Pten基因沉默可抑制子宫内膜细胞增殖
|
摘要
本研究将Pten-siRNA转染至子宫内膜细胞,通过蛋白免疫印迹检测子宫内膜细胞中Pten蛋白的表达情况;通过MTT法检测Pten-siRNA转染组和空质粒转染组(对照组)细胞增殖状态;通过蛋白免疫印迹检测Pten-siRNA转染组和对照组子宫内膜细胞中PCNA、Ki67蛋白表达情况;通过流式细胞仪检测子宫内膜细胞Pten-siRNA转染组和对照组中ROS的水平,以探究Pten基因对子宫内膜癌发生发展的影响。研究表明,Pten-siRNA转染后,子宫内膜细胞中Pten蛋白明显低于对照组;Pten-siRNA转染后,子宫内膜细胞增殖率显著高于对照组(p<0.05);Pten-siRNA转染后,细胞增殖相关蛋白PCNA和Ki67明显高于对照组(p<0.05);Pten-siRNA转染后,子宫内膜细胞中ROS水平明显低于对照组(p<0.05)。本研究初步结论显示Pten-siRNA可降低子宫内膜细胞中Pten蛋白表达,抑制子宫内膜细胞的增殖并增加子宫内膜细胞中ROS水平。
In this research, we transfected Pten-siRNA into endometrial cells, and detected the expression situation of Pten protein in endometrial cells by Western blotting, as well as detected the cell proliferation of endometrial cells in Pten-siRNA-transfected group(Pten-siRNA group) and Vector-NC group(control group) by MTT assay. Furthermore, Western blotting was used to detect the expression situations of PCNA and Ki67 protein in endometrial cells of Pten-siRNA group and Vector-NC group; This study detected the reactive oxygen species(ROS) level of endometrial cells in Pten-siRNA group and Vector-NC group by flow cytometry(FCM) as well, in order to explore the effects of Pten gene on the occurrence and development of endometrial cancer through the study of human endometrial cells. The results showed that after Pten-siRNA transfection, the Pten protein was significantly lower than the control group(p<0.05) and the cell proliferation of endometrial cells was higher than the control group(p<0.05). Meanwhile, the expression of cell proliferation-associated proteins PCNA and Ki67 were significantly higher than control group(p<0.05), and the level of reactive oxygen species(ROS) in endometrial cells was lower than control group(p<0.05). The preliminary conclusions of this study showed that Pten-siRNA could significantly decrease the expression of Pten protein in endometrial cells, inhibit the proliferation of endometrial cells, and improve the ROS level in endometrial cells.
In this research, we transfected Pten-siRNA into endometrial cells, and detected the expression situation of Pten protein in endometrial cells by Western blotting, as well as detected the cell proliferation of endometrial cells in Pten-siRNA-transfected group(Pten-siRNA group) and Vector-NC group(control group) by MTT assay. Furthermore, Western blotting was used to detect the expression situations of PCNA and Ki67 protein in endometrial cells of Pten-siRNA group and Vector-NC group; This study detected the reactive oxygen species(ROS) level of endometrial cells in Pten-siRNA group and Vector-NC group by flow cytometry(FCM) as well, in order to explore the effects of Pten gene on the occurrence and development of endometrial cancer through the study of human endometrial cells. The results showed that after Pten-siRNA transfection, the Pten protein was significantly lower than the control group(p<0.05) and the cell proliferation of endometrial cells was higher than the control group(p<0.05). Meanwhile, the expression of cell proliferation-associated proteins PCNA and Ki67 were significantly higher than control group(p<0.05), and the level of reactive oxygen species(ROS) in endometrial cells was lower than control group(p<0.05). The preliminary conclusions of this study showed that Pten-siRNA could significantly decrease the expression of Pten protein in endometrial cells, inhibit the proliferation of endometrial cells, and improve the ROS level in endometrial cells.
