CDR3Characteristics And Antigen Recognization Of Human Peripheral Blood Vγ9Vδ2T Lymphocytes From Patients With Systemic Lupus Erythematosus

With high incidence and diverse clinical manifestations, autoimmune diseases are lifelong diseases. They are terrible with regard to patients, families and the society, a typical of which is SLE. Etiology and pathogenesis of most autoimmune diseases are not very clear. Specific effective treatment is not available. Only the adrenal cortex hormones and immunosuppressive agents are used to control the disease, and a considerable number of patients are not sensitive to such treatments.T cell immunity plays an important role in anti-tumor, anti-virus, autoimmune diseases and transplantation immunology.γδ T cells, a unique T-cell subsets, not only take part in both specific and non-specific immune responses, but also have unique receptor and biological characteristics. Most importantly, they can recognize the conservative endogenous and stress-induced antigen in the absence of antigen presenting cells, suggesting that γδ T cells may be involved in autoimmune disease pathogenesis. T cell receptors are critical receptors during T cell-specific antigen recognition and specific cellular immune responses. Further researches on T cell immunity depend on clarification T cell receptor gene complex system. In recent decades, although great progress has been made on γδ T cells, structural basis of γδ TCR ligand recognition is still not fully understood. Progresses of γδ TCR ligand identification are fairly slow. To date, only few antigens recognized by human γδ T cells have been defined. To better understand γδ T cell function, a crucial issue is to identify more ligands for γδ T cells. It is also the key point to understand the principles governing γδ TCR specificity and diversity and to reveal the biological functions of γδ T cells.After analysis of peripheral blood γδ T cell receptor CDR3domain structure and comparison of Vγ CDR3and Vδ CDR3region expression characteristics in patients with SLE, we chose synthetic shared and superiority TCR82-CDR3sequence, and used immunofluorescence and enzyme immunoassay techniques to observe binding properties of the peptide and plasma/cells derived from patients with SLE. Then we verified that γδT cells indeed infiltrated in the corresponding organizations in patients with SLE and chose HK-2cell lines as the target cells in the future. Subsequently, a variety of experiments were used to validate that the shared and superiority TCRδ2-CDR3sequence could bind HK-2cell line surface and block the cytotoxity effect of y8T cells to HK-2, indicating that the very TCRδ2-CDR3sequence can identify and combine a particular molecule on HK-2cells surface. Finally, HK-2cell proteins were isolated and the special binding protein was identified by mass spectrum technology. Expression of the possible ligand protein on HK-2cell surface was also verified.Disorder of immuneregulatory is an important factor that can induce autoimmune diseases. The negative regulation of T cells mainly includes three aspects:inhibitory T cells, activated T cell apoptosis and inhibitory receptors of immune cells. In the second part, we observed expression of activation markers, activated receptors and inhibitory receptors on γδ T cells. We hope to enrich the γδ T cell function and clarify whether these unique cells play a role in the pathogenesis of SLE from another point of view.We conducted the experiments and got some results shown as follows:1. We found some clues of Vγ-chain CDR3Region in γδ T cell antigen recognition and confirmed that Vγ chain expression not only involved in γδ T cell infiltration in selective tissues, but also took part in γδ T cell antigen recognition processes;2. There were discrepancies of Vy chain CDR3Region and Vδ chain CDR3Region changes in SLE status compared with healthy donors, which is helpful to understand the biological significance of γδ T cell receptor during antigen recognition processes;3. A dominant TCRδ2-CDR3sequence was shared by different patients with SLE, which confirmed the critical role of CDR3δ in antigen binding specificity of y8TCR. The very synthetic CDR3δ peptide was used as an excellent probe to find new ligands of γδ TCR.4. A possible γδ TCR CDR3bounded protein; 5. A state of low level expression of CD94/NKG2A on peripheral blood γδ T cells and antigen stimulated γδ T cells were detected, which may be a powerful explanation for overactivated γδ T cells in SLE status.