Renal Division, Department of Internal Medicine, National Taiwan University Hospital, College of Medicine, National Taiwan University, Taipei, Taiwan.
Br J Pharmacol. 2010 Aug;160(7):1611-20. doi: 10.1111/j.1476-5381.2010.00837.x.
Renal fibroblasts play a pivotal role in the development of tubulointerstitial fibrosis, a condition highly predictive of progression towards end-stage renal disease. The present study investigated the anti-mitogenic and anti-inflammatory effects of an inhibitor of inosine monophosphate dehydrogenase, mycophenolic acid (MPA) and the mechanisms underlying its action in normal rat kidney fibroblasts (49F cells).
Proliferation of 49F cells was studied by tetrazole 3-(4, 5-dimethylthiazol-2-yl-)-2,5-diphenyltetrazolium bromide (MTT) test, bromodeoxyuridine incorporation and flow cytometry. The cyclins, tumour suppressor genes and phospho-mitogen-activated protein kinases (MAPKs) were semiquantified by immunoblotting. Apoptosis was measured by quantifying the fragmented DNA and the activity of caspase 3. The monocyte chemokine CCL2 was measured by ELISA. The mRNA expression of CCL2 was measured by real-time PCR.
Mycophenolic acid dose-dependently inhibited steady-state proliferation of 49F cells by up-regulation of p21, p27 and p53, in association with a decrease in cyclins D2 and E. Treatment with MPA also triggered apoptosis of 49F cells by activating the caspase 3 cascade. Furthermore, MPA attenuated tumour necrosis factor-alpha-induced CCL2 expression through down-regulation of p38 MAPK, but not that of ERK1/2 or JNK.
The anti-mitogenic and anti-inflammatory effects of MPA were mediated by up-regulation of cell cycle inhibitors and pro-apoptotic signals, and by suppression of p38 MAPK pathway respectively. This dual effect of MPA may form the rationale for animal or clinical trials for the treatment of fibrotic renal diseases.
肾成纤维细胞在肾小管间质纤维化的发展中起着关键作用，肾小管间质纤维化是导致终末期肾病进展的高度预测因素。本研究旨在探讨肌苷单磷酸脱氢酶抑制剂霉酚酸（MPA）对正常大鼠肾成纤维细胞（49F 细胞）的抗有丝分裂和抗炎作用及其作用机制。
通过四唑 3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四唑溴盐（MTT）试验、溴脱氧尿苷掺入和流式细胞术研究 49F 细胞的增殖。通过免疫印迹法半定量测定细胞周期蛋白、肿瘤抑制基因和磷酸化丝裂原活化蛋白激酶（MAPKs）。通过定量检测片段化 DNA 和 caspase 3 的活性来测量细胞凋亡。通过 ELISA 测量单核细胞趋化因子 CCL2 的表达。通过实时 PCR 测量 CCL2 的 mRNA 表达。
霉酚酸通过上调 p21、p27 和 p53，降低细胞周期蛋白 D2 和 E，剂量依赖性地抑制 49F 细胞的稳态增殖。MPA 处理还通过激活 caspase 3 级联反应触发 49F 细胞凋亡。此外，MPA 通过下调 p38 MAPK 而非 ERK1/2 或 JNK 抑制肿瘤坏死因子-α诱导的 CCL2 表达。
MPA 的抗有丝分裂和抗炎作用分别通过上调细胞周期抑制剂和促凋亡信号以及抑制 p38 MAPK 通路来介导。MPA 的这种双重作用可能为治疗纤维化肾脏疾病的动物或临床试验提供依据。
