Amoozadeh Yasaman, Dan Qinghong, Xiao Jenny, Waheed Faiza, Szászi Katalin
Keenan Research Centre for Biomedical Science of the St. Michael's Hospital and Department of Surgery, University of Toronto, Ontario, Canada.
Keenan Research Centre for Biomedical Science of the St. Michael's Hospital and Department of Surgery, University of Toronto, Ontario, Canada
Am J Physiol Cell Physiol. 2015 Jul 1;309(1):C38-50. doi: 10.1152/ajpcell.00388.2014. Epub 2015 May 6.
The inflammatory cytokine tumor necrosis factor-α (TNF-α) is a pathogenic factor in acute and chronic kidney disease. TNF-α is known to alter expression of epithelial tight junction (TJ) proteins; however, the underlying mechanisms and the impact of this effect on epithelial functions remain poorly defined. Here we describe a novel biphasic effect of TNF-α on TJ protein expression. In LLC-PK1 tubular cells, short-term (1-6 h) TNF-α treatment selectively elevated the expression of the channel-forming TJ protein claudin-2. In contrast, prolonged (>8 h) TNF-α treatment caused a marked downregulation in claudin-2 and an increase in claudin-1, -4, and -7. The early increase and the late decrease in claudin-2 expression involved distinct mechanisms. TNF-α slowed claudin-2 degradation through ERK, causing the early increase. This increase was also mediated by the EGF receptor and RhoA and Rho kinase. In contrast, prolonged TNF-α treatment reduced claudin-2 mRNA levels and promoter activity independent from these signaling pathways. Electric Cell-substrate Impedance Sensing measurements revealed that TNF-α also exerted a biphasic effect on transepithelial resistance (TER) with an initial decrease and a late increase. Thus there was a good temporal correlation between TNF-α-induced claudin-2 protein and TER changes. Indeed, silencing experiments showed that the late TER increase was at least in part caused by reduced claudin-2 expression. Surprisingly, however, claudin-2 silencing did not prevent the early TER drop. Taken together, the TNF-α-induced changes in claudin-2 levels might contribute to TER changes and could also play a role in newly described functions of claudin-2 such as proliferation regulation.
炎性细胞因子肿瘤坏死因子-α（TNF-α）是急性和慢性肾脏疾病的致病因素。已知TNF-α可改变上皮紧密连接（TJ）蛋白的表达；然而，其潜在机制以及这种效应上皮功能的影响仍不清楚。在此，我们描述了TNF-α对TJ蛋白表达的一种新型双相效应。在LLC-PK1肾小管细胞中，短期（1-6小时）TNF-α处理选择性地提高了形成通道的TJ蛋白claudin-2的表达。相反，长时间（>8小时）TNF-α处理导致claudin-2明显下调，而claudin-1、-4和-7增加。claudin-2表达的早期增加和晚期减少涉及不同机制。TNF-α通过ERK减缓claudin-2降解，导致早期增加。这种增加也由表皮生长因子受体、RhoA和Rho激酶介导。相反，长时间TNF-α处理降低了claudin-2 mRNA水平和启动子活性，且与这些信号通路无关。细胞-基质阻抗传感测量显示，TNF-α对跨上皮电阻（TER）也有双相效应，最初降低，后期增加。因此，TNF-α诱导的claudin-2蛋白变化与TER变化之间存在良好的时间相关性。事实上，沉默实验表明，后期TER增加至少部分是由claudin-2表达降低引起的。然而，令人惊讶的是，claudin-2沉默并不能阻止早期TER下降。综上所述，TNF-α诱导的claudin-2水平变化可能导致TER变化，也可能在claudin-2新描述的功能（如增殖调节）中发挥作用。
