载脂蛋白A-I模拟肽L-4F减轻过氧化氢诱导的小鼠骨髓内皮祖细胞损伤

 目的: 研究载脂蛋白A-I模拟肽L-4F是否减轻过氧化氢(hydrogen peroxide,H₂O₂)诱导的小鼠骨髓来源内皮祖细胞(endothelial progenitor cells,EPCs)氧化应激损伤及其机制。方法: 通过密度梯度离心法分离小鼠骨髓单个核细胞,条件培养基EGM-2MV诱导分化培养EPCs。对培养EPCs先以PI3K/AKT抑制剂LY294002(30 μmol/L)干预2 h后加入L-4F(75 mg/L)4 h,再加入100 μmol/L H₂O₂ 24 h。MTT 法检测细胞活力,试剂盒检测各实验组培养基超氧化物歧化酶(superoxide dismutase,SOD)和丙二醛(malondialdehyde,MDA)含量,以评价细胞氧化与抗氧化平衡及脂质过氧化程度;流式细胞术Annexin V/PI双染法检测细胞凋亡;免疫印迹法检测p-AKT蛋白水平。结果: L-4F预处理显著抑制了H₂O₂诱导的细胞活力降低、SOD活性下降、MDA含量增加及细胞凋亡。H₂O₂下调p-AKT的蛋白水平,L-4F呈浓度依赖性地上调p-AKT的蛋白水平。LY294002抑制了L-4F减轻EPCs损伤的作用。结论: 载脂蛋白A-I 模拟肽L-4F部分通过PI3K/AKT通路减轻H₂O₂诱导的EPCs氧化应激损伤。

Apolipoprotein A-I mimetic peptide L-4F reduces hydrogen peroxide-induced injury of mouse bone marrow-derived EPCs

AIM: To investigate the protective effect and underlying mechanism of apolipoprotein A-I mimetic peptide L-4F on mouse bone marrow-derived endothelial progenitor cells (EPCs) injured by hydrogen peroxide (H₂O₂). METHODS: The bone marrow-derived mononuclear cells of C57 mice were isolated by density gradient centrifugation and cultured in EGM-2MV medium. EPCs were pretreated with a PI3K inhibitor LY294002 (30 μmol/L) for 2 h, and then incubated with L-4F (75 mg/L) for 4 h, followed by the treatment with 100 μmol/L H₂O₂ for 24 h. MTT assay and flow cytometry with Annexin V/PI staining were used to detect cell viability and apoptosis. The extracellular superoxide dismutase (SOD) andmalondialdehyde (MDA) levels were measured to characterize the balance between oxidation and antioxidation, and lipid peroxidation. The protein level of phosphorylated AKT (p-AKT) was analyzed by Western blot. RESULTS: L-4F restored H₂O₂-induced decrease in EPCs cell viability and apoptosis, and inhibited H₂O₂-induced decrease in SOD activity and increase in MDA level in culture medium. In addition, H₂O₂ inactivated p-AKT, which was significantly restored by L-4F. However, LY294002 inhibited the restoration of L-4F on EPCs impaired by H₂O₂. CONCLUSION: Apolipoprotein A-I mimetic peptide L-4F significantly reduced H₂O₂-induced mouse bone marrow-derived EPC injury partially through stimulating the PI3K/AKT signaling pathway.