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目的观察AnnexinⅡ在氧化型低密度脂蛋白(ox-LDL)诱导泡沫细胞形成中表达的改变及其与细胞内胆固醇酯(CE)/总胆固醇(TC)比值(CE/TC)改变的相关性,从而探讨AnnexinⅡ在ox-LDL诱导泡沫细胞形成中的意义。方法常规培养RAW264.7单核细胞株,待细胞长至90%汇合度时,分别加入不同浓度的ox-LDL(50、100、200 mg/L)孵育24 h,以未经氧化修饰的天然低密度脂蛋白(n-LDL,200 mg/L)为阴性对照,测定细胞内TC和CE含量,油红O染色鉴定泡沫细胞;比色法测定乳酸脱氢酶(LDH)释出量和MTT法检测细胞存活率来观察oxLDL对细胞的毒性作用;RT-PCR和Western blot检测AnnexinⅡmRNA和蛋白的表达水平。结果不同浓度的oxLDL(50、100、200 mg/L)孵育24 h,仅200 mg/L ox-LDL组细胞活力显著降低,LDH释出量显著升高,与n-LDL(200mg/L)组比较差异具有显著性(P<0.01),50 mg/L和100 mg/L ox-LDL对细胞活力和LDH释出量无显著影响(P>0.05);CE/TC比值分别是55.0%±6.7%、61.8%±4.8%和59.6%±5.2%,与n-LDL(200 mg/L)组比较差异均具有显著性(P<0.05),其中浓度为100 mg/L的ox-LDL作用最显著;ox-LDL呈浓度依赖性抑制AnnexinⅡmRNA和蛋白的表达,AnnexinⅡmRNA和蛋白的表达改变与CE/TC比值呈负相关(r_1=-0.9374,P<0.05;r_2=-0.9548,P<0.05)。结论 ox-LDL呈浓度依赖性抑制AnnexinⅡ表达,提示其可能影响细胞内胆固醇自平衡从而诱导泡沫细胞形成。
Objective To observe the changes of Annexin Ⅱ expression in foam cells induced by ox-LDL and its correlation with the change of the ratio of cholesterol / cholesterol (TC / TC) To explore the significance of Annexin II in ox-LDL-induced foam cell formation. Methods RAW264.7 mononuclear cell line was cultured routinely. When the cells reached 90% confluency, the cells were incubated with different concentrations of ox-LDL (50, 100, 200 mg / L) for 24 h. Low-density lipoprotein (n-LDL, 200 mg / L) was used as a negative control to measure the contents of TC and CE in cells and the foam cells by Oil Red O staining. The amount of lactate dehydrogenase (LDH) The cell viability assayed by oxLDL was used to observe the cytotoxicity of oxLDL. The expression of AnnexinⅡmRNA and protein was detected by RT-PCR and Western blot. Results Compared with n-LDL (200mg / L), the cell viability of oxLDL (50,100,200 mg / L) (P <0.01). There was no significant difference in cell viability and LDH release between 50 mg / L and 100 mg / L ox-LDL (P> 0.05). The CE / TC ratios were 55.0% ± 6.7%, 61.8% ± 4.8% and 59.6% ± 5.2%, respectively. The difference was significant (P <0.05) with n-LDL (P <0.05); ox-LDL inhibited AnnexinⅡmRNA and protein expression in a concentration-dependent manner. . Conclusion Ox-LDL can inhibit the expression of Annexin Ⅱ in a concentration-dependent manner, suggesting that ox-LDL may affect the intracellular cholesterol homeostasis and induce the formation of foam cells.