Six1基因在肺癌组织表达水平及RNA干扰抑制其表达后对肺癌细胞生物学特性的影响
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摘要
目的探讨Six1基因在肺癌组织表达及抑制其表达后对肺癌细胞增殖凋亡的影响。方法提取肺癌及癌旁组织蛋白,Western印迹检测Six1的表达; Six1-siRNA转染人肺癌A549细胞,分为空白组和阴性对照组(NC组)、Six1-siRNA组,转染48 h后通过Western印迹检测各组细胞Six1的表达; CCK8检测Six1-siRNA转染A549细胞24～72 h的细胞增殖;流式细胞术检测转染48 h的细胞凋亡率; Western印迹检测增殖相关蛋白细胞增殖核抗原(PCNA)、B细胞淋巴瘤/白血病-2(Bcl-2)家族Bcl-2相关X蛋白(Bax)及Notch1信号通路Notch1和Hes1的蛋白表达。结果 Six1在肺癌组织中的表达显著高于癌旁组织(P<0. 05); Six1-siRNA转染能明显降低A549细胞Six1的表达;与空白组比较,Six1-siRNA组细胞24～72 h的细胞增殖均显著降低,在48 h的细胞凋亡率显著升高,PCNA、Notch1和Hes1蛋白表达均显著降低,Bax蛋白表达显著升高(P<0. 05)。结论 Six1在肺癌组织中高表达,通过RNA干扰抑制其表达可通过下调Notch1信号通路降低肺癌细胞增殖及诱导细胞凋亡,对细胞增殖凋亡的影响方式是下调PCNA和上调Bax蛋白表达。
Objective To investigate Six1 gene expression in lung cancer tissues and the effect of inhibiting its expressiononon proliferation and apoptosis of lung cancer cell. Methods The proteins in lung cancer and adjacent tissues were extracted,and theexpression of Six1 was detected by Western blot; Six1-siRNA was transfected into human A549 lung cancer cells,and the blank andnegative control groups (NC group) were set,the expression of Six1 was detected by Western blot; cell proliferation was detected byCCK8; apoptosis was detected by flow cytometry; PCNA,Bcl-2,Bax and Notch1 protein expressions were detected by Western blot.Results The expression of Six1 in lung cancer tissues was significantly higher than that in paracancerous tissues (P<0.05); Six1-siR-NA transfection significantly reduced the expression of Six1 in A549 cells; compared with that of control group,cell proliferation inSix1-siRNA group was decreased significantly,cell apoptosis rate in 48 h was significantly increased,the expressions of PCNA,Notch1 and Hes1 protein were significantly decreased,and the expression of Bax protein was significantly increased (P<0.05).ConclusionsInhibiting Six1 expression could reduce lung cancer cell proliferation and induce apoptosis by downregulating the Notch1 signaling path-way. The effect on cell proliferation and apoptosis is to down-regulate PCNA and up-regulate the expression of Bax protein.
Objective To investigate Six1 gene expression in lung cancer tissues and the effect of inhibiting its expressiononon proliferation and apoptosis of lung cancer cell. Methods The proteins in lung cancer and adjacent tissues were extracted,and theexpression of Six1 was detected by Western blot; Six1-siRNA was transfected into human A549 lung cancer cells,and the blank andnegative control groups (NC group) were set,the expression of Six1 was detected by Western blot; cell proliferation was detected byCCK8; apoptosis was detected by flow cytometry; PCNA,Bcl-2,Bax and Notch1 protein expressions were detected by Western blot.Results The expression of Six1 in lung cancer tissues was significantly higher than that in paracancerous tissues (P<0.05); Six1-siR-NA transfection significantly reduced the expression of Six1 in A549 cells; compared with that of control group,cell proliferation inSix1-siRNA group was decreased significantly,cell apoptosis rate in 48 h was significantly increased,the expressions of PCNA,Notch1 and Hes1 protein were significantly decreased,and the expression of Bax protein was significantly increased (P<0.05).ConclusionsInhibiting Six1 expression could reduce lung cancer cell proliferation and induce apoptosis by downregulating the Notch1 signaling path-way. The effect on cell proliferation and apoptosis is to down-regulate PCNA and up-regulate the expression of Bax protein.
