MicroRNA-188在胃癌中的表达及对胃癌细胞增殖和凋亡的影响
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摘要
目的:探究microRNA-188在胃癌患者的组织和血液中的表达情况,并研究microRNA-188对人胃癌细胞MGC-803增殖和凋亡的影响及其机制。方法:收集10例胃癌患者的肿瘤组织和癌旁组织,应用PCR方法检测microRNA-545、microRNA-575、microRNA-1269、microRNA-99a、microRNA-452、microRNA-188的表达水平;收集健康志愿者和胃癌患者的血液,应用PCR检测这6种microRNAs的表达水平;应用转染试剂将microRNA-188 mimics、microRNA-188 AMO及其NC转染人胃癌细胞MGC-803, 24 h后应用CCK-8实验和TUNEL染色检测细胞增殖能力和凋亡情况;应用在线网站,预测microRNA-188的靶基因;应用q-PCR和Western blot实验检测microRNA-188对SIX1是否具有调控作用。结果:与癌旁组织相比, microRNA-188在胃癌组织中表达显著降低。与健康人相比,microRNA-188在胃癌患者的血液中显著降低。CCK-8结果显示,microRNA-188 mimics显著抑制胃癌细胞的增殖,而microRNA-188 AMO能够显著提高胃癌细胞的增殖能力。TUNEL染色结果显示,microRNA-188能够促进胃癌细胞发生凋亡,而其AMO能够显著抑制胃癌细胞发生凋亡。q-PCR和Western blot实验结果表明,microRNA-188能够显著降低其靶点SIX1的mRNA和蛋白水平。结论:MicroRNA-188在胃癌患者的组织和血液中的表达水平显著降低,且其通过抑制SIX1发挥调控胃癌细胞增殖和凋亡的作用。
Objective: To investigate the expression of microRNA-188 in tissues and blood of gastric cancer patients, and to study the effect of microRNA-188 on the proliferation and apoptosis of human gastric cancer cell line MGC-803 and its mechanism. Methods: Gastric cancer tissues and adjacent tissues were collected from 10 patients with gastric cancer. Quantitative real time PCR(qPCR) was used to detect the expression levels of microRNA-545, microRNA-575, microRNA-1269, microRNA-99 a, microRNA-452 and microRNA-188. The levels of six microRNAs in blood samples were detected by qPCR in healthy volunteers and patients with gastric cancer. The microRNA-188 mimics, microRNA-188 AMO and their NCs were transfected into human gastric cancer cells MGC-803 using transfection reagent. After 24 h, CCK-8 assay and TUNEL staining were used to detect the cell proliferation and apoptosis. The website online was used to predict the target gene of microRNA-188. qRT-PCR and western blot assays were used to determine whether microRNA-188 had a regulatory effect on SIX1. Results: Compared with adjacent tissues, microRNA-188 was found to markedly decrease in gastric cancer tissues. Compared with healthy volunteers, microRNA-188 was significantly lower in the blood samples of gastric cancer patients. CCK-8 results showed that microRNA-188 mimics significantly inhibited the proliferation of gastric cancer cells, while microRNA-188 AMO could significantly increase the proliferation of gastric cancer cells. TUNEL staining showed that microRNA-188 can promote the apoptosis of gastric cancer cells, and its AMO can significantly inhibit the apoptosis of gastric cancer cells. qRT-PCR and western blot experiments showed that microRNA-188 can significantly reduce the mRNA and protein levels of its target SIX1. Conclusion: The expression level of microRNA-188 is significantly decreased in the tissue and blood of patients with gastric cancer and microRNA-188 plays an important role in regulating the proliferation and apoptosis of gastric cancer cells by inhibiting SIX1.
