The diverse N-glycosylation profiles of CD4 + CD25 -and CD4 + CD25 + T cells in Hashimoto's thyroiditis

Sara Trzos¹Marta Szewczyk¹Paweł Link-Lenczowski²Grzegorz Sokolowski³Malgorzata Trofimiuk-Muldner³Katarzyna Bocian⁴Ewa Pocheć^1*

• ¹Department of Glycoconjugate Biochemistry, Institute of Zoology and Biomedical Research, Faculty of Biology, Jagiellonian University, Kraków, Poland
• ²Department of Medical Physiology, Faculty of Health Sciences, Jagiellonian University Medical College, Krakow, Poland, Jagiellonian University, Kraków, Poland
• ³Department of Endocrinology, Faculty of Medicine, Jagiellonian University Medical College, Krakow, Poland, Jagiellonian University, Kraków, Poland
• ⁴Department of Immunology, Institute of Experimental Zoology, Faculty of Biology, University of Warsaw, Warsaw, Poland, Uniwersytet Warszawski, Warsaw, Poland

Hashimoto's thyroiditis (HT) is one of the most common organ-specific autoimmune diseases, characterized by chronic thyroid gland inflammation. Helper T (Th) CD4 + cells, whose surface receptors are highly glycosylated, are involved in the pathomechanism of HT. Our study aimed to characterize N-glycosylation profiles in two pools of CD4 + T cells, defined by the expression of CD25 + late activation marker (CD4 + CD25 + ) and CD25-negative cells (CD4 + CD25 -) in HT. Two study groups were recruited: HT1 with elevated thyroid autoantibodies and TSH level within the normal range without hypothyroidism, and HT2, hypothyroid HT patients, adequately metabolically controlled while on L-thyroxine replacement therapy, and healthy subjects to the control group (CTR). N-glycans from CD4 + cell proteins, released using N-glycosidase F, were analyzed by MALDI-Tof mass spectrometry. RT-qPCR was used to determine the expression of selected glycogenes. We found significant differences in the glycome of CD4 + CD25 -and CD4 + CD25 + cells. In homeostasis (CTR), a predominance of complex-type glycans was observed in CD4 + CD25 -cells, whereas the oligomannosetype structures prevail in CD4 + CD25 + lymphocytes. In autoimmunity and progressive thyroid dysfunction, the rearrangement of N-glycans in Th cells was observed, in opposite directions in the CD4 + pools. Complex-type structures are replaced by oligomannose forms in CD4 + CD25 -in the HT1 group, while in HT2, a restoration of glycosylation profile to the level of CTR was detected. CD4 + CD25 + cells accelerated complex-type synthesis in HT1, which was normalised in HT2 patients. Changes in the profile of N-linked glycans are partially reflected in the expression of mannosidases and glycosyltransferases. Our study demonstrates for the first time the diverse N-glycosylation profiles in CD4 + CD25 -and CD4 + CD25 + cells, and the rearrangement of N-glycan structures specific for each pool of Th cells in HT. Further studies are needed to determine the functional aspect of the identified N-glycosylation changes during thyroid autoimmunity.