Department of Nephrology, Huadong Hospital Affiliated to Fudan University, Shanghai, People's Republic of China.
Inflamm Res. 2015 Aug;64(8):603-14. doi: 10.1007/s00011-015-0838-5. Epub 2015 Jun 14.
In our previous study, we observed the crosstalk between peroxisome proliferator-activated receptor-γ (PPAR-γ) and angiotensin II in activated renal tubular cells. The present study is aimed to further explore the crosstalk between PPAR-γ and mineralocorticoid receptor (MR) in tumor necrosis factor (TNF)-α activated renal tubular cells.
Human proximal renal tubular epithelial cells HK-2 were cultured with the pre-treatment of PPAR-γ agonist, pioglitazone (5 μM), MR antagonist, eplerenone (5 μM), or their combined treatment, followed by activation with TNF-α (20 ng/ml). In the parallel experiment, PPAR-γ inhibitor GW9662 (25 µM) was used to study the independence of PPAR-γ. Gene expression and protein synthesis of intercellular adhesion molecule-1 (ICAM-1), interleukin-6 (IL-6), MR and PPAR-γ were measured by RT-PCR, ELISA and Western blot, respectively; nuclear factor κB (NF-κB) nuclear translocation activity in the nucleus was examined by EMSA assay.
TNF-α effectively activated HK-2 cells by up-regulating gene expression and protein synthesis of ICAM-1, IL-6 and MR and down-regulating PPAR-γ in a dose-dependent manner. TNF-α also significantly induced NF-κB nuclear translocation in HK-2 cells. Dual treatment of pioglitazone and eplerenone demonstrated synergistic effect on reducing ICAM-1 and IL-6 expression and alleviating NF-κB activation when compared with their monotherapies in TNF-α activated renal tubular cells. PPAR-γ antagonist, GW9662, significantly attenuated protective effect on ICAM-1, IL-6 and PPAR-γ expression by pioglitazone, eplerenone and their combined treatment.
Our data suggest that pioglitazone, in a PPAR-γ-dependent manner, trans-represses MR signaling by suppressing NF-κB activation. MR antagonist also restored PPAR-γ expression. Dual treatment of pioglitazone and eplerenone present better efficacy in attenuating excessive inflammatory response in activated renal tubular cells under stimulation of TNF-α than single treatment.
在我们之前的研究中，我们观察到过氧化物酶体增殖物激活受体-γ（PPAR-γ）与血管紧张素Ⅱ在激活的肾小管细胞中的相互作用。本研究旨在进一步探讨肿瘤坏死因子（TNF）-α激活的肾小管细胞中 PPAR-γ与盐皮质激素受体（MR）之间的相互作用。
用 PPAR-γ激动剂吡格列酮（5 μM）、MR 拮抗剂依普利酮（5 μM）或两者联合预处理人近端肾小管上皮细胞株 HK-2，然后用 TNF-α（20 ng/ml）激活。在平行实验中，使用 PPAR-γ 抑制剂 GW9662（25 μM）来研究 PPAR-γ 的独立性。通过 RT-PCR、ELISA 和 Western blot 分别测量细胞间黏附分子-1（ICAM-1）、白细胞介素-6（IL-6）、MR 和 PPAR-γ 的基因表达和蛋白合成；通过 EMSA 测定核因子κB（NF-κB）在核内的核转位活性。
TNF-α 以剂量依赖性方式有效激活 HK-2 细胞，上调 ICAM-1、IL-6 和 MR 的基因表达和蛋白合成，下调 PPAR-γ。TNF-α 还显著诱导 HK-2 细胞中 NF-κB 的核转位。与 TNF-α 激活的肾小管细胞中单药治疗相比，吡格列酮和依普利酮联合治疗对 ICAM-1 和 IL-6 表达的降低和 NF-κB 激活的缓解具有协同作用。PPAR-γ 拮抗剂 GW9662 显著减弱了吡格列酮、依普利酮及其联合治疗对 ICAM-1、IL-6 和 PPAR-γ 表达的保护作用。
我们的数据表明，吡格列酮以 PPAR-γ 依赖性方式通过抑制 NF-κB 激活来反式抑制 MR 信号。MR 拮抗剂也恢复了 PPAR-γ 的表达。与 TNF-α 刺激下的单一治疗相比，吡格列酮和依普利酮的联合治疗在减轻激活的肾小管细胞中过度炎症反应方面具有更好的疗效。
