abnormality of GPIb/IX complex on the platelet surface [1, 7, 8] (**Table 1**).

If there is bleeding after easy bruising, mucous and oral cavities, menorrhagia, tooth extraction, tonsillectomy, or other surgical interventions, inherited platelet dysfunction should be considered if the platelet count is normal while the bleeding time is long. Firstly, other causes should be investigated by making differential diagnosis of GT and BSS [1, 7]. In this chapter, the definition, etiology, historical process, epidemiology, genetic basis, pathophysiology, clinical findings, diagnosis, differential diagnosis, and treatment approach of GT and BSS will be reviewed according to the current medical literature.

#### **2. Glanzmann thrombasthenia**

#### **2.1 Definition**

GT is an autosomal recessive congenital bleeding disorder characterized by a lack of platelet aggregation due to defect and/or deficiency of α IIbβ 3 integrin. Integrin is a platelet fibrinogen receptor, necessary for platelet aggregation and hemostasis. Patients with this disorder often experience lifelong bleeding episodes involving mucocutaneous membranes [9–13].

#### **2.2 History**

This disease was first described by Swiss pediatrician Eduard Glanzmann in 1918 as "hereditary hemorrhagic thrombasthenia." Braunsteiner and Pakesch, on the other hand, reviewed platelet dysfunctions in 1956, after which they identified thrombasthenia as a hereditary disease characterized by normal size platelets that did not spread

**105**

of the epithelial growth factor [20–23].

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

to the surface and did not support clot retraction. The diagnostic characteristics of GT including the absence of platelet aggregation as a primary feature, were reported in 1964 by Caen et al. has been clearly identified by the classical report on 15 French patients. Those patients without platelet aggregation and no clot retraction were later called type I disease patients and those with absent aggregation but residual clot retraction were called type II disease patients; variant disease was first identified in 1987 [8, 10, 14–16].

GT is an autosomal recessive disease with mutations containing the 17q21 chromosome, especially the ITGA2B or ITGB3 genes. GT results when a patient is homozygous for the same mutation or is a compound heterozygote for different mutations. GT is usually caused by decreased or absent expression of αIIb or β3, abnormalities in protein folding, transport of the integrin subunit causing post-translational defective processing or decreased surface expression, or abnormalities affecting protein function. Other defects change the integrin function by altering the ligand binding pocket (interface between αIIb and β3) that changes the cytoplasmic domain and affects the binding of regulators or locks integrin in active

GT has an increasing incidence in populations where marriage between close relatives is an accepted tradition. The prevalence is estimated to be approximately 1:1,000,000 in the general population. Research shows that women are slightly more frequently affected than men. For example, when 177 patients with GT in Paris were examined, 102 (58%) of the patients were shown to be women. In addition, 12 patients were in the USA, 55 were in Israel and Jordan, and 42 were in South India. Some patients may have mild symptoms and are never detected to have GT, so the true prevalence may be higher than reported. Type I is the most common subtype and accounts for about 78% of patients with GT type II and type III (functional variant in receptor) and accounts for about 14% and 8% of cases [8, 9, 12].

The ITGA2B and ITGB3 genes are found on chromosomes 17q21.31 and 17q21.32, respectively, and are independently expressed. Due to autosomal recessive inheritance, compound heterozygosity is common, except for selected ethnic groups, where homozygosity is more likely due to kinship. A higher percentage of pathogenic variants occur in ITGA2B compared to ITGB3, which consists of 15 exons with 788 amino acids, probably because this gene is larger with 30 exons encoding 1039 amino acids. There is a constantly updated database on the Internet http://sinaicentral.mssm.edu/intranet/research/glanzmann: currently, it contains a list of 558 mutations that lead to GT. In addition, when the data in the database were examined, it was found that 269 patients had homozygous mutations. This shows us that consanguineous marriage is an important feature in the heredity of this disease. Some researchers described that pathogenic, nonsense missense, and splice site variants are commonly observed and large deletions and duplications are rarely observed. Pathogenic missense variants cause the disruption of subunit biosynthesis megakaryocytes or prevention of the exit of pro-αIIbβ3 complexes from endoplasmic reticulum to Golgi device or the cell surface mature complexes. Most of the genetic variants affect the β-propeller region of αIIb and domains of β3

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

**2.3 Etiology**

form [8, 9, 12, 13, 17–20].

**2.4 Epidemiology**

**2.5 Genetic basis**

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

to the surface and did not support clot retraction. The diagnostic characteristics of GT including the absence of platelet aggregation as a primary feature, were reported in 1964 by Caen et al. has been clearly identified by the classical report on 15 French patients. Those patients without platelet aggregation and no clot retraction were later called type I disease patients and those with absent aggregation but residual clot retraction were called type II disease patients; variant disease was first identified in 1987 [8, 10, 14–16].

#### **2.3 Etiology**

*Platelets*

**Table 1.**

procoagulant activity [1–6].

*thrombasthenia and Bernard-Soulier syndrome.*

Affected platelet function

Genetic mutation 17q21 chromosome. ITGA2B or

Pathophysiology Deficiency or dysfunction of the

ITGBS genes

platelet GPIIb/IIIa complex

according to the current medical literature.

involving mucocutaneous membranes [9–13].

**2. Glanzmann thrombasthenia**

**2.1 Definition**

**2.2 History**

change in GPIIb/IIIa on the platelet surface. Fibrinogen binds two or more platelets via GPIIb/IIIa receptors that are structurally altered, resulting in platelet aggregation. After aggregation of platelets, platelet plugs are formed at the damage site. Activation of platelets leads to changes in phospholipids on their surface. These phospholipids enable the activation of some clotting factors and perform platelet

*Comparison of genetic mutation, pathophysiology, and affected platelet function status of Glanzmann* 

Aggregation Adhesion

**Glanzmann thrombasthenia Bernard-Soulier syndrome**

GPIbα, GPIbβ, and GPIX genes

Deficiency or dysfunction of the platelet GpIb/V/IX complex

The problem in any of the functions of platelets creates a tendency for the primary

In GT, the platelet aggregation stage due to deficiency or dysfunction of the platelet GPIIb/IIIa complex cannot take place. BSS is a platelet adhesion disorder due to the absence or abnormality of GPIb/IX complex on the platelet surface [1, 7, 8] (**Table 1**). If there is bleeding after easy bruising, mucous and oral cavities, menorrhagia, tooth extraction, tonsillectomy, or other surgical interventions, inherited platelet dysfunction should be considered if the platelet count is normal while the bleeding time is long. Firstly, other causes should be investigated by making differential diagnosis of GT and BSS [1, 7]. In this chapter, the definition, etiology, historical process, epidemiology, genetic basis, pathophysiology, clinical findings, diagnosis, differential diagnosis, and treatment approach of GT and BSS will be reviewed

GT is an autosomal recessive congenital bleeding disorder characterized by a lack of platelet aggregation due to defect and/or deficiency of α IIbβ 3 integrin. Integrin is a platelet fibrinogen receptor, necessary for platelet aggregation and hemostasis. Patients with this disorder often experience lifelong bleeding episodes

This disease was first described by Swiss pediatrician Eduard Glanzmann in 1918 as "hereditary hemorrhagic thrombasthenia." Braunsteiner and Pakesch, on the other hand, reviewed platelet dysfunctions in 1956, after which they identified thrombasthenia as a hereditary disease characterized by normal size platelets that did not spread

hemostatic plug not to form and therefore to bleed. Platelet dysfunctions can be hereditary or acquired. Although the prevalence of inherited platelet disorders is not fully known, it is a rare disease group and is frequently encountered in approximately between 10000 and 1000000. Glanzmann thrombasthenia (GT) and Bernard-Soulier

syndrome (BSS) are more frequently observed in inherited platelet disorders.

**104**

GT is an autosomal recessive disease with mutations containing the 17q21 chromosome, especially the ITGA2B or ITGB3 genes. GT results when a patient is homozygous for the same mutation or is a compound heterozygote for different mutations. GT is usually caused by decreased or absent expression of αIIb or β3, abnormalities in protein folding, transport of the integrin subunit causing post-translational defective processing or decreased surface expression, or abnormalities affecting protein function. Other defects change the integrin function by altering the ligand binding pocket (interface between αIIb and β3) that changes the cytoplasmic domain and affects the binding of regulators or locks integrin in active form [8, 9, 12, 13, 17–20].

#### **2.4 Epidemiology**

GT has an increasing incidence in populations where marriage between close relatives is an accepted tradition. The prevalence is estimated to be approximately 1:1,000,000 in the general population. Research shows that women are slightly more frequently affected than men. For example, when 177 patients with GT in Paris were examined, 102 (58%) of the patients were shown to be women. In addition, 12 patients were in the USA, 55 were in Israel and Jordan, and 42 were in South India. Some patients may have mild symptoms and are never detected to have GT, so the true prevalence may be higher than reported. Type I is the most common subtype and accounts for about 78% of patients with GT type II and type III (functional variant in receptor) and accounts for about 14% and 8% of cases [8, 9, 12].

#### **2.5 Genetic basis**

The ITGA2B and ITGB3 genes are found on chromosomes 17q21.31 and 17q21.32, respectively, and are independently expressed. Due to autosomal recessive inheritance, compound heterozygosity is common, except for selected ethnic groups, where homozygosity is more likely due to kinship. A higher percentage of pathogenic variants occur in ITGA2B compared to ITGB3, which consists of 15 exons with 788 amino acids, probably because this gene is larger with 30 exons encoding 1039 amino acids. There is a constantly updated database on the Internet http://sinaicentral.mssm.edu/intranet/research/glanzmann: currently, it contains a list of 558 mutations that lead to GT. In addition, when the data in the database were examined, it was found that 269 patients had homozygous mutations. This shows us that consanguineous marriage is an important feature in the heredity of this disease. Some researchers described that pathogenic, nonsense missense, and splice site variants are commonly observed and large deletions and duplications are rarely observed. Pathogenic missense variants cause the disruption of subunit biosynthesis megakaryocytes or prevention of the exit of pro-αIIbβ3 complexes from endoplasmic reticulum to Golgi device or the cell surface mature complexes. Most of the genetic variants affect the β-propeller region of αIIb and domains of β3 of the epithelial growth factor [20–23].

#### **2.6 Pathophysiology**

The main mechanism in the pathophysiology of GT is the qualitative or quantitative disorder of the autosomal recessive platelet surface receptor of GPIIb/IIIa (ITG αIIbβ3). As a result, it results in erroneous platelet aggregation and reduced clot retraction. ITG αIIbβ3 is a large heterodimeric cell transmembrane receptor consisting of a larger αIIb and a smaller P3 subunit. These subunits were not covalently attached to permit bidirectional signal between the cell membrane and extracellular matrix when initiating intracellular signaling pathways. It contains cytoplasmic and transmembrane domains that act as the junction point for intracellular signal molecules and proteins. The activation of ITG αIIbβ3 is provided by the B3 subunit consisting of large disulfide epidermal growth factor (EGF) domains. Calcium binding sites for complex formation and platelet-platelet adhesion are found on the p-propeller region of the αIIb subunit. The receptor head function consisting of binding fibrinogen, VWF, vitronectin, and fibronectin is necessary for platelet communications by regulation of cell migration, platelet aggregation and adhesion, and the formation of a thrombus [24].

The ITGA2B gene, located on chromosome 17q21.31, encodes the platelet GPαIIb, while the gene encoding the glycoprotein subunit IIIa is found in chromosome ITGB3, 17q21.32. Both subunits are collected from the precursors of the endoplasmic reticulum by further processing in the Golgi apparatus. Nurden et al. examined more closely the p-propeller ectodomain mutations of the αIIb subunit. Nurden et al. concluded that a large series of mutations affecting the β-propeller field interrupts calcium binding and has numerous harmful effects on αIIbβ3 expression and function, and causes different types of GT [21, 25, 26].

Homozygous or heterozygous mutations in both gene locations determine the severity of abnormality seen in GT. Mutations can stop subunit production, prevent complex formation, and/or inhibit intracellular trade. When complex build-up is prevented, the subunits of αIIb or β3 are now broken. Now based on the expression and functionality of the subunits, GT is classified into three types: <5% of αIIbβ3 now specifies type I GT; now 5–20% of αIIbβ3 is type II GT; and rarely >20% of residual αIIbβ3 with dysfunctional features make up the variant type GT. Acquired GT is usually the result of autoantibody attack on platelet αIIbβ3 or isoanorbons that inhibit proper function. The production of autoantibodies has been associated with multiple hematological conditions, including immune thrombocytopenic purpura, non-Hodgkin lymphoma, multiple myeloma, myelodysplastic syndrome, hairy cell leukemia, and acute lymphoblastic leukemia, as well as platelet transfusions [21, 24, 26].

#### **2.7 Clinical manifestations**

The most common symptoms of bleeding are purpura, nosebleeds (60–80%), gingival bleeding (20–60%), and menorrhagia (60–90%). Gastrointestinal bleeding in the form of melena or hematochezia is found in 10–20% and intracranial hemorrhage is developed in 1–2%. Mucocutaneous bleeding may occur spontaneously or following minimal trauma. Epistaxis is the most common cause of severe bleeding especially in children. Menorrhagia is quite common in affected women, and there is a higher risk of serious bleeding during menarche due to the prolonged estrogenic effect on the proliferative endometrium that occurs during anovulatory cycles. Bleeding complications during pregnancy are rare; however, there is a high risk of obstetric bleeding during and after birth. Hematuria and spontaneous hemarthrosis have been described in some cases, but are generally not part of the bleeding phenotype. Currently, specific cuts could

**107**

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

not be identified to define a positive bleeding score. Although the types of bleeding are consistent among individuals, the degree of bleeding is quite variable. The severity of bleeding (except for menorrhagia and pregnancy-related bleeding)

The diagnosis of GT is often not noticed, because many platelet disorders share common clinical and laboratory features. GT should be remembered in the differential diagnosis of medical history (insidious or bleeding episodes or severe bleeding after minor trauma), family history (consanguinity). In order to diagnose GT, it is necessary to choose the appropriate laboratory tests. A normal platelet count on a on a routine blood smear does not exclude the diagnosis of GT. Because patients with GT usually do not show any abnormalities in the number of platelets, complete blood count may be normal or show iron deficiency. Prothrombin time and activated partial thromboplastin time may be normal if bleeding time is prolonged; further

In the evaluation of peripheral blood smear with light microscopy, normal platelet count and normal granular size should be. If the bleeding is severe and/ or chronic, patients may have a red cell distribution width that increases with low hemoglobin, microcytosis, and secondary iron deficiency. Other abnormalities of

Prothrombin time (PT), activated thromboplastin time (aPTT), and fibrinogen values are usually normal unless a patient is evaluated in a significant acute bleeding

Platelet function analyzer PFA-100 provides a measure of platelet function under reduced platelets. Very long closing times (>300 s) show GT but are heat-specific. Some other disorders such as severe von Willebrand disease, Bernard Soulier syndrome, and afibrinogenemia may produce the same result. A normal PFA-100 reveals a very high negative predictive value for GT and generally excludes this diagnosis [27].

Light transmission aggregometry (LTA) is widely accepted as the gold standard diagnostic tool for evaluating platelet function. Although this test provides specific data, LTA is very time-consuming and dependent on staff and requires the use of

Although it can be performed using whole blood samples and lower volumes, there is insufficient evidence to support equivalent sensitivity and reproducibility

the complete blood count (CBC) suggest an alternative diagnosis [20].

environment and there is no evidence of consumption coagulopathy [20].

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

decreases with age [8, 20].

investigations should be done [24].

*2.8.1 Complete blood count*

*2.8.2 Coagulation screening tests*

*2.8.3 Platelet function screening tests*

*2.8.4 Platelet light transmission aggregometry*

*2.8.5 Whole blood impedance aggregometry*

experienced laboratories [24].

compared to LTA [28].

Let us examine the laboratory methods in detail.

**2.8 Diagnosis**

not be identified to define a positive bleeding score. Although the types of bleeding are consistent among individuals, the degree of bleeding is quite variable. The severity of bleeding (except for menorrhagia and pregnancy-related bleeding) decreases with age [8, 20].

