目的 分析橄榄苦苷对HK-2细胞缺糖缺氧损伤的保护作用并探讨其可能的作用机制.方法 实验于2017年1-4月在武汉大学人民医院中心实验室进行,体外培养HK-2细胞,应用无糖无血清培养基和三气培养箱建立缺糖缺氧模型,将细胞分为对照组、单纯缺氧24 h复氧3 h组(IR组)和缺氧24 h复氧3 h加不同浓度(10 μmol/L、50 μmol/L、100 μmol/L、200 μmol/L)的橄榄苦苷干预组.用CCK-8检测各组细胞的A450值,与对照组比较计算细胞的存活率,Hoechst染色检测各组细胞的凋亡情况,Western Blot法测定B淋巴细胞瘤-2(B-cell lymphoma-2,Bcl-2)、Bcl-2相关x蛋白(Bcl-2 associated X protein,Bax)的蛋白表达量.结果 橄榄苦苷随浓度升高能够提高HK-2细胞在缺糖缺氧环境下的细胞存活率,100 μmol/L组存活率最高,但至200 μmol/L组存活率呈降低趋势;Hoechst染色显示橄榄苦苷可减少缺糖缺氧损伤引起的细胞凋亡,100 μmol/L组凋亡率最低;Western blot 结果显示橄榄苦苷能够增加细胞Bcl-2蛋白的表达,降低Bax蛋白的表达.结论 橄榄苦苷对HK-2细胞的缺糖缺氧损伤具有一定的保护作用,其作用机制可能与其抑制细胞凋亡有关.%Objective To investigate the protective effect of oleuropein and its possible mechanism on the glucose deprivation hypoxia injury of HK-2 cells.Methods From January 2017 to April 2017, the experiment was conducted at the Central Laboratory of Renmin Hospital of Wuhan University.HK-2 cells were cultured in vitro, using sugar free serum free culture medium and three gas incubators to establish the model of oxygen glucose deprivation, the cells were divided into control group, hypoxia 24 h reoxygenation 3 h group (IR group) and 24 h of hypoxia reoxygenation for 3 h with different concentrations (10 μmol/L, 50 μmol/L, 100 μmol/L, 200 μmol/L) of oleuropein in intervention group.With the detected CCK-8 A450 value, compared with the normal group the survival rate calculated cells, the apoptosis cells were detected with Hoechst staining, Western Blot assay of B lymphocyte tumor 2 (B cell lymphoma 2, Bcl-2), Bcl-2 X protein (Bcl-2 associated X protein, Bax) expression of protein.Results With the increase of concentration of oleuropein can improve the survival rate of HK2 cells in OGD environment, 100 μmol/L group showed the highest survival rate, but the survival rate in 200 μmol/L group decreased;Hoechst staining showed that oleuropein could reduce apoptosis induced by OGD injury, lower apoptosis rate, the apoptosis rate of 100 μmol/L group was the lowest;Western blot results showed that oleuropein could increase the expression of Bcl-2 protein and decrease the expression of Bax.Conclusion It has a protective effect on glucose deprivation and hypoxia injury in HK-2 cells, and its mechanism may be related to its inhibition of apoptosis.
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