五步蛇毒AAVC-I对人口腔鳞癌细胞的增殖抑制作用  

Effect of Agkistrodon acutus venom AAVC-I on the proliferation of human oral squamous-cell carcinoma cells

作　　者：廖国奇 方静 王振杰 李红艳 张根葆[2] 孔天翰 汪胜松 LIAN Guo-qi;FANG Jing;WANG Zhen-jie;LI Hong-yan;ZHANG Gen-bao;KONG Tian-han;WANG Sheng-song(Qimen Institute of Snakebite of Anhui Province,Qimen City,Anhui Province,245699,China;Institute of snake venom,wannan medical college,Wuhu City,Anhui Province,241002,China;Sanqibio-pharmaceutical technology co,LTD,Qimen City,Anhui Province,245699,China)

机构地区：[1]安徽省祁门蛇伤研究所,安徽祁门245699 [2]皖南医学院蛇毒研究所,安徽芜湖241002 [3]安徽省黄山市三祈生物医药科技有限公司,安徽祁门245699

出　　处：《蛇志》2020年第3期273-276,共4页Journal of Snake

基　　金：安徽省重点研究与开发计划项目(1704a0802146);黄山市科技计划项目(2017KN-21)。

摘　　要：目的观察祁门五步蛇毒抑瘤组分I(Agkistrodon acutus venom component-I,AAVC-I)对人口腔鳞癌Tca8113细胞的增殖抑制及凋亡的影响。方法以离子交换和凝胶过滤法从安徽祁门五步蛇粗毒中分离纯化五步蛇毒AAVC-I。常规培养口腔鳞癌Tca8113细胞,观察正常细胞形态学。实验分为对照组(加入等量细胞培养液)和实验组(分别加入AAVC-I 2.5、5.0、10.0μg/ml)。采用MTT法检测不同浓度(2.5、5.0、10.0μg/ml)的AAVC-I对Tca8113细胞24、48及72 h的增殖抑制率,倒置显微镜观察不同浓度AAVC-I处理后的细胞形态变化,透射电镜及AO/EB双荧光染色观察AAVC-I对细胞凋亡的影响,并与对照组进行比较。结果与对照组比较,实验组AAVC-I对Tca8113细胞增殖具有明显抑制作用(P<0.05);AAVC-I处理后,倒置显微镜下可见明显细胞凋亡现象,电镜下可见细胞核染色质、胞质浓缩及凋亡小体;AO/EB双荧光染色可见细胞膜完整,细胞核致密浓染,染成黄绿色荧光的早期凋亡细胞及细胞膜破损,细胞核浓聚及偏移,呈橘红色荧光的晚期凋亡细胞。结论祁门五步蛇毒AAVC-I能抑制人口腔鳞癌Tca8113细胞生长增殖,诱导其凋亡。Objective To study the effects of AAVC-I from Agkistrodon acutus venom on the growth and apoptosis of human oral squamous-cell carcinoma Tca8113 cells.Methods Isolation and purification of anti-tumor component I(AAVC-I)from Agkistrodon acutus venom of Qimen in Anhui province were carried out by ion exchange and gel filtration.Human oral squamous-cell carcinoma Tca8113 cells were conventionally cultured and the normal morphology of Tca8113 was observed.The experiment was divided into control group and experimental group.AAVC-I concentration is(2.5、5.0、10.0)μg/ml.The change of cell morphology were observed by inverted microscope after the different concentration of AAVC-I treatment.The effect of AAVC-I on cell apoptosis was observed by transmission electron microscope and acridine orange and ethidium bromide flurescence staining method(AO/EB).Results The proliferation of Tca8113 was inhibited significantly by experimental group AAVC-I compared with control group(P<0.05).After treating with AAVC-I,we could observe the phenomena of apoptotic body formation and the condensed nuclear chromatin and cytoplasm by electron microscope and significant cell apoptosis by inverted microscope.By AO/EB fluorescence staining,early stage apoptotic cells of intact cell membrane and hyperchromatic cell nucleus could be stained yellow-Green and late stage apoptotic cells of damaged cell membrane and nuclear concentration and migration could be stained orange.Conclusion AAVC-I can inhibit the proliferation of Tca8113 and induce apoptosis of Tca8113.

关 键 词：五步蛇毒 人口腔鳞癌细胞 增殖抑制 细胞凋亡 

分 类 号：R739.8[医药卫生—肿瘤] R996.3[医药卫生—临床医学]