摘要
目的:研究壳聚糖氧化石墨烯载体(CS-GO)负载冬凌草甲素(CS-GO-oridonin)对人肺癌细胞A549增殖及凋亡的影响。方法:以A549细胞为对象,采用CCK-8法检测经不同质量浓度的CS-GO(3、6、12、24、48μg/mL)和负载了等质量冬凌草甲素的CS-GO-oridonin(3、6、12、24、48μg/mL,以冬凌草甲素质量计,下同)作用后的细胞存活率,并计算CS-GO-oridonin的半数抑制浓度(IC_(50));采用显微镜观察CS-GO和CS-GO-oridonin(均为32μg/mL)作用2、4、10、24 h后的细胞形态,并采用荧光标记法观察细胞对CS-GO、冬凌草甲素、CS-GO-oridonin(均为32μg/mL)的摄取情况;采用流式细胞术观察不同质量浓度CS-GO(16、32、64μg/mL)和CS-GO-oridonin(16、32、64μg/mL)作用后细胞的凋亡情况及细胞中活性氧(ROS)的含量,采用Westernblot法检测细胞中抗凋亡相关蛋白[髓样细胞白血病1(Mcl-1)、Bax、Bak]蛋白的表达情况。结果:经不同质量浓度CS-GO作用后,细胞的存活率仍不低于90%;经不同质量浓度CS-GO-oridonin作用后,细胞存活率呈下降趋势,且显著低于相同质量浓度的CS-GO组(P<0.01);CS-GO-oridonin的IC_(50)为32.61μg/mL。经CS-GO作用后,细胞形态未见明显改变;经CS-GO-oridonin作用后,细胞出现皱缩、成团脱落、悬浮物增多等现象;当细胞摄取冬凌草甲素和CS-GO-oridonin后,其荧光均较摄取CS-GO的细胞有所增强。与空白组比较,CS-GO 16、32、64μg/mL组细胞的凋亡率和细胞中ROS含量、凋亡相关蛋白的表达水平均无显著变化(P>0.05);而CS-GO-oridonin 16、32、64μg/mL组细胞的凋亡率、细胞中ROS含量和Bax、Bak蛋白的表达水平均显著升高,Mcl-1蛋白的表达水平均显著降低,且上述指标水平均显著高于或低于同质量浓度CS-GO组(P<0.05)。结论:CS-GO不会影响A549细胞的增殖及凋亡;CS-GOoridonin对细胞有明显的抑制和促凋亡作用,这种作用可能与增加细胞中ROS的产生、调控凋亡相关蛋白的表达有关。
OBJECTIVE:To study the effects of chitosan graphene oxide carrier(CS-GO)loaded with oridonin(CS-GOoridonin)on the proliferation and apoptosis of human lung cancer A549 cells.METHODS:Taking A549 cells as objects,the survival rate of cells were detected by CCK-8 method after treated with different concentrations of CS-GO(3,6,12,24,48μg/m L)and CS-GO-oridonin loaded with same mass of oridonin(3,6,12,24,48μg/mL,by the weight of oridonin,the same below).IC_(50) of CS-GO-oridonin was calculated.The cell morphology were observed by microscope after treated with CS-GO and CS-GO-oridonin(both 32μg/mL)for 2,4,10,24 h.The uptake of CS-GO,oridonin,CS-GO-orionin(all 32μg/m L)by cells was observed with fluorescence labeling method.The apoptosis of cells and the content of ROS were observed by flow cytometry after treated with different concentrations of CS-GO(16,32,64μg/mL)and CS-GO-oridonin(16,32,64μg/mL).The expression of anti-apoptosis related proteins(Mcl-1,Bax and Bak)were detected by Western blot.RESULTS:After treated with different concentrations of CS-GO,the survival rate of cells was still above 90%;after treated with different concentrations of CS-GO-oridonin,the survival rate of cells showed a downward trend,and was significantly lower than that of CS-GO group(P<0.01);IC_(50) of CS-GO-oridonin was 32.61μg/mL.After CS-GO treatment,the cell morphology did not change significantly;after CS-GO-oridonin treatment,the cells shrunk and fell off in clusters,and the suspended matter increased;the fluorescence of oridonin and CS-GO-orionin taken up by cells was enhanced than CS-GO.Compared with blank group,there was no significant change in the apoptosis rate,the content of ROS and the expression of apoptosis-related protein in 16,32,64μg/mL CS-GO groups(P>0.05);apoptosis rate,the content of ROS,the protein expression of Bax and Bak in 16,32,64μg/mL CSGO-oridonin groups were increased significantly,while the protein expression of Mcl-1 were decreased significantly.Above indexes were significantly higher or lower than
作者
谢谭芳
朱小勇
黄天衍
XIE Tanfang;ZHU Xiaoyong;HUANG Tianyan(School of Pharmacy,Guangxi University of TCM/Key Laboratory of Common Technology of Traditional Chinese Medicine Preparation in Universities of Guangxi/Modern Chinese Medicine Preparation Engineering Technology Research Center,Nanning 530022,China)
出处
《中国药房》
CAS
北大核心
2021年第13期1589-1595,共7页
China Pharmacy
基金
广西高校中青年教师基础能力提升项目(No.2017KY0292)
广西高校中药制剂共性技术研发重点实验室课题(No.70-ZJGX2017003)。