引文
1 Nie F,Sun M,Yang J,et al.Long noncoding RNA ANRIL promotes non-small cell lung cancer cell proliferation and inhibits apoptosis by silencing KLF2 and P21 expression[J].Mol Cancer Ther,2015;14(1):268-77.
2 Wang L,Liu H.microRNA-188 is downregulated in oral squamous cell carcinoma and inhibits proliferation and invasion by targeting SIX1[J].Tumor Biol,2016;37(3):4105-13.
3 Xu H,Zhang Y,Pe1a MM,et al.Six1 promotes colorectal cancer growth and metastasis by stimulating angiogenesis and recruiting tumor-associated macrophages[J].Carcinogenesis,2017;38(3):281-92.
4 Zeng J,Wei M,Shi R,et al.MiR-204-5p/Six1 feedback loop promotes epithelial-mesenchymal transition in breast cancer.[J].Tumor Biol,2016;37(2):2729-35.
5 Sun SH,Liu D,Deng YT,et al.SIX1 coordinates with TGFβsignals to induce epithelial-mesenchymal transition in cervical cancer[J].Oncol Lett,2016;12(2):1271-8.
6 Xin X,Li Y,Yang X.SIX1 is overexpressed in endometrial carcinoma and promotes the malignant behavior of cancer cells through ERK and AKT signaling[J].Oncol Lett,2016;12(5):3435-40.
7 Du P,Zhao J,Wang J,et al.Sine oculis homeobox homolog 1 regulates mitochondrial apoptosis pathway via caspase-7 in gastric cancer cells[J].J Cancer,2017;8(4):636-45.
8 Tian T,Li A,Lu H,et al.Six1 promotes glioblastoma cell proliferation and invasion by upregulation of connective tissue growth factor[J].Am J Cancer Res,2015;5(5):1823-30.
9蔡少鑫,陈炆颖,王乐,等.Six1蛋白对在结肠癌细胞株HT-29增殖能力影响的实验研究[J].福建医药杂志,2016;38(2):81-3.
10 Xu H,Zhang Y,Pirisi L,et al.Abstract 3868:Six1 overexpression promotes malignant progression in models of cervical and colon cancer[J].Cancer Res,2014;74(19 Supplement):3868.
11 Liu H,Liu F,Wei A,et al.Inhibition of Six1 promotes apoptosis,suppresses proliferation,and migration of osteosarcoma cells[J].Tumor Biol,2014;35(3):1925-31.
12李天客,陈中,包阳,等.RNA干扰SBF2对人口腔癌细胞株SCC15增殖、凋亡及侵袭的影响[J].中国老年学杂志,2016;36(21):5260-2.
13 Zhao JS,Fang MX,Guo QY,et al.Hydroxysafflor yellow A inhibits VSMCs proliferation via PCNA and MEK-ERK1/2[J].Chin Pharmacol Bull,2015;31(7):984-8.
14久太,冯喜英,安文静,等.肺腺癌中锌指E-盒结合同源异型盒蛋白-1、-2和增殖细胞核抗原表达及意义[J].中国老年学杂志,2015;35(20):5829-31.
15 Jafarinejad-Farsangi S,Farazmand A,Mahmoudi M,et al.MicroRNA-29a induces apoptosis via increasing the Bax:Bcl-2 ratio in dermal fibroblasts of patients with systemic sclerosis[J].Autoimmunity,2015;48(6):369-78.
16 Li C,Wu X,Sun R,et al.Croton tiglium extract induces apoptosis via Bax/Bcl-2 pathways in human lung cancer A549 cells[J].Asian Pac J Cancer Prev,2016;17(11):4893-8.
17 Baumgart A,Mazur PK,Anton M,et al.Opposing role of Notch1 and Notch2 in a Kras G12D-driven murine non-small cell lung cancer model[J].Oncogene,2015;34(5):578-88.
18 Song M,Yin Y,Zhang J,et al.MiR-139-5p inhibits migration and invasion of colorectal cancer by downregulating AMFR and NOTCH1[J].Protein Cell,2014;5(11):851-61.
