IIRC-5, Clinical Biochemistry & Natural Product Research Lab, Department of Biosciences, Integral University, Lucknow, 226026, U.P., India.
Stem Cell Research Center, Department of Hematology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, U.P., India.
Life Sci. 2019 Oct 15;235:116823. doi: 10.1016/j.lfs.2019.116823. Epub 2019 Aug 30.
Advanced glycation end products (AGEs) trigger intracellular reactive oxygen species (ROS) generation, activation of receptor for AGEs (RAGE) expression/functionality and RAGE-associated signalling pathways which influence the diabetic-cum-atherosclerotic complications, whereas, the atherosclerosis progression is greatly influenced by hepatic β-Hydroxy-β-methyl-glutaryl-Co-A reductase (HMG-R) activity. The present report was premeditated to uncover the regulatory role of HMG-R inhibitors and ezetimibe (EZ) in attenuating the LDL-AGEs-induced pathogenicity via targeting cellular-ROS and RAGE-associated signalling in HEK-293 cells.
The MTT assay was used to assess either the cytotoxic or cytoprotective impact of each HMG-R inhibitors, EZ, and LDL-AGEs, whereas, quantification of ROS was performed by DCFDA method. The qRT-PCR was used to detect the mRNA level of RAGE, neuropilin-1 (NRP-1) and other RAGE-associated genes like MMP-2, NF-κB, and TGFβ-1.
The HMG-R inhibitors do not exert any cytotoxicity in HEK-293 cells, whereas, and LDL-AGEs negatively affected the cell viability of HEK-293 cells. However, viability of LDL-AGEs-treated HEK-293was markedly retained after simultaneous treatment with our test inhibitors. Further, DCFDA staining showed that LDL-AGEs-induced ROS was also suppressed upon treatment with our test inhibitors in HEK-293 cells. qRT-PCR analysis reflected that these inhibitors suppress the RAGE, NF-κB, TGFβ-1, and MMP-2 expression, whereas, the NRP-1 was up-regulated by these compounds in LDL-AGEs-exposed HEK-293 cells.
The above pharmacological effects signify that HMG-R inhibitors and EZ (alone or in combination) may implied in the treatment of AGEs-induced oxidative stress and tissue damage in diabetic complications via targeting intracellular-ROS, NRP-1 functionality and RAGE-associated genes i.e. NF-κB, TGFβ-1, and MMP-2.
晚期糖基化终产物（AGEs）触发细胞内活性氧（ROS）的产生，激活 AGEs 受体（RAGE）的表达/功能以及 RAGE 相关信号通路，从而影响糖尿病合并动脉粥样硬化的并发症，而动脉粥样硬化的进展则受肝β-羟-β-甲基戊二酸单酰辅酶 A 还原酶（HMG-R）活性的极大影响。本报告旨在揭示 HMG-R 抑制剂和依折麦布（EZ）通过靶向细胞内 ROS 和 RAGE 相关信号通路，在减轻 LDL-AGEs 诱导的致病性方面的调节作用。
MTT 测定法用于评估每种 HMG-R 抑制剂、EZ 和 LDL-AGEs 的细胞毒性或细胞保护作用，而 DCFDA 法用于定量 ROS。qRT-PCR 用于检测 RAGE、神经纤毛蛋白-1（NRP-1）和其他 RAGE 相关基因（如 MMP-2、NF-κB 和 TGFβ-1）的 mRNA 水平。
HMG-R 抑制剂在 HEK-293 细胞中没有表现出任何细胞毒性，而 LDL-AGEs 则对 HEK-293 细胞的细胞活力产生负面影响。然而，在用我们的测试抑制剂同时处理后，LDL-AGEs 处理的 HEK-293 细胞的活力明显得到保留。此外，DCFDA 染色显示，在用我们的测试抑制剂处理后，LDL-AGEs 诱导的 ROS 也得到了抑制。qRT-PCR 分析反映，这些抑制剂抑制 RAGE、NF-κB、TGFβ-1 和 MMP-2 的表达，而在 LDL-AGEs 暴露的 HEK-293 细胞中，NRP-1 则被这些化合物上调。
上述药理作用表明，HMG-R 抑制剂和 EZ（单独或联合）可能通过靶向细胞内 ROS、NRP-1 功能和 RAGE 相关基因（即 NF-κB、TGFβ-1 和 MMP-2），用于治疗 AGEs 诱导的糖尿病并发症中的氧化应激和组织损伤。
