伊立替康对RAW 264.7巨噬细胞增殖和细胞凋亡的影响
发布时间：2019-06-28 11:44
【摘要】：目的分析抗肿瘤药伊立替康(CPT-11)对RAW 264.7巨噬细胞增殖及细胞凋亡的影响,并探讨其作用机制。方法WST-1法检测CPT-11对RAW 264.7细胞增殖的作用,倒置显微镜观察CPT-11对RAW 264.7细胞形态的影响,Hoechst33342染细胞核观察RAW 264.7细胞的核形态,流式细胞术检测亚二倍体(凋亡峰)的变化,免疫印迹法检测Caspase-3活化、PARP及细胞周期相关蛋白的变化。结果 CPT-11能明显抑制RAW 264.7细胞增殖,并呈时间和剂量依赖性。在CPT-11作用下RAW 264.7细胞体积增大,细胞核增大,部分细胞发生核碎裂(细胞凋亡),并且原本贴壁生长、具有伪足的细胞变圆脱落。流式细胞术结果也显示,CPT-11可诱导RAW 264.7细胞产生亚二倍体峰(凋亡细胞),且细胞凋亡率呈浓度依赖性上升。同时,处于G2/M期细胞也明显增多,且呈浓度依赖性。免疫印迹分析显示,CPT-11可活化Caspase-3,使PARP水平下降,同时上调细胞周期蛋白Cyclin B1,下调Cyclin D1的表达。结论伊立替康通过诱导细胞周期阻滞而抑制RAW 264.7细胞增殖,通过激活Caspase-3通路诱导细胞凋亡。
[Abstract]:Objective to analyze the effect of CPT-11 on the proliferation and apoptosis of RAW 264.7 macrophages and to explore its mechanism. Methods the effect of CPT-11 on the proliferation of RAW 264.7 cells was detected by WST-1 assay, the effect of CPT-11 on the morphology of RAW 264.7 cells was observed by inverted microscope, the nuclear morphology of RAW 264.7 cells was observed by Hoechst33342 staining, the changes of subdiploid (apoptosis peak) were detected by flow cytometry, and the activation of Caspase-3, PARP and cell cycle related proteins were detected by immunoblotting. Results CPT-11 could significantly inhibit the proliferation of RAW 264.7 cells in a time-and dose-dependent manner. Under the action of CPT-11, the volume of RAW 264.7 cells increased, the nucleus increased, some cells broke into nucleus (apoptosis), and the cells with pseudopod became round and exfoliated. The results of flow cytometry also showed that CPT-11 could induce subdiploid peak (apoptotic cells) of RAW 264.7 cells, and the apoptosis rate increased in a concentration-dependent manner. At the same time, the number of cells in G _ 2 鈮，
本文编号：2507277
[Abstract]:Objective to analyze the effect of CPT-11 on the proliferation and apoptosis of RAW 264.7 macrophages and to explore its mechanism. Methods the effect of CPT-11 on the proliferation of RAW 264.7 cells was detected by WST-1 assay, the effect of CPT-11 on the morphology of RAW 264.7 cells was observed by inverted microscope, the nuclear morphology of RAW 264.7 cells was observed by Hoechst33342 staining, the changes of subdiploid (apoptosis peak) were detected by flow cytometry, and the activation of Caspase-3, PARP and cell cycle related proteins were detected by immunoblotting. Results CPT-11 could significantly inhibit the proliferation of RAW 264.7 cells in a time-and dose-dependent manner. Under the action of CPT-11, the volume of RAW 264.7 cells increased, the nucleus increased, some cells broke into nucleus (apoptosis), and the cells with pseudopod became round and exfoliated. The results of flow cytometry also showed that CPT-11 could induce subdiploid peak (apoptotic cells) of RAW 264.7 cells, and the apoptosis rate increased in a concentration-dependent manner. At the same time, the number of cells in G _ 2 鈮，
本文编号：2507277
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