引文
Carnero A.,Blanco-Aparicio C.,Renner O.,Link W.,and Leal J.F.,2008,The PTEN/PI3K/AKT signalling pathway in cancer,therapeutic implications,Curr.Cancer Drug Targets,8(3):187-198
de Araujo W.M.,Robbs B.K.,Bastos L.G.,de Souza W.F.,Vidal F.C.,Viola J.P.,and Morgado-Diaz J.A.,2016,PTEN overexpression cooperates with lithium to reduce the malignancy and to increase cell death by apoptosis via PI3K/Akt suppression in colorectal cancer cells,J.Cell Biochem.,117(2):458-469
Duchrow M.,Schluter C.,Key G.,Zámborszky J.,and Szüts D.,1995,Cell proliferation-associated nuclear antigen defined by antibody Ki-67:a new kind of cell cycle-maintaining proteins,Arch.Immunol.Ther.Exp.(Warsz),43(2):117-121
Garg K.,and Soslow R.A.,2014,Endometrial carcinoma in women aged 40 years and younger,Arch.Pathol.Lab.Med.,138(3):335-342
Gervai J.Z.,Galicza J.,Szeltner Z.,Zámborszky J.,and Szütsa D.,2017,A genetic study based on PCNA-ubiquitin fusions reveals no requirement for PCNA polyubiquitylation in DNAdamage tolerance,DNA Repair(Amst),54:46-54
Hollander M.C.,Blumenthal G.M.,and Dennis P.A.,2011,PTENloss in the continuum of common cancers,rare syndromes and mouse models,Nat.Rev.Cancer,11(4):289-301
Hrycay E.G.,and Bandiera S.M.,2015,Involvement of cytochrome P450 in reactive oxygen species formation and cancer,Adv.Pharmacol.,74:35-84
Khalid A.,Hussain T.,Manzoor S.,Manzoor S.,Saalim M.,and Khaliq S.,2017,PTEN:a potential prognostic marker in virus-induced hepatocellular carcinoma,Tumour Biol.,39(6):1010428317705754
Kitson S.,Ryan N.,and MacKintosh M.L.,2018,Interventions for weight reduction in obesity to improve survival in women with endometrial cancer,Cochrane Database Syst.Rev.,2:Cd012513
Li J.,Yen C.,Liaw D.,Podsypanina K.,Bose S.,Wang S.I.,Puc J.,Miliaresis C.,Rodgers L.,McCombie R.,Bigner S.H.,Giovanella B.C.,Ittmann M.,Tycko B.,Hibshoosh H.,Wigler M.H.,and Parsons R.,1997,PTEN,a putative protein tyrosine phosphatase gene mutated in human brain,breast,and prostate cancer,Science,275(5308):1943-1947
Marinescu A.,Stepan A.E.,Margaritescu C.,Marinescu A.M.,Z觍voi R.E.,Simionescu C.E.,and Niculescu M.,2016,P53,p16 and Ki67 immunoexpression in cutaneous squamous cell carcinoma and its precursor lesions,Rom.J.Morphol.Embryol.,57(2):691-696
McAlpine J.,Leon-Castillo A.,and Bosse T.,2018,The rise of a novel classification system for endometrial carcinoma,Integration of Molecular Subclasses,244(5):538-549
Reczek C.R.,and Chandel N.S.,2018,ROS promotes cancer cell survival through calcium signaling,Cancer Cell,33(6):949-951
Rhei E.,Kang L.,Bogomolniy F.,Federici M.G.,Borgen P.I.,and Boyd J.,1997,Mutation analysis of the putative tumor suppressor gene PTEN/MMAC1 in primary breast carcinomas,Cancer Res.,57(17):3657-3659
Teng D.H.,Hu R.,Lin H.,Iliev D.,Frye C.,Swedlund B.,Hansen K.L.,Vinson V.L.,Gumpper K.L.,Ellis L.,El-Naggar A.,Frazier M.,Jasser S.,Langford L.A.,Lee J.,Mills G.B.,Pershouse M.A.,Pollack R.E.,Tornos C.,Troncoso P.,Yung W.K.,Fujii G.,Berson A.,and Steck P.A.,1997,MMAC1/PTEN mutations in primary tumor specimens and tumor cell lines,Cancer Res.,57(23):5221-5225
Wright J.D.,Ruiz M.P.,and Chen L.,2018,Changes in surgical volume and outcomes over time for women undergoing hysterectomy for endometrial cancer,Obstet.Gynecol,1(3):235-239
Xu W.,Yang Z.,Zhou S.F.,Ling H.,Li Z.,Li M.,and Hu D.,2014,Posttranslational regulation of phosphatase and tensin homolog(PTEN)and its functional impact on cancer behaviors,Drug Des.Devel.Ther.,8:1745-1751
Zhang X.,Yin B.,Zhu F.,Huang G.,and Li H.A.,2015,PTENtranslational isoform has PTEN-like activity,Chin.J.Cancer Res.,27(5):524-532
Zhu Y.,Matsumura Y.,Velayutham M.,Foley L.M.,Hitchens T.K.,and Wagner W.R.,2018,Reactive oxygen species scavenging with a biodegradable,thermally responsive hydrogel compatible with soft tissue injection,Biomaterials,177:98-112
