High glucose induces renal tubular epithelial injury via Sirt1/NF-kappaB/microR-29/Keap1 signal pathway

Journal of Translational Medicine, Nov 2015

Objective Diabetic nephropathy (DN) is a serious complication that commonly confronted by diabetic patients. A common theory for the pathogenesis of this renal dysfunction in diabetes is cell injury, inflammation as well as oxidative stress. In this content, the detailed molecular mechanism underlying high glucose induced renal tubular epithelial injury was elaborated. Methods An in vivo rat model of diabetes by injecting streptozotocin (STZ) and an in vitro high glucose incubated renal tubular epithelial cell (HK-2) model were used. Expression levels of Keap1, nuclear Nrf2 and p65 were determined by western blotting. Level of microR-29 (miR-29) was assessed using quantitative RT-PCR. Combination of p65 and miR-29 promotor was assessed using chromatin immunoprecipitation. Keap1 3′-UTR activity was detected using luciferase reporter gene assay. Cell viability was determined using MTT assay. Results In diabetic rat, miR-29 was downregulated and its expression is negatively correlated with both of serum creatinine and creatinine clearance. In high glucose incubated HK-2 cell, deacetylases activity of Sirt1 was attenuated that leads to decreased activity of nuclear factor kappa B (NF-κB). NF-κB was demonstrated to regulate miR-29 expression by directly binding to its promotor. The data of luciferase assay showed that miR-29 directly targets to Keap1 mRNA. While high glucose induced down regulation of miR-29 contributed to enhancement of Keap1 expression that finally reduced Nrf2 content by ubiquitinating Nrf2. Additionally, overexpression of miR-29 effectively relieved high glucose-reduced cell viability. Conclusion High glucose induces renal tubular epithelial injury via Sirt1/NF-κB/microR-29/Keap1 signal pathway.

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High glucose induces renal tubular epithelial injury via Sirt1/NF-kappaB/microR-29/Keap1 signal pathway

Zhou et al. J Transl Med High glucose induces renal tubular epithelial injury via Sirt1/NF-kappaB/microR-29/ Keap1 signal pathway Ling Zhou 3 Dey‑u Xu 3 Wen‑gang Sha 3 Lei Shen 3 Guoy‑uan Lu 3 Xia Yin 2 Ming‑jun Wang 0 1 0 Department of Rheumatology, The First Affiliated Hospital of Soochow University , 188 shizi Rd., Suzhou 215006 , People's Republic of China 1 Department of Rheu‐ matology, The First Affiliated Hospital of Soochow University , 188 shizi Rd., Suzhou 215006 , People's Republic of China 2 Department of Endocrinology, The First Affiliated Hospital of Soochow University , Suzhou 215006 , China 3 Department of Nephrology, The First Affiliated Hospital of Soochow Univer‐ sity , Suzhou 215006 , China Objective: Diabetic nephropathy (DN) is a serious complication that commonly confronted by diabetic patients. A common theory for the pathogenesis of this renal dysfunction in diabetes is cell injury, inflammation as well as oxidative stress. In this content, the detailed molecular mechanism underlying high glucose induced renal tubular epithelial injury was elaborated. Methods: An in vivo rat model of diabetes by injecting streptozotocin (STZ) and an in vitro high glucose incubated renal tubular epithelial cell (HK‑ 2) model were used. Expression levels of Keap1, nuclear Nrf2 and p65 were determined by western blotting. Level of microR‑ 29 (miR‑ 29) was assessed using quantitative RT‑ PCR. Combination of p65 and miR‑ 29 promotor was assessed using chromatin immunoprecipitation. Keap1 3′‑ UTR activity was detected using luciferase reporter gene assay. Cell viability was determined using MTT assay. Results: In diabetic rat, miR‑ 29 was downregulated and its expression is negatively correlated with both of serum creatinine and creatinine clearance. In high glucose incubated HK‑ 2 cell, deacetylases activity of Sirt1 was attenuated that leads to decreased activity of nuclear factor kappa B (NF‑ κB). NF‑ κB was demonstrated to regulate miR‑ 29 expression by directly binding to its promotor. The data of luciferase assay showed that miR‑ 29 directly targets to Keap1 mRNA. While high glucose induced down regulation of miR‑ 29 contributed to enhancement of Keap1 expression that finally reduced Nrf2 content by ubiquitinating Nrf2. Additionally, overexpression of miR‑ 29 effectively relieved high glucose‑ reduced cell viability. Conclusion: High glucose induces renal tubular epithelial injury via Sirt1/NF‑ κB/microR‑ 29/Keap1 signal pathway. Acetyl‑ p65; HK‑ 2; miR‑ 29; Nrf2; Serum creatinine - Background Diabetes mellitus is a common metabolic disorder which is associated with chronic complications such as angiopathy, retinopathy, and peripheral neuropathy. It was recognized that diabetes also can lead to nephropathy. Since then, studies in experimental models and in patients observed structural abnormalities in vascular and glomerular [1]. However, tubular cells are proved to be primary targets of hyperglycemia which is the major cause for renal injury and previous evidence indicated that chronic exposure to elevated blood glucose levels contributes to the organic pathologic changes in clinical diabetes nephropathy (DN) [2, 3]. Thus searching for therapy targets might be benefit for concurrency of DN. In previous study, high glucose induced inflammatory response as well as activated inflammation related signal pathway. Sirt1, a class III histone deacetylase, is recognized as an important regulator in many high glucoserelated inflammatory diseases [4]. For example, Sirt1 activator blunting pro-inflammatory pathways in mice fed a high fat, high calorie diet [5]. Especially, Sirt1 and its modulation of the acetylation status of the p65 subunit of nuclear factor kappa B (NF-κB) play an important role in regulating the inflammatory and apoptotic responses © 2015 Zhou et al. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. in cells or tissues [6]. As an essential nuclear transcription factor, NF-κB is commonly activated by wide variety of cell-tress stimuli including obesity, oxidative stress, as well as stimulated by hyperglycemia. Accumulated evidence pointed out that NF-κB was upregulated in diabetic rat kidneys [7] and its inhibition contributed to significant amelioration of DN [8]. Although observation has showed that master regulator of inflammation of NF-κB is essential for pathological process of DN, the detailed downstream intracellular signal transduction is not yet clear. MicroRN (...truncated)


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Ling Zhou, De-yu Xu, Wen-gang Sha, Lei Shen, Guo-yuan Lu, Xia Yin, Ming-jun Wang. High glucose induces renal tubular epithelial injury via Sirt1/NF-kappaB/microR-29/Keap1 signal pathway, Journal of Translational Medicine, 2015, pp. 352, 13, DOI: 10.1186/s12967-015-0710-y