Thakar Sumukh, Bhattacharyya Ritobrata, Katakia Yash Tushar, Jayakumar Hariharan, Chakraborty Srinjoy, Ramakrishnan Shyam Kumar, Thakkar Niyati Pandya, Dey Smita, Chowdhury Shibasish, Majumder Syamantak
Department of Biological Sciences, Birla Institute of Technology and Science Pilani, Pilani Campus, Pilani, Rajasthan, India.
Department of Biological Sciences, Birla Institute of Technology and Science Pilani, Pilani Campus, Pilani, Rajasthan, India.
Int J Biol Macromol. 2025 Jul;318(Pt 1):144963. doi: 10.1016/j.ijbiomac.2025.144963. Epub 2025 Jun 4.
Endothelial senescence has been linked with several cardiovascular diseases. Dysregulation of TRF2 and TERF2IP, proteins of the shelterin complex, causes cellular senescence. However, it is still unknown whether exposure to hyperglycemia alters levels of proteins of the shelterin complex, further dictating the senescent phenotype of endothelial cells (EC). In this study, we observed elevated protein levels of p21 and p53 in EC upon exposure to Intermittent High Glucose(IHG). Furthermore, exposure to hyperglycemia increased levels of TERF2IP and TRF2 proteins. Interestingly, we observed a robust induction in the p65 protein level as well as it's phosphorylation upon intermittent hyperglycemia challenge. ChIP-qPCR analysis revealed enhanced H3K4me3 enrichment in the gene promoters of p65, TERF2IP, and TRF2. Inhibition of H3K4me3 deposition either by pharmacological inhibitor or siRNA-mediated knockdown of MLL2 reduced the expression of p65, TERF2IP and TRF2 and restored the levels of senescence markers, namely p53 and p21. Interestingly, pharmacological inhibition of NF-κB signaling reduced the expression of TERF2IP and TRF2 levels, reversing the levels of p53 and p21. Co-immunoprecipitation and co-localization analysis revealed an association between nuclear p65 and MLL2 proteins in EC exposed to hyperglycemia. Knockdown of either TERF2IP or TRF2 impaired intermittent hyperglycemia-induced p53 and p21 expression and associated endothelial senescence.
内皮细胞衰老与多种心血管疾病有关。端粒保护蛋白复合体中的TRF2和TERF2IP蛋白失调会导致细胞衰老。然而，暴露于高血糖环境是否会改变端粒保护蛋白复合体中蛋白质的水平，进而决定内皮细胞（EC）的衰老表型，目前仍不清楚。在本研究中，我们观察到内皮细胞在暴露于间歇性高糖（IHG）后，p21和p53蛋白水平升高。此外，暴露于高血糖环境会增加TERF2IP和TRF2蛋白的水平。有趣的是，我们观察到在间歇性高血糖刺激下，p65蛋白水平及其磷酸化水平均有显著诱导。染色质免疫沉淀-定量聚合酶链反应（ChIP-qPCR）分析显示，p65、TERF2IP和TRF2基因启动子中的H3K4me3富集增强。通过药物抑制剂或小干扰RNA（siRNA）介导的MLL2敲低抑制H3K4me3沉积，可降低p65、TERF2IP和TRF2的表达，并恢复衰老标志物p53和p21的水平。有趣的是，对核因子κB（NF-κB）信号通路的药物抑制可降低TERF2IP和TRF2的水平，逆转p53和p21的水平。免疫共沉淀和共定位分析显示，在暴露于高血糖环境的内皮细胞中，核p65与MLL2蛋白之间存在关联。敲低TERF2IP或TRF2会损害间歇性高血糖诱导的p53和p21表达以及相关的内皮细胞衰老。
