Abstract

Muscular dystrophies are a diverse group of inherited muscle disorders with a wide range of clinical manifestations from a severe form with early onset and early death to adult forms with later onset and minimal clinical manifestation that do not affect life-span. Overlapping clinical symptoms and the multitude of genes that need to be analyzed for an accurate characterization make the diagnosis hard. In next-generation sequencing era, a lot of used assay in molecular diagnostics must be taken into consideration for muscular dystrophy diagnosis. However, for more accurate diagnosis, muscle protein expressions analysis may have prognostic value. In this chapter, we present the most important clinical and laboratory findings in the most common forms of muscular dystrophies and molecular diagnostic approaches for a more accurate diagnosis.

Keywords: muscular dystrophy, multiplex Western blot, immunofluorescence, MLPA, hrMCA, dystrophin, calpain 3, DMD gene, CAPN3, genetic diagnosis

### 1. Introduction

Muscular dystrophies (MD) are an inherited group of genetic disorders clinically characterized by progressive muscular weakness and wasting [1] and reduced skeletal muscle mass until their destruction due to a primary defect in the muscle cell. To date, there are known more than 30 different forms of MD with specific signs, symptoms, and genetic basis but sharing common histological features like variation in fiber shape and size and the presence of degeneration and regenerating fibers and connective tissue proliferation [2]. The diseases are distinguished from one another by the age of onset, muscles affected, as well as rate of disease progression [3]. While for some forms of MD, the initial symptoms manifested begin with childhood and have a rapid progression of muscle weakness causing the death of the patients around the age of 20 years, the other forms debut later in adulthood [4, 5] and have a slow rate of progression and an almost normal lifetime [6, 7]. Also, heart disease and mental retardation accompany some types of MD [8, 9], suggesting a different pathogenesis of the disease. It also found that there are subtypes of MD that share similar clinical manifestations and different genetic defects with similar clinical manifestation [10, 11].

However, the progress made in the past 33 years, since the first protein involved in a type of muscular dystrophy was discovered, leads to identify a large number of the genes as well as novel proteins involved in these muscle disorders [12].

For a rapid and an accurate diagnosis improvement in analysis, methods have become a necessity. The combination of clinical signs with muscle histopathology and protein and genetic analyses becomes the diagnostic gold standard for these disorders. Nevertheless, for many patients with yet unidentified muscular dystrophy, the diagnosis continues to be challenging.

involvement of skeletal and cardiac muscle in different degree [24] and include Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD), involving mostly skeletal muscles, and DMD-associated dilated cardiomyopathy (DCM), affecting preferentially the myocardium. The clinical picture in males with dystrophinopathies ranges from mild to severe. The mild forms consist of high level of CK in serum and muscle cramps with myoglobinuria. The severe forms include progressive muscle diseases (Duchenne muscle dystrophy, DMD, Becker muscle

In DMD, affected boys are clinically normal at birth. The onset of clinical features is in early childhood with delayed motor milestones, including delayed independent walking, with a mean age of walking of 18 months, and difficulties in standing up from the floor. The most characteristic clinical features of DMD are general motor delays (42%) and gait problems including persistent toe walking and flat-footedness (30%) [25]. Patients with DMD have a waddling gait and difficulties in climbing stairs, running, jumping, and standing up due to proximal weakness [26]. They rise from a supine position using their arms (Gower maneuver). The boys have hypertrophic and firm calf muscles [27]. The disease is rapidly progressive, at the age of 12 years, most boys being wheelchair bound [25]. Subsequently, the function of upper extremity will be lost, and, by the age of 14–15 years, cardiomyopathy is a common feature [26]; during the teenage years, the patients will require assisted ventilation [27]. Progressive cardiomyopathy and respiratory complications represent the most common causes of death in patients with DMD. Typically, the death occurs by 30 years of age, but currently the life expectancy of these patients has been improved through an improved management of cardiore-

Intellectual disability can be present in up to 27% of boys with DMD, and 44% of patients have learning disability [28]. Other psychiatric disorders reported in these patients include attention-deficit hyperactivity disorder (ADHD) (32%), anxiety

BMD is a milder form of muscular dystrophy characterized by skeletal muscle weakness with a later onset and a preservation of the gait for a longer period (age of 40–50 years) [25]. The patients present usually with high serum CK concentration, calf muscle hypertrophy, muscle cramps, myalgia, or with muscle weakness in the pelvic and shoulder girdles [26]. Cardiomyopathy is a common complication of BMD, the mean age of diagnosis being around 14, 6 years [28]. Heart failure represents the most common cause of death in BMD, at an age of mid-40s [23, 24].

EDMD is a muscular disorder with different inheritance patterns: X-linked recessive or autosomal dominant or autosomal recessive [25, 26]. The clinical picture includes the classical triad: (i) early joint contractures, (ii) slowly progressive muscle weakness and wasting in humeroperoneal distribution (upper arm, lower legs), and (iii) cardiac disease (atrial-ventricular conduction anomalies, atrial arrhythmias) [27]. Usually, the clinical presentation is characterized by Achilles contractures with toe walking in childhood. Later, arm weakness and elbow contractures develop, associated with biceps or triceps wasting with sparing of deltoid

dystrophy, BMD).

spiratory function [28].

11

(27%), and autism spectrum disorder (15%) [29].

2.1.2 Emery-Dreyfus muscular dystrophy (EDMD)

2.1.1.2 Becker muscle dystrophy (BMD)

2.1.1.1 Duchenne muscular dystrophy (DMD)

Clinical and Molecular Diagnosis in Muscular Dystrophies

DOI: http://dx.doi.org/10.5772/intechopen.85339

In this book chapter, we draw attention on clinical manifestation and the most important laboratory investigations such us muscle histopathology, protein analysis, and genetic tests that can help in distinguishing between different forms of muscular dystrophy and could lead to an accurate diagnosis.

