甲状腺乳头状癌肿瘤干细胞增殖、凋亡、侵袭与miR-93的影响
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摘要
背景:前期研究发现,mi R-93作为保护因子而非致病因子参与甲状腺腺瘤及其应激胰岛素抵抗的发生和发展。目的:研究mi R-93对甲状腺乳头状癌肿瘤干细胞增殖、凋亡及侵袭能力的影响。方法:采用流式细胞技术从人甲状腺乳头状癌细胞TPC-1中分选CD44+CD24-肿瘤干细胞与非CD44+CD24-细胞,检测两种细胞中mi R-93基因的表达。取CD44+CD24-肿瘤干细胞,分别转染mi R-93模拟物与mi RNA模拟物对照,采用CCK-8法、流式细胞技术、Transwell小室检测细胞增殖、凋亡、侵袭能力,采用Western Blot检测细胞增殖蛋白Ki67、凋亡蛋白Bax及Bcl-2、侵袭标志蛋白MMP-2及MMP-9的表达。结果与结论:(1)CD44+CD24-细胞中的mi R-93表达明显低于非CD44+CD24-细胞(P<0.05);(2)转染mi R-93模拟物的CD44+CD24-肿瘤干细胞的增殖速度明显慢于转染mi RNA模拟物对照组(P<0.05),细胞增殖蛋白Ki67表达明显低于转染mi RNA模拟物对照组(P<0.05);(3)转染mi R-93模拟物的CD44+CD24-肿瘤干细胞的凋亡率明显高于转染mi RNA模拟物对照组(P<0.05),促凋亡蛋白Bax表达高于转染mi RNA模拟物对照组(P<0.05),抗凋亡蛋白Bcl-2表达低于转染mi RNA模拟物对照组(P<0.05);(4)转染mi R-93模拟物的CD44+CD24-肿瘤干细胞的侵袭能力、侵袭标志蛋白MMP-2及MMP-9表达均明显低于转染mi RNA模拟物对照组(P<0.05);(5)结果表明,mi R-93可抑制甲状腺乳头状癌肿瘤干细胞的增殖及侵袭能力,促进其凋亡。
BACKGROUND: Previous studies have found that mi R-93 acts as a protective factor rather than a virulence factor involved in the occurrence and development of thyroid adenomas and stress insulin resistance. OBJECTIVE: To study the effect of mi R-93 on the proliferation, apoptosis and invasion of tumor stem cells in thyroid papillary carcinoma. METHODS: Flow cytometry was used to sort CD44+CD24-cells(cancer stem cells) and CD44+CD24-cells in human thyroid cancer cells TPC-1. The expression levels of mi R-93 gene were detected in these two kinds of cells. CD44+CD24-cancer stem cells were transfected with mi R-93 mimics and mi RNA mimics. Then, cell proliferation, apoptosis and invasion were detected through cell counting kit-8, flow cytometry, Transwell assay, respectively. Western blot assay was also used to measure expression of Ki67, Bax, Bcl-2, matrix metalloproteinases 2 and 9. RESULTS AND CONCLUSION: The expression of mi R-93 in CD44+CD24-cells was significantly lower than that in non-CD44+CD24-cells(P < 0.05). CD44+CD24-tumor stem cells transfected with mi R-93 mimics exhibited lower proliferation(P < 0.05) and lower Ki67 expression than those transfected with mi RNA mimics(P < 0.05). Compared with the mi RNA mimics group, higher apoptotic rate, higher Bax expression and lower Bcl-2 expression were detected in the mi R-93 mimics group(P < 0.05). Cell invasion ability and expression levels of matrix metalloproteinases 2 and 9 were also lower in the CD44+CD24-cancer stem cells transfected with mi R-93 mimics than those transfected with mi RNA mimics(P < 0.05). Our experimental findings suggest that mi R-93 can inhibit the proliferation and invasion of tumor stem cells in thyroid papillary carcinoma and promote cell apoptosis.
