机构地区:[1]东南大学附属中大医院普外科,南京210009
出 处:《实用肿瘤学杂志》2008年第5期401-405,共5页Practical Oncology Journal
基 金:国家自然科学基金资助项目(NO.30500491)
摘 要:目的探讨CDC25B1基因在人胰腺癌细胞株Patu8988中的表达情况,并将真核表达重组载体pcDNA-CDC25B1转染至细胞中,观察CDC25B1基因对细胞生物学行为的影响,以了解它在细胞周期及胰腺肿瘤细胞恶性转化方面的作用,为研究胰腺癌的发病机制与治疗提供理论依据。方法利用Lipofectamine2000将CDC25B1转染至人胰腺癌细胞株Patu8988中,通过RT-PCR和Westernblot在mRNA和蛋白水平验证质粒转染成功,通过MTT实验、侵袭实验和细胞周期检测,观察CDC25B1对细胞生物学行为的影响。结果RT-PCR发现在胰腺癌细胞株Patu8988和转染空质粒的细胞株中,CDC25B1在mRNA水平表达很弱;转染pcDNA-CDC25B1组中CDC25B1高表达。Westernblot发现在胰腺癌细胞株Patu8988中,CDC25B蛋白有少量表达;转染pcDNA-CDC25B1组CDC25B蛋白高表达。MTT实验显示,转染pcDNA-CDC25B1组的细胞生长在72h受到明显抑制(P(0.05)。侵袭实验显示,转染pcDNA-CDC25B1的细胞其穿透Matrigel胶的能力明显增强(P(0.05)。流式细胞计量术显示,转染了pcDNA-CDC25B1组中,G1期的细胞明显增多(P(0.05);同时转染了pcDNA-CDC25B1组中,G2期的细胞明显减少,与未处理组比较有统计学意义(P(0.05)。结论在胰腺癌细胞株Patu8988中的CDC25B1转录水平很弱,存在少量CDC25B蛋白表达。转染的CDC25B1基因提高CDC25B1蛋白的表达,使细胞停滞在G1期,对细胞的生长有抑制作用,但能够增强细胞的侵袭能力;CDC25B1可能不是通过单一的调控细胞周期而实现其对细胞的影响,还存在其它多种途径。Objective To investigate the expression of CDC25B1 in the pancreatic cen line Pa- tu8988, and evaluate the effect of CDC25B1 on the biological behaviors of pancreatic cancer ceil line. Methods CDC25B1 was transfected into Patu8988 through liposome-mediated approach. In order to detect the success of transfection, CDC25B1 mRNA expression level in cell line was assayed by reverse transcript polymerase chain reaction (RT-PCR) , and CDC25B protein expression level was assayed by Western blot. The MTT assay, invasion assay and flow cytometry were then carried out to investigate the effect of CDC25B1 on the biological behaviors of celt line Patu8988. Result High level of CDC25B1 mRNA and protvin expression were found in the group transfected with pcDNA-CDC25B1, while only weak mRNA and protein expression of CDC25B1 were found in the groups of non-treated and transfected with plasmid pcDNA. MTT assay showed that growth rate was significantly inhibited at 72h in group transfected with pcDNA-CDC25B1 comparing with that of the other two groups (P 〈 0.05 ). However, transfected cells showed significantly stronger invasive capability comparing with that of the other two groups ( P 〈0.05 ). The flow cytometry revealed transfected cells had a higher ratio in the G1 stage comparing with the groups of the non-treated and transfected with plasmid pcDNA ( P 〈0.05 ) and also had a lower ratio in the G2 stage. Conclusion In the human pancreatic cancer eell line Patu8988, the mRNA expression level of CDC25B1 and the protein expression level of CDC25B are weak. The cells are retained in the G1 stage and the capability of proliferation is inhibited by the CDC25B1. On the other hand, transfected pcDNA-CDC25B1 promotes the capability of the invasion in Patu8988. There might be some other ways that CDC25B1 participates in controlling the biological behaviors of pancreatic cancer cell line.
关 键 词:CDC25B基因 基因转染 胰腺癌细胞株Patu8988
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