摘要
采用MTT检测法、Annexin-V/PI双染流式细胞仪检测法及细胞形态学观察法,研究了植物型杀虫剂胡椒碱对白纹伊蚊Aedes albopictusC6/36细胞株的细胞毒性、诱导细胞凋亡与坏死的关系、Fas蛋白的表达、细胞生长状态改变及受损细胞的恢复。结果表明:胡椒碱浓度在0.035mmol/L以上时可以诱发C6/36细胞形态学改变,在倒置显微镜下表现为细胞呈多形性,细胞胀大或皱缩,细胞间隙增宽,大片细胞聚集成团,脱落、崩解、死亡。用胡椒碱处理24h后,对C6/36细胞的半数毒性浓度(IC50)为0.32mmol/L,且毒性作用强度随着药物浓度增加而增强。Annexin-V/PI双染法检测结果显示药物浓度在0.28mmol/L以上诱导的细胞凋亡明显,药物浓度为0.56mmol/L时细胞凋亡率达19.4%,而药物引起的细胞总死亡率为72.7%;Fas蛋白表达在药物浓度0.28mmol/L以上时有所上调,但不明显。高浓度胡椒碱(0.56mmol/L)分别作用于C6/36细胞24和48h,在经历一段生长停滞后,24h组受损细胞均可缓慢恢复,而48h组细胞则出现不可逆性的损伤。由此认为:胡椒碱对C6/36细胞株有毒性作用,但药物诱导的细胞凋亡在细胞毒性作用机制中不占主导地位;胡椒碱对C6/36细胞的作用有时间依赖性,延长作用时间,药物对细胞的毒性作用增强。
We observed cytotoxicity, apoptosis, the expression of Fas protein, growth state and damaged cells produced by botanical insecticide piperine on C6/36 cell strain of Aedes albopictus recovery of using MTT, Annexin-V/PI double staining and inverted microscope observation. The results indicated that piperine caused the morphological change in the treated cells at the concentration of above 0.035 mmol/L. The cells showed pantomorphia under the inverted microscope. They became swollen or shrunk, and the space between cells was widened. A large number of cells assembled into conglobations, and then ablated, disaggregated and died. The ICso of C6/36 cell 24 hours after treatment by piperine was 0.32 mmol/L. Moreover, the cytotoxicity increased with the raising of concentration of piperine. The results of Annexin-V/PI double staining showed that the apoptosis was apparent in treatments of the piperine concentration above 0.28 mmol/L, while the rate of apoptosis was 19.4 % in the treatement of the piperine concentration of 0.56 mmol/L. The expression of Fas protein increased in the treatments of the piperine concentration above 0.28 mmol/L but not apparently, and the total death rate was 72.7%. Treated with high concentration of piperine （0.56 mmol/L） for 24 hours and 48 hours respectively, following a period of growth retardation, the cells of 24-hour-group could recover slowly while the 48-hour-group showed inconvertibility. The results suggest that there is time dependence of the effect on C6/36 by piperine. Apoptosis, however, is not the main mechanism. The cytotoxicity would be increased with the treating time prolonged.
出处
《昆虫学报》
CAS
CSCD
北大核心
2007年第5期481-487,共7页
Acta Entomologica Sinica
基金
国家自然科学基金项目(39970654)