## **2.8 Diagnosis**

*Platelets*

**2.6 Pathophysiology**

The main mechanism in the pathophysiology of GT is the qualitative or quantitative disorder of the autosomal recessive platelet surface receptor of GPIIb/IIIa (ITG αIIbβ3). As a result, it results in erroneous platelet aggregation and reduced clot retraction. ITG αIIbβ3 is a large heterodimeric cell transmembrane receptor consisting of a larger αIIb and a smaller P3 subunit. These subunits were not covalently attached to permit bidirectional signal between the cell membrane and extracellular matrix when initiating intracellular signaling pathways. It contains cytoplasmic and transmembrane domains that act as the junction point for intracellular signal molecules and proteins. The activation of ITG αIIbβ3 is provided by the B3 subunit consisting of large disulfide epidermal growth factor (EGF) domains. Calcium binding sites for complex formation and platelet-platelet adhesion are found on the p-propeller region of the αIIb subunit. The receptor head function consisting of binding fibrinogen, VWF, vitronectin, and fibronectin is necessary for platelet communications by regulation of cell migration, platelet aggregation

The ITGA2B gene, located on chromosome 17q21.31, encodes the platelet GPαIIb, while the gene encoding the glycoprotein subunit IIIa is found in chromosome ITGB3, 17q21.32. Both subunits are collected from the precursors of the endoplasmic reticulum by further processing in the Golgi apparatus. Nurden et al. examined more closely the p-propeller ectodomain mutations of the αIIb subunit. Nurden et al. concluded that a large series of mutations affecting the β-propeller field interrupts calcium binding and has numerous harmful effects on αIIbβ3 expression and function, and causes different types of GT [21, 25, 26].

Homozygous or heterozygous mutations in both gene locations determine the severity of abnormality seen in GT. Mutations can stop subunit production, prevent complex formation, and/or inhibit intracellular trade. When complex build-up is prevented, the subunits of αIIb or β3 are now broken. Now based on the expression and functionality of the subunits, GT is classified into three types: <5% of αIIbβ3 now specifies type I GT; now 5–20% of αIIbβ3 is type II GT; and rarely >20% of residual αIIbβ3 with dysfunctional features make up the variant type GT. Acquired GT is usually the result of autoantibody attack on platelet αIIbβ3 or isoanorbons that inhibit proper function. The production of autoantibodies has been associated with multiple hematological conditions, including immune thrombocytopenic purpura, non-Hodgkin lymphoma, multiple myeloma, myelodysplastic syndrome, hairy cell leukemia, and acute lymphoblastic leukemia, as

The most common symptoms of bleeding are purpura, nosebleeds (60–80%),

gingival bleeding (20–60%), and menorrhagia (60–90%). Gastrointestinal bleeding in the form of melena or hematochezia is found in 10–20% and intracranial hemorrhage is developed in 1–2%. Mucocutaneous bleeding may occur spontaneously or following minimal trauma. Epistaxis is the most common cause of severe bleeding especially in children. Menorrhagia is quite common in affected women, and there is a higher risk of serious bleeding during menarche due to the prolonged estrogenic effect on the proliferative endometrium that occurs during anovulatory cycles. Bleeding complications during pregnancy are rare; however, there is a high risk of obstetric bleeding during and after birth. Hematuria and spontaneous hemarthrosis have been described in some cases, but are generally not part of the bleeding phenotype. Currently, specific cuts could

and adhesion, and the formation of a thrombus [24].

well as platelet transfusions [21, 24, 26].

**2.7 Clinical manifestations**

**106**

The diagnosis of GT is often not noticed, because many platelet disorders share common clinical and laboratory features. GT should be remembered in the differential diagnosis of medical history (insidious or bleeding episodes or severe bleeding after minor trauma), family history (consanguinity). In order to diagnose GT, it is necessary to choose the appropriate laboratory tests. A normal platelet count on a on a routine blood smear does not exclude the diagnosis of GT. Because patients with GT usually do not show any abnormalities in the number of platelets, complete blood count may be normal or show iron deficiency. Prothrombin time and activated partial thromboplastin time may be normal if bleeding time is prolonged; further investigations should be done [24].

Let us examine the laboratory methods in detail.

### *2.8.1 Complete blood count*

In the evaluation of peripheral blood smear with light microscopy, normal platelet count and normal granular size should be. If the bleeding is severe and/ or chronic, patients may have a red cell distribution width that increases with low hemoglobin, microcytosis, and secondary iron deficiency. Other abnormalities of the complete blood count (CBC) suggest an alternative diagnosis [20].

### *2.8.2 Coagulation screening tests*

Prothrombin time (PT), activated thromboplastin time (aPTT), and fibrinogen values are usually normal unless a patient is evaluated in a significant acute bleeding environment and there is no evidence of consumption coagulopathy [20].

## *2.8.3 Platelet function screening tests*

Platelet function analyzer PFA-100 provides a measure of platelet function under reduced platelets. Very long closing times (>300 s) show GT but are heat-specific. Some other disorders such as severe von Willebrand disease, Bernard Soulier syndrome, and afibrinogenemia may produce the same result. A normal PFA-100 reveals a very high negative predictive value for GT and generally excludes this diagnosis [27].

### *2.8.4 Platelet light transmission aggregometry*

Light transmission aggregometry (LTA) is widely accepted as the gold standard diagnostic tool for evaluating platelet function. Although this test provides specific data, LTA is very time-consuming and dependent on staff and requires the use of experienced laboratories [24].

### *2.8.5 Whole blood impedance aggregometry*

Although it can be performed using whole blood samples and lower volumes, there is insufficient evidence to support equivalent sensitivity and reproducibility compared to LTA [28].

The best way to fully diagnose GT is by mutation analysis. The genomic DNA sequence of 45 exons containing the αIIb and 3 unit should be investigated together with the junctions of the ITGB3 and ITGA2B gene and established mutations should be confirmed by a second DNA sample analysis. Genetic analysis is clinically useful for confirming the diagnosis, identifying carriers at risk, reproductive risk counseling for a particular couple/family, and definitive prenatal or preimplantation genetic diagnosis. Consequently, the diagnosis of GT involves the presence of normal platelet count (typically at the lower end of normal), long bleeding time, and long PFA time [24].

### **2.9 Differential diagnosis**

Leukocyte adhesion deficiency type III, RASGRP2-related platelet dysfunction, BSS, Hermansky-Pudlak syndrome, von Willebrand disease, Medich platelet syndrome, Scott syndrome, and Acquired Glanzmann thrombasthenia are among the differential diagnoses [20].

#### **2.10 Treatment**

A gradual treatment standard is applied in GT treatment. The first treatment for mild bleeding is local measures including local compression, cauterization, stitching, or ice therapy. The treatment applied in case of unresponsiveness to these treatments or in heavier bleeding is antifibrinolytic therapy first, followed by platelet transfusion, and recombinant active factor VII (rFVIIa) if bleeding persists. Platelet concentrates may be single-donor and HLA-matched due to the risk of developing alloantibodies against the platelet glycoproteins, αIIbβ3, or αIIbβ3, and/or the HLA antigens. Platelet concentrates may be repeatedly transfused. If HLA-matched platelets are not found, patients should be given leukocyte-reduced platelets. This has been shown to reduce the rate of HLA immunization. Patients with severe bleeding cases should continue to receive platelet transfusion for 48 h until bleeding ceases and wound healing occurs in operated cases. These patients should be trained to avoid over-the-counter drugs that increase the risk of bleeding, such as nonsteroidal anti-inflammatories and aspirin products. Prescription drugs that may affect hemostasis should be carefully monitored [9, 24, 25, 29, 30].

Let us examine the treatment of GT according to the frequently observed conditions.

#### *2.10.1 Treatment of minor to moderate bleeding*

Local measures and/or antifibrinolytic drugs can stop mild to moderate bleeding. Local measures include compression, gelatin sponges, fibrin sealants, and topical thrombin. Antifibrinolytic agents include epsilon aminocaproic acid and tranexamic acid. Since both agents can be given orally or intravenously, they have been used successfully in the treatment of nosebleeds, bleeding gums, and menorrhagia, as well as prophylaxis before tooth extraction and other minor surgical procedures. Antifibrinolytic agents, such as tranexamic acid, can be used as a mouthwash for gingival bleeding. Antifibrinolytic use in cases of hematuria should be avoided due to the risk of a clot in the urinary tract and should be used with caution in patients undergoing procedures at high risk of thrombosis [26].

#### *2.10.2 Treatment in epistaxis*

One of the most common bleeding symptoms in GT patients is epistaxis. Local compression to epistaxis, application of tampons to the nose, topical thrombin,

**109**

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

antifibrinolytics, and a combination of these may respond. If bleeding persists, further treatments with platelets transfusion and/or rFVIIa should be given. Antifibrinolytic agents, nasal cautery, rFVIIa, and nasal packing with synthetic materials may be used to control bleeding. If these treatments fail nasal packing with salt pork strips may be successfully used for life-threatening nasal hemorrhage

Antifibrinolytic agents should be first-line therapy to control menorrhagia. If it fails, hormone supplementation either progesterone alone or progesterone with estrogen may be given. A continuous estrogen-progestin oral contraceptive agent or intramuscular depot medroxyprogesterone acetate regimen given every 3 months in women with GT has been used successfully. It can be tried on hormonal intrauterine devices to reduce bleeding. Severe menorrhagia, which may be seen in many women with GT, can be treated with high-dose conjugated estrogen intravenously for 24–48 h and later by following with a combination of high doses of oral estrogen-progestin. Intensive menstrual bleeding does not always respond to typical treatment. rFVIIa has been utilized with anecdotal success in GT when anti-fibrinolytics and platelet transfusions did not control excessive menorrhagia. In addition, surgical treatments such as hysterectomy or endometrial ablation in

Pregnant women with GT have high complications and are best managed in a specialized center with a multidisciplinary team. Although most complications are associated with bleeding and occur during delivery, treatment of pregnant GT patients should start in the prenatal period. According to the recommendations in the guidelines, platelet transfusions, or rFVIIa in combination with an antifibrinolytic can be used as a prophylaxis for vaginal delivery. A systematic review of 35 pregnant women with GT showed that hemorrhage during or after delivery is common and severe, and occurred up to 20 days postpartum. If patients were not given any platelet transfusions as prophylaxis, they were more likely to experience postpartum hemorrhage (63% versus 38%). The use of rFVIIa as prophylaxis was documented in three pregnancies, and it did not prevent hemorrhage in those cases. A study showed that maternal platelet alloantibodies were documented in 16 pregnancies, and plasma exchange successfully reduced the alloantibody titer in one case. Four of the 16 cases resulted in neonatal deaths, 3 of which resulted from intracranial hemorrhage between 24- and 31-weeks' gestation. One study reported successful use of rFVIIa for permanent postpartum hemorrhage. In one study, the patient was followed up with the diagnosis of GT and 18 units of random platelet concentrates, 6 units of apheresis platelet concentrates, and 2 units of erythrocyte suspension were given in the peripartum period. Although various forms of treatment have been reported about the treatment of obstetric bleeding occurring during and after birth of women pregnant with GT, there is no consensus on the most appropriate treatment. Further studies on this subject are needed [21, 26, 32].

Platelet transfusion allows partial correction of functional defect in patients with GT. Platelet transfusion is the standard prophylaxis when local precautions and/or antifibrinolytics cannot control bleeding and the patient is undergoing

treatment-resistant severe menorrhagia are therapeutic options [24].

*2.10.4 Treatment of postpartum hemorrhage*

*2.10.5 Role of transfusions in the therapy of GT*

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

in a child with GT [6, 31].

*2.10.3 Treatment of menorrhagia*

#### *Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

antifibrinolytics, and a combination of these may respond. If bleeding persists, further treatments with platelets transfusion and/or rFVIIa should be given. Antifibrinolytic agents, nasal cautery, rFVIIa, and nasal packing with synthetic materials may be used to control bleeding. If these treatments fail nasal packing with salt pork strips may be successfully used for life-threatening nasal hemorrhage in a child with GT [6, 31].

#### *2.10.3 Treatment of menorrhagia*

*Platelets*

**2.9 Differential diagnosis**

the differential diagnoses [20].

**2.10 Treatment**

conditions.

*2.10.1 Treatment of minor to moderate bleeding*

The best way to fully diagnose GT is by mutation analysis. The genomic DNA sequence of 45 exons containing the αIIb and 3 unit should be investigated together with the junctions of the ITGB3 and ITGA2B gene and established mutations should be confirmed by a second DNA sample analysis. Genetic analysis is clinically useful for confirming the diagnosis, identifying carriers at risk, reproductive risk counseling for a particular couple/family, and definitive prenatal or preimplantation genetic diagnosis. Consequently, the diagnosis of GT involves the presence of normal platelet count (typically at the lower end of normal), long bleeding time, and long PFA time [24].

Leukocyte adhesion deficiency type III, RASGRP2-related platelet dysfunction, BSS, Hermansky-Pudlak syndrome, von Willebrand disease, Medich platelet syndrome, Scott syndrome, and Acquired Glanzmann thrombasthenia are among

A gradual treatment standard is applied in GT treatment. The first treatment for mild bleeding is local measures including local compression, cauterization, stitching, or ice therapy. The treatment applied in case of unresponsiveness to these treatments or in heavier bleeding is antifibrinolytic therapy first, followed by platelet transfusion, and recombinant active factor VII (rFVIIa) if bleeding persists. Platelet concentrates may be single-donor and HLA-matched due to the risk of developing alloantibodies against the platelet glycoproteins, αIIbβ3, or αIIbβ3, and/or the HLA antigens. Platelet concentrates may be repeatedly transfused. If HLA-matched platelets are not found, patients should be given leukocyte-reduced platelets. This has been shown to reduce the rate of HLA immunization. Patients with severe bleeding cases should continue to receive platelet transfusion for 48 h until bleeding ceases and wound healing occurs in operated cases. These patients should be trained to avoid over-the-counter drugs that increase the risk of bleeding, such as nonsteroidal anti-inflammatories and aspirin products. Prescription drugs that may affect hemostasis should be carefully monitored [9, 24, 25, 29, 30]. Let us examine the treatment of GT according to the frequently observed

Local measures and/or antifibrinolytic drugs can stop mild to moderate bleeding. Local measures include compression, gelatin sponges, fibrin sealants, and topical thrombin. Antifibrinolytic agents include epsilon aminocaproic acid and tranexamic acid. Since both agents can be given orally or intravenously, they have been used successfully in the treatment of nosebleeds, bleeding gums, and menorrhagia, as well as prophylaxis before tooth extraction and other minor surgical procedures. Antifibrinolytic agents, such as tranexamic acid, can be used as a mouthwash for gingival bleeding. Antifibrinolytic use in cases of hematuria should be avoided due to the risk of a clot in the urinary tract and should be used with cau-

One of the most common bleeding symptoms in GT patients is epistaxis. Local compression to epistaxis, application of tampons to the nose, topical thrombin,

tion in patients undergoing procedures at high risk of thrombosis [26].

**108**

*2.10.2 Treatment in epistaxis*

Antifibrinolytic agents should be first-line therapy to control menorrhagia. If it fails, hormone supplementation either progesterone alone or progesterone with estrogen may be given. A continuous estrogen-progestin oral contraceptive agent or intramuscular depot medroxyprogesterone acetate regimen given every 3 months in women with GT has been used successfully. It can be tried on hormonal intrauterine devices to reduce bleeding. Severe menorrhagia, which may be seen in many women with GT, can be treated with high-dose conjugated estrogen intravenously for 24–48 h and later by following with a combination of high doses of oral estrogen-progestin. Intensive menstrual bleeding does not always respond to typical treatment. rFVIIa has been utilized with anecdotal success in GT when anti-fibrinolytics and platelet transfusions did not control excessive menorrhagia. In addition, surgical treatments such as hysterectomy or endometrial ablation in treatment-resistant severe menorrhagia are therapeutic options [24].

#### *2.10.4 Treatment of postpartum hemorrhage*

Pregnant women with GT have high complications and are best managed in a specialized center with a multidisciplinary team. Although most complications are associated with bleeding and occur during delivery, treatment of pregnant GT patients should start in the prenatal period. According to the recommendations in the guidelines, platelet transfusions, or rFVIIa in combination with an antifibrinolytic can be used as a prophylaxis for vaginal delivery. A systematic review of 35 pregnant women with GT showed that hemorrhage during or after delivery is common and severe, and occurred up to 20 days postpartum. If patients were not given any platelet transfusions as prophylaxis, they were more likely to experience postpartum hemorrhage (63% versus 38%). The use of rFVIIa as prophylaxis was documented in three pregnancies, and it did not prevent hemorrhage in those cases. A study showed that maternal platelet alloantibodies were documented in 16 pregnancies, and plasma exchange successfully reduced the alloantibody titer in one case. Four of the 16 cases resulted in neonatal deaths, 3 of which resulted from intracranial hemorrhage between 24- and 31-weeks' gestation. One study reported successful use of rFVIIa for permanent postpartum hemorrhage. In one study, the patient was followed up with the diagnosis of GT and 18 units of random platelet concentrates, 6 units of apheresis platelet concentrates, and 2 units of erythrocyte suspension were given in the peripartum period. Although various forms of treatment have been reported about the treatment of obstetric bleeding occurring during and after birth of women pregnant with GT, there is no consensus on the most appropriate treatment. Further studies on this subject are needed [21, 26, 32].

