EVIDENCE FOR INVOLVEMENT OF PROTEASOME COMPLEX

白介素1(IL-1)被认为是一种效应分子 自身免疫性糖尿病的(-)细胞破坏。IL-1对胰岛素的抑制作用 胰腺(-)细胞通过刺激其表达分泌 诱导型一氧化氮合酶(INOS)，可产生一氧化氮。 IL-1还可诱导可诱导的同种异构体 环氧合酶(COX-2)与促炎性细胞过度产生 前列腺素及其前体花生四烯酸 用同位素稀释质谱仪测定。当前的研究 蛋白水解酶(S)参与信号通路的特征 抑制IL-1诱导的iNOS和COX-2的表达。生化与分子 使用大鼠胰岛素瘤(-)细胞系RINm5F进行研究。 丝氨酸蛋白酶抑制剂N(-对甲苯磺基-L-赖氨酸氯甲基酮) (TLCK)和蛋白酶体复合体(26S)抑制剂MG 132被抑制 IL-1诱导一氧化氮氧化产物亚硝酸盐的形成 由iNOS以浓度依赖的方式产生。 TLCK和MG 132也能抑制iNOS基因的表达 MRNA和蛋白质水平。TLCK和MG 132也抑制IL-1 诱导COX-2酶活性(PGE2形成)和COX-2基因 在信使核糖核酸和蛋白质水平表达。在人类的胰岛中， 蛋白酶体抑制剂MG 132也抑制了 INOS和COX-2酶活性、亚硝酸盐和前列腺素E_2的乘积 分别进行了分析。这些发现表明，黄曲霉毒素的抑制作用 TLCK和MG 132对iNOS和COX-2表达的影响先于转录。 转录因子，核因子(B)，是激活a 细胞因子诱导的酶的数量，并被评估为可能的 IL-1诱导共表达所需的蛋白酶作用部位 INOS和COX-2。TLCK和MG 132均抑制IL-1诱导 由胰岛和RINm5F细胞激活核因子(B)和降解1(B)。 这些结果表明，蛋白水解酶的激活是一种早期信号 IL-1诱导的(-细胞功能抑制)事件。

Interleukin-1( (IL-1) has been implicated as an effector molecule of (-cell destruction in autoimmune diabetes. IL-1 inhibits insulin secretion from pancreatic (-cells by stimulating the expression of inducible nitic oxice synthase (iNOS) that generates nitric oxide. IL-1 also induces co-expression of the inducible isoform of cyclooxygenase (COX-2) and overproduction of proinflammatory prostaglandins and their precursor arachidonic acid, as demonstrated by isotope dilution mass spectrometry. The current studies characterize the involvement of protease(s) in the signaling pathway of IL-1-induced iNOS and COX-2 expression. Biochemical and molecular studies were performed using the rat insulinoma (-cell line, RINm5F. A serine protease inhibitor, N(-p-tosyl-L-lysine chloromethyl ketone (TLCK) and a proteasome complex (26S) inhibitor, MG 132, inhibited Il-1-induced nitrite formation, an oxidation product of nitric oxide produced by iNOS, in a concentration-dependen t manner. Both TLCK and MG 132 also inhibited iNOS gene expression at the level of mRNA and protein. TLCK and MG 132 also inhibited IL-1 induced COX-2 enzyme activity (PGE2 formation) and COX-2 gene expression at the level of mRNA and protein. In human islets, the proteasome inhibitor, MG 132, also inhibited the formation of the products of iNOS and COX-2 enzyme activity, nitrite and PGE2 respectively. These findings suggested that the inhibitory action of TLCK and MG 132 on iNOS and COX-2 expression precedes transcription. The transcription factor, NF(B, is essential for activation of a number of cytokine-inducible enzymes and was evaluated as a possible site of protease action necessary for IL-1 induced co-expression of iNOS and COX-2. TLCK and MG 132 inhibited both IL-1 induced activation of NF(B and degradatin of 1(B( by islets and RINm5F cells. These results implicate protease activation as an early signaling event in IL-1-induced inhibition of (-cell function.