口服免疫耐受大鼠脾淋巴细胞对肾小球系膜细胞中NF-κBp65活性的影响

The effect of splenic lymphocytes from oral immune tolerance rats on activity of NF-κB p65 in glomerular mesangial cells

目的 :探讨口服免疫耐受大鼠脾淋巴细胞对肾小球系膜细胞中NF κBp6 5活性的影响。方法 :将 10只 6～ 8wk龄雌性Wistar大鼠随机分为两组 ,每组 5只。一组用Fx1A抗原灌胃诱导其产生免疫耐受 ,另一组用PBS灌胃作为对照组。 2wk后 ,杀鼠取脾、分离淋巴细胞进行抗原特异性淋巴细胞增殖实验。制备免疫耐受大鼠脾淋巴细胞培养上清 ,用夹心ELISA法测定其中IL 10的水平。将体外培养的系膜细胞用高水平的胰岛素 ,同时加入大鼠的脾淋巴细胞培养上清作用 2 4h。用免疫组化染色及夹心ELISA法检测系膜细胞中NF κBp6 5的活性。结果 :与对照组大鼠相比较 ,口服免疫耐受组大鼠抗原特异性脾淋巴细胞的增殖明显受到抑制 ,其脾淋巴细胞培养上清中IL 10的水平明显升高 (P <0 .0 1)。在体外实验中发现 ,口服免疫耐受大鼠脾淋巴细胞培养上清 ,可抑制胰岛素刺激的系膜细胞内NF κBp6 5的活性 (P <0 .0 5 )。结论 :口服免疫耐受大鼠的脾淋巴细胞 ,可能通过其分泌的IL 10抑制系膜细胞内NF κBp6

AIM: To explore the effect of splenic lymphocytes from oral immune tolerance rats on NF-κB p65 activity in glomerular (mesangial) cells. METHODS: 10 female Wistar rats with (6-8 weeks) of age were divided randomly into two groups, 5 rats for each group. The rats of one group were given through intragastric feeding with 2 mL Fx1A antigen to induce oral immune tolerance, and the rats of the other group only with PBS (as control group). Two weeks later, the rats were sacrificed, and then their spleens were excised and lymphocytes were separated routinely for antigen-specific lymphocytes reaction. The culture supernatants of splenic lymphocytes from oral immune tolerance rats were prepared. The IL-10 level in the supernatants was detected by sandwich ELISA. After the cultured mesangial cells were stimulated with high dose of insulin, the culture supernatants of rat’s splenic lymphocytes were added to insulin-treated mesangial cells. Twenty-four hours later, the NF-κB p65 activity in mesangial cells was detected by immunohistochemical staining and sandwich ELISA. RESULTS: As compared with control group, the proliferation reaction of splenic lymphocytes from oral immune tolerance rats was inhibited obviously, and IL-10 level in splenic lymphocyte culture supernatants from the tolerance rats rose markedly(P<0.01). (NF-κB) p65 activity induced by insulin in mesangial cells was inhibited by culture superatants of splenic lymphocytes from oral immune toterance rats(P<0.05). CONCLUSION: NF-κB p65 activity in mesangial cells may be inhibited by IL-10 secreted by splenic lymphocytes from the oral immune tolerance rats.