Liang Guibin, He Zhihui, Peng Huaihuai, Zeng Menghao, Zhang Xuefeng
Department of Critical Care Medicine, the Third Xiangya Hospital, Central South University, Changsha, China.
Department of Critical Care Medicine, the Second Xiangya Hospital, Central South University, Changsha, China.
Tob Induc Dis. 2023 Oct 3;21:122. doi: 10.18332/tid/170581. eCollection 2023.
Endothelial progenitor cells (EPCs) are the main source of endothelial cells. The senescence of EPCs is involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). Cigarette smoke extract (CSE) can directly induce the dysfunction and increased expression of senescence-related markers in EPCs cultured in vitro. Histone acetyltransferase p300 is a transcriptional activator, and its changes can lead to cell senescence. The present study investigated whether CSE can induce the senescence of EPCs by upregulating p300.
EPCs were isolated from bone marrow of C57BL/6J mice by density gradient centrifugation. The p300 inhibitor C646 and agonist CTPB were used to interfere with EPCs, cell cycle and apoptosis were detected by flow cytometry, the proportion of senile cells was counted by β-galactosidase staining, the protein expression of p300, H4K12, Cyclin D1, TERT and Ki67 were detected by western blot.
Compared with the control group, the cell cycle of CSE group and CTPB group were blocked, the apoptosis rate and early apoptosis rate were increased, the proportion of senile cells counted by β-galactosidase staining was increased, the expression of p300 and H4K12 protein were increased, the expression of Cyclin D1, TERT and Ki67 protein were decreased. C646 could partly alleviate the damages caused by CSE.
CSE may promote the apoptosis and senescence of EPCs by upregulating the expression of p300 and H4K12 protein, thus preventing the transition of EPCs from G1 phase to S phase, affecting telomerase synthesis, and reducing EPCs proliferation.
内皮祖细胞（EPCs）是内皮细胞的主要来源。EPCs的衰老参与慢性阻塞性肺疾病（COPD）的发病机制。香烟烟雾提取物（CSE）可直接诱导体外培养的EPCs功能障碍及衰老相关标志物表达增加。组蛋白乙酰转移酶p300是一种转录激活因子，其变化可导致细胞衰老。本研究探讨CSE是否可通过上调p300诱导EPCs衰老。
通过密度梯度离心从C57BL/6J小鼠骨髓中分离EPCs。使用p300抑制剂C646和激动剂CTPB干预EPCs，采用流式细胞术检测细胞周期和凋亡，通过β-半乳糖苷酶染色计数衰老细胞比例，采用蛋白质免疫印迹法检测p300、H4K12、细胞周期蛋白D1、端粒酶逆转录酶（TERT）和Ki67的蛋白表达。
与对照组相比，CSE组和CTPB组细胞周期阻滞，凋亡率和早期凋亡率升高，β-半乳糖苷酶染色计数的衰老细胞比例增加，p300和H4K12蛋白表达增加，细胞周期蛋白D1、TERT和Ki67蛋白表达降低。C646可部分减轻CSE所致损伤。
CSE可能通过上调p300和H4K12蛋白表达促进EPCs凋亡和衰老，从而阻止EPCs从G1期向S期转变，影响端粒酶合成，降低EPCs增殖。
