SUPPRESSION OF EXPERIMENTAL AUTOIMMUNE UVEORETINITIS BY NEUROPEPTIDE GENE-TRANSFECT-CELLS

探讨降钙素基因相关肽（CGRP）培养的成熟树突状细胞（DC）对小鼠实验性自身免疫性葡萄膜视网膜炎（EAU）的抑制作用。用GM-CSF培养C57BL/6小鼠骨髓细胞6天，用autoMACS纯化分化的CD11^+未成熟DC。将未成熟DC用LPS （1 μg/ml）培养过夜，然后在CGRP或TGF-B2存在下用人光间受体类视黄酸结合蛋白（IRBP）残基1-20 （hIRBP-p）培养。用弗氏佐剂和百日咳毒素免疫hIRBP-p诱导小鼠EAU。同时，将经hirbp -p免疫的小鼠分别静脉注射CGRP或TGF-B2培养的成熟DC。免疫后第19天测定hirbp -p特异性延迟超敏反应（DH）。免疫后第21天处死动物，病理评估EAU程度。通过注射CGRP或TGF-B2和hIRBP-p培养的成熟DC， EAU和抗原特异性DH明显受到抑制。对照组（15只EAU小鼠中13只）的发生率为87%，TGF-B2培养组（15只EAU小鼠中9只）的发生率为60%，CGRP培养组（18只EAU小鼠中5只）的发生率为28%。在接下来的实验中，我们采用电穿孔法制备了CGRP基因转染- dc。结果表明，CGRP基因转染- dc可抑制EAU，抑制IRBP-p的DH特异性。结果表明，CGRP和CGRP基因转染DC培养的成熟DC对EAU的发展具有免疫抑制作用，且比TGF-B2培养的更有效。

To investigate the role of the suppression in murine experimental autoimmune uveoretinitis (EAU) by mature dendritic cells (DC) cultured with calcitonin gene-related peptide (CGRP). Bone marrow cells derived from C57BL/6 mice were cultured with GM-CSF for six days, and differentiated CD11^+ immature DC were purified by autoMACS. Immature DC were further cultured with LPS (1 μg/ml) overnight, and then with human interphotoreceptor retinoid binding protein (IRBP) residues 1-20 (hIRBP-p) in the presence of CGRP or TGF-B2. EAU was induced by immunization with hIRBP-p in Freund' s adjuvant and pertussis toxin in mice. At the same time, hIRBP-p-immunized mice were injected the mature DC cultured with CGRP or TGF-B2 intravenously. On day 19 after immunization, hIRBP-p-specific delayed hypersensitivity (DH) were measured. On day 21 after immunization, animals were sacrificed, and assessed the extent of EAU pathologically. EAU and antigen-specific DH was predominantly suppressed by injection of mature DC cultured with CGRP or TGF-B2 and hIRBP-p. Incidence of EAU on control group was 87% (13 of 15 EAU mice), TGF-B2 culture group was 60% (9 of 15 EAU mice), and CGRP culture group was 28% (5 of 18 EAU mice). On the next experiments, we prepared CGRP gene-transfect-DC by electroporation methods. As the results, CGRP gene-transfect-DC can suppress EAU, and suppress DH spepific for IRBP-p. It was demonstrated that mature DC cultured with CGRP and CGRP gene-transfect-DC play an immunosuppressive role in development of EAU, which were more effective than those cultured with TGF-B2.