Медицинская иммунология (Sep 2024)

The immune response formed by the introduction of a DNA vector containing a cDNA fragment of the human thyroid-stimulating hormone receptor gene into transgenic mice

  • A. V. Zubkov,
  • L. G. Butova,
  • N. S. Kuzmina,
  • I. V. Yakovleva,
  • N. F. Gavrilova,
  • A. A. Kornoukhova,
  • A. V. Sidorov,
  • B. V. Khraim Wael,
  • V. V. Kuzmenko,
  • I. V. Zubkova,
  • V. V. Fadeev

DOI
https://doi.org/10.15789/1563-0625-TIR-16841
Journal volume & issue
Vol. 26, no. 5
pp. 941 – 952

Abstract

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The number of patients with autoimmune thyroid diseases (Graves’disease, Hashimoto’sthyroiditis) is increasing globally. The most important part in the diagnosis of Graves’ disease (GD) is the detection of autoantibodies to the thyrotropin receptor (TSHR) in Graves’ patients’ sera. For the differential diagnosis of antibodies to thyroid antigens, it is promising to use tests based on monoclonal antibodies to TSHR, which can be obtained not only as a result of immunization with native or recombinant TSHR protein, but also through DNA immunization with genetically engineered constructs containing fragments of the TSHR gene. Based on mRNA we isolated from the thyroid tissue in GD, a number of fragments of the thyrotropin receptor gene were cloned, suitable for DNA immunization of animals. The purpose of this work is to evaluate the immunogenic properties of one of the constructed vectors, pVAX1-TSHR (1160), in a mouse model. The successful inclusion of the extracellular domain gene fragment of the human TSHR (1160), which was transfected into CHO cells as a part of the pVAX1 vector was confirmed by immunoblotting and ELISA. The immune response formed to the injection of the pVAX1 vector into BALB/c mice, containing a fragment of the human TSHR gene, was detected in different versions of ELISA. Immunization of animals with the DNA vector pVAX1-TSHR according to an experimentally selected scheme was effective for the formation of mouse splenocytes, secreting antibodies to TSHR, which were used for successful hybridization. This was confirmed by the results of determining antibody production to TSHR in murine blood sera. The level of antibody production remained high (titer more than 1:10.000) at the 8th week of the experiment. As a result of selection of individual clones according to the criteria of proliferative activity and stability of antibody production, the most stable cultures secreting mAbs against TSHR were selected.

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