Objective: To investigate the expression of microRNA-188 in tissues and blood of gastric cancer patients, and to study the effect of microRNA-188 on the proliferation and apoptosis of human gastric cancer cell line MGC-803 and its mechanism. Methods: Gastric cancer tissues and adjacent tissues were collected from 10 patients with gastric cancer. Quantitative real time PCR(qPCR) was used to detect the expression levels of microRNA-545, microRNA-575, microRNA-1269, microRNA-99 a, microRNA-452 and microRNA-188. The levels of six microRNAs in blood samples were detected by qPCR in healthy volunteers and patients with gastric cancer. The microRNA-188 mimics, microRNA-188 AMO and their NCs were transfected into human gastric cancer cells MGC-803 using transfection reagent. After 24 h, CCK-8 assay and TUNEL staining were used to detect the cell proliferation and apoptosis. The website online was used to predict the target gene of microRNA-188. qRT-PCR and western blot assays were used to determine whether microRNA-188 had a regulatory effect on SIX1. Results: Compared with adjacent tissues, microRNA-188 was found to markedly decrease in gastric cancer tissues. Compared with healthy volunteers, microRNA-188 was significantly lower in the blood samples of gastric cancer patients. CCK-8 results showed that microRNA-188 mimics significantly inhibited the proliferation of gastric cancer cells, while microRNA-188 AMO could significantly increase the proliferation of gastric cancer cells. TUNEL staining showed that microRNA-188 can promote the apoptosis of gastric cancer cells, and its AMO can significantly inhibit the apoptosis of gastric cancer cells. qRT-PCR and western blot experiments showed that microRNA-188 can significantly reduce the mRNA and protein levels of its target SIX1. Conclusion: The expression level of microRNA-188 is significantly decreased in the tissue and blood of patients with gastric cancer and microRNA-188 plays an important role in regulating the proliferation and apoptosis of gastric cancer cells by inhibiting SIX1.
引文
1 Sammarco G,Gadaleta CD,Zuccalà V,et al.Tumor-associated macrophages and mast cells positive to tryptase are correlated with angiogenesis in surgically-treated gastric cancer patients[J].Int J Mol Sci,2018,19(4):E1176.
2 Xue BY,Lu QY,Massie L,et al.Grape seed procyanidin extract against lung cancer:the role of microrna-106b,bioavailability,and bioactivity[J].Oncotarget,2018,9(21):15579-15590.
3 Viczenczova C,Kura B,Egan Benova T,et al.Irradiation-induced cardiac connexin-43 and mir-21 responses are hampered by treatment with atorvastatin and aspirin[J].Int J Mol Sci,2018,19(4):E1128.
4 Ma M,Zhao JX,Wu QF,et al.MiRNA-545 negatively regulates the oncogenic activity of EMS1 in gastric cancer[J].Cancer Med,2018,7(6):2452-2462.
5 Wang HW,Yan CH,Shi XD,et al.MicroRNA-575 targets BLID to promote growth and invasion of non-small cell lung cancer cells[J].FEBS Lett,2015,589(7):805-811.
6 Bao M,Song YJ,Xia JJ,et al.MiR-1269 promotes cell survival and proliferation by targeting tp53 and caspase-9 in lung cancer[J].Onco Targets Ther,2018,11:1721-1732.
7 Yin H,Ma J,Chen L,Piao S,Zhang Y,et al.MiR-99a Enhances the Radiation Sensitivity of Non-Small Cell Lung Cancer by Targeting mTOR[J].Cell Physiol Biochem,2018,46:471-481.
8 Li HY,Wu Y,Li PX.MicroRNA-452 suppresses pancreatic cancer migration and invasion by directly targeting B-cell-specific Moloney murine leukemia virus insertion site 1[J].Oncol Lett,2017,14(3):3235-3242.
9 Wu JB,Lv Q,He J,et al.MicroRNA-188 suppresses G1/S transition by targeting multiple cyclin/CDK complexes[J].Cell Commun Signal,2014,12:66.
10 Wang H,Chen SH,Kong P,et al.Increased expression of miR-330-3p:a novel independent indicator of poor prognosis in human breast cancer[J].Eur Rev Med Pharmacol Sci,2018,22(6):1726-1730.
11 Zheng ZX,Bao F,Chen XH,et al.MicroRNA-330-3p expression indicates good prognosis and suppresses cell proliferation by targeting bmi-1 in osteosarcoma[J].Cell Physiol Biochem,2018,46(2):442-450.
12 Wei CH,Wu G,Cai Q,et al.Correction to:MicroRNA-330-3p promotes cell invasion and metastasis in non-small cell lung cancer through GRIA3 by activating MAPK/ERK signaling pathway[J].J Hematol Oncol,2018,11(1):6.
13 Meng H,Wang K,Chen XD,et al.MicroRNA-330-3p functions as an oncogene in human esophageal cancer by targeting programmed cell death 4[J].Am J Cancer Res,2015,5(3):1062-1075.
