NECROSIS AND APORTOSIS OF HUMAN PANCREATIC CANCER CELL LINES-FOR THE IMPROVEMENT OF ADJUVANT CHEMOTHERAPY FOR PANCREATIC CANCER

本文对比研究了50Gy射线照射、依托泊苷和叔丁基氢过氧化氢对人胰腺癌细胞株的凋亡和坏死的影响。本实验以人胰腺癌细胞株MIA paa2、PANC-1和人胆囊癌细胞株GB-D1为实验对象。用台盼蓝拒染法检测细胞死亡，漂浮细胞和贴壁细胞分别计数细胞数。50Gy射线照射后5d，3种细胞系60%以上的细胞死亡，1um的蛋白质合成抑制剂放线菌酮可明显保护这些细胞。Papanicolaou染色显示，3种不同细胞系均可见典型的凋亡核碎裂和凋亡小体。PANC-1和GB-D1的DNA呈梯状条带，而MIA PACA2的DNA呈拖尾状。加入10微米依托泊苷后，MIA PACA2上再次显示DNA涂片图案。流式细胞仪检测显示照射后24小时细胞周期完全停滞于G2-M期，依托泊苷处理后细胞向G2-M期聚集，而叔丁基氢过氧化氢诱导的细胞坏死未见细胞周期延迟或停滞。结果表明，大剂量X射线照射后的细胞死亡可能是一种与常规坏死完全不同的细胞凋亡。

Apoptosis and necrosis of human pancreatic cancer cell lines were comparatively investigated after 50 Gy irradiation, etoposide and tert-butylhydroperoxide. Human pancreatic cancer cell lines, MIA paca2 and Panc-1, and a human gall bladder cancer cell line, GB-d1 were used in the experiment. Cell deaths were determined by trypan blue exclusion test with cell number counting separately both on the floating cells and on the attached cells. Over 60% of the cells from three cell lines were dead 5 days after 50 Gy irradiation and those were clearly protected by the addition of 1 uM cycloheximide, a protein synthesis inhibitor. Typical apoptotic nuclear fragmentations and apoptotic bodies were also observed on 3 different cell lines by Papanicolaou staining. Electrophoresis revealed DNA ladder pattern on Panc-1 and GB-d1, however, DNA from MIA paca2 showed smear pattern. Again, DNA smear pattern was shown on MIA paca2 after the addition of 10 uM etoposide. With the flowcytometry, complete G2-M arrest 24 hr after 50 Gy irradiation and the accumulation to G2-M after the treatment of etoposide were shown while any cell cycle delay or arrest was not observed on necrosis induced by the treatment of tert-butylhydroperoxide. The results indicated that cell deaths after high dose X-ray irradiation may be apoptosis which is completely different from conventional necrosis.