### 2. A general approach to the diagnosis of muscular dystrophy

The complexity and similarity of clinical manifestation of these conditions represent a challenge for getting an accurate diagnosis for patients. A complete diagnosis involved clinical examination and patient's medical history, blood tests (creatine kinase and serum transaminase levels), electromyography, muscle biopsy examination, and genetic tests [13–15].

Muscle biopsy had an important role in muscular dystrophy diagnosis and still provides essential information for diagnosis. Although in clinical observations, family history, muscle biopsy, and biochemical tests such as serum creatine kinase (CK) are still important tools for muscular dystrophy diagnosis, protein analysis and genetic study have an increasing importance in accurate establishing a diagnosis.

For several years, until the discovery of other muscle proteins, dystrophin was the only protein studied to establish a diagnosis of muscular dystrophy. It is also used today in the differential diagnosis between Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). In the last 10 years, genes involved in various types of autosomal recessive muscular dystrophy (LGMD) as well as in congenital muscle dystrophies have been identified [16–18].

Differentiation between recessive muscle dystrophies is much more difficult to achieve on the basis of clinical criteria because of phenotypic variability [19], different starting age of onset [20, 21], and a variable progression rate [22].

Identification of protein defect by immunohistochemistry and Western blotting allows firm and specific diagnosis in a wide variety of muscular dystrophies. However, while immunohistochemistry is very useful in identifying abnormal expression of primary protein deficiency, in genetically inherited recessive diseases, it is less useful for identifying primary defect in dominant diseases.

### 2.1 Clinical manifestations and symptoms for most common forms of muscular dystrophy

The patient's medical history and clinical examination allow the doctor to identify the signs and the specific symptoms of the diseases. A complete examination should include evaluation of movement and difficulty controlling movement, gait abnormalities, muscle strength, and the presence of weakness pattern, and also identification of the muscle groups affected.

While the most common sign for different types of muscular dystrophy is the progressive muscle weakness, the other features like age of disease onset, muscle group affected, and rate of progression are specific for each type of muscular dystrophy [21, 23].

#### 2.1.1 Dystrophinopathies

Dystrophinopathies are recessive X-linked disorders caused by mutation in dystrophin gene [1]. Currently, they are recognized as a spectrum of disease with

and protein and genetic analyses becomes the diagnostic gold standard for these disorders. Nevertheless, for many patients with yet unidentified muscular dystro-

In this book chapter, we draw attention on clinical manifestation and the most important laboratory investigations such us muscle histopathology, protein analysis, and genetic tests that can help in distinguishing between different forms of

The complexity and similarity of clinical manifestation of these conditions represent a challenge for getting an accurate diagnosis for patients. A complete diagnosis involved clinical examination and patient's medical history, blood tests (creatine kinase and serum transaminase levels), electromyography, muscle biopsy

Muscle biopsy had an important role in muscular dystrophy diagnosis and still provides essential information for diagnosis. Although in clinical observations, family history, muscle biopsy, and biochemical tests such as serum creatine kinase (CK) are still important tools for muscular dystrophy diagnosis, protein analysis and genetic study have an increasing importance in accurate establishing a diagnosis. For several years, until the discovery of other muscle proteins, dystrophin was the only protein studied to establish a diagnosis of muscular dystrophy. It is also used today in the differential diagnosis between Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). In the last 10 years, genes involved in various types of autosomal recessive muscular dystrophy (LGMD) as well as in

Differentiation between recessive muscle dystrophies is much more difficult to

Identification of protein defect by immunohistochemistry and Western blotting allows firm and specific diagnosis in a wide variety of muscular dystrophies. However, while immunohistochemistry is very useful in identifying abnormal expression of primary protein deficiency, in genetically inherited recessive diseases, it is

2.1 Clinical manifestations and symptoms for most common forms of muscular

The patient's medical history and clinical examination allow the doctor to identify the signs and the specific symptoms of the diseases. A complete examination should include evaluation of movement and difficulty controlling movement, gait abnormalities, muscle strength, and the presence of weakness pattern, and also

While the most common sign for different types of muscular dystrophy is the progressive muscle weakness, the other features like age of disease onset, muscle group affected, and rate of progression are specific for each type of muscular

Dystrophinopathies are recessive X-linked disorders caused by mutation in dys-

trophin gene [1]. Currently, they are recognized as a spectrum of disease with

achieve on the basis of clinical criteria because of phenotypic variability [19], different starting age of onset [20, 21], and a variable progression rate [22].

phy, the diagnosis continues to be challenging.

Muscular Dystrophies

examination, and genetic tests [13–15].

dystrophy

dystrophy [21, 23].

10

2.1.1 Dystrophinopathies

muscular dystrophy and could lead to an accurate diagnosis.

congenital muscle dystrophies have been identified [16–18].

less useful for identifying primary defect in dominant diseases.

identification of the muscle groups affected.

2. A general approach to the diagnosis of muscular dystrophy

involvement of skeletal and cardiac muscle in different degree [24] and include Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD), involving mostly skeletal muscles, and DMD-associated dilated cardiomyopathy (DCM), affecting preferentially the myocardium. The clinical picture in males with dystrophinopathies ranges from mild to severe. The mild forms consist of high level of CK in serum and muscle cramps with myoglobinuria. The severe forms include progressive muscle diseases (Duchenne muscle dystrophy, DMD, Becker muscle dystrophy, BMD).
