探讨Her 2靶向重组的caspase6融合蛋白对骨肉瘤SOSP9607细胞的促凋亡作用。方法: 将抗Her 2单链抗体基因e23sFv与绿脓杆菌外毒素PE的转膜结构域基因（PEⅡ）和活性caspase6 基因连接，构建成Immunocasp6（e23sFvPEⅡcasp6）基因，将其克隆入真核表达载体pCMV中，转染SOSP9607细胞，间接免疫荧光法检测目的基因表达和细胞形态学变化，通过Annexin V染色、流式细胞术、MTT法检测来观察其促肿瘤细胞凋亡的作用。结果：转染SOSP9607细胞后，间接免疫荧光染色检测出caspase6的表达，SOSP9607细胞出现明显的核浓缩、碎裂；电镜观察到细胞质浓缩，胞内空泡，核染色质浓集等典型的凋亡特征；MTT法检测发现细胞的增殖明显被抑制；Annexin V染色流式细胞术检测可见明显的凋亡细胞,凋亡率为31.4%，较对照组细胞(凋亡率为6.0%)有非常显著的增加（P<0.01）。 结论： 重组Immunocasp6基因可以在转染的SOSP9607细胞中表达，并诱导细胞发生凋亡。

To investigate the proapoptotic effect of Her 2targeted recombinant caspase6 fusion protein on osteosarcoma SOSP9607 cells. Methods: Recombinant immunocasp6 was generated by fusing the genes of a signal peptide, a singlechain Her 2 antibody (e23sFv), a PEA translocation domain (PEA aa253364), and an active caspase6. The constructed immunocasp6 gene（e23sFvPEⅡcasp6） was cloned into pCMV plasmid and transfected into SOSP9607 cells. The target gene expression and the morphology of the transfected cells were observed by fluorescence and electron microscopy, and the proapoptotic effect of the recombinant gene was analyzed by Annexin VFITC staining, flow cytometry, and MTT assay. Results： The e23sFvPEⅡcasp6 fusion protein was detected in the cytoplasm of transfected SOSP9607 cells. The transfected cells presented the typical characteristics of apoptosis as detected by electron microscopy (cytoplasm concentration, chromatin condensation, vacuolation). Annexin VFITC staining revealed that the percentage of apoptotic cells in the transfectants of immunocasp6 genes was 31.4％,compared with 6.0％ in the control cells. MTT assay showed that the proliferation of immunocasp6 genetransfected SOSP9607 cells was much lower than that of non or mocktransfected cells. Conclusion： Her 2 targeted recombinant immunocasp6 fusion protein can express in SOSP9607 cells and induce apoptosis of SOSP9607 cells.

R73-362

国家自然科学基金重点项目（No.30330610）；国家自然科学基金项目（No. 30471988）；中国博士后基金项目（No.2005038259）