HPK1在乳腺癌组织中的表达及HPK1过表达对人乳腺癌MCF-7细胞增殖、凋亡的影响

摘要

Aim:To detect the expression of hematopoietic progenitor kinase 1(HPK1) in carcinoma tissue of human in-vasive ductal breast carcinoma-not otherwise specified and breast carcinoma cells , and to investigate the effects of HPK 1 over-expression on proliferation and apoptosis of human breast carcinoma cells by constructing HPK 1 lentiviral vector .Methods:The expression level of HPK 1 protein in carcinoma tissue and paired adjacent tissue from 48 cases of breast carcinoma was de-tected by Western blot .The expression level of HPK1 in MCF10A and MCF-7 cells was detected by Western blot and RT-PCR.The HPK1 sequence was amplified by PCR , lentivirus recombinant vector pCDH-HPK1-puro was constructed , virus was packaged and MCF-7 cells were transfected ( overexpression group ) .The MCF-7 cells infected with empty vector virus was used as control group .The expression level of HPK 1 protein and mRNA was detected by Western blot and RT-PCR, respec-tively.The cell proliferation ability, cell apoptosis and cell cycle were detected by MTT assay and flow cytometry , respective-ly.Results:The expression level of HPK1 protein was lower in breast carcinoma tissue compared with adjacent tissue (P=0.036).The level of HPK1 protein and mRNA was lower in MCF-7 cells, compared with that of MCF10A cells(P<0.05). Compared with the control group, the expression level of HPK1 in the overexpression group was up-regulated, cell prolifera-tion ability was decreased significantly , while the apoptosis rate and the proportion of G 0/G1 were significantly increased ( P<0.05).Conclusion:The expression of HPK1 in both breast carcinoma tissue and cell line is low , and overexpression of HPK1 could inhibit proliferation and induce apoptosis and block the cells in G 0/G1 phase.%目的:检测造血祖细胞激酶1(HPK1)在人非特异型浸润性乳腺癌组织及乳腺癌细胞中的表达,并构建HPK1慢病毒载体以探究HPK1过表达对人乳腺癌细胞增殖、凋亡的影响.方法:采用Western blot检测48例乳腺癌组织和配对癌旁组织中HPK1蛋白的表达,采用Western blot、RT-PCR法检测正常乳腺上皮细胞(MCF10A)和乳腺癌MCF-7细胞中HPK1的表达水平.PCR扩增HPK1序列,构建慢病毒pCDH-HPK1-puro重组载体,包装病毒、转染MCF-7细胞(过表达组),以感染空载体病毒的MCF-7细胞作为对照组,分别采用Western blot、RT-PCR检测2组细胞HPK1的表达水平,MTT法检测细胞增殖能力的变化,流式细胞术检测细胞凋亡及细胞周期.结果:与癌旁组织相比,HPK1蛋白在乳腺癌组织中低表达(P=0.036).与MCF10A细胞相比,HPK1蛋白及mRNA在MCF-7细胞中低表达;与对照组相比,过表达组中HPK1蛋白及mRNA的表达水平上调,细胞增殖能力下降,细胞凋亡率升高,G0/G1期细胞比例升高(P均<0.05).结论:HPK1在乳腺癌组织及细胞中低表达,HPK1过表达可抑制MCF-7细胞增殖和诱导凋亡,并使细胞周期阻断在G0/G1期.