Infection by Cx43 adenovirus increased chemotherapy sensitivity in human gastric cancer BGC-823 cells: not involving in induction of cell apoptosis
Infection by Cx43 adenovirus increased chemotherapy sensitivity in human gastric cancer BGC-823 cells: not involving in induction of cell apoptosis
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DOI：
10.1016/j.gene.2015.08.052
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发表时间：
2015-12-15
期刊：
影响因子：
3.5
通讯作者：
Zhang, Xufeng
中科院分区：
文献类型：
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作者：
Liu, Dan;Zhou, Hongfeng;Zhang, Xufeng
背景与目的：人胃癌BGC-823细胞中Connexin43（Cx43）的基础表达较低。本研究将重组Cx43腺病毒质粒载体转染BGC-823细胞，探讨Cx43表达对细胞增殖、化疗敏感性、集落形成能力、侵袭能力和凋亡的影响。此外，我们还检测了Pgp、Cx43以及凋亡相关蛋白（bcl-2、box、caspase3和caspase 9）的表达。方法：采用MTT法检测Cx43转染前后BGC-823细胞的增殖情况。采用MTT法检测Cx43感染对化疗（包括阿霉素、氟尿嘧啶、奥沙利铂）敏感性的影响。采用Western blotting分析Cx43腺病毒感染前后BGC-823细胞中Pgp、Cx43以及凋亡相关蛋白（bcl-2、bax、caspase-3和caspase-9）的表达水平。通过RT-PCR测定Cx43感染前后的MDR表达。通过侵袭室检测侵袭能力。采用流式细胞术检测Cx43腺病毒感染对BGC-823细胞凋亡的影响。结果：Cx43腺病毒感染后，BGC-823细胞集落形成率和侵袭能力下降。流式细胞术结果显示细胞凋亡无显着增加。 MTT检测数据显示，感染Cx43腺病毒后，细胞增殖能力下降，对化疗药物（包括阿霉素、氟尿嘧啶、奥沙利铂）的敏感性增加。 Western blotting分析结果显示，Cx43表达水平升高，Pgp表达水平降低，bc1-2、bax、caspase3和caspase 9变化不显着。 RT-PCR显示编码Pgp的基因MDR1基因的表达显着下降(p < 0.05)。结论：Cx43-IRES2-EGFP重组腺病毒载体感染人胃癌BGC-823细胞。 Cx43 感染的 BGC-823 细胞的集落形成、侵袭能力和细胞增殖均下降，而化疗敏感性增加。 Cx43 表达的增加伴随着 Pgp 表达和 MDR1 m RNA 水平的降低。然而，凋亡相关蛋白（bc1-2、bax、caspase3 和 caspase 9）和细胞凋亡增加不显着。所有结果均表明Cx43可能负向调控胃癌的发生发展、侵袭和转移，但与肿瘤细胞凋亡无明显关系。 (C) 2015 Elsevier B.V. 保留所有权利。
Background and objective: There is a lower basal expression of Connexin43 (Cx43) in human gastric cancer BGC-823 cells. In the present study, BGC-823 cells were transfected with recombinant Cx43 adenovirus plasmid vector, and we explored the influences of Cx43 expression on cell proliferation, chemo-sensitivity, colony forming ability, invasion ability and apoptosis. Moreover, we also determined the expression of Pgp, Cx43, as well as apoptosis-related proteins (bcl-2, box, caspase3 and caspase 9).Methods: MTT assay was performed to determine the proliferation of BGC-823 cells before and after Cx43 transfection. The influences of Cx43 infection on sensitivity of chemotherapy (including Doxorubicin, fluorouracil, oxaliplatin) were detected by MTT assay. Expression levels of Pgp, Cx43, as well as apoptosis-related proteins (bcl-2, bax, caspase-3 and caspase-9) in BGC-823 cells were determined by Western blotting analysis before and after the infection with Cx43 adenovirus. MDR expression was determined by RT-PCR before and after Cx43 infection. Invasive ability was detected by invasion chamber. Influence of Cx43 adenovirus infection on apoptosis of BGC-823 cells was determined by flow cytometry.Results: After infection by Cx43 adenovirus, colony forming rate and invasive ability of BGC-823 cells were decreased. Flow cytometry results revealed that cell apoptosis were insignificantly increased. The data of MTT assay revealed that infection with Cx43 adenovirus, cell proliferation ability decreased and sensitivity to chemotherapy drugs (including doxorubicin, fluorouracil, oxaliplatin) increased. Results of Western blotting analysis revealed that increasing expression levels of Cx43, decreasing expression levels of Pgp, and insignificant changes of bc1-2, bax, caspase3 and caspase 9 were detected. RT-PCR revealed the expression of MDR1 gene, the gene encoding Pgp, decreased significantly (p < 0.05).Conclusion: The human gastric cancer BGC-823 cells were infected with Cx43-IRES2-EGFP recombinant adenovirus vector. Colony formation, invasive ability and cell proliferation all decreased, whereas chemo-sensitivity increased in Cx43 infected BGC-823 cells. The increasing Cx43 expression was accompanied by decreasing Pgp expression and MDR1 m RNA levels. However, apoptosis-related proteins (bc1-2, bax, caspase3 and caspase 9) and cell apoptosis increased insignificantly. All results demonstrated that Cx43 may be negatively regulated the development, invasion and metastasis of gastric cancers, however, it had no obvious relationship with tumor cell apoptosis. (C) 2015 Elsevier B.V. All rights reserved.