#### *2.10.5 Role of transfusions in the therapy of GT*

Platelet transfusion allows partial correction of functional defect in patients with GT. Platelet transfusion is the standard prophylaxis when local precautions and/or antifibrinolytics cannot control bleeding and the patient is undergoing

major surgery. It is not uncommon for patients with severe bleeding after trauma or delivery to require multiple platelet transfusions. Multiple platelet transfusions can be performed if necessary. An important risk associated with platelet transfusion is the possibility of developing isoantibodies. Up to 30% of patients develop anti-GPIIb/IIIa or anti-HLA antibodies after platelet transfusion. Platelet alloimmunization can lead to relative or absolute platelet refractory, causing rapid destruction of platelets and therapeutic failure of platelet transfusions. For this reason, platelet transfusions should be reserved for only major surgeries, life-threatening bleeding, and significant bleeding that does not respond to the above measure. When possible, platelet concentrates should be single-donor derived and HLA-matched. If HLA-matched platelets are not available, patients should be given leukocytereduced platelets because this has been shown to reduce the rate of HLA immunization. Transfusions in women of reproductive age should ideally be avoided as the antibodies can cross the placenta and affect the fetus [13, 21, 22, 33–35].

#### *2.10.6 Use of rFVIIa in GT*

Treatment of rFVIIa in a GT patient was successfully used for severe and uncontrolled bleeding in a 2-year-old child in 1996 for the first time. The worldwide use of rFVIIa continued afterward, and it was observed that most patients with GT were effective in successfully controlling bleeding. But it was also observed that it was not effective in all GT patients. The mechanism of rFVIIa is not fully delineated. It is thought are poorly attached to the surface of platelets and increase the activation of factor IX and X, thereby increasing thrombin production. Increased amount of thrombin increases platelet adhesion and supports platelet aggregation, including those not containing GPIIb/IIIa [6, 25, 33].

High success rates and relatively low risks associated with the use of rFVIIa as a treatment or prevention of bleeding in GT patients have yielded good results, especially in those who are refractory to platelet transfusion or have antiplatelet antibodies. HLA-compatible platelets have been used in the past and have been recommended as prophylaxis for major surgical procedures, including cesarean section. rFVIIa can be used to completely prevent platelet transfusion, which will reduce the risk of platelet alloimmunization in case of life-threatening bleeding when local measures and antifibrinolytics fail. The optimal dosage for use in GT patients has not been established. However, the recommended dose is bolus injections of 90 mcg/kg intravenously 3 times a day or every 2 h until bleeding stops, followed by one or more maintenance doses [6, 8, 21, 24, 33, 36, 37].

The adverse or thromboembolic events have not been reported in patients given the rFVIIa bolus. The incidence of thrombotic events is not known in GT patients treated with rFVIIa. Controlled clinical trials are needed to further assess risk [26, 36].

A UK study showed that rFVIIa was successful in 71% of patients treated within 12 h of onset, but only after 12 h, only 18% of patients responded to rFVIIa. Therefore, rFVIIa should be administered as early as possible in bleeding episodes. Minor surgeries in GT patients have been successfully treated by rFVIIa prophylaxis without the need for platelet transfusion. rFVIIa prophylaxis used is recommended by the United Kingdom Hemophilia Centre Doctors' Organization for minor surgical prophylaxis including dental extractions [6, 26, 33].

#### *2.10.7 Other treatments*

Desmopressin (DDAVP) causes VWF, FVIII, and tissue plasminogen activator to be released into the plasma. Although DDAVP is successful in treating other platelet disorders, there is little data to support its use in GT patients [26].

**111**

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

tranexamic acid, and embolization, but responded to bevacizumab [25].

Hematopoietic stem cell transplantation (HSCT) provides a treatment for patients with severe, recurrent bleeding episodes and resistant cases to platelet transfusion due to platelet alloantibodies. There is currently no clearly defined algorithm for transplantation in GT, and HSCT is rarely used for GT. The first successful bone marrow transplantion in GT was performed in a 4-year-old child with anti-GPIIb/IIIa antibodies in 1985. It has been reported in the literature that successful stem cell transplantation has been performed in 19 severe GT patients [26, 33, 39].

Gene therapy is very promising for GT patients to provide a treatment with significant progress using different techniques, vectors, and model organisms [40–42].

BSS is a rare autosomal recessive platelet dysfunction that is characterized by a low levels, absence, or dysfunction of the GpIb/V/IX complex on the platelet

adhesion, abnormal prothrombin consumption, and low-surviving large platelets. Mucocutaneous hemorrhages such as purpura, epistaxis, oral mucosa bleeding, GIS bleeding, and menorrhagia are generally seen in BSS as in other platelet function

BSS with autosomal recessive transition was first described by Bernard and Soulier in 1948 as congenital bleeding disorder characterized by thrombocytopenia

Mutations in GP1BA [GPIbα], GP1BB (GPIbß), and GP9 (GPIX) cause BSS. There of the four genes encode for the subunita of the GP Ib-IX-V complex. This key platelet receptor constituted of four subunits, GPIbα, GPIbß, GPIX, and GP5 (GPV), which included in the ratio 2:4:2:1 in endoplasmic reticulum. They because mature in Golgi apparatus before localizing at the cell surface. The GPIb-IX-V complex can attach to von Willebrand factor, fitting together like a lock and its key. Von Willebrand factor is located on the inside surface of blood vessels when there is an injury. These platelets form clots, plugging holes in the blood vessels to help stop bleeding. Due to the specified conditions occurring in BSS, clot formation is

is characterized by decreased platelet

Rituximab (anti-CD20) is a human-mouse chimeric monoclonal antibody that targets the B cell CD20 antigen. Successful treatment has been reported for acquired GT patients. Multiple case reports have demonstrated the efficacy of rituximab in patients with treatment-resistant GT and bleeding symptoms or ecchymosis [38]. Bevacizumab (Avastin) is an anti-VEGF antibody used in combination with chemotherapy in various cancers. A single case report in the literature documented success using bevacizumab in a patient with type I GT who had severe, recurrent GI bleeding due to angiodysplasia. The patient was resistant to platelet transfusion,

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

*2.10.8 Future therapy*

**3.1 Definition**

disorders [43, 44].

and large platelets [45].

**3.2 History**

**3.3 Etiology**

**3. Bernard-Soulier syndrome**

surface. BSS thrombocytopenia <20,000/mm3

impaired and excessive bleeding occurs [46–50].

#### *Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

Rituximab (anti-CD20) is a human-mouse chimeric monoclonal antibody that targets the B cell CD20 antigen. Successful treatment has been reported for acquired GT patients. Multiple case reports have demonstrated the efficacy of rituximab in patients with treatment-resistant GT and bleeding symptoms or ecchymosis [38].

Bevacizumab (Avastin) is an anti-VEGF antibody used in combination with chemotherapy in various cancers. A single case report in the literature documented success using bevacizumab in a patient with type I GT who had severe, recurrent GI bleeding due to angiodysplasia. The patient was resistant to platelet transfusion, tranexamic acid, and embolization, but responded to bevacizumab [25].

Hematopoietic stem cell transplantation (HSCT) provides a treatment for patients with severe, recurrent bleeding episodes and resistant cases to platelet transfusion due to platelet alloantibodies. There is currently no clearly defined algorithm for transplantation in GT, and HSCT is rarely used for GT. The first successful bone marrow transplantion in GT was performed in a 4-year-old child with anti-GPIIb/IIIa antibodies in 1985. It has been reported in the literature that successful stem cell transplantation has been performed in 19 severe GT patients [26, 33, 39].

### *2.10.8 Future therapy*

*Platelets*

*2.10.6 Use of rFVIIa in GT*

those not containing GPIIb/IIIa [6, 25, 33].

major surgery. It is not uncommon for patients with severe bleeding after trauma or delivery to require multiple platelet transfusions. Multiple platelet transfusions can be performed if necessary. An important risk associated with platelet transfusion is the possibility of developing isoantibodies. Up to 30% of patients develop anti-GPIIb/IIIa or anti-HLA antibodies after platelet transfusion. Platelet alloimmunization can lead to relative or absolute platelet refractory, causing rapid destruction of platelets and therapeutic failure of platelet transfusions. For this reason, platelet transfusions should be reserved for only major surgeries, life-threatening bleeding, and significant bleeding that does not respond to the above measure. When possible, platelet concentrates should be single-donor derived and HLA-matched. If HLA-matched platelets are not available, patients should be given leukocytereduced platelets because this has been shown to reduce the rate of HLA immunization. Transfusions in women of reproductive age should ideally be avoided as the

antibodies can cross the placenta and affect the fetus [13, 21, 22, 33–35].

Treatment of rFVIIa in a GT patient was successfully used for severe and uncontrolled bleeding in a 2-year-old child in 1996 for the first time. The worldwide use of rFVIIa continued afterward, and it was observed that most patients with GT were effective in successfully controlling bleeding. But it was also observed that it was not effective in all GT patients. The mechanism of rFVIIa is not fully delineated. It is thought are poorly attached to the surface of platelets and increase the activation of factor IX and X, thereby increasing thrombin production. Increased amount of thrombin increases platelet adhesion and supports platelet aggregation, including

High success rates and relatively low risks associated with the use of rFVIIa as a treatment or prevention of bleeding in GT patients have yielded good results, especially in those who are refractory to platelet transfusion or have antiplatelet antibodies. HLA-compatible platelets have been used in the past and have been recommended as prophylaxis for major surgical procedures, including cesarean section. rFVIIa can be used to completely prevent platelet transfusion, which will reduce the risk of platelet alloimmunization in case of life-threatening bleeding when local measures and antifibrinolytics fail. The optimal dosage for use in GT patients has not been established. However, the recommended dose is bolus injections of 90 mcg/kg intravenously 3 times a day or every 2 h until bleeding stops,

The adverse or thromboembolic events have not been reported in patients given the rFVIIa bolus. The incidence of thrombotic events is not known in GT patients treated with rFVIIa. Controlled clinical trials are needed to further assess risk [26, 36]. A UK study showed that rFVIIa was successful in 71% of patients treated within 12 h of onset, but only after 12 h, only 18% of patients responded to rFVIIa. Therefore, rFVIIa should be administered as early as possible in bleeding episodes. Minor surgeries in GT patients have been successfully treated by rFVIIa prophylaxis without the need for platelet transfusion. rFVIIa prophylaxis used is recommended by the United Kingdom Hemophilia Centre Doctors' Organization for minor surgical

Desmopressin (DDAVP) causes VWF, FVIII, and tissue plasminogen activator to be released into the plasma. Although DDAVP is successful in treating other platelet

followed by one or more maintenance doses [6, 8, 21, 24, 33, 36, 37].

disorders, there is little data to support its use in GT patients [26].

prophylaxis including dental extractions [6, 26, 33].

**110**

*2.10.7 Other treatments*

Gene therapy is very promising for GT patients to provide a treatment with significant progress using different techniques, vectors, and model organisms [40–42].

## **3. Bernard-Soulier syndrome**

### **3.1 Definition**

BSS is a rare autosomal recessive platelet dysfunction that is characterized by a low levels, absence, or dysfunction of the GpIb/V/IX complex on the platelet surface. BSS thrombocytopenia <20,000/mm3 is characterized by decreased platelet adhesion, abnormal prothrombin consumption, and low-surviving large platelets. Mucocutaneous hemorrhages such as purpura, epistaxis, oral mucosa bleeding, GIS bleeding, and menorrhagia are generally seen in BSS as in other platelet function disorders [43, 44].

### **3.2 History**

BSS with autosomal recessive transition was first described by Bernard and Soulier in 1948 as congenital bleeding disorder characterized by thrombocytopenia and large platelets [45].

## **3.3 Etiology**

Mutations in GP1BA [GPIbα], GP1BB (GPIbß), and GP9 (GPIX) cause BSS. There of the four genes encode for the subunita of the GP Ib-IX-V complex. This key platelet receptor constituted of four subunits, GPIbα, GPIbß, GPIX, and GP5 (GPV), which included in the ratio 2:4:2:1 in endoplasmic reticulum. They because mature in Golgi apparatus before localizing at the cell surface. The GPIb-IX-V complex can attach to von Willebrand factor, fitting together like a lock and its key. Von Willebrand factor is located on the inside surface of blood vessels when there is an injury. These platelets form clots, plugging holes in the blood vessels to help stop bleeding. Due to the specified conditions occurring in BSS, clot formation is impaired and excessive bleeding occurs [46–50].

## **3.4 Epidemiology**

The incidence of BSS is estimated to be 1 in 1 million live births, but is likely to be higher since it is often misdiagnosed [50]. In a study with 97 BSS patients in Iran, consanguineous marriage was reported in 81% of the cases' families [51].

## **3.5 Genetic basis**

BSS occurs as a result of homozygous or compound heterozygous mutations that affect the expression of genes encoding GPIbα, GPIbß, and GPIX proteins. Two types of mutations have been reported in the GP Ib-IX-V complex. The first one is biallelic mutations, often homozygous mutations. It is characterized by a severe decrease or absence of the GP Ib-IX-V complex. To date, more than 50 biallelic mutations have been identified in the GPIbα, GPIbß, and GP9 gene. In a few cases, there is a compound heterozygous mutation. Most of the mutations identified are missense and nonsense mutations. Most BSS mutations occur in the GPIbα gene, and most of these mutations lead to a decrease in GPIbα expression on the platelet surface, and some to a loss of function. GPIbα is connected to GPIbß by disulfide bond. These are connected by noncovalent bonds with GPIX and GPV. GPV is the proteolytic subunit in this complex, and its extracellular part is destroyed by GPIbαbound thrombin activates platelets. As a result, mutations in GP1BA, GPIbß, and GP9 in humans generally lead to a decrease in the total expression of the GP Ib-IX-V complex on the platelet surface and the disease occurs [6, 43, 44, 50, 52].

### **3.6 Pathophysiology**

Platelets play a critical role in normal primary hemostasis and clot formation. There are specific GP receptors on the platelet membrane, which function in platelet adhesion, activation, and aggregation. The GPIb-IX-V receptor complex is responsible for platelet adhesion through its interaction with von Willebrand factor on the exposed subendothelium. The GPIb-IX-V receptor complex is composed of four transmembrane polypeptide subunits-disulfide-linked alpha and beta subunits of GPIb, and noncovalently bound subunits GPIX and GPV. The platelets of BSS cases lack or have a dysfunctional GPIb-IX-V receptor. This results in defective adhesion to the subendothelium. The dysfunctional platelets found in BSS can result from one of several different glycoprotein mutations such as missense, nonsense, or deletion mutations of the GPIbα, GPIbβ, or GPIX genes [53].

## **3.7 Clinical manifestations**

As with other inherited platelet disorders, BSS can manifest with a tendency to bleed in early childhood. Mucocutaneous bleeding is seen predominantly. Easy bruising, purpura, epistaxis, bleeding gums, menorrhagia, and excessive bleeding after surgery or trauma are common symptoms. Menorrhagia is an important problem for female BSS patients. Prolonged menstruation may be the first symptom to help diagnose BSS in some patients. Although the severity of bleeding is associated with a genetic defect that affects receptor function and platelet count, it is highly variable in patients with the same mutations. Although bleeding sites are well defined for BSS, it is difficult to predict the severity of bleeding in patients with BSS. In some cases, no serious bleeding is observed and diagnosis may not be established until adulthood. Other genetic differences and acquired conditions affecting hemostasis are thought to affect the severity of bleeding in these patients,

**113**

**Figure 1.**

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

lets is variable. The platelet count typically ranges from 30 to 200 × 103

Giant platelets are seen in peripheral blood smear (**Figure 1**). In order to the differential diagnosis of other giant platelet syndromes, leukocyte counts and morphology should be carefully examined. Skin bleeding time and PFA-100 closure time are found to be prolonged. Routine coagulation tests should be found normal. Prothrombin consumption and thrombin generation tests are found markedly decreased because of the defective binding of FXI and thrombin. Results of platelet aggregation studies are pathognomonic for BSS. In vitro platelet aggregation studies characteristically indicate that aggregation with ristocetin failed and responded slowly with low doses of thrombin. Flow cytometric analysis of platelet also show characteristic for BSS normal binding with CD41 (GPIIb) and CD61 (GPIIIa) antibodies, but defective binding with CD42a (GPIX), CD42b (GP Ib), CD42c (GP Ib), and CD42d (GPV) antibodies suggest BSS. Immunoblotting after separating components of the GP Ib-IX-V complex with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) may describe the defective fragments but needs specialized interpretation. Also, in recent years, most families are offered molecular

genetic testing to identify which gene carries the mutations [6, 53, 56–59].

platelet syndrome are among the differential diagnoses [52, 53].

