Generation of recombinant, enzymatically active human thyroid peroxidase and its recognition by antibodies in the sera of patients with Hashimoto's thyroiditis.

Area of Science:

• Biochemistry
• Immunology
• Molecular Biology

Background:

• Thyroid peroxidase (TPO) is a critical enzyme in thyroid hormone synthesis.
• Thyroid microsomal antigen (TMA) is a primary target in autoimmune thyroid diseases like Hashimoto's thyroiditis.
• The precise role and structure of TPO within TMA require further elucidation.

Purpose of the Study:

• To express enzymatically active human TPO in a nonthyroidal cell line.
• To confirm TPO as a major component of the thyroid microsomal antigen.
• To investigate the immunoreactivity of recombinant TPO with anti-microsomal antibodies.

Main Methods:

• Stable transfection of Chinese hamster ovary (CHO) cells with human TPO cDNA.
• Flow cytometry (FACS) analysis to detect cell surface TPO expression.
• Western blot analysis of particulate antigen from transfected cells using Hashimoto's sera.

Main Results:

• Enzymatically active human TPO was successfully expressed on the surface of CHO cells.
• Western blot analysis revealed TPO-specific bands (approx. 200 kD and 110 kD) recognized by Hashimoto's sera.
• Antibody reactivity correlated with anti-microsomal antibody titers, and reducing conditions altered band patterns.

Conclusions:

• Functionally active human TPO can be expressed in eukaryotic nonthyroidal cells.
• These findings demonstrate that TPO is a major component of the thyroid microsomal antigen.
• This provides a valuable tool for studying autoimmune thyroid diseases.