新城疫病毒对人类大肠癌细胞的杀伤作用研究

目的：研究NDV（F48E9和La Sota株）体外感染人类大肠癌细胞导致细胞死亡的方式。方法：实验采用处于对数生长期的人类大肠癌细胞。首先采用MTT法测定NDV对人类大肠癌细胞的杀伤活性。当细胞铺满30%培养瓶时,向细胞中加入10、20和40倍稀释的新城疫病毒,37℃感染1小时,继续培养24和48小时后进行进一步分析。通过倒置显微镜、HE染色、AO-EB荧光染色、透射电镜和琼脂糖凝胶电泳法从形态学和生物化学角度判断细胞死亡方式。结果：MTT法表明NDV可体外杀伤人类大肠癌细胞,且强毒株F48E9的杀伤活性强于弱毒株La Sota。细胞感染病毒后24小时即可表现出明显的病理变化：通过倒置显微镜和HE染色可见细胞融合及多核巨细胞,AO-EB荧光染色可见橙黄色荧光聚集于核膜边缘,透射电镜表现出典型的细胞凋亡形态学改变,琼脂糖凝胶电泳可见典型DNA ladder。上述变化在细胞感染病毒48小时后更加明显。结论：NDV可通过凋亡方式诱导人类大肠癌细胞死亡,且强毒株F48E9的杀伤活性高于弱毒株La Sota。

THE EFFECT OF NEWCASTLE DISEASE VIRUS (NDV) ON THE HUMAN COLORECTAL CANCER CELLS

Objective：To study the in vitro cytoidal infection of NDV（strain F48E9 and La Sota） to human colorectal cancer cells,and analyze the manner of cell death.Methods The human colorectal cancer cells which were growing in logarithmic propagation was used in the test.First,MTT assay was adopted to determine the activity of NDV that kils human colorectal cancer cells.The cells in 30% confluent were treated by adding NDV in 1 to 10、20 and 40 dilution kept culturing for another 1 hour at 37℃ for infection,then the cells were cultured for another 24 to 48 hours for further analysis.Reverse microscope,HE dye,AO-EB fluorescence dye,transmission electron microscope and agarose gel electrophoresis were used to determine the cell death manner in morphologic and biochemisty.Result： MTT assay showed that NDV could kill human colorectal cancer cells in vitro,the velogenic strain demonstrated stronger cytocidal infection than that of the lentogenic strain.Obvious cell pathological changes were observed 24 hours after infection in those cells positive with virus： cell fusion and coenocytic cell could be observed through reverse microscope and HE staining,orange fluorescence congregating in the edge of karyotheca was seen in AO-EB stained cells,transmission electron microscope revealed the typical morphologic changes of apoptosis,DNA ladder was also detected by agarose gel electrophoresis.The phenomenon mentioned above became more evident 48 hours later.Conclusion NDV could induced human colorectal cancer cells death in vitro via apoptosis,and the killing effect of velogenic strain F48E9 was stronger than that of the lentogenic strain La Sota.