BACKGROUND: Previous studies have found that mi R-93 acts as a protective factor rather than a virulence factor involved in the occurrence and development of thyroid adenomas and stress insulin resistance. OBJECTIVE: To study the effect of mi R-93 on the proliferation, apoptosis and invasion of tumor stem cells in thyroid papillary carcinoma. METHODS: Flow cytometry was used to sort CD44+CD24-cells(cancer stem cells) and CD44+CD24-cells in human thyroid cancer cells TPC-1. The expression levels of mi R-93 gene were detected in these two kinds of cells. CD44+CD24-cancer stem cells were transfected with mi R-93 mimics and mi RNA mimics. Then, cell proliferation, apoptosis and invasion were detected through cell counting kit-8, flow cytometry, Transwell assay, respectively. Western blot assay was also used to measure expression of Ki67, Bax, Bcl-2, matrix metalloproteinases 2 and 9. RESULTS AND CONCLUSION: The expression of mi R-93 in CD44+CD24-cells was significantly lower than that in non-CD44+CD24-cells(P < 0.05). CD44+CD24-tumor stem cells transfected with mi R-93 mimics exhibited lower proliferation(P < 0.05) and lower Ki67 expression than those transfected with mi RNA mimics(P < 0.05). Compared with the mi RNA mimics group, higher apoptotic rate, higher Bax expression and lower Bcl-2 expression were detected in the mi R-93 mimics group(P < 0.05). Cell invasion ability and expression levels of matrix metalloproteinases 2 and 9 were also lower in the CD44+CD24-cancer stem cells transfected with mi R-93 mimics than those transfected with mi RNA mimics(P < 0.05). Our experimental findings suggest that mi R-93 can inhibit the proliferation and invasion of tumor stem cells in thyroid papillary carcinoma and promote cell apoptosis.
引文
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[13]陈鑫,韩芦芦,姜忠,等.mi R-93对神经母细胞瘤细胞侵袭和迁移的影响及机制研究[J].中华小儿外科杂志,2015,36(12):930-935.
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[20]李峻,陈劼,孙雪梅,等.重型再障患者中医证型与外周血micro RNA、淋巴细胞亚群及细胞因子的相关性研究[J].江苏中医药,2017,49(9):25-27,30.
[21]Zhang YT.Effects of micro RNA-129 and its target gene c-Fos on proliferation and apoptosis of hippocampal neurons in rats with epilepsy via the MAPK signaling pathway.J Cell Physiol.2017 Nov 30.doi:10.1002/jcp.26297.[Epub ahead of print]
[22]Raghuwanshi S,Gutti U,Kandi R,et al.Micro RNA-9promotes cell proliferation by regulating RUNX1 expression in human megakaryocyte development.Cell Prolif.2018;51(1):e12414.
[23]Lin H,Ewing LE,Koturbash I,et al.Micro RNAs as biomarkers for liver injury:Current knowledge,challenges and future prospects.Food Chem Toxicol.2017;110:229-239.
[24]宋军营,李晔,白洁.micro RNAs在糖代谢和脂代谢中的作用[J].中国老年学杂志,2013,33(21):5488-5490.
[25]乔德辉.Micro RNA与甲状腺乳头状癌发生、发展的研究进展[J].重庆医学,2017,46(21):2995-2998.
[26]姜兴莲,张徽,陈晏林,等.micro RNA-221及白细胞介素-17在甲状腺乳头状癌中的表达及相关性[J].中华病理学杂志,2017,46(3):160-165.
[27]刘洋,卢秀波,耿祖仕,等.微小RNA-21在不同甲状腺组织中的表达及其与乳头状癌临床特征间的关系[J].中华实验外科杂志,2013,30(11):2391-2393.
[28]卢冠铭,潘运龙,李震东,等.mi R-93表达与甲状腺腺瘤手术应激胰岛素抵抗患者的关联性[J].上海交通大学学报:医学版,2014,34(12):1749-1752.
[2]Zhang L,Guo X,Zhang L,et al.SLC34A2 regulates mi R-25-Gsk3βsignaling pathway to affect tumor progression in gastric cancer stem cell-like cells.Mol Carcinog.2018;57(3):440-450.
[3]李远强,张坤,孙丽丹.肿瘤干细胞靶向给药系统的研究进展[J].海南医学,2017,28(2):260-262.
[4]Li Y,Chen T,Zhu J,et al.High ALDH activity defines ovarian cancer stem-like cells with enhanced invasiveness and EMT progress which are responsible for tumor invasion.Biochem Biophys Res Commun.2018;495(1):1081-1088.
[5]王凌翔,吴正春,文宇,等.肿瘤干细胞的光动力治疗研究进展[J].激光生物学报,2017,26(1):1-8.
