【摘要】 目的:探讨环磷腺苷效应元件结合蛋白(c AMP-response element binding protein,CREB)及磷酸化CREB(p-CREB)在前列腺癌细胞增殖中的作用。方法:采用组织芯片技术检测人正常前列腺、前列腺增生、不同分级前列腺癌组织中CREB及p-CREB的表达水平;免疫荧光及Western blotting检测人正常前列腺基质永生化细胞WPMY-1、前列腺癌细胞PC3及LNCap中CREB及p-CREB的表达水平;构建重组人源CREB质粒,并转染PC3及LNCap细胞,Western blotting及CCK-8试剂盒检测转染效率及细胞增殖水平的变化。结果:CREB及p-CREB在前列腺癌组织中表达率明显高于正常前列腺(96%vs 50%,88%vs 40%,P<0.05)和前列腺增生组织(96%vs 80%,88%vs 60%,P<0.05);CREB及p-CREB蛋白水平在前列腺癌PC3及LNCap细胞中表达量显著高于正常前列腺基质永生化细胞WPMY-1[PC3细胞:(5.10±0.62)vs(2.31±0.40)、(4.31±0.54)vs(2.02±0.38);LNCap细胞:(7.5±0.83)vs(2.31±0.40)、(6.15±0.69)vs(2.02±0.38),均P<0.05)];转染重组质粒CREB可上调前列腺癌细胞PC3及LNCap中增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)的表达(均P<0.05),且转染后的PC3及LNCap细胞增殖水平明显增加(P<0.05)。结论:CREB及p-CREB在前列腺癌组织、体外培养前列腺癌细胞PC3及LNCap中高表达,并可上调PC3及LNCap中增殖相关基因PCNA的表达,提高细胞增殖水平。更多还原

【Abstract】 Objective: To investigate roles of c AMP-response element binding protein( CREB) and phosphorglated CREB( p-CREB) in proliferation of prostate cancer cells. Methods: Tissue microarray technology was adopted to examine the expression levels of CREB and p-CREB in normal prostate tissue,hyperplasia prostate tissue and prostate cancer tissues at different grades. Immuno fluorescence staining and Western blotting were performed to detect the expression levels of CREB and p-CREB in human normal prostatic stromal immortalized WPMY-1 cell line,prostate cancer PC3 and LNCap cell lines. Recombinant human plasmid CREB was constructed and transfected into the PC3 and LNCap cells. Efficiency of the transfection and proliferation changes of the cells were verified by Western blotting and CCK-8 kit respectively. Results: Expression rates of CREB and p-CREB in prostate cancer tissue were significantly higher than those in normal prostate tissue( 96% vs 50% and 88% vs 40%,P < 0. 05) and hyperplasia prostate tissue( 96% vs 80% and88% vs 60%,P < 0. 05). Expression levels of CREB and p-CREB proteins in the prostate cancer PC3 and LNCap cell lines were significantly higher than those in the normal prostatic stromal immortalized WPMY-1 cell line [( 5. 10 ± 0. 62 vs2. 31 ± 0. 40),( 7. 5 ± 0. 83 vs 2. 31 ± 0. 40),( 4. 31 ± 0. 54 vs 0. 02 ± 0. 38),( 6. 15 ± 0. 69 vs 2. 02 ± 0. 38),all P <0. 05]. Transfection with recombined CREB plasmid could up-regulate the expression of proliferating cell nuclear antigen( PCNA) in the prostate tumor PC3 and LNCap cells( all P < 0. 05),and proliferation levels of the PC3 and LNCap cells significantly increased after the transfection( P < 0. 05). Conclusion: The CREB and p-CREB were highly expressed in human prostate cancer tissues and,the prostate cancer PC3 and LNCap cells cultured in vitro. In addition,the CREB and p-CREB could up-regulate the expressions of proliferation-related gene PCNA,and raise proliferation level of the cells.更多还原