19 Deng F,Yang ZF,Sun CQ.The role of Notch1 genes in lung cancer A594 cells and the impact on chemosensitivity[J].Eur Rev Med Pharmacol Sci,2017;21(11):2659-64.
2 Wang L,Liu H.microRNA-188 is downregulated in oral squamous cell carcinoma and inhibits proliferation and invasion by targeting SIX1[J].Tumor Biol,2016;37(3):4105-13.
3 Xu H,Zhang Y,Pe1a MM,et al.Six1 promotes colorectal cancer growth and metastasis by stimulating angiogenesis and recruiting tumor-associated macrophages[J].Carcinogenesis,2017;38(3):281-92.
4 Zeng J,Wei M,Shi R,et al.MiR-204-5p/Six1 feedback loop promotes epithelial-mesenchymal transition in breast cancer.[J].Tumor Biol,2016;37(2):2729-35.
5 Sun SH,Liu D,Deng YT,et al.SIX1 coordinates with TGFβsignals to induce epithelial-mesenchymal transition in cervical cancer[J].Oncol Lett,2016;12(2):1271-8.
6 Xin X,Li Y,Yang X.SIX1 is overexpressed in endometrial carcinoma and promotes the malignant behavior of cancer cells through ERK and AKT signaling[J].Oncol Lett,2016;12(5):3435-40.
7 Du P,Zhao J,Wang J,et al.Sine oculis homeobox homolog 1 regulates mitochondrial apoptosis pathway via caspase-7 in gastric cancer cells[J].J Cancer,2017;8(4):636-45.
8 Tian T,Li A,Lu H,et al.Six1 promotes glioblastoma cell proliferation and invasion by upregulation of connective tissue growth factor[J].Am J Cancer Res,2015;5(5):1823-30.
9蔡少鑫,陈炆颖,王乐,等.Six1蛋白对在结肠癌细胞株HT-29增殖能力影响的实验研究[J].福建医药杂志,2016;38(2):81-3.
10 Xu H,Zhang Y,Pirisi L,et al.Abstract 3868:Six1 overexpression promotes malignant progression in models of cervical and colon cancer[J].Cancer Res,2014;74(19 Supplement):3868.
11 Liu H,Liu F,Wei A,et al.Inhibition of Six1 promotes apoptosis,suppresses proliferation,and migration of osteosarcoma cells[J].Tumor Biol,2014;35(3):1925-31.
12李天客,陈中,包阳,等.RNA干扰SBF2对人口腔癌细胞株SCC15增殖、凋亡及侵袭的影响[J].中国老年学杂志,2016;36(21):5260-2.
13 Zhao JS,Fang MX,Guo QY,et al.Hydroxysafflor yellow A inhibits VSMCs proliferation via PCNA and MEK-ERK1/2[J].Chin Pharmacol Bull,2015;31(7):984-8.
14久太,冯喜英,安文静,等.肺腺癌中锌指E-盒结合同源异型盒蛋白-1、-2和增殖细胞核抗原表达及意义[J].中国老年学杂志,2015;35(20):5829-31.
15 Jafarinejad-Farsangi S,Farazmand A,Mahmoudi M,et al.MicroRNA-29a induces apoptosis via increasing the Bax:Bcl-2 ratio in dermal fibroblasts of patients with systemic sclerosis[J].Autoimmunity,2015;48(6):369-78.
16 Li C,Wu X,Sun R,et al.Croton tiglium extract induces apoptosis via Bax/Bcl-2 pathways in human lung cancer A549 cells[J].Asian Pac J Cancer Prev,2016;17(11):4893-8.
17 Baumgart A,Mazur PK,Anton M,et al.Opposing role of Notch1 and Notch2 in a Kras G12D-driven murine non-small cell lung cancer model[J].Oncogene,2015;34(5):578-88.
18 Song M,Yin Y,Zhang J,et al.MiR-139-5p inhibits migration and invasion of colorectal cancer by downregulating AMFR and NOTCH1[J].Protein Cell,2014;5(11):851-61.
19 Deng F,Yang ZF,Sun CQ.The role of Notch1 genes in lung cancer A594 cells and the impact on chemosensitivity[J].Eur Rev Med Pharmacol Sci,2017;21(11):2659-64.