Li Yiqun, Zhang Jiarong, Zhu Yanli
Department of Nephrology, Dingxi Municipal People's Hospital, No. 22, Anding Road, Dingxi, 743000, China.
Clin Exp Nephrol. 2024 Dec;28(12):1221-1231. doi: 10.1007/s10157-024-02536-0. Epub 2024 Jul 30.
Methyltransferase 14 (METTL14) mediated N6-methyladenine (m6A) RNA methylation and progestin and AdipoQ receptor family member 3 (PAQR3) are reported to be involved in diabetic nephropathy (DN) progression. Here, we explored whether the effects of PAQR3 on DN was associated with METTL14-induced m6A and their relationship with macrophage-related exosomes in DN progression.
Human glomerular endothelial cells (GECs) were incubated in high glucose (HG) condition to mimic DN condition in vitro. Exosomes were isolated from M1 macrophages and co-cultured with GECs. qRT-PCR and western blotting detected the levels of genes and proteins. Cell functions were determined using cell counting kit-8 assay and flow cytometry. ELISA analysis detected inflammatory factors, and oxidative stress was evaluated by measuring reactive oxygen species and malondialdehyde. The m6A modification profile was determined by methylated RNA immunoprecipitation assay and the interaction was verified by dual-luciferase reporter assay.
HG elevated PAQR3 expression levels in GECs. PAQR3 silencing reversed HG-induced viability arrest, apoptosis, inflammatory response, and oxidative stress. M1 macrophage co-culture could suppress HG-induced GEC injury. PAQR3 was packaged into M1 macrophage-derived exosomes, and M1 macrophages regulated HG-induced GEC injury by secreting PAQR3 into cells via exosomes. Mechanistically, METTL14 induced PAQR3 m6A modification. METTL14 was enriched in M1 macrophage-derived exosomes. METTL14 knockdown in M1 macrophage-derived exosomes protected GEC from HG-induced viability arrest, apoptosis, inflammation and oxidative stress by regulating PAQR3.
Exosomal METTL14 derived from M1 macrophages promoted HG-induced apoptosis, inflammation and oxidative stress in GECs by mediating PAQR3 m6A modification.
据报道，甲基转移酶14（METTL14）介导的N6-甲基腺嘌呤（m6A）RNA甲基化以及孕激素和脂联素受体家族成员3（PAQR3）参与糖尿病肾病（DN）的进展。在此，我们探讨了PAQR3对DN的影响是否与METTL14诱导的m6A相关，以及它们在DN进展中与巨噬细胞相关外泌体的关系。
将人肾小球内皮细胞（GECs）置于高糖（HG）条件下培养，以在体外模拟DN状态。从M1巨噬细胞中分离出外泌体，并与GECs共培养。采用qRT-PCR和蛋白质印迹法检测基因和蛋白质水平。使用细胞计数试剂盒-8法和流式细胞术测定细胞功能。酶联免疫吸附测定（ELISA）分析检测炎症因子，通过测量活性氧和丙二醛评估氧化应激。通过甲基化RNA免疫沉淀测定法确定m6A修饰谱，并通过双荧光素酶报告基因测定法验证相互作用。
HG升高了GECs中PAQR3的表达水平。PAQR3沉默逆转了HG诱导的活力停滞、凋亡、炎症反应和氧化应激。M1巨噬细胞共培养可抑制HG诱导的GEC损伤。PAQR3被包装到M1巨噬细胞衍生的外泌体中，并且M1巨噬细胞通过外泌体将PAQR3分泌到细胞中来调节HG诱导的GEC损伤。机制上，METTL14诱导PAQR3的m6A修饰。METTL14在M1巨噬细胞衍生的外泌体中富集。敲低M1巨噬细胞衍生外泌体中的METTL14可通过调节PAQR3保护GEC免受HG诱导的活力停滞、凋亡、炎症和氧化应激。
源自M1巨噬细胞的外泌体METTL14通过介导PAQR3的m6A修饰促进HG诱导的GECs凋亡、炎症和氧化应激。
