亚甲蓝对体外培养C6细胞凋亡的实验研究

Methylene blue trial research for apoptosis of rat C6 glioma

目的研究亚甲蓝(MB)对体外培养C6胶质瘤细胞的促凋亡作用及其对Caspase3mRNA水平的影响。初步探讨MB抗胶质瘤的作用机制,并为MB作为肿瘤的辅助治疗提供理论和实验依据。方法以0.1μg/ml MB处理0、6、12、24小时的C6细胞;以0μg/ml、0.05μg/ml、0.1μg/ml、0.15μg/ml MB处理24小时的C6细胞,通过流式细胞仪检测C6细胞的凋亡率;通过RT-PCR进行凋亡相关基因Caspase-3mRNA水平的检测。结果经流式细胞术检测,0.1μg/ml MB作用24小时后细胞大部分集中在G0/G1期,出现典型的凋亡峰。以同一浓度(0.1μg/ml)MB处理不同时间的C6细胞;以不同浓度MB处理同一时间(24小时)的C6细胞,经RT-PCR证实Caspase-3mRNA随MB作用时间延长和浓度增高而升高,各组间均有显著差异(P<0.01)。结论MB对C6细胞具有诱导凋亡的作用,此过程中伴有Caspase-3的升高,且Caspase-3的升高与MB作用时间的延长和作用浓度的增高相一致。提示MB可能通过Caspase-3途径诱导细胞凋亡,且MB的抗胶质瘤作用有明显的时间和浓度依赖性。

Objective To research that MB has the function of inducing apoptosis on C6 glioma and it can effect the level of Caspase-3 mRNA. It could confirm the resistect glioma action of MB and provide us with theory and the test foundation that MB could be the additional therapy to tumor. Methods In this test ,C6 glioma cell were affected for 0,6,12,24 hours by 0.1μg/ ml MB and affected by 0,0.05,0.1,0.15 μg/ ml MB for 24 hours, the cells cycle analysis was determined by flow cytometric;assessing the expression of the caspase-3 mRNA by RT-PCR. Results We find that the typical apoptosis peak appears after the C6 glioma cell affected for 24 hours by 0. 1μg/ml MB form flow cytometric;It is certified by RT-PCR that caspase-3 mRNA ascend along with the time and the concentration of MB ,and the there was statistically signitlcant difference（ P 〈 0.01 ）. Conlusion It suggest that MB could induce the apoptosis of C6 gloma through the way of caspase-3 and the expression of caspase-3 mRNA concide with the time and concentration of MB.