*Giant platelet appearance in peripheral blood smear in Bernard-Soulier syndrome.*

GT, idiopathic thrombocytopenic purpura (ITP), von Willebrand disease, May-Hegglin anomaly, and other inherited giant platelet disorders, for example, gray

BSS treatment is generally supportive. Platelet transfusion is used to treat when surgery is needed or when there is a risk of life-threatening bleeding. The patient may develop antiplatelet antibodies due to the presence of glycoproteins Ib/IX/V,

studies related to this need to be done. Heterozygotes may not have signs of bleeding, but giant platelets may appear in peripheral blood smear [6, 43, 46, 50, 54, 55].

Although thrombocytopenia is generally observed in BSS, the number of plate-

/μL.

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

**3.8 Diagnosis**

**3.9 Differential diagnosis**

**3.10 Treatment**

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

studies related to this need to be done. Heterozygotes may not have signs of bleeding, but giant platelets may appear in peripheral blood smear [6, 43, 46, 50, 54, 55].

## **3.8 Diagnosis**

*Platelets*

**3.4 Epidemiology**

**3.5 Genetic basis**

**3.6 Pathophysiology**

genes [53].

**3.7 Clinical manifestations**

The incidence of BSS is estimated to be 1 in 1 million live births, but is likely to be higher since it is often misdiagnosed [50]. In a study with 97 BSS patients in Iran,

BSS occurs as a result of homozygous or compound heterozygous mutations that affect the expression of genes encoding GPIbα, GPIbß, and GPIX proteins. Two types of mutations have been reported in the GP Ib-IX-V complex. The first one is biallelic mutations, often homozygous mutations. It is characterized by a severe decrease or absence of the GP Ib-IX-V complex. To date, more than 50 biallelic mutations have been identified in the GPIbα, GPIbß, and GP9 gene. In a few cases, there is a compound heterozygous mutation. Most of the mutations identified are missense and nonsense mutations. Most BSS mutations occur in the GPIbα gene, and most of these mutations lead to a decrease in GPIbα expression on the platelet surface, and some to a loss of function. GPIbα is connected to GPIbß by disulfide bond. These are connected by noncovalent bonds with GPIX and GPV. GPV is the proteolytic subunit in this complex, and its extracellular part is destroyed by GPIbαbound thrombin activates platelets. As a result, mutations in GP1BA, GPIbß, and GP9 in humans generally lead to a decrease in the total expression of the GP Ib-IX-V

consanguineous marriage was reported in 81% of the cases' families [51].

complex on the platelet surface and the disease occurs [6, 43, 44, 50, 52].

Platelets play a critical role in normal primary hemostasis and clot formation.

There are specific GP receptors on the platelet membrane, which function in platelet adhesion, activation, and aggregation. The GPIb-IX-V receptor complex is responsible for platelet adhesion through its interaction with von Willebrand factor on the exposed subendothelium. The GPIb-IX-V receptor complex is composed of four transmembrane polypeptide subunits-disulfide-linked alpha and beta subunits of GPIb, and noncovalently bound subunits GPIX and GPV. The platelets of BSS cases lack or have a dysfunctional GPIb-IX-V receptor. This results in defective adhesion to the subendothelium. The dysfunctional platelets found in BSS can result from one of several different glycoprotein mutations such as missense, nonsense, or deletion mutations of the GPIbα, GPIbβ, or GPIX

As with other inherited platelet disorders, BSS can manifest with a tendency to bleed in early childhood. Mucocutaneous bleeding is seen predominantly. Easy bruising, purpura, epistaxis, bleeding gums, menorrhagia, and excessive bleeding after surgery or trauma are common symptoms. Menorrhagia is an important problem for female BSS patients. Prolonged menstruation may be the first symptom to help diagnose BSS in some patients. Although the severity of bleeding is associated with a genetic defect that affects receptor function and platelet count, it is highly variable in patients with the same mutations. Although bleeding sites are well defined for BSS, it is difficult to predict the severity of bleeding in patients with BSS. In some cases, no serious bleeding is observed and diagnosis may not be established until adulthood. Other genetic differences and acquired conditions affecting hemostasis are thought to affect the severity of bleeding in these patients,

**112**

Although thrombocytopenia is generally observed in BSS, the number of platelets is variable. The platelet count typically ranges from 30 to 200 × 103 /μL. Giant platelets are seen in peripheral blood smear (**Figure 1**). In order to the differential diagnosis of other giant platelet syndromes, leukocyte counts and morphology should be carefully examined. Skin bleeding time and PFA-100 closure time are found to be prolonged. Routine coagulation tests should be found normal. Prothrombin consumption and thrombin generation tests are found markedly decreased because of the defective binding of FXI and thrombin. Results of platelet aggregation studies are pathognomonic for BSS. In vitro platelet aggregation studies characteristically indicate that aggregation with ristocetin failed and responded slowly with low doses of thrombin. Flow cytometric analysis of platelet also show characteristic for BSS normal binding with CD41 (GPIIb) and CD61 (GPIIIa) antibodies, but defective binding with CD42a (GPIX), CD42b (GP Ib), CD42c (GP Ib), and CD42d (GPV) antibodies suggest BSS. Immunoblotting after separating components of the GP Ib-IX-V complex with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) may describe the defective fragments but needs specialized interpretation. Also, in recent years, most families are offered molecular genetic testing to identify which gene carries the mutations [6, 53, 56–59].

## **3.9 Differential diagnosis**

GT, idiopathic thrombocytopenic purpura (ITP), von Willebrand disease, May-Hegglin anomaly, and other inherited giant platelet disorders, for example, gray platelet syndrome are among the differential diagnoses [52, 53].

### **3.10 Treatment**

BSS treatment is generally supportive. Platelet transfusion is used to treat when surgery is needed or when there is a risk of life-threatening bleeding. The patient may develop antiplatelet antibodies due to the presence of glycoproteins Ib/IX/V,

which are present on the transfused platelets but absent from the patient's own platelets. Although some publications have suggested that patients should receive platelets from human leukocyte antigen-matched donors in order to avoid alloimmunization, this is not currently a widely accepted strategy. Antifibrinolytic agents such as p-aminocaproic acid or tranexamic acid may be useful for mucosal bleeding. rFVIIa has been reported to reduce bleeding times in patients with BSS. Desmopressin has been found to shorten bleeding episodes for some patients. A test dose should be used to determine those patients who will benefit. Stem cell transplantation has been successfully used to treat two children with BSS who had severe, life-threatening bleeding episodes. Transplantation should be considered in severe disorders when the patients have developed antiplatelet antibodies. Patients with BSS should be counseled about the importance of preventing even minor trauma as well as avoiding aspirin-containing medications and other platelet antagonists [52, 53, 60].

## **4. General recommendations for GT, BSS, and other inherited diseases**


## **5. Conclusion**

Genetic defects of the blood platelet membrane glycoproteins, GPIIb-IIIa (CD41/CD61) and GPIb-IX-V (CD42) are the origin of several rare bleeding disorders, the best known of which are GT and BSS. GT results in defective or absence of GPIIbIIIa. As a result of this, patients with GT are unable to undergo platelet aggregation, a critical step in stemming blood flow. Either gene can be affected and mutations leading to lack of expression or to expression of poorly functional forms have been identified. BSS occurs due to defective or absence of GPIb-IX-V. As a result of this, platelets from patients with BSS are unable to adhere to the damaged vessel wall at high-shear stress and also have a reduced platelet response to thrombin.

Since GT and BSS are rare diseases, diagnosis of patients can be delayed. When diagnosed early, patients will be able to prevent bleeding that may occur due to protective measures. If there is bleeding after easy bruising, mucous and oral cavities, menorrhagia, tooth extraction, tonsillectomy, or other surgical interventions, GT or BSS should be considered among the differential diagnoses. Although GT cannot be diagnosed with routine laboratory tests, BSS is suspected in the presence of thrombocytopenia and giant platelet. Detailed examination is required for a definitive diagnosis. Treatment includes local measures, platelet infusion, rFVIIa,

**115**

**Author details**

Düzce, Turkey

Muhammet Mesut Nezir Engin

provided the original work is properly cited.

Department of Child Health and Diseases, Faculty of Medicine, Düzce University,

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

\*Address all correspondence to: doktormesut@hotmail.com

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

and other treatments. Although there is no permanent treatment for now, research is still ongoing. For this, it is more important for patients to avoid situations that

The author would like to thank Prof. Dr. Kenan KOCABAY, who has contributed greatly to his specialty education in medicine and has helped write this book

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

may increase their tendency to bleed.

**Acknowledgments**

chapter.

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

and other treatments. Although there is no permanent treatment for now, research is still ongoing. For this, it is more important for patients to avoid situations that may increase their tendency to bleed.

## **Acknowledgments**

*Platelets*

antagonists [52, 53, 60].

required.

**5. Conclusion**

to thrombin.

which are present on the transfused platelets but absent from the patient's own platelets. Although some publications have suggested that patients should receive platelets from human leukocyte antigen-matched donors in order to avoid alloimmunization, this is not currently a widely accepted strategy. Antifibrinolytic agents such as p-aminocaproic acid or tranexamic acid may be useful for mucosal bleeding. rFVIIa has been reported to reduce bleeding times in patients with BSS. Desmopressin has been found to shorten bleeding episodes for some patients. A test dose should be used to determine those patients who will benefit. Stem cell transplantation has been successfully used to treat two children with BSS who had severe, life-threatening bleeding episodes. Transplantation should be considered in severe disorders when the patients have developed antiplatelet antibodies. Patients with BSS should be counseled about the importance of preventing even minor trauma as well as avoiding aspirin-containing medications and other platelet

**4. General recommendations for GT, BSS, and other inherited diseases**

1.Should pay attention to dental health by brushing your teeth regularly.

2.Avoid sports activities with potential trauma (wrestling, boxing etc.).

3.Should not use salicylate and nonsteroid drugs that affect platelet function.

4.Oral contraceptives should be considered in patients with hypermenorrhoea.

5.It should be vaccinated against hepatitis A and B since blood products may be

6.The patient should carry a small information card describing the condition,

Genetic defects of the blood platelet membrane glycoproteins, GPIIb-IIIa (CD41/CD61) and GPIb-IX-V (CD42) are the origin of several rare bleeding disorders, the best known of which are GT and BSS. GT results in defective or absence of GPIIbIIIa. As a result of this, patients with GT are unable to undergo platelet aggregation, a critical step in stemming blood flow. Either gene can be affected and mutations leading to lack of expression or to expression of poorly functional forms have been identified. BSS occurs due to defective or absence of GPIb-IX-V. As a result of this, platelets from patients with BSS are unable to adhere to the damaged vessel wall at high-shear stress and also have a reduced platelet response

Since GT and BSS are rare diseases, diagnosis of patients can be delayed. When diagnosed early, patients will be able to prevent bleeding that may occur due to protective measures. If there is bleeding after easy bruising, mucous and oral cavities, menorrhagia, tooth extraction, tonsillectomy, or other surgical interventions, GT or BSS should be considered among the differential diagnoses. Although GT cannot be diagnosed with routine laboratory tests, BSS is suspected in the presence of thrombocytopenia and giant platelet. Detailed examination is required for a definitive diagnosis. Treatment includes local measures, platelet infusion, rFVIIa,

blood group, and what to do in an emergency.

**114**

The author would like to thank Prof. Dr. Kenan KOCABAY, who has contributed greatly to his specialty education in medicine and has helped write this book chapter.

## **Author details**

Muhammet Mesut Nezir Engin Department of Child Health and Diseases, Faculty of Medicine, Düzce University, Düzce, Turkey

\*Address all correspondence to: doktormesut@hotmail.com

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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management of haemostatic defects in children and adults. British Journal of Haematology. 2005;**130**(1):3-10

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[29] Grainger JD, Thachil J, Will AM. How we treat the platelet glycoprotein defects; Glanzmann thrombasthenia and Bernard Soulier syndrome in children and adults. British Journal of Haematology. 2018;**182**(5):621-632

[30] Temizkan RC, Engin MMN, Türen B, Kocabay K. Evaluation of pediatric patients with gastrointestinal bleeding following a single dose ingestion of NSAID. Archives of Emergency Medicine and Intensive

[31] Blickstein D, Dardik R, Rosenthal E, et al. Acquired thrombasthenia due to inhibitory effect of glycoprotein IIbIIIa autoantibodies. The Israel Medical Association Journal. 2014;**16**(5):307-310

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[33] Bennett JS. Structure and function of the platelet integrin alphaIIbbeta3. The Journal of Clinical Investigation.

Nurden P, Heilig R, Nurden AT. Natural history of platelet antibody formation against alphaIIbbeta3 in a French cohort of Glanzmann thrombasthenia patients. Haemophilia. 2012;**18**(3):e201-e209

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2017;**15**(10):2045-2052

Care. 2020;**3**(1):01-05

2014;**52**:208-211

2005;**115**(12):3363-3369

[34] Fiore M, Firah N, Pillois X,

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

[19] Humphreys I, Saraiya S, Belenky W,

[20] Botero JP, Lee K, Branchford BR, Bray PF, Freson K, Lambert MP. Glanzmann thrombasthenia: Genetic

Haematologica. 2020;**105**(4):888-894

[21] Nurden AT, Fiore M, Nurden P, Pillois X. Glanzmann thrombasthenia: A review of ITGA2B and ITGB3 defects with emphasis on variants, phenotypic variability, and mouse models. Blood.

[22] Nurden AT, Pillois X. ITGA2B and ITGB3 gene mutations associated with Glanzmann thrombasthenia. Platelets.

[23] Richards S, Aziz N, Bale S, et al. Standards and guidelines for the interpretation of sequence variants: A joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genetics in Medicine. 2015;**17**(5):405-424

[24] Solh T, Botsford A, Solh M. Glanzmann's thrombasthenia:

Blood Medicine. 2015;**6**:219-227

2002;**119**(4):901-904

2012;**20**(10):1102

[25] Stevens RF, Meyer S. Fanconi and Glanzmann: The men and their works. British Journal of Haematology.

[26] Fiore M, Nurden AT, Nurden P, Seligsohn U. Clinical utility gene card for: Glanzmann thrombasthenia. European Journal of Human Genetics.

[27] Harrison P. The role of PFA-100 testing in the investigation and

Pathogenesis, diagnosis, and current and emerging treatment options. Journal of

basis and clinical correlates.

2011;**118**(23):59966005

2018;**29**(1):98101

Dworkin J. Nasal packing with strips of cured pork as treatment for uncontrollable epistaxis in a patient with Glanzmann thrombasthenia. The Annals of Otology, Rhinology, and Laryngology. 2011;**120**(11):732-736

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

[19] Humphreys I, Saraiya S, Belenky W, Dworkin J. Nasal packing with strips of cured pork as treatment for uncontrollable epistaxis in a patient with Glanzmann thrombasthenia. The Annals of Otology, Rhinology, and Laryngology. 2011;**120**(11):732-736

[20] Botero JP, Lee K, Branchford BR, Bray PF, Freson K, Lambert MP. Glanzmann thrombasthenia: Genetic basis and clinical correlates. Haematologica. 2020;**105**(4):888-894

[21] Nurden AT, Fiore M, Nurden P, Pillois X. Glanzmann thrombasthenia: A review of ITGA2B and ITGB3 defects with emphasis on variants, phenotypic variability, and mouse models. Blood. 2011;**118**(23):59966005

[22] Nurden AT, Pillois X. ITGA2B and ITGB3 gene mutations associated with Glanzmann thrombasthenia. Platelets. 2018;**29**(1):98101

[23] Richards S, Aziz N, Bale S, et al. Standards and guidelines for the interpretation of sequence variants: A joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genetics in Medicine. 2015;**17**(5):405-424

[24] Solh T, Botsford A, Solh M. Glanzmann's thrombasthenia: Pathogenesis, diagnosis, and current and emerging treatment options. Journal of Blood Medicine. 2015;**6**:219-227

[25] Stevens RF, Meyer S. Fanconi and Glanzmann: The men and their works. British Journal of Haematology. 2002;**119**(4):901-904

[26] Fiore M, Nurden AT, Nurden P, Seligsohn U. Clinical utility gene card for: Glanzmann thrombasthenia. European Journal of Human Genetics. 2012;**20**(10):1102

[27] Harrison P. The role of PFA-100 testing in the investigation and management of haemostatic defects in children and adults. British Journal of Haematology. 2005;**130**(1):3-10

[28] Al Ghaithi R, Drake S, Watson SP, Morgan NV, Harrison P. Comparison of multiple electrode aggregometry with lumi-aggregometry for the diagnosis of patients with mild bleeding disorders. Journal of Thrombosis and Haemostasis. 2017;**15**(10):2045-2052

[29] Grainger JD, Thachil J, Will AM. How we treat the platelet glycoprotein defects; Glanzmann thrombasthenia and Bernard Soulier syndrome in children and adults. British Journal of Haematology. 2018;**182**(5):621-632

[30] Temizkan RC, Engin MMN, Türen B, Kocabay K. Evaluation of pediatric patients with gastrointestinal bleeding following a single dose ingestion of NSAID. Archives of Emergency Medicine and Intensive Care. 2020;**3**(1):01-05

[31] Blickstein D, Dardik R, Rosenthal E, et al. Acquired thrombasthenia due to inhibitory effect of glycoprotein IIbIIIa autoantibodies. The Israel Medical Association Journal. 2014;**16**(5):307-310

[32] Ateş S, Molla T, Özkal F, Çetin G, Başaranoğlu G, Dane B. Peripartum management of a pregnant with Glanzmann's thrombasthenia: A case report. The Medical Bulletin of Haseki. 2014;**52**:208-211

[33] Bennett JS. Structure and function of the platelet integrin alphaIIbbeta3. The Journal of Clinical Investigation. 2005;**115**(12):3363-3369

[34] Fiore M, Firah N, Pillois X, Nurden P, Heilig R, Nurden AT. Natural history of platelet antibody formation against alphaIIbbeta3 in a French cohort of Glanzmann thrombasthenia patients. Haemophilia. 2012;**18**(3):e201-e209

[35] Siddiq S, Clark A, Mumford A. A systematic review of the management

**116**

*Platelets*

**References**

2005;**1**(2):31-40

[1] Kaptan K. Functional platelet disorders. Turkiye Klinikleri Journal of Internal Medical Sciences.

of Rare Diseases. 2016;**1**:10. DOI:

[12] Iqbal I, Farhan S, Ahmed N. Glanzmann thrombasthenia: A clinicopathological profile. Journal of the College of Physicians and Surgeons–

Pakistan. 2016;**26**(8):647-650

of Glanzmann thrombasthenia. Transfusion Medicine Reviews.

