Souza Kleber L A, Gurgul-Convey Ewa, Elsner Matthias, Lenzen Sigurd
Hannover Medical School, Institute of Clinical Biochemistry, 30625 Hannover, Germany.
J Endocrinol. 2008 Apr;197(1):139-50. doi: 10.1677/JOE-07-0638.
Pro-inflammatory cytokines cause beta-cell dysfunction and death. The aim of this study was to investigate the interactions between different pro- and anti-inflammatory cytokines and their effects on apoptotic beta-cell death pathways. Insulin-producing RINm5F cells were exposed to different combinations of cytokines. Gene expression analyses of manganese superoxide dismutase (MnSOD) and inducible nitric oxide synthase (iNOS) were performed by real-time RT-PCR. Cell viability was measured by the MTT assay, NFkappaB activation using a SEAP reporter gene assay, protein expression by western blotting and caspase-3 activity using the DEVD cleavage method. IL-1beta, tumour necrosis factor alpha (TNFalpha) and a combination of all three pro-inflammatory cytokines increased while IFNgamma alone did not affect NFkappaB activity and iNOS gene and protein expression. Interestingly, the anti-inflammatory cytokines IL-4, IL-13 and IL-10 decreased IL-1beta-stimulated NFkappaB activation and iNOS expression. IL-1beta, TNFalpha and the pro-inflammatory cytokine combination also increased MnSOD gene and protein expression. But IL-4, IL-13 and IL-10 did not affect MnSOD expression and did not modulate IL-1beta-stimulated MnSOD expression. Caspase-3 activity was increased by IL-1beta and the pro-inflammatory cytokine combination, and to a lesser extent by TNFalpha. In contrast, IFNgamma had no effect on caspase-3 activity. IL-4, IL-13 and IL-10 decreased caspase-3 activity and increased viability of insulin-producing cells treated with pro-inflammatory cytokines. The anti-inflammatory cytokines counteracted the cytotoxic effects of pro-inflammatory cytokines in insulin-producing cells. This was achieved through the reduction of nitrosative stress. Thus, a balance between the anti-inflammatory and the pro-inflammatory cytokines is of crucial importance for the prevention of pancreatic beta-cell destruction.
促炎细胞因子会导致β细胞功能障碍和死亡。本研究的目的是调查不同促炎和抗炎细胞因子之间的相互作用及其对β细胞凋亡死亡途径的影响。将产生胰岛素的RINm5F细胞暴露于不同细胞因子组合中。通过实时逆转录聚合酶链反应（RT-PCR）对锰超氧化物歧化酶（MnSOD）和诱导型一氧化氮合酶（iNOS）进行基因表达分析。通过MTT法测定细胞活力，使用SEAP报告基因测定法检测核因子κB（NFκB）激活情况，通过蛋白质印迹法检测蛋白质表达，并使用DEVD切割法检测半胱天冬酶-3活性。白细胞介素-1β（IL-1β）、肿瘤坏死因子α（TNFα）以及所有三种促炎细胞因子的组合可增加NFκB活性，而单独的γ干扰素（IFNγ）不影响NFκB活性以及iNOS基因和蛋白质表达。有趣的是，抗炎细胞因子IL-4、IL-13和IL-10可降低IL-1β刺激的NFκB激活和iNOS表达。IL-1β、TNFα以及促炎细胞因子组合也可增加MnSOD基因和蛋白质表达。但IL-4、IL-13和IL-10不影响MnSOD表达，也不调节IL-1β刺激的MnSOD表达。IL-1β和促炎细胞因子组合可增加半胱天冬酶-3活性，TNFα在较小程度上也可增加该活性。相比之下，IFNγ对半胱天冬酶-3活性无影响。IL-4、IL-13和IL-10可降低半胱天冬酶-3活性，并增加经促炎细胞因子处理的产生胰岛素细胞的活力。抗炎细胞因子抵消了促炎细胞因子对产生胰岛素细胞的细胞毒性作用。这是通过减少亚硝化应激实现的。因此，抗炎细胞因子和促炎细胞因子之间的平衡对于预防胰腺β细胞破坏至关重要。