14 Medina-Villaamil V,Martínez-Breijo S,Portela-Pereira P,et al.Circulating MicroRNAs in blood of patients with prostate cancer[J].Actas Urol Esp,2014,38(10):633-639.
15 Rider CF,Yamamoto M,Günther OP,et al.Effects of controlled diesel exhaust and allergen exposure on microRNA and gene expression in humans.modulation of lung inflammatory markers associated with asthma[J].Ann Am Thorac Soc,2018,15(Supplement_2):S130-S131.
16 Li Q,Zeng FP,Lyu N,et al.Highly sensitive and specific electrochemical biosensor for microRNA-21 detection by coupling catalytic hairpin assembly with rolling circle amplification[J].Analyst,2018,143(10):2304-2309.
17 Uddin A,Chakraborty S.Role of miRNAs in lung cancer[J].J Cell Physiol,2018,20.
18 Liu XF,Xu YX,Han LP,et al.Reassessing the potential of myb-targeted anti-cancer therapy[J].J Cancer,2018,9(7):1259-1266.
19 Song B,Lin HX,Dong LL,et al.MicroRNA-338 inhibits proliferation,migration,and invasion of gastric cancer cells by the Wnt/β-catenin signaling pathway[J].Eur Rev Med Pharmacol Sci,2018,22(5):1290-1296.
20 Feng R,Beeharry MK,Lu S,et al.Down-regulated serum miR-126 is associated with aggressive progression and poor prognosis of gastric cancer[J].Cancer Biomark,2018,2 2(1):119-126.
21 Gan XN,Gan TQ,He RQ,et al.Clinical significance of high expression of miR-452-5p in lung squamous cell carcinoma[J].Oncol Lett,2018,15(5):6418-6430.
22 Zhao LL,Ni X,Zhao LL,et al.MiroRNA-188 acts as tumor suppressor in non-small-cell lung cancer by targeting MAP3K3[J].Mol Pharm,2018,15(4):1682-1689.
23 Du BW,Wang Z,Zhang X,et al.MicroRNA-545 suppresses cell proliferation by targeting cyclin D1 and CDK4 in lung cancer cells[J].PLoS One,2014,9(2):e88022.
24 Wang YJ,Liu WT,Liu YD,et al.Long noncoding RNA H19 mediates LCoR to impact the osteogenic and adipogenic differentiation of mBMSCs in mice through sponging miR-188[J].J Cell Physiol,2018,233(9):7435-7446.
25 Zheng Y,Liu H,Kong Y.MiR-188 promotes senescence of lineage-negative bone marrow cells by targeting MAP3K3 expression[J].FEBS Lett,2017,591(15):2290-2298.
26 Wang LL,Liu HC.MicroRNA-188 is downregulated in oral squamous cell carcinoma and inhibits proliferation and invasion by targeting SIX1[J].Tumour Biol,2016,37(3):4105-4113.
2 Xue BY,Lu QY,Massie L,et al.Grape seed procyanidin extract against lung cancer:the role of microrna-106b,bioavailability,and bioactivity[J].Oncotarget,2018,9(21):15579-15590.
3 Viczenczova C,Kura B,Egan Benova T,et al.Irradiation-induced cardiac connexin-43 and mir-21 responses are hampered by treatment with atorvastatin and aspirin[J].Int J Mol Sci,2018,19(4):E1128.
4 Ma M,Zhao JX,Wu QF,et al.MiRNA-545 negatively regulates the oncogenic activity of EMS1 in gastric cancer[J].Cancer Med,2018,7(6):2452-2462.
5 Wang HW,Yan CH,Shi XD,et al.MicroRNA-575 targets BLID to promote growth and invasion of non-small cell lung cancer cells[J].FEBS Lett,2015,589(7):805-811.
6 Bao M,Song YJ,Xia JJ,et al.MiR-1269 promotes cell survival and proliferation by targeting tp53 and caspase-9 in lung cancer[J].Onco Targets Ther,2018,11:1721-1732.
7 Yin H,Ma J,Chen L,Piao S,Zhang Y,et al.MiR-99a Enhances the Radiation Sensitivity of Non-Small Cell Lung Cancer by Targeting mTOR[J].Cell Physiol Biochem,2018,46:471-481.