[14] Glanzmann E. Hereditare

J Kinderkranken. 1918;**88**:113

[15] Braunsteiner H, Pakesch F. Thrombocytoasthenia and thrombocytopathia. Old names and new diseases. Blood. 1956;**11**:965-976

[16] Caen JP, Castaldi PA, Lecrec JC, Inceman S, Larrieu MJ, Probst M, et al. Glanzmann's thrombasthenia. I. Congenital bleeding disorders with long bleeding time and normal platelet count. American Journal of Medicine.

[17] Nurden AT, Freson K, Seligsohn U.

Haemophilia. 2012;**18**(Suppl 4):154-160

Inherited platelet disorders.

[18] Vogel ER, VanOosten SK, Holman MA, et al. Cigarette smoke enhances proliferation and extracellular matrix deposition by human fetal airway smooth muscle. American Journal of Physiology. Lung Cellular and Molecular Physiology.

2014;**307**(12):978-986

hamorrhagische thrombasthenie. Ein Beitrag zur Pathologie der Blutplattchen.

2016;**30**(2):92-99

1966;**44**:4

[13] Poon MC, Di Minno G, d'Oiron R, Zotz R. New insights into the treatment

[11] Doherty D, Singleton E, Byrne M, Ryan K, O'Connell NM, O'Donnell JS, et al. Missed at first Glanz: Glanzmann thrombasthenia initially misdiagnosed as Von Willebrand disease. Transfusion and Apheresis Science. 2019;**58**(1):58-60

10.1186/1750-1172-1-10

[2] Fışgın T, Koca D. Inherited platelet disorders. Turkish Children's Journal of

Hematology/Oncology Clinics of North

Hematology. 2011;**5**(1):1-10

America. 2007;**21**(4):66384

[3] Neunert CE, Journeycake JM. Congenital platelet disorders.

[4] Streif W, Knöfler R, Eberl W. Inherited disorders of platelet function

in pediatric clinical practice: A diagnostic challenge. Klinische Pädiatrie. 2010;**222**:203-208

[5] Cattaneo M. Inherited plateletbased bleeding disorders. Journal of Thrombosis and Haemostasis.

disorders including Glanzmann thrombasthenia and Bernard-Soulier syndrome. Hematology. American Society of Hematology. Education Program. 2013;**2013**:268-275

Alloimmunization in congenital deficiencies of platelet surface glycoproteins: Focus on Glanzmann's thrombasthenia and Bernard–Soulier's syndrome. Seminars in Thrombosis and Hemostasis. 2018;**44**(06):604-614

[8] George JN, Caen JP, Nurden AT. Glanzmann's thrombasthenia: The spectrum of clinical disease. Blood.

[9] Krause KA, Graham BC. Glanzmann Thrombasthenia. StatPearls Publishing. 2020. Available from: https://www.ncbi.

nlm.nih.gov/books/NBK538270/

[10] Nurden AT. Glanzmann thrombasthenia. Orphanet Journal

[7] Poon MC, d'Oiron R.

1990;**75**(07):1383-1395

[6] Diz-Kucukkaya R. Inherited platelet

2003;**1**:1628-1636

and outcomes of pregnancy in Glanzmann thrombasthenia. Haemophilia. 2011;**17**(5):e858-e869

[36] Nurden AT, Ruan J, Pasquet KM, et al. A novel 196Leu to Pro substitution in the beta3 subunit of the alphaIIbbeta3 integrin in a patient with a variant form of Glanzmann thrombasthenia. Platelets. 2002;**13**(2):101-111

[37] Cattaneo M, Cerletti C, Harrison P, et al. Recommendations for the standardization of light transmission aggregometry: A consensus of the working party from the platelet physiology subcommittee of SSC/ISTH. Journal of Thrombosis and Haemostasis. 2013;**11**:1183-1189

[38] Morel-Kopp MC, Melchior C, Chen P, et al. A naturally occurring point mutation in the beta3 integrin MIDAS-like domain affects differently alphavbeta3 and alphaIIIbbeta3 receptor function. Thrombosis and Haemostasis. 2001;**86**(6):1425-1434

[39] Nurden AT, Pillois X, Wilcox DA. Glanzmann thrombasthenia: State of the art and future directions. Seminars in Thrombosis and Hemostasis. 2013;**39**(6):642-655

[40] Wilcox DA, Olsen JC, Ishizawa L, et al. Megakaryocyte-targeted synthesis of the integrin beta(3)-subunit results in the phenotypic correction of Glanzmann thrombasthenia. Blood. 2000;**95**(12):3645-3651

[41] Fang J, Hodivala-Dilke K, Johnson BD, et al. Therapeutic expression of the platelet-specific integrin, alphaIIbbeta3, in a murine model for Glanzmann thrombasthenia. Blood. 2005;**106**(8):2671-2679

[42] Sullivan SK, Mills JA, Koukouritaki SB, et al. High-level transgene expression in induced pluripotent stem cell-derived megakaryocytes: Correction of

Glanzmann thrombasthenia. Blood. 2014;**123**(5):753-757

[43] Lambert MP, Poncz M. Inherited platelet disorders. In: Orkin SH, Fisher DE, Ginsburg D, Look AT, Lux SE, Nathan DG, editors. Nathan and Oski's Hematology and Oncology of Infancy and Childhood. 8th ed. Philadelphia: Elsevier Saunders; 2015. pp. 1167-1203

[44] Tokgöz H, Çalışkan Ü. Clinical and genotypic findings in patients with Bernard Soulier syndrome: A single center experience. Turkish Journal of Pediatric Disease. 2017;**1**:51-55

[45] Bernard J, Soulier JP. Sur une nouvelle variete de dystrophie thrombocytaire-hemorragipare congenitale. La Semaine des Hôpitaux de Paris. 1948;**24**:3217-3222

[46] López JA, Andrews RK, Afshar-Kharghan V, Berndt MC. Bernard-Soulier syndrome. Blood. 1998;**91**:4397-4418

[47] Luo SZ, Mo X, Afshar-Kharghan V, Srinivasan S, López JA, Li R. Glycoprotein Ibalpha forms disulfide bonds with 2 glycoprotein Ibbeta subunits in the resting platelet. Blood. 2007;**109**(603):609

[48] Ulsemer U, Strassel C, Baas M-J, Salamero J, Chasserot-Golaz S, Cazenave J-P, et al. Biosynthesis and post-translational processing of normal and mutant platelet glycoprotein GPIb-IX. The Biochemical Journal. 2001;**358**:295-303

[49] Li R, Emsley J. The organizing principle of the platelet glycoprotein Ib-IX-V complex. Journal of Thrombosis and Haemostasis. 2013;**11**:605-614

[50] Savoia A, Kunishima S, De Rocco D, Zieger B, Rand ML, Pujol-Moix N, et al. Spectrum of the mutations in Bernard-Soulier syndrome. Human Mutation. 2014;**35**(9):1033-1045

**119**

860-880

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia…*

*DOI: http://dx.doi.org/10.5772/intechopen.93299*

[51] Toogeh G, Keyhani M, Sharifian R, et al. A study of Bernard-Soulier syndrome in Tehran, Iran. Archives of Iranian Medicine. 2010;**13**(6):549-551

[52] Balduini CL, Savoia A. Genetics of familial forms of thrombocytopenia. Human Genetics. 2012;**131**:1821-1832

[53] Pham A, Wang J. Bernard-Soulier syndrome: An inherited platelet disorder. Archives of Pathology & Laboratory Medicine. 2007;**131**(12):1834-1836

[54] Andrews RK, Berdt MC. Bernard-Soulier syndrome: An update. Seminars

in Thrombosis and Hemostasis.

[55] Farhan S, Iqbal I, Ahmed N. Bernard Soulier syndrome: 10 years' experience at a tertiary care hospital. Pakistan Journal of Medical Sciences.

[56] Bolton-Maggs PHB, Chalmers EA, Collins PW, et al. A review of inherited platelet disorders with guidelines for their management on behalf of the UKHCDO. British Journal of Haematology. 2006;**135**(5):603-633

[57] Balduini CL, Pecci A, Noris P. Diagnosis and management of inherited

[58] Harrison P, Mackie I, Mumford A, et al. Guidelines for the laboratory investigation of heritable disorders of platelet function. British Journal of Haematology. 2011;**155**(1):30-44

[59] Miller JL. Glycoprotein analysis for the diagnostic evaluation of platelet disorders. Seminars in Thrombosis and

[60] Balduini CL, Iolascon A, Savoia A. Inherited thrombocytopenias: From genes to therapy. Hema. 2002;**87**:

Hemostasis. 2009;**35**(2):224232

thrombocytopenias. Seminars in Thrombosis and Hemostasis.

2013;**39**(2):161-171

2013;**39**(6):656-662

2019;**35**(3):705-708

*Bleeding Disorders Associated with Abnormal Platelets: Glanzmann Thrombasthenia… DOI: http://dx.doi.org/10.5772/intechopen.93299*

[51] Toogeh G, Keyhani M, Sharifian R, et al. A study of Bernard-Soulier syndrome in Tehran, Iran. Archives of Iranian Medicine. 2010;**13**(6):549-551

*Platelets*

and outcomes of pregnancy in Glanzmann thrombasthenia. Haemophilia. 2011;**17**(5):e858-e869

Platelets. 2002;**13**(2):101-111

[37] Cattaneo M, Cerletti C,

2013;**11**:1183-1189

2001;**86**(6):1425-1434

2013;**39**(6):642-655

2000;**95**(12):3645-3651

2005;**106**(8):2671-2679

[42] Sullivan SK, Mills JA,

Koukouritaki SB, et al. High-level transgene expression in induced pluripotent stem cell-derived megakaryocytes: Correction of

[41] Fang J, Hodivala-Dilke K,

of the platelet-specific integrin, alphaIIbbeta3, in a murine model for Glanzmann thrombasthenia. Blood.

Harrison P, et al. Recommendations for the standardization of light

[38] Morel-Kopp MC, Melchior C, Chen P, et al. A naturally occurring point mutation in the beta3 integrin MIDAS-like domain affects differently alphavbeta3 and alphaIIIbbeta3 receptor function. Thrombosis and Haemostasis.

[39] Nurden AT, Pillois X, Wilcox DA. Glanzmann thrombasthenia: State of the art and future directions. Seminars in Thrombosis and Hemostasis.

[40] Wilcox DA, Olsen JC, Ishizawa L, et al. Megakaryocyte-targeted synthesis of the integrin beta(3)-subunit results in the phenotypic correction of Glanzmann thrombasthenia. Blood.

Johnson BD, et al. Therapeutic expression

transmission aggregometry: A consensus of the working party from the platelet physiology subcommittee of SSC/ISTH. Journal of Thrombosis and Haemostasis.

[36] Nurden AT, Ruan J, Pasquet KM, et al. A novel 196Leu to Pro substitution in the beta3 subunit of the alphaIIbbeta3 integrin in a patient with a variant form of Glanzmann thrombasthenia.

Glanzmann thrombasthenia. Blood.

[43] Lambert MP, Poncz M. Inherited platelet disorders. In: Orkin SH, Fisher DE, Ginsburg D, Look AT, Lux SE, Nathan DG, editors. Nathan and Oski's Hematology and Oncology of Infancy and Childhood. 8th ed. Philadelphia: Elsevier Saunders; 2015.

[44] Tokgöz H, Çalışkan Ü. Clinical and genotypic findings in patients with Bernard Soulier syndrome: A single center experience. Turkish Journal of Pediatric Disease. 2017;**1**:51-55

[45] Bernard J, Soulier JP. Sur une nouvelle variete de dystrophie thrombocytaire-hemorragipare congenitale. La Semaine des Hôpitaux

de Paris. 1948;**24**:3217-3222

[46] López JA, Andrews RK, Afshar-Kharghan V, Berndt MC. Bernard-Soulier syndrome. Blood.

Srinivasan S, López JA, Li R.

[47] Luo SZ, Mo X, Afshar-Kharghan V,

Glycoprotein Ibalpha forms disulfide bonds with 2 glycoprotein Ibbeta subunits in the resting platelet. Blood.

[48] Ulsemer U, Strassel C, Baas M-J, Salamero J, Chasserot-Golaz S, Cazenave J-P, et al. Biosynthesis and post-translational processing of normal and mutant platelet glycoprotein GPIb-IX. The Biochemical Journal.

[49] Li R, Emsley J. The organizing principle of the platelet glycoprotein Ib-IX-V complex. Journal of Thrombosis and Haemostasis. 2013;**11**:605-614

[50] Savoia A, Kunishima S, De Rocco D, Zieger B, Rand ML, Pujol-Moix N, et al. Spectrum of the mutations in Bernard-Soulier syndrome. Human Mutation.

1998;**91**:4397-4418

2007;**109**(603):609

2001;**358**:295-303

2014;**35**(9):1033-1045

2014;**123**(5):753-757

pp. 1167-1203

**118**

[52] Balduini CL, Savoia A. Genetics of familial forms of thrombocytopenia. Human Genetics. 2012;**131**:1821-1832

[53] Pham A, Wang J. Bernard-Soulier syndrome: An inherited platelet disorder. Archives of Pathology & Laboratory Medicine. 2007;**131**(12):1834-1836

[54] Andrews RK, Berdt MC. Bernard-Soulier syndrome: An update. Seminars in Thrombosis and Hemostasis. 2013;**39**(6):656-662

[55] Farhan S, Iqbal I, Ahmed N. Bernard Soulier syndrome: 10 years' experience at a tertiary care hospital. Pakistan Journal of Medical Sciences. 2019;**35**(3):705-708

[56] Bolton-Maggs PHB, Chalmers EA, Collins PW, et al. A review of inherited platelet disorders with guidelines for their management on behalf of the UKHCDO. British Journal of Haematology. 2006;**135**(5):603-633

[57] Balduini CL, Pecci A, Noris P. Diagnosis and management of inherited thrombocytopenias. Seminars in Thrombosis and Hemostasis. 2013;**39**(2):161-171

[58] Harrison P, Mackie I, Mumford A, et al. Guidelines for the laboratory investigation of heritable disorders of platelet function. British Journal of Haematology. 2011;**155**(1):30-44

[59] Miller JL. Glycoprotein analysis for the diagnostic evaluation of platelet disorders. Seminars in Thrombosis and Hemostasis. 2009;**35**(2):224232

[60] Balduini CL, Iolascon A, Savoia A. Inherited thrombocytopenias: From genes to therapy. Hema. 2002;**87**: 860-880

**121**

**Chapter 6**

**Abstract**

**1. Introduction**

150 × 10<sup>9</sup>

complications.

*Bernhard Resch*

Thrombocytopenia in Neonates

ogy and causes of NT and recommendations regarding platelet transfusions.

