Nee Larine, Tuite Niamh, Ryan Michael P, McMorrow Tara
School of Biomolecular and Biomedical Science, UCD Conway Institute, University College Dublin, Dublin, Ireland.
Nephron Exp Nephrol. 2007;107(2):e73-86. doi: 10.1159/000108645. Epub 2007 Sep 20.
Renal cells such as mesangial cells are known to secrete metalloproteinases that are capable of degrading the constituents of the glomerular basement membrane (GBM). Disruption of the GBM via cytokine-induced alterations in matrixmetalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) may be an important mechanism in the renal disease process. In renal disease, both resident renal cells and infiltrating immune cells are capable of secreting pro-inflammatory cytokines including tumour necrosing factor-alpha (TNFalpha) and interleukin-1 beta (IL-1 beta). In this study, we examine the potential of these cytokines to alter levels of MMPs and TIMPs in human mesangial cells.
The T-HMC human mesangial cell line was cultured in RPMI 1640 containing 5% serum. Cells at confluency were serum starved for 24 h prior to exposure to TNF-alpha (0.1-100 ng/ml) or IL-1 beta (0.1-100 ng/ml) or a combination of both for 48 h. Activity of MMP-9 was examined by gelatin zymography and TIMP-1 expression was analysed by Western blotting.
TNF-alpha but not IL-1 beta resulted in a dose-dependent increase in the latent form of MMP-9 and a decrease in TIMP-1 production. Co-treatment with IL-1 beta had no effect on the induction of MMP-9 but increased the inhibition of TIMP-1 in the presence of TNF-alpha. Inhibition of PKC provided evidence of the importance of this pathway in mediating the TNF-alpha-induced suppression of TIMP-1. Activation of the ERK 1/2 MAPK mediated both the upregulation of MMP-9 and the inhibition of TIMP-1 following TNF-alpha treatment. p38 MAPK activation was also found to be involved in the TNF-alpha-stimulated MMP-9.
The cytokine TNF-alpha causes different effects on human mesangial MMP-9 and TIMP-1 expression which are mediated through the TNF-RI, and the different signalling pathways of PKC, ERK 1/2 and p38 MAPK. This suggests an important role for pro-inflammatory cytokines in renal disease progression.
已知诸如系膜细胞等肾细胞会分泌能够降解肾小球基底膜（GBM）成分的金属蛋白酶。细胞因子诱导的基质金属蛋白酶（MMPs）和金属蛋白酶组织抑制剂（TIMPs）的改变导致GBM破坏可能是肾脏疾病进程中的一个重要机制。在肾脏疾病中，驻留肾细胞和浸润免疫细胞均能够分泌促炎细胞因子，包括肿瘤坏死因子-α（TNFα）和白细胞介素-1β（IL-1β）。在本研究中，我们检测了这些细胞因子改变人系膜细胞中MMPs和TIMPs水平的可能性。
将T-HMC人系膜细胞系培养于含5%血清的RPMI 1640培养基中。汇合的细胞在暴露于TNF-α（0.1 - 100 ng/ml）或IL-1β（0.1 - 100 ng/ml）或两者组合之前血清饥饿24小时，持续48小时。通过明胶酶谱法检测MMP-9的活性，通过蛋白质印迹法分析TIMP-1的表达。
TNF-α而非IL-1β导致MMP-9潜伏形式呈剂量依赖性增加以及TIMP-1产生减少。IL-1β联合处理对MMP-9的诱导无影响，但在存在TNF-α的情况下增加了对TIMP-1的抑制。蛋白激酶C（PKC）的抑制提供了该途径在介导TNF-α诱导的TIMP-1抑制中的重要性的证据。细胞外信号调节激酶1/2（ERK 1/2）丝裂原活化蛋白激酶（MAPK）的激活介导了TNF-α处理后MMP-9的上调和TIMP-1的抑制。还发现p38 MAPK激活参与了TNF-α刺激的MMP-9。
细胞因子TNF-α对人系膜MMP-9和TIMP-1表达产生不同影响，这些影响通过TNF-RI以及PKC、ERK 1/2和p38 MAPK的不同信号通路介导。这表明促炎细胞因子在肾脏疾病进展中起重要作用。
