Herbal Research Laboratory, Food, Drug & Chemical Toxicology Group, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Vishvigyan Bhawan 31, Mahatma Gandhi Marg, Lucknow, 226001, Uttar Pradesh, India.
Academy of Scientific and Innovative Research, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Lucknow, 226001, Uttar Pradesh, India.
Mol Cell Biochem. 2021 Oct;476(10):3681-3699. doi: 10.1007/s11010-021-04177-3. Epub 2021 May 31.
Impaired PI3K/Akt signaling (insulin resistance) and poor glycemic control (hyperglycemia) are the major risk factors involved in the progression of diabetic nephropathy (DN). This study was designed to identify factors influencing cell survival during DN. We found that high glucose exposure in renal proximal tubular cells (NRK52E) upregulated PHLPP1, an Akt phosphatase (Ser473), causing suppression in Akt and IGF1β phosphorylation leading to inhibition in insulin signaling pathway. Results demonstrate that sustained activation of PHLPP1 promoted nuclear retention of FoxO1 by preventing its ubiquitination via Mdm2, an Akt/ Nrf2-dependent E3 ligase. Thus, enhanced FoxO1 nuclear stability caused aberration in renal gluconeogenesis and activated apoptotic cascade. Conversely, gene silencing of PHLPP1-enhanced Nrf2 expression and attenuated FoxO1 regulated apoptosis compared to hyperglycemic cells. Mechanistic aspects of PHLPP1-Nrf2/FoxO1 signaling were further validated in STZ-nicotinamide-induced type 2 diabetic Wistar rats. Importantly, we observed via immunoblotting and dual immunocytochemical studies that treatment of Morin (2',3,4',5,7-Pentahydroxyflavone) during diabetes significantly augmented FoxO1 nuclear exclusion, resulting in its ubiquitination via Akt-Nrf2/Mdm2 pathway. Furthermore, lowering of PHLPP1 expression by Morin also prevented FoxO1/Mst1-mediated apoptotic signaling in vitro and in vivo. Morin treatment under the experimental conditions, effectively decreased blood glucose levels, ameliorated insulin resistance, alleviated oxidative stress and attenuated renal apoptosis in diabetic rats comparable to metformin thereby exhibiting tremendous potential against renal complications of diabetes. These novel results further acclaim that inhibition of PHLPP1/FoxO1-Mdm2 axis is critical in the pathogenesis of diabetic nephropathy.
PI3K/Akt 信号转导受损（胰岛素抵抗）和血糖控制不佳（高血糖）是糖尿病肾病（DN）进展的主要危险因素。本研究旨在确定影响 DN 期间细胞存活的因素。我们发现，高糖暴露可使肾近端小管细胞（NRK52E）中 PHLPP1（一种 Akt 磷酸酶（Ser473））上调，从而抑制 Akt 和 IGF1β 的磷酸化，导致胰岛素信号通路受到抑制。结果表明，PHLPP1 的持续激活通过阻止 Akt/Nrf2 依赖性 E3 连接酶 Mdm2 对其泛素化，促进 FoxO1 的核保留。因此，增强的 FoxO1 核稳定性导致肾脏糖异生的异常和凋亡级联的激活。相反，与高糖细胞相比，PHLPP1 基因沉默增强了 Nrf2 的表达并减弱了 FoxO1 调节的凋亡。PHLPP1-Nrf2/FoxO1 信号的机制方面在 STZ-烟酰胺诱导的 2 型糖尿病 Wistar 大鼠中得到了进一步验证。重要的是，我们通过免疫印迹和双重免疫细胞化学研究观察到，在糖尿病期间用 Morin（2'，3'，4'，5'，7-五羟基黄酮）治疗可显著增加 FoxO1 的核排斥，从而通过 Akt-Nrf2/Mdm2 途径对其进行泛素化。此外，Morin 降低 PHLPP1 的表达也可防止 FoxO1/Mst1 介导的体外和体内凋亡信号。在实验条件下用 Morin 治疗可有效降低血糖水平，改善胰岛素抵抗，减轻氧化应激并减轻糖尿病大鼠的肾细胞凋亡，与二甲双胍相当，从而表现出对糖尿病肾脏并发症的巨大潜力。这些新的结果进一步证明，抑制 PHLPP1/FoxO1-Mdm2 轴在糖尿病肾病的发病机制中至关重要。
