**3.1 Autoantibodies**

The major markers of Graves' Disease include TSHR-directed autoantibodies (TRAb). TSHR belongs to the family of glycoprotein hormone receptors and stimulates adenyl cyclase (cAMP) through G protein. The cAMP activates all the functions of thyroid cells, e.g. thyroglobulin synthesis, functioning of iodine pump, activity of thyroid peroxidase and release of hormones. Thyrotropin is a physiological agonist for the receptor.

Three types of TSH receptor antibodies can be distinguished:


In rare cases, both TSI and TBI are present or they are changed, one into the other, due to treatment. In clinical studies, the level of TRAb can be assessed using methods based on receptor binding (the so called binding assays), used to detect antibodies in the blood that compete with TSH for binding to its receptor (TSH binding inhibitory immunoglobulins, TBII or TRAb). However, in this way it is impossible to differentiate between their biological, stimulating or blocking activity.

The newest method, the so called bioassay (biological tests), is not routinely applied due to high costs. It can be used to determine the level of cAMP production after TSI binding to TSH receptor. The method is also used to monitor thyroid opthalmopathy and in the case of doubtful diagnosis of Graves' Disease due to borderline or negative TRAb values. Currently, no system is available to measure the activity of neutral antibodies. In our patients, TSI and TBI were determined with a

#### *Graves' Disease*

biological test using Chinese hamster ovary cells with an embedded gene encoding the TSH receptor and with the luciferase system. TSI from a sample obtained from a patient binds to the receptor and via cAMP triggers the production of luciferase, thereby initiating a light reaction. The emission is measured with a luminometer. The measured values are compared to the reference values and in this way the presence of antibodies is confirmed or denied. The cut-off point in the analysis using a Thyretain TSI bioassay is the sample-to-reference ratio of more than 140% for the stimulating antibodies, and the degree of inhibition over 40% for the blocking antibodies.

Patients with hyperthyroidism and concomitant Graves' opthalmopathy showed significantly higher values of TSI/TSAb before and during treatment as compared to hyperthyroid patients without opthalmopathy (**Figure 4**). Patients in the prepubertal age had higher levels of TSI/TSAb than those in the pubertal age. Girls had significantly higher values than boys (p < 0.02) (**Figures 5** and **6**).

Moreover, the comparison of the proportion of positive values of TSI/TSAb vs. TBII in the whole group of patients with untreated Graves' Disease and

#### **Figure 4.**

*Levels of thyroid stimulating immunoglobulins (TSAb): (A) untreated children; (B) children treated with Graves' Disease and with or without ophthalmopathy.*

#### **Figure 5.**

*Levels of thyroid stimulating immunoglobulins (TSAb) in children with Graves' Disease before and during adolescence.*

*Hyperthyroidism in Children DOI: http://dx.doi.org/10.5772/intechopen.97444*

#### **Figure 6.**

*Levels of thyroid stimulating immunoglobulins (TSAb) in females and males with Graves' Disease.*

#### **Figure 7.**

*Comparison of positive values of TSI/TSAb to TBII levels in all patients with Graves' Disease – untreated and with Graves' Disease and ophthalmopathy.*

**Figure 8.** *ROC curves for TSI and TBII levels in patients with Graves' Disease; (A) untreated; (B) treated.*

Graves' Disease with concomitant opthalmopathy revealed significantly higher values for TSAb than TBII in each group (**Figure 7**). The ROC curves for TSI and TBII in the above mentioned group of patients with Graves' Disease are presented in **Figure 8**.

Graves' Disease is also characterized by the presence of autoantibodies directed against various components of the thyroid, i.e. antibodies against thyroid peroxidase (anti-TPO), thyroglobulin (anti-TG). However, they are not highly specific and sensitive in the detection of this disease [15–17].
