Ding Weixing, Peng Huijuan, Tian Jianyou, Wang Siyan
Pain Department, Qujing Second People's Hospital, No. 289 Qilin Xi Road, Qilin District, Qujing, Yunnan, 655009, China.
Anesthesiology Department, Qujing Second People's Hospital, No. 289 Qilin Xi Road, Qilin District, Qujing, Yunnan, 655009, China.
Heliyon. 2023 Sep 21;9(9):e20285. doi: 10.1016/j.heliyon.2023.e20285. eCollection 2023 Sep.
This study was performed to explore the role of Re in liver IRI progression. Hypoxia and reperfusion (H/R) treated human embryo liver cell line (L-02) was used to establish a liver IRI model.
Cell behaviors were detected using CCK-8, flow cytometry and TUNEL staining assays. The m6A content was detected using m6A dot blot assay. RT-qPCR and western blot assays were used to assessed the relative mRNA and protein levels. MeRIP assay was conducted to determine the m6A levels of P53. The relationship between METTL3 and P53 was demonstrated using RIP and dual-luciferase reporter assays.
The results showed that Re treatment significantly decreased the cell apoptosis and promoted the cell viability in the H/R treated L-02 cells. Besides, H/R treatment increased the METTL3 and m6A levels in the L-02 cells, and Re treatment decreased them. Additionally, METTL3 overexpression reversed the role of Re in the H/R treated L-02 cells. Mechanistically, METTL3 overexpression enhanced the m6A levels and mRNA stability and expressions of P53. The combination of METTL3 and P53 was further confirmed.
In conclusion, this study demonstrated that Re treatment relieved the H/R induced injury in the L-02 cells through decreasing the METTL3 levels. METTL3 enhanced the mRNA stability and expressions of P53 through m6A modification. Re-METTL3-P53 axis might a new direction for the treatment of liver IRI in the future.
本研究旨在探讨白藜芦醇（Re）在肝脏缺血再灌注损伤（IRI）进展中的作用。采用缺氧复氧（H/R）处理的人胚胎肝细胞系（L-02）建立肝脏IRI模型。
使用CCK-8、流式细胞术和TUNEL染色法检测细胞行为。采用m6A斑点印迹法检测m6A含量。运用RT-qPCR和蛋白质免疫印迹法评估相关mRNA和蛋白质水平。进行甲基化RNA免疫沉淀（MeRIP）分析以确定P53的m6A水平。通过RNA免疫沉淀（RIP）和双荧光素酶报告基因检测法证明甲基转移酶样蛋白3（METTL3）与P53之间的关系。
结果显示，白藜芦醇处理显著降低了H/R处理的L-02细胞的凋亡率并提高了细胞活力。此外，H/R处理增加了L-02细胞中METTL3和m6A的水平，而白藜芦醇处理则降低了它们。此外，METTL3过表达逆转了白藜芦醇在H/R处理的L-02细胞中的作用。机制上，METTL3过表达增强了m6A水平以及P53的mRNA稳定性和表达。进一步证实了METTL3与P53的结合。
总之，本研究表明白藜芦醇处理通过降低METTL3水平减轻了H/R诱导的L-02细胞损伤。METTL3通过m6A修饰增强了P53的mRNA稳定性和表达。Re-METTL3-P53轴可能是未来治疗肝脏IRI的新方向。
