The Immunosuppressive And Protective Ability Of HAAT For Beta Cells Transplantation In Diabetes

Objective： The investigate the immunosuppressive and protective effect of human alpha1-antitrypsin (hAAT) in the beta cell transplantation of diabetes. We constructed an insulinomacell line, transfected with hAAT, namely NIT-hAAT cells.Methods: The expression of hAAT was detected by Western Blotting. NIT-hAAT cells weretransplanted into the subrenal capsule of diabetes mice. In order to observe the effect of distincttreatment in diabetes, we detect the blood glucose, serum insulin level and survival time of micefrom different transplantation groups. These cells viability and the expression of hAAT gene intransplant parts were determined by glucose tolerance test and RT-PCR, respectively. Besides,Observe the immune protection of hAAT in vivo, the leves of pro-inflammatory cytokines(TNF-α, IFN-γ, IL-1β and IL-2) and anti-inflammatory cytokines (IL-4) in serum or transplantparts were detected by ELISA and RT-PCR, respectively. The biopsy of tissues was stained byhematoxylin and eosin (HE). The CCK-8kit was used to determine the spleen lymphocyteproliferation ability, in which the percentage of Th17/Treg cell in CD4+cells were detected byflow cytometry. The DCs from marrow of mice were isolated, cultured and detected the cellphenotype by flow cytometry, while the leves of IL-10and IL-12from supernatant wereanalysed by ELISA.Results: After NIT-hAAT cells transplantation, the blood glucose levels of mice declinedeffectively and maintained to be normal. The expression of hAAT can prolong the survival timeof graft. Transplantation with NIT-hAAT cells can enhance the survival rate of diabetes mice(P<0.05). At the beginning of trsplantation, the expression of hAAT was stable and protected theviability of NIT-hAAT cells. Moreover, hAAT can inhibit the synthesis of inflammatorycytokines, and promote anti-inflammatory cytokine secretion, so that it could preven theoccurrence of non-specific inflammation effectively. The result of pathology test with HEstaining confirmed that the hAAT can significantly reduce graft inflammation. Besides, hAATcould significantly inhibit the proliferation of spleen lymphocytes, and additionally make a greatimpact on the balance of Th17/Treg cells and the mature of DCs. Therefore, hAAT had immuneprotection in β cells transplantation. Conclusions: NIT-hAAT cells expressing hAAT can inhibit the immune rejection and prolongthe survival time of β cells.