人乳腺癌细胞RECK基因的表达及检测

The Expression and Determination of RECK Gene in Breast Cancer Cells

目的:研究人乳腺癌细胞株RECK基因的表达,阐明RECK基因表达水平与乳腺癌细胞侵袭力的关系。方法:采用transwell法,测定HBL-100,MCF-7和MDA-MB-435S三株人乳腺(癌)细胞的体外侵袭能力,同时应用免疫细胞化学技术,Westernblotting和RT-PCR技术分别检测三株细胞株MMP-2,MMP-9和RECK基因蛋白表达水平,及RECK基因mRNA转录丰度。结果:三株细胞中,MDA-MB-435S侵袭力最高(425.20±54.09),HBL-100最低(101.00±63.88),MCF-7居中,为(239.00±91.39),三组细胞的侵袭力比较差异均有显著性(P<0.01)。RECK基因在HBL-100细胞株中蛋白表达水平最高(免疫细胞化学值:3188.24±894.86;Westernblotting值:3.32±0.25),在MDA-MB-435S细胞株中不表达,MCF-7为(免疫细胞化学值:1058.92±336.53,P<0.01;Westernblotting值:2.23±0.59,P<0.05)。且在免疫细胞化学中,RECK基因的蛋白表达与MMP-2,MMP-9的表达水平成负相关。RECK基因在HBL-100细胞株中mRNA水平最高(2.32±0.02),在MDA-MB-435S细胞株中不表达(P<0.001)。结论:RECK基因表达水平与乳腺癌细胞体外侵袭力及MMP-2,MMP-9的表达水平成明显负相关。

Objective： To investigate the expression of the RECK gene in the human breast （cancer） cell lines and to determine the relationship between the expression level of RECK gene and invasive capacity of the breast cancer cell lines. Methods： The invasive capacity of breast （cancer） cell lines, including HBL-100, MCF-7 and MDA-MB-435S, were determined by Transwell method. The protein expressive levels of RECK, MMP-2 and MMP-9 gene in these three cell lines were measured by immunocytochemical method. And the expressions of RECK in gene and protein level were measured by Western Bloting and RT-PCR technique in breast （cancer） cell lines respectively. Results： The invasive capacity of breast （cancer） cell lines was as follows： the MDA-MB-435S was the highest （425.20±54.09）, the MCF-7 the moderate （239.00±91.39） and the HBL-100 （101.00±63.88） the lowest. There were significant differences in the invasive capacity among the 3 groups （P〈0.01）. The protein expression level of RECK gene in HBL-100 cell line was high （IC： 3188.24±894.86; WB： 3.32±0.25）, but no expression was found in MDA-MB-435S cell line. The value of MCF-7 was as follows, i.e. IC： 1058.92±336.53, （P〈0.01） and WB： 2.23±0.59, （P〈0.05）. Moreover, the expression of RECK gene was negatively correlated with that of the MMP-2, MMP-9 gene. The mRNA level of RECK gene in the HBL-100 cell line was the highest （2.32±0.02）, and no expression of the RECK was found in the MDA-MB-435S cell line （P〈0.001）. Conclusion： There was a significant negative correlation between the expressive level of RECK gene and the invasive capacity in vitro as well as expressive level of the MMP-2, MMP-9 gene.