[6]曹颖,傅士龙.宫颈癌与肿瘤干细胞的研究进展[J].江苏医药,2017,43(5):356-359.
[7]邢越.甲状腺癌干细胞研究进展[J].岭南现代临床外科,2013,13(5):458-460.
[8]李景,付焱,肖光雄,等.乳腺癌干细胞向血管内皮细胞分化及血管形成的实验研究[J].西安交通大学学报:医学版,2017,38(2):251-256,265.
[9]郑仁东,刘超.甲状腺肿瘤干细胞的研究进展[J].国际内分泌代谢杂志,2015,35(4):261-263.
[10]Guanzon D,Iljas JD,Rice GE,et al.Using a Next-Generation Sequencing Approach to Profile Micro RNAs from Human Origin.Methods Mol Biol.2018;1710:203-217.
[11]Mellis D,Caporali A.Micro RNA-based therapeutics in cardiovascular disease:screening and delivery to the target.Biochem Soc Trans.2018;46(1):11-21.
[12]杨帆,王微,袁璐,等.micro RNA-93过表达促进A172脑胶质瘤细胞增殖并抑制其凋亡[J].细胞与分子免疫学杂志,2014,30(4):342-345,350.
[13]陈鑫,韩芦芦,姜忠,等.mi R-93对神经母细胞瘤细胞侵袭和迁移的影响及机制研究[J].中华小儿外科杂志,2015,36(12):930-935.
[14]Xue Y,Lv J,Xu P,et al.Identification of micro RNAs and genes associated with hyperandrogenism in the follicular fluid of women with polycystic ovary syndrome.J Cell Biochem.2017 Nov 28.doi:10.1002/jcb.26531.[Epub ahead of print]
[15]Paszek S,Gab?o N,Barna?E,et al.Dysregulation of micro RNAs in triple-negative breast cancer.Ginekol Pol.2017;88(10):530-536.
[16]许坚吉,王爽,寇卜心,等.肿瘤基因治疗的研究进展[J].中华全科医学,2017,15(4):655-658.
[17]张积太.IGF-1甲基化和胎盘micro RNA表达与巨大儿发生的关联性研究[D].温州:温州医科大学,2015.
[18]许婷,肖国宏,杨洁.Micro RNA在卵泡生长发育中的研究进展[J].实用医学杂志,2015,31(2):319-321.
[19]孙晨光,陈明,阮峥,等.利用斑马鱼模型研究mi R-191对髓系造血分化的调控[J].中国实验血液学杂志,2017,25(4):1172-1177.
[20]李峻,陈劼,孙雪梅,等.重型再障患者中医证型与外周血micro RNA、淋巴细胞亚群及细胞因子的相关性研究[J].江苏中医药,2017,49(9):25-27,30.
[21]Zhang YT.Effects of micro RNA-129 and its target gene c-Fos on proliferation and apoptosis of hippocampal neurons in rats with epilepsy via the MAPK signaling pathway.J Cell Physiol.2017 Nov 30.doi:10.1002/jcp.26297.[Epub ahead of print]
[22]Raghuwanshi S,Gutti U,Kandi R,et al.Micro RNA-9promotes cell proliferation by regulating RUNX1 expression in human megakaryocyte development.Cell Prolif.2018;51(1):e12414.
[23]Lin H,Ewing LE,Koturbash I,et al.Micro RNAs as biomarkers for liver injury:Current knowledge,challenges and future prospects.Food Chem Toxicol.2017;110:229-239.
[24]宋军营,李晔,白洁.micro RNAs在糖代谢和脂代谢中的作用[J].中国老年学杂志,2013,33(21):5488-5490.
[25]乔德辉.Micro RNA与甲状腺乳头状癌发生、发展的研究进展[J].重庆医学,2017,46(21):2995-2998.
[26]姜兴莲,张徽,陈晏林,等.micro RNA-221及白细胞介素-17在甲状腺乳头状癌中的表达及相关性[J].中华病理学杂志,2017,46(3):160-165.
[27]刘洋,卢秀波,耿祖仕,等.微小RNA-21在不同甲状腺组织中的表达及其与乳头状癌临床特征间的关系[J].中华实验外科杂志,2013,30(11):2391-2393.
[28]卢冠铭,潘运龙,李震东,等.mi R-93表达与甲状腺腺瘤手术应激胰岛素抵抗患者的关联性[J].上海交通大学学报:医学版,2014,34(12):1749-1752.