de Araujo W.M.,Robbs B.K.,Bastos L.G.,de Souza W.F.,Vidal F.C.,Viola J.P.,and Morgado-Diaz J.A.,2016,PTEN overexpression cooperates with lithium to reduce the malignancy and to increase cell death by apoptosis via PI3K/Akt suppression in colorectal cancer cells,J.Cell Biochem.,117(2):458-469
Duchrow M.,Schluter C.,Key G.,Zámborszky J.,and Szüts D.,1995,Cell proliferation-associated nuclear antigen defined by antibody Ki-67:a new kind of cell cycle-maintaining proteins,Arch.Immunol.Ther.Exp.(Warsz),43(2):117-121
Garg K.,and Soslow R.A.,2014,Endometrial carcinoma in women aged 40 years and younger,Arch.Pathol.Lab.Med.,138(3):335-342
Gervai J.Z.,Galicza J.,Szeltner Z.,Zámborszky J.,and Szütsa D.,2017,A genetic study based on PCNA-ubiquitin fusions reveals no requirement for PCNA polyubiquitylation in DNAdamage tolerance,DNA Repair(Amst),54:46-54
Hollander M.C.,Blumenthal G.M.,and Dennis P.A.,2011,PTENloss in the continuum of common cancers,rare syndromes and mouse models,Nat.Rev.Cancer,11(4):289-301
Hrycay E.G.,and Bandiera S.M.,2015,Involvement of cytochrome P450 in reactive oxygen species formation and cancer,Adv.Pharmacol.,74:35-84
Khalid A.,Hussain T.,Manzoor S.,Manzoor S.,Saalim M.,and Khaliq S.,2017,PTEN:a potential prognostic marker in virus-induced hepatocellular carcinoma,Tumour Biol.,39(6):1010428317705754
Kitson S.,Ryan N.,and MacKintosh M.L.,2018,Interventions for weight reduction in obesity to improve survival in women with endometrial cancer,Cochrane Database Syst.Rev.,2:Cd012513
Li J.,Yen C.,Liaw D.,Podsypanina K.,Bose S.,Wang S.I.,Puc J.,Miliaresis C.,Rodgers L.,McCombie R.,Bigner S.H.,Giovanella B.C.,Ittmann M.,Tycko B.,Hibshoosh H.,Wigler M.H.,and Parsons R.,1997,PTEN,a putative protein tyrosine phosphatase gene mutated in human brain,breast,and prostate cancer,Science,275(5308):1943-1947
Marinescu A.,Stepan A.E.,Margaritescu C.,Marinescu A.M.,Z觍voi R.E.,Simionescu C.E.,and Niculescu M.,2016,P53,p16 and Ki67 immunoexpression in cutaneous squamous cell carcinoma and its precursor lesions,Rom.J.Morphol.Embryol.,57(2):691-696
McAlpine J.,Leon-Castillo A.,and Bosse T.,2018,The rise of a novel classification system for endometrial carcinoma,Integration of Molecular Subclasses,244(5):538-549
Reczek C.R.,and Chandel N.S.,2018,ROS promotes cancer cell survival through calcium signaling,Cancer Cell,33(6):949-951
Rhei E.,Kang L.,Bogomolniy F.,Federici M.G.,Borgen P.I.,and Boyd J.,1997,Mutation analysis of the putative tumor suppressor gene PTEN/MMAC1 in primary breast carcinomas,Cancer Res.,57(17):3657-3659
Teng D.H.,Hu R.,Lin H.,Iliev D.,Frye C.,Swedlund B.,Hansen K.L.,Vinson V.L.,Gumpper K.L.,Ellis L.,El-Naggar A.,Frazier M.,Jasser S.,Langford L.A.,Lee J.,Mills G.B.,Pershouse M.A.,Pollack R.E.,Tornos C.,Troncoso P.,Yung W.K.,Fujii G.,Berson A.,and Steck P.A.,1997,MMAC1/PTEN mutations in primary tumor specimens and tumor cell lines,Cancer Res.,57(23):5221-5225
Wright J.D.,Ruiz M.P.,and Chen L.,2018,Changes in surgical volume and outcomes over time for women undergoing hysterectomy for endometrial cancer,Obstet.Gynecol,1(3):235-239
Xu W.,Yang Z.,Zhou S.F.,Ling H.,Li Z.,Li M.,and Hu D.,2014,Posttranslational regulation of phosphatase and tensin homolog(PTEN)and its functional impact on cancer behaviors,Drug Des.Devel.Ther.,8:1745-1751
Zhang X.,Yin B.,Zhu F.,Huang G.,and Li H.A.,2015,PTENtranslational isoform has PTEN-like activity,Chin.J.Cancer Res.,27(5):524-532
Zhu Y.,Matsumura Y.,Velayutham M.,Foley L.M.,Hitchens T.K.,and Wagner W.R.,2018,Reactive oxygen species scavenging with a biodegradable,thermally responsive hydrogel compatible with soft tissue injection,Biomaterials,177:98-112
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |