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目的探讨过表达钠氢交换体1(NHE1)对RAW264.7细胞三磷酸腺苷结合盒转运体A1(ABCA1)蛋白表达的影响。方法采用Ad NHE1腺病毒感染RAW264.7细胞,Western blot法检测NHE1-EGFP融合蛋白的表达,激光共聚焦显微镜技术检测NHE1-EGFP融合蛋白细胞内定位,酸负载p H回复检测NHE1活性,Western blot法检测NHE1-EGFP融合蛋白对RAW264.7细胞ABCA1蛋白水平及钙蛋白酶(calpain)活性的影响,加入calpain抑制剂N-乙酰基-L-亮氨酰-L-亮氨酰-L-正亮氨酸(ALLN),Western blot法检测肝脏X受体激动剂TO-901317诱导ABCA1蛋白表达水平的影响。结果 Ad NHE1腺病毒感染RAW264.7细胞后,高表达NHE1-EGFP融合蛋白,定位在细胞质及细胞膜。NHE1-EGFP融合蛋白可降低ABCA1蛋白的表达水平并提高calpain活性,而calpain抑制剂ALLN能阻断ABCA1蛋白表达水平降低。结论 NHE1过表达通过上调calpain活性而降低ABCA1蛋白表达水平。
Objective To investigate the effect of overexpression of sodium hydrogen exchange 1 (NHE1) on the expression of adenosine triphosphate binding cassette transporter A1 (ABCA1) in RAW264.7 cells. METHODS: The RAW264.7 cells were infected with Ad NHE1 adenovirus and the expression of NHE1-EGFP fusion protein was detected by Western blot. The intracellular localization of NHE1-EGFP fusion protein was detected by laser scanning confocal microscopy. The NHE1 activity was detected by acid load p H, Method to detect the effect of NHE1-EGFP fusion protein on ABCA1 protein level and calpain activity in RAW264.7 cells, adding calpain inhibitor N-acetyl-L-leucyl-L-leucyl-L- (ALLN) and Western blot were used to detect the effect of liver X receptor agonist TO-901317 on ABCA1 protein expression. Results After infected with Ad NHE1 adenovirus, RAW264.7 cells were highly expressed NHE1-EGFP fusion protein and localized in the cytoplasm and cell membrane. NHE1-EGFP fusion protein can reduce the expression of ABCA1 protein and increase calpain activity, while calpain inhibitor ALLN can block the expression of ABCA1 protein. Conclusion NHE1 overexpression decreases ABCA1 protein expression by up-regulating calpain activity.