Zhao Jin, Wang Di, Zhao Hong, Lin Wei
The Fifth Department,Medical College of Northwest Minorities University, Lanzhou, Gansu, 730000, PR China.
Ai Zheng. 2003 Jul;22(7):719-24.
BACKGROUND & OBJECTIVE: Clostridium difficile is recognized as a frequent cause of antibiotic-induced diarrhea. This study was designed to investigate whether Clostridium difficile toxin A might induce apoptosis on human hepatoma cell line SMMC7721 and African green monkey kidney Vero cells.
Highly purified toxin A was obtained by bovine thyroglobulin affinity purification followed by ion exchange chromatography on Q sepharose. The apoptosis induction of toxin A was examined on SMMC7721 cells with Vero cells as a control. Inhibition of proliferation was measured by MTT assay. Morphological assessment of apoptosis was performed with fluorescence and electronic microscopy. DNA fragmentation was observed by agarose gel electrophoresis. Cell cycle distribution was analyzed by flow cytometry.
Toxin A (0.293-4.690 mg x L(-1) ) inhibited proliferation of SMMC7721 and Vero cells in a time- and concentration-dependent manner. Morphological changes of typical apoptosis showed that SMMC7721 had a higher percentage of apoptosis than Vero cells. Agarose gel electrophoresis of DNA from SMMC7721 treated with toxin A for 48 hours revealed a "ladder" pattern.
Clostridium difficile toxin A induced apoptosis of SMMC7721 and Vero cells. The apoptosis induction on SMMC7721 cells was more effective than that on Vero cells.
艰难梭菌被认为是抗生素相关性腹泻的常见病因。本研究旨在探讨艰难梭菌毒素A是否能诱导人肝癌细胞系SMMC7721和非洲绿猴肾Vero细胞发生凋亡。
通过牛甲状腺球蛋白亲和纯化，随后在Q琼脂糖上进行离子交换色谱法获得高度纯化的毒素A。以Vero细胞为对照，检测毒素A对SMMC7721细胞的凋亡诱导作用。采用MTT法检测细胞增殖抑制情况。用荧光显微镜和电子显微镜对凋亡进行形态学评估。通过琼脂糖凝胶电泳观察DNA片段化。采用流式细胞术分析细胞周期分布。
毒素A（0.293 - 4.690 mg·L⁻¹）以时间和浓度依赖性方式抑制SMMC7721和Vero细胞的增殖。典型凋亡的形态学变化显示，SMMC7721细胞的凋亡率高于Vero细胞。用毒素A处理48小时后的SMMC7721细胞DNA琼脂糖凝胶电泳呈现“梯状”图谱。
艰难梭菌毒素A可诱导SMMC7721和Vero细胞凋亡。毒素A对SMMC7721细胞的凋亡诱导作用比对Vero细胞更有效。
