壳聚糖氧化石墨烯负载冬凌草甲素对A549细胞增殖及凋亡的影响
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篇名: | 壳聚糖氧化石墨烯负载冬凌草甲素对A549细胞增殖及凋亡的影响 |
TITLE: | Effects of Oridonin-loaded Chitosan Graphene Oxide on the Proliferation and Apoptosis of A 549 Cells |
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(IC50);采用显微镜观察CS-GO和CS-GO-oridonin(均为32μg/mL)作用2、4、10、24h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IC50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-GO16、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-oridonin16、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-GO-oridonin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。 |
ABSTRACT: | OBJECTIVE:To study the effects of chitosan graphene oxide car rier(CS-GO)loaded with oridonin (CS-GO- oridonin)on the proliferation and apoptosis of human lung cancer A 549 cells. METHODS :Taking A 549 cells as objects ,the survival rate of cells were detected by CCK- 8 method after treated with different concentrations of CS-GO (3,6,12,24,48 μg/mL)and CS-GO-oridonin loaded with same mass of oridonin (3,6,12,24,48 μg/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 32 μg/mL)for 2,4,10,24 h. The uptake of CS-GO ,oridonin,CS-GO-orionin(all 32 μg/mL)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 (16,32,64 μg/mL)and CS-GO-oridonin (16,32,64 μg/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);IC50 of CS-GO-oridonin was 32.61 μg/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 16,32,64 μg/mL CS-GO groups(P>0.05);apoptosis rate ,the content of ROS ,the protein expression of Bax and Bak in 16,32,64 μg/mL CS- GO-oridonin groups were increased significantly ,while the protein expression of Mcl- 1 were decreased significantly. Above indexes were significantly high er or lower than the same concentration CS-GO group (P<0.05). CONCL USIONS:CS-GO dose not affect the proliferation and apoptosis of A 549 cells;CS-GO-oridonin has obvious inhibition and apoptosis promoting effect on cells ,which may be related to increasing ROS production and regulating the expression of apoptosis related proteins. |
期刊: | 2021年第32卷第13期 |
作者: | 谢谭芳,朱小勇,黄天衍 |
AUTHORS: | XIE Tanfang ,ZHU Xiaoyong ,HUANG Tianyan |
关键字: | 壳聚糖氧化石墨烯载体;冬凌草甲素;A549细胞;增殖;凋亡 |
KEYWORDS: | Chitosan graphene oxide carrier ;Oridonin;A549 cells;Proliferation;Apoptosis |
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