Thrombocytopenia, defined as a platelet count <150,000/μL or below

populations. Thrombocytopenia implicates an increased risk of bleeding, and is associated with significant morbidity mainly due to intracranial hemorrhage. As a result, it is important to identify infants at risk, and if indicated (see below recommendations for platelet transfusion), to initiate therapy to prevent

/L, occurs more often during the neonatal period than in any other

Recently our research group on neonatal infectious diseases and epidemiology at the Medical University of Graz, Austria, published retrospectively collected data on neonatal thrombocytopenia (NT) [1]. Of 371 neonates diagnosed as having NT, 312 (84.1%) had early-onset NT (EOT) and 59 (15.9%) had late-onset NT (LOT) defined as NT before or after 72 hours of age, respectively. The degree of NT was defined as mild, platelet counts of 100,000–150,000/μL; moderate, counts of 50,000–<100,000/μL; severe, counts of 30,000–<50,000/μL; and very severe, counts of <30,000/μL, according to the description of Wiedmeier et al. [2]. The majority—nearly three-fourth of the cohort—had mild (33%) to moderate (38%) NT; only 14% had severe and 15% very severe NT [1]. Seventy-six percent of the neonates were born preterm and this rate was approximately the same for

**Keywords:** neonatal thrombocytopenia, prematurity, platelet transfusion, alloimmune thrombocytopenia, early- and late-onset thrombocytopenia, severity,

intracranial hemorrhage, bleeding, incidence, mortality

Thrombocytopenia defined as platelet count below 150,000/μL is not an uncommon event at the neonatal intensive care unit (NICU). In our region we calculated a prevalence of nearly 2 of 1000 live births. Early-onset neonatal thrombocytopenia (NT) occurring within the first 72 hours of life is more common than late-onset NT. Preterm infants are affected more often than term infants and bacterial infection is the most common diagnosis associated with NT. There are a lot of maternal, perinatal, and neonatal causes associated with NT and complications include bleedings with potentially life-threatening intracranial hemorrhage. Alloimmune thrombocytopenia (NAIT) often presents with severe thrombocytopenia (<30,000/ μL) in otherwise healthy newborns and needs careful evaluation regarding HPA-1a antigen status and HLA typing. Platelet transfusions are needed in severe NT and threshold platelet counts might be at ≤25,000/μL irrespective of bleeding or not. Immune mediated NT recovers within 2 weeks with a good prognosis when there happened no intracranial hemorrhage. This short review gives an overview on etiol-

## **Chapter 6** Thrombocytopenia in Neonates

*Bernhard Resch*

## **Abstract**

Thrombocytopenia defined as platelet count below 150,000/μL is not an uncommon event at the neonatal intensive care unit (NICU). In our region we calculated a prevalence of nearly 2 of 1000 live births. Early-onset neonatal thrombocytopenia (NT) occurring within the first 72 hours of life is more common than late-onset NT. Preterm infants are affected more often than term infants and bacterial infection is the most common diagnosis associated with NT. There are a lot of maternal, perinatal, and neonatal causes associated with NT and complications include bleedings with potentially life-threatening intracranial hemorrhage. Alloimmune thrombocytopenia (NAIT) often presents with severe thrombocytopenia (<30,000/ μL) in otherwise healthy newborns and needs careful evaluation regarding HPA-1a antigen status and HLA typing. Platelet transfusions are needed in severe NT and threshold platelet counts might be at ≤25,000/μL irrespective of bleeding or not. Immune mediated NT recovers within 2 weeks with a good prognosis when there happened no intracranial hemorrhage. This short review gives an overview on etiology and causes of NT and recommendations regarding platelet transfusions.

**Keywords:** neonatal thrombocytopenia, prematurity, platelet transfusion, alloimmune thrombocytopenia, early- and late-onset thrombocytopenia, severity, intracranial hemorrhage, bleeding, incidence, mortality

## **1. Introduction**

Thrombocytopenia, defined as a platelet count <150,000/μL or below 150 × 10<sup>9</sup> /L, occurs more often during the neonatal period than in any other populations. Thrombocytopenia implicates an increased risk of bleeding, and is associated with significant morbidity mainly due to intracranial hemorrhage. As a result, it is important to identify infants at risk, and if indicated (see below recommendations for platelet transfusion), to initiate therapy to prevent complications.

Recently our research group on neonatal infectious diseases and epidemiology at the Medical University of Graz, Austria, published retrospectively collected data on neonatal thrombocytopenia (NT) [1]. Of 371 neonates diagnosed as having NT, 312 (84.1%) had early-onset NT (EOT) and 59 (15.9%) had late-onset NT (LOT) defined as NT before or after 72 hours of age, respectively. The degree of NT was defined as mild, platelet counts of 100,000–150,000/μL; moderate, counts of 50,000–<100,000/μL; severe, counts of 30,000–<50,000/μL; and very severe, counts of <30,000/μL, according to the description of Wiedmeier et al. [2]. The majority—nearly three-fourth of the cohort—had mild (33%) to moderate (38%) NT; only 14% had severe and 15% very severe NT [1]. Seventy-six percent of the neonates were born preterm and this rate was approximately the same for

either EOT or LOT (76 and 77%, respectively). The percentage of extremely low gestational age newborns (ELGAN, below 28 weeks of gestational age) was 20% in total. The incidence of NT in preterm infants was 4% (282/6964) in our population during the years 1990–2012. Thus, we calculated the prevalence of NT as being 1.8/1000 live births in our region (Southern Styria with around 200,000 live births during the study period) [1].

A total of 40 neonates (10.8%) died; 36 (90%) had EOT, and 4 (10%) had LOT; and 30 (75%) neonates were still thrombocytopenic at the time of death. Interestingly, bleeding signs were significantly associated with mortality in our study. On the other hand, severity of NT was not associated with mortality. Only cutaneous bleedings were found to be associated with severity of NT. The mean duration of NT was significantly longer in case of LOT compared to EOT (8.9 vs. 16.8 days; p < 0.001); and the duration was positively correlated with severity of NT. At least we found that platelet transfusion did not shorten the duration of NT [1].

## **2. Etiology and causes of neonatal thrombocytopenia**

Thrombocytopenia is present in 1–5% of newborns at birth, and severe thrombocytopenia defined as platelet count below 50,000/μL occurs in 0.1–0.5% [3]. But thrombocytopenia is more common in neonates needing intensive care at the neonatal intensive care unit (NICU) with rates up to 50%. Every fifth newborn is at risk at the NICU to develop thrombocytopenia, and 8% of preterm and 6% of term infants are at risk for severe thrombocytopenia [4].

Main mechanisms of thrombocytopenia include increased platelet consumption and/or sequestration, and often neonatal thrombocytopenia is of multifactorial origin. Thus, on the one hand there is rapid consumption like in case of necrotizing enterocolitis (NEC) and on the other hand slow recovery by impaired platelet production.

There are a lot of maternal, perinatal, and neonatal causes that might be associated with the occurrence of NT. **Figure 1** shows the main features and causes of NT by separating early- and late-onset thrombocytopenia [5].

**123**

did more sporadic causes of NT.

*Data are given as number (%).*

**Table 1.**

*Turkey [7]).*

result in prolonged courses of NT [1, 9, 10].

*Thrombocytopenia in Neonates*

371 cases 23-year

Chrom. aberration

Metab. disorders

*DOI: http://dx.doi.org/10.5772/intechopen.92857*

period

Own data on neonatal diagnoses associated with thrombocytopenia retrospectively collected over 23 years compared to data from Tunisia and Turkey—in order to give a broader view on dominant causes and diagnoses—are shown in **Table 1** [1]. Main diagnoses were early- and late-onset sepsis, and intrauterine growth restriction. Other dominant features like asphyxia and NEC differed between centers as

*EOS: early-onset sepsis, LOS: late-onset sepsis, NEC: necrotizing enterocolitis, HDN: hemolytic disease of the newborn, CMV: cytomegalovirus, MPD: myeloproliferative disease, NAIT: neonatal alloimmune thrombocytopenia, K-M-syndrome: Kasabach-Merritt syndrome, DIC: disseminated intravasal coagulation, IUGR: intrauterine growth* 

*Neonatal diagnoses associated with thrombocytopenia from three studies (Austria [1], Tunisia [6], and* 

*restriction, GDM: gestational diabetes mellitus, SLE: systemic lupus erythematodes.*

**Austria [1] 2018 Tunisia [] 2016 Turkey [] 2013**

112 episodes 4-year

EOS 128 (34) LOS 29 (19) Sepsis 45 (34) Asphyxia 95 (25) IUGR 26 (17) IUGR 25 (19) LOS 47 (13) EOS 23 (15) Preeclampsia 13 (9.7) NEC 16 (4.1) DIC 22 (14) Maternal thr. 6 (4.5)

HDN 9 (2.4) Unexplained 15 (9.6) Hydrops fetalis 4 (3.0) CMV 9 (2.4) Cong. rubella 6 (3.8) Perinatal asphyxia 4 (3.0) MPD 6 (1.6) Asphyxia 5 (3.2) Ablatio placenta 4 (3.0) NAIT 4 (1.0) NEC 3 (1.9) Drug + sepsis 4 (3.0) K-M-syndrome 2 (0.5) Tris 21 2 (1.3) Rh incompatibility 4 (3.0)

Thrombosis 2 (0.5) Exchange transf. 2 (1.3) Maternal ITP 3 (2.2)

period

15 (3.9) PIH 18 (12) Drug 5 (3.7)

2 (0.5) CMV 2 (1.3) NEC 3 (2.2)

Toxoplasmosis 1 (0.7) Congenital anomaly 2 (1.5) Neonatal lupus 1 (0.7) Tris 21 2 (1.5) Maternal ITP 1 (0.7) NAIT 2 (1.5)

134 cases 5-year

HELLP syndrome 2 (1.5) Metab. disorders 2 (1.5) Maternal GDM 2 (1.5) Neonatal jaundice 1 (0.8) Mother with SLE 1 (0.8)

period

An association of NT with bacterial infection is well known. Rates have been reported of 30% [7], 36% [8], and 47% [1]. And severe NT and very severe NT were commonly associated with sepsis [1, 8]. Late-onset NT often is reported to

Birth asphyxia is a common diagnosis associated with NT and reported up to 25% [1, 7, 8]. NEC is a morbidity often complicated by NT. Resch et al. reported a rate of 4.1% [1] and that was twice as high as reported elsewhere [7, 8]. Other associations, including chromosomal anomalies, metabolic disorders, and thromboses,

**Figure 1.**

*Etiology and causes of early- and late-onset thrombocytopenia.*

*Thrombocytopenia in Neonates*


*DOI: http://dx.doi.org/10.5772/intechopen.92857*

*Platelets*

during the study period) [1].

either EOT or LOT (76 and 77%, respectively). The percentage of extremely low gestational age newborns (ELGAN, below 28 weeks of gestational age) was 20% in total. The incidence of NT in preterm infants was 4% (282/6964) in our population during the years 1990–2012. Thus, we calculated the prevalence of NT as being 1.8/1000 live births in our region (Southern Styria with around 200,000 live births

A total of 40 neonates (10.8%) died; 36 (90%) had EOT, and 4 (10%) had LOT; and 30 (75%) neonates were still thrombocytopenic at the time of death. Interestingly, bleeding signs were significantly associated with mortality in our study. On the other hand, severity of NT was not associated with mortality. Only cutaneous bleedings were found to be associated with severity of NT. The mean duration of NT was significantly longer in case of LOT compared to EOT (8.9 vs. 16.8 days; p < 0.001); and the duration was positively correlated with severity of NT. At least we

Thrombocytopenia is present in 1–5% of newborns at birth, and severe thrombocytopenia defined as platelet count below 50,000/μL occurs in 0.1–0.5% [3]. But thrombocytopenia is more common in neonates needing intensive care at the neonatal intensive care unit (NICU) with rates up to 50%. Every fifth newborn is at risk at the NICU to develop thrombocytopenia, and 8% of preterm and 6% of term

Main mechanisms of thrombocytopenia include increased platelet consumption and/or sequestration, and often neonatal thrombocytopenia is of multifactorial origin. Thus, on the one hand there is rapid consumption like in case of necrotizing enterocolitis (NEC) and on the other hand slow recovery by impaired platelet

There are a lot of maternal, perinatal, and neonatal causes that might be associated with the occurrence of NT. **Figure 1** shows the main features and causes of NT

found that platelet transfusion did not shorten the duration of NT [1].

**2. Etiology and causes of neonatal thrombocytopenia**

infants are at risk for severe thrombocytopenia [4].

by separating early- and late-onset thrombocytopenia [5].

*Etiology and causes of early- and late-onset thrombocytopenia.*

**122**

**Figure 1.**

production.

*Data are given as number (%).*

*EOS: early-onset sepsis, LOS: late-onset sepsis, NEC: necrotizing enterocolitis, HDN: hemolytic disease of the newborn, CMV: cytomegalovirus, MPD: myeloproliferative disease, NAIT: neonatal alloimmune thrombocytopenia, K-M-syndrome: Kasabach-Merritt syndrome, DIC: disseminated intravasal coagulation, IUGR: intrauterine growth restriction, GDM: gestational diabetes mellitus, SLE: systemic lupus erythematodes.*

#### **Table 1.**

*Neonatal diagnoses associated with thrombocytopenia from three studies (Austria [1], Tunisia [6], and Turkey [7]).*

Own data on neonatal diagnoses associated with thrombocytopenia retrospectively collected over 23 years compared to data from Tunisia and Turkey—in order to give a broader view on dominant causes and diagnoses—are shown in **Table 1** [1]. Main diagnoses were early- and late-onset sepsis, and intrauterine growth restriction. Other dominant features like asphyxia and NEC differed between centers as did more sporadic causes of NT.

An association of NT with bacterial infection is well known. Rates have been reported of 30% [7], 36% [8], and 47% [1]. And severe NT and very severe NT were commonly associated with sepsis [1, 8]. Late-onset NT often is reported to result in prolonged courses of NT [1, 9, 10].

Birth asphyxia is a common diagnosis associated with NT and reported up to 25% [1, 7, 8]. NEC is a morbidity often complicated by NT. Resch et al. reported a rate of 4.1% [1] and that was twice as high as reported elsewhere [7, 8]. Other associations, including chromosomal anomalies, metabolic disorders, and thromboses, ranged between 0.5 and 3.9% in the literature [1, 6–8]. Von Lindern et al. [8] reported on a 10% rate of hemolytic disease of the newborn (HDN), which was four times higher compared to the rate of 2.4% reported by Resch et al. [1].

In preterm infants, NT is rarely diagnosed at low rates between 4 and 12% [1, 10]. Most studies report higher rates of NT ranging between 22 and 35% [2, 3, 8, 11–13], and highest rates (53–70%) have been reported from developing countries [14, 15]. Own data revealed that 75% of a cohort of thrombocytopenic neonates were preterm neonates. The association between NT and prematurity or low birth weight is well documented [3, 6–8, 11, 16, 17]. In this context small-forgestational age (SGA) is a well-known risk factor for developing NT [11, 16, 17], and rates have been reported as high as 30–53% [1, 17, 18].

## **3. Pathomechanisms of neonatal thrombocytopenia**

#### **3.1 Immune-mediated thrombocytopenia**

One possibility of low platelet counts is increased destruction that is observed in several neonatal conditions. Placental crossing of maternal antibodies is the cause of NT in case of immune-mediated NT, which destroys neonatal platelets. Immunemediated processes are very common causes of neonatal thrombocytopenia, and the antibodies responsible may be autoantibodies, drug-dependent antibodies, or alloantibodies. The mechanism behind is an interaction with platelet membrane antigens or the formation of immune complexes, which can bind to reticuloendothelial cell Fc receptors. As a result platelets become cleared from blood vessels [19].

#### *3.1.1 Neonatal alloimmune thrombocytopenia (NAIT)*

In NAIT, fetal platelets contain an antigen inherited from the father that the mother lacks. The mother produces antiplatelet antibodies from the immunoglobulin G (IgG)-type against the platelet antigen during pregnancy that is recognized as being foreign. Thereafter IgG antibodies cross the placenta and destroy fetal platelets that express the paternal antigen [20].

#### *3.1.2 Neonatal autoimmune thrombocytopenia*

It is mediated by maternal autoantibodies that react with both maternal and fetal platelets. This occurs in maternal autoimmune disorders, including immune thrombocytopenia purpura (ITP) and systemic lupus erythematosus (SLE) [20].

#### *3.1.3 Drug-induced immune thrombocytopenia*

Drug-induced immune thrombocytopenia is typically caused by platelet destruction from maternal drug-dependent antibodies and, rarely, by neonatal antibodies. Bone marrow suppression also can result in thrombocytopenia due to decreased platelet production. Neonatal drug-induced immune thrombocytopenia is usually caused by maternal drug-dependent antibodies formed after drug exposure to the mother during pregnancy [19, 20]. Maternal antibodies can cross the placenta and affect fetal and neonatal platelets. This mechanism is similar to that seen in mothers with primary immune thrombocytopenia purpura (ITP). Drugs associated with maternal drug-mediated platelet destruction include quinine, quinidine, trimethoprim-sulfamethoxazole, vancomycin, penicillin, rifampin, carbamazepine, phenytoin, valproic

**125**

*Thrombocytopenia in Neonates*

agulant [20].

**3.2 Thrombopoiesis**

production [22].