8 Li HY,Wu Y,Li PX.MicroRNA-452 suppresses pancreatic cancer migration and invasion by directly targeting B-cell-specific Moloney murine leukemia virus insertion site 1[J].Oncol Lett,2017,14(3):3235-3242.
9 Wu JB,Lv Q,He J,et al.MicroRNA-188 suppresses G1/S transition by targeting multiple cyclin/CDK complexes[J].Cell Commun Signal,2014,12:66.
10 Wang H,Chen SH,Kong P,et al.Increased expression of miR-330-3p:a novel independent indicator of poor prognosis in human breast cancer[J].Eur Rev Med Pharmacol Sci,2018,22(6):1726-1730.
11 Zheng ZX,Bao F,Chen XH,et al.MicroRNA-330-3p expression indicates good prognosis and suppresses cell proliferation by targeting bmi-1 in osteosarcoma[J].Cell Physiol Biochem,2018,46(2):442-450.
12 Wei CH,Wu G,Cai Q,et al.Correction to:MicroRNA-330-3p promotes cell invasion and metastasis in non-small cell lung cancer through GRIA3 by activating MAPK/ERK signaling pathway[J].J Hematol Oncol,2018,11(1):6.
13 Meng H,Wang K,Chen XD,et al.MicroRNA-330-3p functions as an oncogene in human esophageal cancer by targeting programmed cell death 4[J].Am J Cancer Res,2015,5(3):1062-1075.
14 Medina-Villaamil V,Martínez-Breijo S,Portela-Pereira P,et al.Circulating MicroRNAs in blood of patients with prostate cancer[J].Actas Urol Esp,2014,38(10):633-639.
15 Rider CF,Yamamoto M,Günther OP,et al.Effects of controlled diesel exhaust and allergen exposure on microRNA and gene expression in humans.modulation of lung inflammatory markers associated with asthma[J].Ann Am Thorac Soc,2018,15(Supplement_2):S130-S131.
16 Li Q,Zeng FP,Lyu N,et al.Highly sensitive and specific electrochemical biosensor for microRNA-21 detection by coupling catalytic hairpin assembly with rolling circle amplification[J].Analyst,2018,143(10):2304-2309.
17 Uddin A,Chakraborty S.Role of miRNAs in lung cancer[J].J Cell Physiol,2018,20.
18 Liu XF,Xu YX,Han LP,et al.Reassessing the potential of myb-targeted anti-cancer therapy[J].J Cancer,2018,9(7):1259-1266.
19 Song B,Lin HX,Dong LL,et al.MicroRNA-338 inhibits proliferation,migration,and invasion of gastric cancer cells by the Wnt/β-catenin signaling pathway[J].Eur Rev Med Pharmacol Sci,2018,22(5):1290-1296.
20 Feng R,Beeharry MK,Lu S,et al.Down-regulated serum miR-126 is associated with aggressive progression and poor prognosis of gastric cancer[J].Cancer Biomark,2018,2 2(1):119-126.
21 Gan XN,Gan TQ,He RQ,et al.Clinical significance of high expression of miR-452-5p in lung squamous cell carcinoma[J].Oncol Lett,2018,15(5):6418-6430.
22 Zhao LL,Ni X,Zhao LL,et al.MiroRNA-188 acts as tumor suppressor in non-small-cell lung cancer by targeting MAP3K3[J].Mol Pharm,2018,15(4):1682-1689.
23 Du BW,Wang Z,Zhang X,et al.MicroRNA-545 suppresses cell proliferation by targeting cyclin D1 and CDK4 in lung cancer cells[J].PLoS One,2014,9(2):e88022.
24 Wang YJ,Liu WT,Liu YD,et al.Long noncoding RNA H19 mediates LCoR to impact the osteogenic and adipogenic differentiation of mBMSCs in mice through sponging miR-188[J].J Cell Physiol,2018,233(9):7435-7446.
25 Zheng Y,Liu H,Kong Y.MiR-188 promotes senescence of lineage-negative bone marrow cells by targeting MAP3K3 expression[J].FEBS Lett,2017,591(15):2290-2298.
26 Wang LL,Liu HC.MicroRNA-188 is downregulated in oral squamous cell carcinoma and inhibits proliferation and invasion by targeting SIX1[J].Tumour Biol,2016,37(3):4105-4113.