反义封闭NGAL基因表达对SHEEC食管癌细胞微丝骨架的影响

为了研究反义封闭NGAL基因表达对SHEEC食管癌细胞微丝骨架以及肿瘤细胞生物学行为的影响，以不同长度NGAL基因片段反义表达载体和硫代修饰反义寡核苷酸单链片段转染SHEEC食管癌细胞，通过G418筛选,建立一系列旨在封闭SHEEC食管癌细胞NGAL基因表达的亚细胞克隆.在细胞内F-肌动蛋白(F-actin)及DNA荧光双标记基础上，通过流式细胞术、激光共聚焦显微镜扫描术等技术手段检测封闭反义NGAL基因表达后， SHEEC食管癌细胞中F-actin和DNA含量、F-actin形态结构以及肿瘤细胞生物学行为的变化特征.结果显示，反义封闭NGAL基因表达后，SHEEC食管癌细胞F-actin的含量明显降低，与永生化食管上皮细胞SHEE相近，但细胞分裂增殖指数未见明显变化.表明反义封闭NGAL基因表达对SHEEC食管癌细胞的微丝骨架有明显影响，而对SHEEC食管癌细胞的分裂增殖影响不明显.激光共聚焦显微镜扫描观测显示，反义封闭NGAL基因表达可使SHEEC食管癌细胞F-actin分布均匀，F-actin小体减少，细胞间连接重新建立，结构较紧密，主要形态结构特征与SHEE细胞趋于一致.提示反义封闭NGAL基因表达可对SHEEC食管癌细胞的微丝骨架F-actin产生明显影响，推测癌细胞的微丝骨架F-actin可能是NGAL基因在SHEEC食管癌细胞中发挥功能的一种作用环节.

Antisense Blocking of NGAL Gene Expression Affects The Microfilament Cytoskeleton in SHEEC Esophageal Cancer Cells

In order to study the effect of microfilament cytoskeleton on SHEEC esophageal cancer cells and the biological activity variation of cancer cells caused by antisense blocking of NGAL gene expression, making use of antisense expressing vector of different length NGAL gene segments and antisense oligonucleotide segments of thio-modification to transfer the SHEEC cancer cells of esophagus, a series of subcellular clones aiming at the blocking of NGAL gene expression of SHEEC esophageal cancer cells were established by means of G418 selection. On the basis of the fluorescent dual-labeling of F-actin and DNA within cells and after antisense blocking of NGAL gene expression, the content of F-actin and DNA, the F-actin cytoskeleton structure in SHEEC cell and the variational characteristics of biological action on the tumor cell were tested by use of flow cytometry and confocal scanning laser microscope (CSLM). The results showed that F-actin level in the SHEEC cells was obviously reduced after antisense blocking and it was similar to that in SHEE cells, but no obvious change of cellular division index was found. This indicates that the blocking of NGAL gene expression has an obvious effect on the microfilament cytoskeleton of SHEEC cells, but no obvious influence on SHEEC cell proliferation. With the technique of CSLM, it is revealed that the blocking of NGAL gene expression could make the cellular F-actin cytoskeleton of SHEEC cell uniformly distributed, F-actin body evidently decreased, the joint structure of the cell reestablished, the cellular structure more compact, and the main structure characteristics of SHEEC cell and those of F-actin cytoskeleton structures of SHEE cell consistently approached. The result suggests that SHEEC cellular F-actin cytoskeleton of the esophagus cancer could be obviously affected by antisense blocking of NGAL gene expression, and it is conjectured that microfilament cytoskeleton F-actin in cancer cell could be a function key of NGAL gene that plays a role in SHEEC cancer cell.