Study of the inhibitory effects on malignant progression and anticancer drug sensitivity of human squamous cell carcinoma by intensifying cell to cell communication

苹果酸酯（二异丙基，1,3 -二硫代1-2-酰基柠檬酸酯，MT）在临床上被用作肝保护剂。由于我们注意到MT诱导培养的血管内皮细胞分化，我们检测了MT对口腔鳞状细胞癌细胞（SAS, Ca9-22, HSC-2,-3,-4）侵袭的抑制作用，并分析了其增强内皮细胞和肿瘤细胞对细胞粘附的机制。在添加10%胎牛血清的DMEM和D/F-12混合培养基中培养5株SCC细胞系和大鼠肺内皮细胞。利用相差显微镜计数RLE单层下形成的菌落/cm2数，测定SCC细胞的侵袭能力。细胞间连接通讯通过刮擦加载法检测。与刮边耦合的细胞百分比由染料转移细胞的数量决定。电镜下观察RLE单层的形态学变化。western blotting分析MT处理RLE后Connexin43的表达。在RLE单层体外侵袭实验中，经MT （10ng/ml, 24h）处理的肿瘤细胞的侵袭不受影响；然而，经MT处理后，三种细胞系的侵袭均明显受到抑制（p< 0.05）。01)。经MT处理的RLE单层经电镜观察，形成了类似缝隙结的结构。经MT处理的RLE单层细胞间通讯增加（82）。5%)，与未接受MT治疗的患者相比（32%）。5%)。一种连接相关蛋白cnnexin43也在MT处理的RLE和SAS中升高。在SAS肿瘤的体内生长中，MT+CDDP处理的裸鼠与CDDP处理的裸鼠相比，原发肿瘤的大小减小。结论：MT通过增强血管内皮细胞间的粘附抑制肿瘤细胞的侵袭，通过改变肿瘤细胞间的通讯增强抗癌药物敏感性。少

Malatolate (Diisoprppyl, 1, 3-dithio1-2-ylidenemalonate, MT) is clinically used as a hepatoprotective agent. Because we noticed that MT induced the differetiation of cultured vascular endothelial cell, we examined the inhibitory effects of MT on invasion of oral squamous cell carcinoma cell (SCC) lines (SAS, Ca9-22, HSC-2,-3,-4) and analyzed its mechanisms of intensifying endothelial and tumor cell to cell adhesion. Five SCC cell lines and rat lung endothelial (RLE) cells were grown in a mixed medium of DMEM and D/F-12 supplemented 10% fetal bovine serum. The invasion capacity of the SCC cells was measured by counting the number of colonies /cm2 formed under the RLE monolayer using a phase contrast microscope. Intercellular junctional communication was examined by the scrape-loading assay. The percentage of cells coupled with the scrape edge was determined by the number of dye-transferred cells. Morphological changes in RLE monolayer observed with electron microscope. Connexin43 expres … More sion of RLE treated with MT was analyzed by western blotting. In an in vitro invasion assay using a RLE monolayer, invasion of tumor cells which had been treated with MT (10ng/ml, 24h) was not affected; however, when had been treated with MT, invasion was significantly inhibited in three cell lines (p<0. 01). Electron-microscopical examination of the RLE monolayer treated with MT showed the development of gap junction like structure. The cell to cell communication increased in RLE monolayers treated with MT(82. 5%), compared to non-treated with MT(32. 5%). A junction associated protein, cnnexin43 was also elevated at RLE and SAS treated with MT. In vivo growth of SAS tumor, primary tumor size was reduced in MT+CDDP treated nude mice compared with CDDP treated nude mice. Conclusion: These findings suggests that MT suppressed tumor cell invasion by intensifying the cell-cell adhesion of vascular endothelium and that enhanced anticancer drug sensitivity by modifying tumor cell communication. Less