*DOI: http://dx.doi.org/10.5772/intechopen.92857*

*3.1.4 Non-immune drug-induced NT*

*3.2.1 The homeostasis of TPO levels*

induced thrombocytosis [22].

by suppression of platelet production [20].

acid, ceftriaxone, ibuprofen, mirtazapine, oxaliplatin, suramin, GP IIb/IIIa inhibitors (e.g., abciximab, tirofiban, eptifibatide), and heparin. Some drugs may cause thrombocytopenia at the initial exposure without prior sensitization. This commonly occurs with the glycoprotein IIb/IIIa inhibitors and has also been seen with other drugs, such as vancomycin and piperacillin [21]. Rarely, platelet destruction can be caused by neonatal drug-dependent antibodies, such as seen in heparin-induced thrombocytopenia (HIT). HIT antibodies can promote thrombosis by inducing platelet activation. Patients with suspected HIT require immediate institution of a non-heparin antico-

Many drugs used as chemotherapy cause thrombocytopenia by bone marrow suppression (*non-immune drug induced NT*). Antibiotics, such as linezolid, daptomycin, and valacyclovir can also cause moderate thrombocytopenia in some patients

Platelet production—thrombopoiesis—is initiated by a thrombopoietic stimulus, and the most important stimulant is the chemokine thrombopoietin (TPO) besides several cytokines and chemokines that are also involved in the process (e.g., IL-3, IL-6, IL-11, GM-CSF, stromal cell-derived factor-1 and fibroblast growth factor 4) [22]. TPO promotes the proliferation of megakaryocyte progenitors and the maturation of megakaryocytes. These mature megakaryocytes are at least responsible for

The homeostasis of TPO levels is regulated by the thrombopoietin c-Mpl (myeloproliferative leukemia protein) receptor-mediated uptake and destruction of the hormone with the aim to have steady-state amounts of hepatic TPO. When bound to the platelet c-Mpl receptors, the hormone gets removed from the circulation and blood levels are reduced. In case of inflammatory processes IL-6 is released from macrophages and fibroblasts (via TNF-α) and enhances hepatic TPO

Another phenomenon adding to the steady-state model of TPO regulation is the physiological response to severe thrombocytopenia of bone marrow stromal cells, which under normal circumstances produce low amounts of TPO-mRNA but increase transcription markedly in case of thrombocytopenia [22]. IL-6, stimulated by inflammation processes, leads to increased levels of TPO resulting in reactive thrombocytosis; and TPO is now confirmed as the final mediator of inflammation-

There are similarities and differences between neonatal and adult thrombocytopenia. Plasma TPO concentrations are known to be higher in healthy neonates compared to healthy adults. But in NT TPO levels are lower even when adult thrombocytopenia has the same degree [19]. Interestingly, megakaryocyte progenitors of neonates have a higher proliferative potential than those of adults resulting in larger megakaryocytes. Neonatal megakaryocyte progenitors are more sensitive to TPO both in vitro and in vivo than adult progenitors [19]. Cells are present both in the bone marrow and the peripheral blood of neonates in contrast to adult ones that are

almost exclusively present in the bone marrow [19] (**Figure 2**).

generation and release of new platelets into the blood vessels [22].

#### *Thrombocytopenia in Neonates DOI: http://dx.doi.org/10.5772/intechopen.92857*

*Platelets*

ranged between 0.5 and 3.9% in the literature [1, 6–8]. Von Lindern et al. [8]

times higher compared to the rate of 2.4% reported by Resch et al. [1].

and rates have been reported as high as 30–53% [1, 17, 18].

**3. Pathomechanisms of neonatal thrombocytopenia**

**3.1 Immune-mediated thrombocytopenia**

*3.1.1 Neonatal alloimmune thrombocytopenia (NAIT)*

platelets that express the paternal antigen [20].

*3.1.2 Neonatal autoimmune thrombocytopenia*

*3.1.3 Drug-induced immune thrombocytopenia*

reported on a 10% rate of hemolytic disease of the newborn (HDN), which was four

One possibility of low platelet counts is increased destruction that is observed in several neonatal conditions. Placental crossing of maternal antibodies is the cause of NT in case of immune-mediated NT, which destroys neonatal platelets. Immunemediated processes are very common causes of neonatal thrombocytopenia, and the antibodies responsible may be autoantibodies, drug-dependent antibodies, or alloantibodies. The mechanism behind is an interaction with platelet membrane antigens or the formation of immune complexes, which can bind to reticuloendothelial cell Fc receptors. As a result platelets become cleared from blood vessels [19].

In NAIT, fetal platelets contain an antigen inherited from the father that the mother lacks. The mother produces antiplatelet antibodies from the immunoglobulin G (IgG)-type against the platelet antigen during pregnancy that is recognized as being foreign. Thereafter IgG antibodies cross the placenta and destroy fetal

It is mediated by maternal autoantibodies that react with both maternal and fetal platelets. This occurs in maternal autoimmune disorders, including immune thrombocytopenia purpura (ITP) and systemic lupus erythematosus (SLE) [20].

Drug-induced immune thrombocytopenia is typically caused by platelet destruction from maternal drug-dependent antibodies and, rarely, by neonatal antibodies. Bone marrow suppression also can result in thrombocytopenia due to decreased platelet production. Neonatal drug-induced immune thrombocytopenia is usually caused by maternal drug-dependent antibodies formed after drug exposure to the mother during pregnancy [19, 20]. Maternal antibodies can cross the placenta and affect fetal and neonatal platelets. This mechanism is similar to that seen in mothers with primary immune thrombocytopenia purpura (ITP). Drugs associated with maternal drug-mediated platelet destruction include quinine, quinidine, trimethoprim-sulfamethoxazole, vancomycin, penicillin, rifampin, carbamazepine, phenytoin, valproic

In preterm infants, NT is rarely diagnosed at low rates between 4 and 12% [1, 10]. Most studies report higher rates of NT ranging between 22 and 35% [2, 3, 8, 11–13], and highest rates (53–70%) have been reported from developing countries [14, 15]. Own data revealed that 75% of a cohort of thrombocytopenic neonates were preterm neonates. The association between NT and prematurity or low birth weight is well documented [3, 6–8, 11, 16, 17]. In this context small-forgestational age (SGA) is a well-known risk factor for developing NT [11, 16, 17],

**124**

acid, ceftriaxone, ibuprofen, mirtazapine, oxaliplatin, suramin, GP IIb/IIIa inhibitors (e.g., abciximab, tirofiban, eptifibatide), and heparin. Some drugs may cause thrombocytopenia at the initial exposure without prior sensitization. This commonly occurs with the glycoprotein IIb/IIIa inhibitors and has also been seen with other drugs, such as vancomycin and piperacillin [21]. Rarely, platelet destruction can be caused by neonatal drug-dependent antibodies, such as seen in heparin-induced thrombocytopenia (HIT). HIT antibodies can promote thrombosis by inducing platelet activation. Patients with suspected HIT require immediate institution of a non-heparin anticoagulant [20].

## *3.1.4 Non-immune drug-induced NT*

Many drugs used as chemotherapy cause thrombocytopenia by bone marrow suppression (*non-immune drug induced NT*). Antibiotics, such as linezolid, daptomycin, and valacyclovir can also cause moderate thrombocytopenia in some patients by suppression of platelet production [20].

## **3.2 Thrombopoiesis**

Platelet production—thrombopoiesis—is initiated by a thrombopoietic stimulus, and the most important stimulant is the chemokine thrombopoietin (TPO) besides several cytokines and chemokines that are also involved in the process (e.g., IL-3, IL-6, IL-11, GM-CSF, stromal cell-derived factor-1 and fibroblast growth factor 4) [22]. TPO promotes the proliferation of megakaryocyte progenitors and the maturation of megakaryocytes. These mature megakaryocytes are at least responsible for generation and release of new platelets into the blood vessels [22].

## *3.2.1 The homeostasis of TPO levels*

The homeostasis of TPO levels is regulated by the thrombopoietin c-Mpl (myeloproliferative leukemia protein) receptor-mediated uptake and destruction of the hormone with the aim to have steady-state amounts of hepatic TPO. When bound to the platelet c-Mpl receptors, the hormone gets removed from the circulation and blood levels are reduced. In case of inflammatory processes IL-6 is released from macrophages and fibroblasts (via TNF-α) and enhances hepatic TPO production [22].

Another phenomenon adding to the steady-state model of TPO regulation is the physiological response to severe thrombocytopenia of bone marrow stromal cells, which under normal circumstances produce low amounts of TPO-mRNA but increase transcription markedly in case of thrombocytopenia [22]. IL-6, stimulated by inflammation processes, leads to increased levels of TPO resulting in reactive thrombocytosis; and TPO is now confirmed as the final mediator of inflammationinduced thrombocytosis [22].

There are similarities and differences between neonatal and adult thrombocytopenia. Plasma TPO concentrations are known to be higher in healthy neonates compared to healthy adults. But in NT TPO levels are lower even when adult thrombocytopenia has the same degree [19]. Interestingly, megakaryocyte progenitors of neonates have a higher proliferative potential than those of adults resulting in larger megakaryocytes. Neonatal megakaryocyte progenitors are more sensitive to TPO both in vitro and in vivo than adult progenitors [19]. Cells are present both in the bone marrow and the peripheral blood of neonates in contrast to adult ones that are almost exclusively present in the bone marrow [19] (**Figure 2**).

#### *3.2.2 Clinical conditions and their pathomechanisms*

*Chronic intrauterine hypoxia* is commonly observed when associated with placental insufficiency due to all conditions of pregnancy-induced hypertension (hypertension alone or pre-eclampsia or hypertension-elevated liver enzymes-low platelet counts—HELLP—syndrome, and gestational or maternal diabetes) and is usually manifested by fetal intrauterine growth restriction and hematological abnormalities including NT. The pathomechanisms behind are not completely understood but lower levels of megakaryocyte progenitors have been found that increased during normalization of NT [23].

The hematopoietic microenvironment plays a significant role in chronic hypoxia-induced suppression of megakaryocytopoiesis, and, not astonishingly, preterm infants' megakaryocyte progenitors are more vulnerable to ischemic insults than progenitors from term neonates or adults [24].

Overall, observations demonstrated that thrombopoiesis is up-regulated in *neonatal sepsis and/or NEC*, but this effect can also be down-regulated resulting in "hypoproliferation" [19]. Platelet factor 4 is a potent inhibitor of megakaryocyte proliferation that is released from activated platelets during severe sepsis. This regulates neonatal megakaryocytopoiesis negatively [25].

In *HIV-associated thrombocytopenia*, evidence that splenic platelet sequestration decreased platelet production had been observed despite larger megakaryocyte mass [26]. In neonates, ineffective platelet production was described as being the main mechanism of HIV-associated thrombocytopenia [27]. The mechanisms in other *congenital infections* of the TORCH complex mostly remain to be speculative despite their common association with NT [20].

#### **4. Neonatal alloimmune thrombocytopenia (NAIT)**

Harrington et al. first described in 1953 two infants born with severe thrombocytopenia to mothers with normal platelet counts [28]. Both newborns recovered despite severe bleeding and other complications after 2 and 8 weeks, respectively.

**127**

*Thrombocytopenia in Neonates*

bocytopenia (NAIT).

mother and father.

*DOI: http://dx.doi.org/10.5772/intechopen.92857*

by HLA-DR antigen B3\*0101 positivity [31].

This immunological disease now is well described as neonatal alloimmune throm-

In 1962 a maternal antibody against a platelet alloantigen was detected causing NAIT [29]. This platelet alloantigen determined as PlA1 was the cause of platelet destruction in two of the newborns reported in the study by Shulman et al. [29]. Later, PlA1 was found to be identical to an antigen called Zwa [30] and now is known as human platelet antigen 1a (HPA-1a). Over the following years several other platelet-specific antigens were detected as being able to induce maternal immunization during pregnancy with subsequent fetal platelet destruction, thus, being an important complication of pregnancy with diagnostic and therapeutic challenges [31]. The incidence of NAIT calculated from large studies on women negative for HPA-1a lies in between 1 in 1000–2000 HPA-1a positive newborns [32–34]. The incidence of HPA-1a negative phenotype is about 2.5%; and one-third is at high risk to get immunized in case of a HPA-1a positive fetus, and this association is triggered

The main problem of NAIT is that it can lead to serious bleedings including intracranial hemorrhage and death. In full term infants it is the leading cause of intracranial hemorrhage [35]. Other clinical findings are petechiae or purpura associated with very low platelet counts without any explanation (after exclusion of bacterial and viral infection—TORCH complex, or disseminated intravascular coagulation). A previous history of NAIT results in more severe disease. Around 10–20% of the newborns have intracranial hemorrhages, and the vast majority of 80% occurs already before birth. After birth the greatest risk of bleeding is in the time span of the first 4 days of life. Untreated, NT resolves within 2–3 weeks [35]. Even in mildly affected infants serological investigation including ABO, HPA and HLA typing (further details are beyond the scope of this chapter) is indicated because results can be critical for effective management of future pregnancies. For the most informative evaluation, it is important to study blood samples from both

A systematic review on incidence and consequences of NAIT reported on 6 of 21 studies (full text analysis) from initial 768 studies [36]. Nearly 60,000 newborns were screened, with severe thrombocytopenia in 89 cases (0.15%); and NAIT was diagnosed in 24 of these 89 newborns (27%) resulting in an incidence of 1:2500. Six newborns (25%) had diagnosis of intracranial hemorrhage and most likely of antenatal origin. Hence, intracranial hemorrhage due to NAIT occurred in 1:10–11,000 newborns [36, 37]. This is the most severe complication having a 1–7% risk of death. Survivors are known to have sequelae including mental retardation, cerebral palsy, cortical blindness and seizures in 7–26% of pregnancies [38]. In contrast to ABO- or Rh-incompatibility, immunization occurs often during the first pregnancy. Severity of NAIT is associated with parity (second pregnancy often more severe), HPA-a1 antibodies level, outcome of a former pregnancy, and the type of alloimmunization (HPA-1a more severe than HPA-5b) and the HLA type: homozygote HPA-1b and negative for HLA DRB3\*0101 leads to no NAIT with negative predictive value of 99.6%, but homozygote HPA-1b plus HLA DRB3\*0101 positivity leads to NAIT with a positive predictive value of 35% [39] Interestingly, antenatal IVIG is again more effective depending on HLA status [31]. Also of importance is

At first the medical history should be looked for a previous neonate with throm-

previous history of intracranial hemorrhage of uncertain origin [40]. Is the mother

/L and/or

bocytopenia of unknown origin and/or having platelet count below 50 × 109

the risk of recurrent intracranial hemorrhage of being 80–90%.

**4.1 How to proceed in case of suspected NAIT?**

#### *Thrombocytopenia in Neonates DOI: http://dx.doi.org/10.5772/intechopen.92857*

*Platelets*

**Figure 2.**

*3.2.2 Clinical conditions and their pathomechanisms*

than progenitors from term neonates or adults [24].

despite their common association with NT [20].

regulates neonatal megakaryocytopoiesis negatively [25].

**4. Neonatal alloimmune thrombocytopenia (NAIT)**

normalization of NT [23].

*A simplified model of TPO regulation.*

*Chronic intrauterine hypoxia* is commonly observed when associated with placental insufficiency due to all conditions of pregnancy-induced hypertension (hypertension alone or pre-eclampsia or hypertension-elevated liver enzymes-low platelet counts—HELLP—syndrome, and gestational or maternal diabetes) and is usually manifested by fetal intrauterine growth restriction and hematological abnormalities including NT. The pathomechanisms behind are not completely understood but lower levels of megakaryocyte progenitors have been found that increased during

The hematopoietic microenvironment plays a significant role in chronic hypoxia-induced suppression of megakaryocytopoiesis, and, not astonishingly, preterm infants' megakaryocyte progenitors are more vulnerable to ischemic insults

Overall, observations demonstrated that thrombopoiesis is up-regulated in *neonatal sepsis and/or NEC*, but this effect can also be down-regulated resulting in "hypoproliferation" [19]. Platelet factor 4 is a potent inhibitor of megakaryocyte proliferation that is released from activated platelets during severe sepsis. This

In *HIV-associated thrombocytopenia*, evidence that splenic platelet sequestration decreased platelet production had been observed despite larger megakaryocyte mass [26]. In neonates, ineffective platelet production was described as being the main mechanism of HIV-associated thrombocytopenia [27]. The mechanisms in other *congenital infections* of the TORCH complex mostly remain to be speculative

Harrington et al. first described in 1953 two infants born with severe thrombocytopenia to mothers with normal platelet counts [28]. Both newborns recovered despite severe bleeding and other complications after 2 and 8 weeks, respectively.

**126**

This immunological disease now is well described as neonatal alloimmune thrombocytopenia (NAIT).

In 1962 a maternal antibody against a platelet alloantigen was detected causing NAIT [29]. This platelet alloantigen determined as PlA1 was the cause of platelet destruction in two of the newborns reported in the study by Shulman et al. [29]. Later, PlA1 was found to be identical to an antigen called Zwa [30] and now is known as human platelet antigen 1a (HPA-1a). Over the following years several other platelet-specific antigens were detected as being able to induce maternal immunization during pregnancy with subsequent fetal platelet destruction, thus, being an important complication of pregnancy with diagnostic and therapeutic challenges [31].

The incidence of NAIT calculated from large studies on women negative for HPA-1a lies in between 1 in 1000–2000 HPA-1a positive newborns [32–34]. The incidence of HPA-1a negative phenotype is about 2.5%; and one-third is at high risk to get immunized in case of a HPA-1a positive fetus, and this association is triggered by HLA-DR antigen B3\*0101 positivity [31].

The main problem of NAIT is that it can lead to serious bleedings including intracranial hemorrhage and death. In full term infants it is the leading cause of intracranial hemorrhage [35]. Other clinical findings are petechiae or purpura associated with very low platelet counts without any explanation (after exclusion of bacterial and viral infection—TORCH complex, or disseminated intravascular coagulation). A previous history of NAIT results in more severe disease. Around 10–20% of the newborns have intracranial hemorrhages, and the vast majority of 80% occurs already before birth. After birth the greatest risk of bleeding is in the time span of the first 4 days of life. Untreated, NT resolves within 2–3 weeks [35].

Even in mildly affected infants serological investigation including ABO, HPA and HLA typing (further details are beyond the scope of this chapter) is indicated because results can be critical for effective management of future pregnancies. For the most informative evaluation, it is important to study blood samples from both mother and father.

A systematic review on incidence and consequences of NAIT reported on 6 of 21 studies (full text analysis) from initial 768 studies [36]. Nearly 60,000 newborns were screened, with severe thrombocytopenia in 89 cases (0.15%); and NAIT was diagnosed in 24 of these 89 newborns (27%) resulting in an incidence of 1:2500. Six newborns (25%) had diagnosis of intracranial hemorrhage and most likely of antenatal origin. Hence, intracranial hemorrhage due to NAIT occurred in 1:10–11,000 newborns [36, 37]. This is the most severe complication having a 1–7% risk of death. Survivors are known to have sequelae including mental retardation, cerebral palsy, cortical blindness and seizures in 7–26% of pregnancies [38]. In contrast to ABO- or Rh-incompatibility, immunization occurs often during the first pregnancy.

Severity of NAIT is associated with parity (second pregnancy often more severe), HPA-a1 antibodies level, outcome of a former pregnancy, and the type of alloimmunization (HPA-1a more severe than HPA-5b) and the HLA type: homozygote HPA-1b and negative for HLA DRB3\*0101 leads to no NAIT with negative predictive value of 99.6%, but homozygote HPA-1b plus HLA DRB3\*0101 positivity leads to NAIT with a positive predictive value of 35% [39] Interestingly, antenatal IVIG is again more effective depending on HLA status [31]. Also of importance is the risk of recurrent intracranial hemorrhage of being 80–90%.

#### **4.1 How to proceed in case of suspected NAIT?**

At first the medical history should be looked for a previous neonate with thrombocytopenia of unknown origin and/or having platelet count below 50 × 109 /L and/or previous history of intracranial hemorrhage of uncertain origin [40]. Is the mother

thrombocytopenic, one should evaluate for maternal anti-platelet auto-antibodies or a history of immune thrombocytopenia. Is the mother not thrombocytopenic, maternal and paternal platelet antigen typing and maternal platelet HPA antibody testing should be done. In case of incompatibility at HPA loci (1–6, 9, 15) and presence of specific maternal HPA-antibody, diagnosis of NAIT is given. Are there no incompatibilities and no anti-platelet antibodies or only nonspecific antibodies of the mother present, then no further evaluation is necessary besides maternal antibodies against paternal platelets are positive (preferred at 30 weeks' gestation). A third variant is a positive incompatibility without maternal anti-HPA antibodies (provided no reaction with paternal platelets), then there is no further evaluation necessary [40].

During pregnancy, strategies using IVIG and corticosteroids have been successful. The success rate with IVIG alone was reported to be as high as 98.7% and is in line with a Cochrane analysis reporting 97.3% [37, 41]

## **5. Complications of neonatal thrombocytopenia**

#### **5.1 Bleedings**

The prevalence of hemorrhages in thrombocytopenic neonates is approximately 20–30% according to the literature [16, 42]. The risk of hemorrhage is associated with lower gestational age, definite causes of thrombocytopenia, and the severity of concomitant morbidities [8, 9, 43].

An observational study including 169 neonates with severe NT identified severe sepsis and NEC as the most common diagnoses associated with bleeding neonates [35]. In those neonates with mild or no hemorrhage, the most common cause of severe NT has been documented as being intrauterine growth restriction and maternal pregnancy-induced hypertension [44].

A causal link between thrombocytopenia and intracranial hemorrhage is not known. Interestingly, platelet transfusions could not reduce the risk of intracranial bleedings [1, 45]. The majority of preterm neonates with severe intraventricular hemorrhage (IVH) becomes thrombocytopenic during the course of bleeding, thus, thrombocytopenia might not be the cause of IVH [46, 47]. Additionally, considering IVH as a multifactorial event, it seems highly unlikely that an isolated low platelet count leads to bleeding [44].

Another point of interest is the fact that comparable rates of IVH have been reported independent of the severity of NT [17]. In contrast, cutaneous bleeding conditions have been associated with the severity of NT [1, 46], and the prevalence of skin bleeding in thrombocytopenic neonates has been reported as being as high as 81% [48].

#### **5.2 Mortality**

The association of increased mortality rates with increasing numbers of platelet tranfusions mainly reflects the severity of the underlying disease or condition, for example, extremely low gestational age newborn. These infants are known to be at high risk for severe IVH that again is associated with high risk of death. Additionally, the more severe NT is the higher is the rate of mortality, and some data suggest that NT contributes to mortality rather than simply being a measure of disease severity [5]. Three studies from the USA, the UK, and Mexico reported higher mortality rates in neonates who had received platelet transfusions compared with those who had not [49–51]. The direct effects of platelet transfusions are questionable, as specific effects have not been properly evaluated, and the influence of

**129**

**Table 2.**

*Thrombocytopenia in Neonates*

volume of platelet transfusions.

50 × 109

is given in **Table 2**.

**Cut-off value thrombocyte counts**

<20 × 109

20–29 × 109

30–49 × 109

50–99 × 109

100–150 × 109

*DOI: http://dx.doi.org/10.5772/intechopen.92857*

preexisting morbidity is difficult to evaluate [44]. But as shown by Curley et al.

ing transfusion is of more benefit than earlier transfusion at cut-off level below

/L. Again the observation of Kenton et al. [53] is of interest who did not find an improvement in NEC-associated mortality with an increasing number or

Platelet transfusions are commonly used in preterm infants with NT at different

An overview of recommendations at different thresholds of thrombocyte counts

**Recommendation Authors**

Bleeding neonates Murray et al. [62]; Roberts et al. [4, 44];

Bleeding neonates Blanchette et al. [54]; Roberts et al. [44, 59];

Bleeding neonates Roberts et al. [4, 44, 59]; Murray et al. [62];

Chakravorty et al. [57]; Carr et al. [58]

Blanchette et al. [55]; Roberts et al. [59]; Calhoun et al. [60]; Sola-Visner et al. [61]; Murray [62]; Gibson et al. [56]; Carr et al. [58]; Sparger et al. [63]

Chakravorty et al. [57]

Blanchette et al. [54, 55]; Roberts et al. [59]; Sola-Visner et al. [61]; Calhoun et al. [60]; Murray [62]; Roberts et al. [4]; Sparger et al. [63]

Gibson et al. [56]; Chakravorty et al. [57]

Sola-Visner et al. [61]; Sparger et al. [63]

/L All neonates (prophylactic) Blanchette et al. [54, 55]; Gibson et al. [56];

/L Non-bleeding sick preterm Blanchette et al. [54, 55]; Roberts et al. [59]

/L No recommendations —

/L Non-bleeding term and preterm

/L Preterm infants (unstable neonate,

infant

first week of life, surgery or invasive procedures)

*Recommendations for platelet transfusions depending on different thrombocyte counts.*

threshold values. In 2018, a milestone study prospectively investigated whether platelet transfusion should be given at platelet-count thresholds of 50,000/μL (high-threshold group) or 25,000/μL (low-threshold group) [52]. In this multicenter trial, preterm infants born at less than 34 weeks of gestation in whom severe thrombocytopenia was diagnosed were included and randomly assigned to high or low platelet transfusion groups. The primary outcome was death or new major bleeding within 28 days after randomization. Of 660 infants (median birth weight, 740 g; and median gestational age, 26.6 weeks), 90% of the infants (296 of 328 infants) of the high-threshold group received at least one platelet transfusion, as compared with 53% (177 of 331 infants) in the low-threshold group. A new major bleeding episode or death happened in 26% of the infants in the high-threshold group and in 19% in the low-threshold group (odds ratio, 1.57; 95% confidence interval 1.06–2.32; p = 0.02). Most exciting, there was no significant difference between corresponding rates of serious adverse events (25 vs. 22%) [51].

/L before indicat-

[52] (see below), waiting until platelets have fallen below 25 × 109

**6. Recommendations for treatment with platelet transfusions**

#### *Thrombocytopenia in Neonates DOI: http://dx.doi.org/10.5772/intechopen.92857*

*Platelets*

**5.1 Bleedings**

as 81% [48].

**5.2 Mortality**

concomitant morbidities [8, 9, 43].

nal pregnancy-induced hypertension [44].

platelet count leads to bleeding [44].

thrombocytopenic, one should evaluate for maternal anti-platelet auto-antibodies or a history of immune thrombocytopenia. Is the mother not thrombocytopenic, maternal and paternal platelet antigen typing and maternal platelet HPA antibody testing should be done. In case of incompatibility at HPA loci (1–6, 9, 15) and presence of specific maternal HPA-antibody, diagnosis of NAIT is given. Are there no incompatibilities and no anti-platelet antibodies or only nonspecific antibodies of the mother present, then no further evaluation is necessary besides maternal antibodies against paternal platelets are positive (preferred at 30 weeks' gestation). A third variant is a positive incompatibility without maternal anti-HPA antibodies (provided no reaction with paternal platelets), then there is no further evaluation necessary [40]. During pregnancy, strategies using IVIG and corticosteroids have been successful. The success rate with IVIG alone was reported to be as high as 98.7% and is in

The prevalence of hemorrhages in thrombocytopenic neonates is approximately 20–30% according to the literature [16, 42]. The risk of hemorrhage is associated with lower gestational age, definite causes of thrombocytopenia, and the severity of

An observational study including 169 neonates with severe NT identified severe sepsis and NEC as the most common diagnoses associated with bleeding neonates [35]. In those neonates with mild or no hemorrhage, the most common cause of severe NT has been documented as being intrauterine growth restriction and mater-

A causal link between thrombocytopenia and intracranial hemorrhage is not known. Interestingly, platelet transfusions could not reduce the risk of intracranial bleedings [1, 45]. The majority of preterm neonates with severe intraventricular hemorrhage (IVH) becomes thrombocytopenic during the course of bleeding, thus, thrombocytopenia might not be the cause of IVH [46, 47]. Additionally, considering IVH as a multifactorial event, it seems highly unlikely that an isolated low

Another point of interest is the fact that comparable rates of IVH have been reported independent of the severity of NT [17]. In contrast, cutaneous bleeding conditions have been associated with the severity of NT [1, 46], and the prevalence of skin bleeding in thrombocytopenic neonates has been reported as being as high

The association of increased mortality rates with increasing numbers of platelet

tranfusions mainly reflects the severity of the underlying disease or condition, for example, extremely low gestational age newborn. These infants are known to be at high risk for severe IVH that again is associated with high risk of death. Additionally, the more severe NT is the higher is the rate of mortality, and some data suggest that NT contributes to mortality rather than simply being a measure of disease severity [5]. Three studies from the USA, the UK, and Mexico reported higher mortality rates in neonates who had received platelet transfusions compared with those who had not [49–51]. The direct effects of platelet transfusions are questionable, as specific effects have not been properly evaluated, and the influence of

line with a Cochrane analysis reporting 97.3% [37, 41]

**5. Complications of neonatal thrombocytopenia**

**128**

preexisting morbidity is difficult to evaluate [44]. But as shown by Curley et al. [52] (see below), waiting until platelets have fallen below 25 × 109 /L before indicating transfusion is of more benefit than earlier transfusion at cut-off level below 50 × 109 /L. Again the observation of Kenton et al. [53] is of interest who did not find an improvement in NEC-associated mortality with an increasing number or volume of platelet transfusions.

## **6. Recommendations for treatment with platelet transfusions**

Platelet transfusions are commonly used in preterm infants with NT at different threshold values. In 2018, a milestone study prospectively investigated whether platelet transfusion should be given at platelet-count thresholds of 50,000/μL (high-threshold group) or 25,000/μL (low-threshold group) [52]. In this multicenter trial, preterm infants born at less than 34 weeks of gestation in whom severe thrombocytopenia was diagnosed were included and randomly assigned to high or low platelet transfusion groups. The primary outcome was death or new major bleeding within 28 days after randomization. Of 660 infants (median birth weight, 740 g; and median gestational age, 26.6 weeks), 90% of the infants (296 of 328 infants) of the high-threshold group received at least one platelet transfusion, as compared with 53% (177 of 331 infants) in the low-threshold group. A new major bleeding episode or death happened in 26% of the infants in the high-threshold group and in 19% in the low-threshold group (odds ratio, 1.57; 95% confidence interval 1.06–2.32; p = 0.02). Most exciting, there was no significant difference between corresponding rates of serious adverse events (25 vs. 22%) [51].

**Cut-off value thrombocyte counts Recommendation Authors** <20 × 109 /L All neonates (prophylactic) Blanchette et al. [54, 55]; Gibson et al. [56]; Chakravorty et al. [57]; Carr et al. [58] 20–29 × 109 /L Non-bleeding term and preterm infant Blanchette et al. [55]; Roberts et al. [59]; Calhoun et al. [60]; Sola-Visner et al. [61]; Murray [62]; Gibson et al. [56]; Carr et al. [58]; Sparger et al. [63] Bleeding neonates Murray et al. [62]; Roberts et al. [4, 44]; Chakravorty et al. [57] 30–49 × 109 /L Preterm infants (unstable neonate, first week of life, surgery or invasive procedures) Blanchette et al. [54, 55]; Roberts et al. [59]; Sola-Visner et al. [61]; Calhoun et al. [60]; Murray [62]; Roberts et al. [4]; Sparger et al. [63]

An overview of recommendations at different thresholds of thrombocyte counts is given in **Table 2**.

## Bleeding neonates Blanchette et al. [54]; Roberts et al. [44, 59]; Gibson et al. [56]; Chakravorty et al. [57] 50–99 × 109 /L Non-bleeding sick preterm Blanchette et al. [54, 55]; Roberts et al. [59] Bleeding neonates Roberts et al. [4, 44, 59]; Murray et al. [62]; Sola-Visner et al. [61]; Sparger et al. [63] 100–150 × 109 /L No recommendations —

#### **Table 2.**

*Recommendations for platelet transfusions depending on different thrombocyte counts.*

## **7. Conclusions**

In conclusion, neonatal thrombocytopenia is a common problem at the neonatal intensive care unit. In most cases, it is a mild to moderate, self-limited entity. In severe cases, immune-mediated disease has to be suspected warranting prompt diagnosis and careful management. The threshold for platelet transfusions better should be at thrombocyte counts of 25–30,000/µL due to recent data reporting on a reduced mortality rate using a more restrictive transfusion regimen.

## **Acknowledgements**

I want to thank my daughter Elisabeth who published our own data in 2018 and inspired me to write this chapter.

## **Conflict of interest**

The author declares no conflict of interest.

## **Author details**

Bernhard Resch Division of Neonatology, Department of Pediatrics, Medical University of Graz, Graz, Austria

\*Address all correspondence to: bernhard.resch@medunigraz.at

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**131**

*Thrombocytopenia in Neonates*

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**7. Conclusions**

**Acknowledgements**

**Conflict of interest**

**Author details**

Bernhard Resch

Graz, Austria

inspired me to write this chapter.

The author declares no conflict of interest.

In conclusion, neonatal thrombocytopenia is a common problem at the neonatal

I want to thank my daughter Elisabeth who published our own data in 2018 and

Division of Neonatology, Department of Pediatrics, Medical University of Graz,

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

\*Address all correspondence to: bernhard.resch@medunigraz.at

provided the original work is properly cited.

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**135**

Section 3

Platelet Application
