PTPN9的表达下调通过抑制STAT3活化促进结直肠癌细胞凋亡
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摘要
目的:探讨PTPN9对结直肠癌细胞生长和存活的影响及其机制。方法:建立稳定PTPN9高表达的细胞系,使用Real-timePCR检测其内源性变达,使用细胞集落实验及Caspase-3、Caspase-9,检测其细胞活力及凋亡情况。同时我们抑制了细胞中PTPN9的表达,使用实时PCR来验证敲低效率,应用100μM H_2O_2诱导凋亡模型,利用CCK-8测定来确定细胞活力,Caspase-9和Cas-pase-3测定检测其凋亡情况。最后我们应用Western blot技术,检测PTPN9抑制STAT3通路,来调控细胞凋亡。结果:PTPN9的表达在结直肠癌组织中下调。PTPN9的高表达减慢细胞生长和集落的形成,从而诱导结直肠癌细胞凋亡。相反,PTPN9低表达促进细胞生长和存活。此外,PTPN9负责调控STAT3的活化,并在结直肠癌中抑制核易位,并且通过抑制STAT3通路,来抑制PTPN9低表达对细胞凋亡的影响。结论:PTPN9在结直肠癌组织中通过抑制STAT3的活化而抑制细胞生长和存活。
Objective: To investigate the effect of Protein tyrosine phosphatase non-receptor type 9(PTPN9) on the growth and survival of colorectal cancer cells and its mechanism. Methods: A stable cell line with high expression of PTPN9 was established, the endogenous expression of PTPN9 was detected by Real-time PCR. Cell colony assay, Caspase-3 and Caspase-9 activity were used to detect cell viability and apoptosis. At the same time, we inhibited the expression of PTPN9 in the cells. Real-time PCR was used to verify knockdown efficiency. 100 μM H_2O_2-induced apoptosis model, CCK-8 assay was used to determine cell viability, Caspase-9 and Caspase-3 assay to detect the apoptosis. Finally, we applied Western blot technology to detect PTPN9 inhibition of STAT3 pathway to regulate apoptosis. Results: The expression of PTPN9 was frequently down-regulated in the tissues of colorectal cancer as compared with their adjacent normal tissues. Overexpression of PTPN9 mitigated cell growth and colony formation and induced cell apoptosis in colorectal cancer. Conversely, PTPN9 knockdown promoted cell growth and survival. Moreover, PTPN9 negatively regulated the activation of STAT3 and depressed its nuclear translocation in colorectal cancer. And the effects of PTPN9 knockdown on cell apoptosis were attenuated by inhibition of the STAT3 pathway. Conclusions: PTPN__ inhibited the growth and survival of colorectal cancer cells via repressing the activation of STAT3.
Objective: To investigate the effect of Protein tyrosine phosphatase non-receptor type 9(PTPN9) on the growth and survival of colorectal cancer cells and its mechanism. Methods: A stable cell line with high expression of PTPN9 was established, the endogenous expression of PTPN9 was detected by Real-time PCR. Cell colony assay, Caspase-3 and Caspase-9 activity were used to detect cell viability and apoptosis. At the same time, we inhibited the expression of PTPN9 in the cells. Real-time PCR was used to verify knockdown efficiency. 100 μM H_2O_2-induced apoptosis model, CCK-8 assay was used to determine cell viability, Caspase-9 and Caspase-3 assay to detect the apoptosis. Finally, we applied Western blot technology to detect PTPN9 inhibition of STAT3 pathway to regulate apoptosis. Results: The expression of PTPN9 was frequently down-regulated in the tissues of colorectal cancer as compared with their adjacent normal tissues. Overexpression of PTPN9 mitigated cell growth and colony formation and induced cell apoptosis in colorectal cancer. Conversely, PTPN9 knockdown promoted cell growth and survival. Moreover, PTPN9 negatively regulated the activation of STAT3 and depressed its nuclear translocation in colorectal cancer. And the effects of PTPN9 knockdown on cell apoptosis were attenuated by inhibition of the STAT3 pathway. Conclusions: PTPN__ inhibited the growth and survival of colorectal cancer cells via repressing the activation of STAT3.
引文
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[14]Bu Y,Su F,Wang X,et al.Protein tyrosine phosphatase PTPN9 regulates erythroid cell development through STAT3 dephosphorylation in zebrafish[J].J Cell Sci,2014,127:2761-2770
[15]Tonks NK.Protein tyrosine phosphatases:from genes,to function,to disease[J].Nat Rev Mol Cell Biol,2006,7:833-846
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[17]Hu B,Yan X,Liu F,et al.Downregulated Expression of PTPN9 Contributes to Human Hepatocellular Carcinoma Growth and Progression[J].Pathol Oncol Res,2016,22:555-565
[18]Ray S,Zhao Y,Jamaluddin M,et al.Inducible STAT3 NH2 terminal mono-ubiquitination promotes BRD4 complex formation to regulate apoptosis[J].Cell Signal,2014,26:1445-1455
[19]Wang P,Xue Y,Han Y,et al.The STAT3-binding long noncoding RNA lnc-DC controls human dendritic cell differentiation[J].Science,2014,344:310-313
[20]Inghirami G,Chiarle R,Simmons WJ,et al.New and old functions of STAT3:a pivotal target for individualized treatment of cancer[J].Cell Cycle,2005,4:1131-1133
[2]Brenner H,Kloor M,Pox CP.Colorectal cancer[J].Lancet,2014,383:1490-1502
[3]Ryoo HD,Bergmann A.The role of apoptosis-induced proliferation for regeneration and cancer[J].Cold Spring Harb Perspect Biol,2012,4:a008797
[4]Zhao S,Sedwick D,Wang Z.Genetic alterations of protein tyrosine phosphatases in human cancers[J].Oncogene,2015,34:3885-3894
[5]LabbéDP,Hardy S,Tremblay ML.Protein tyrosine phosphatases in cancer:friends and foes![J].Prog Mol Biol Transl Sci,2012,106:253-306
[6]Zhu Z,Liu Y,Li K,et al.Protein tyrosine phosphatase receptor U(PT-PRU)is required for glioma growth and motility[J].Carcinogenesis,2014,35:1901-1910
[7]Julien SG,DubéN,Read M,et al.Protein tyrosine phosphatase 1B deficiency or inhibition delays ErbB2-induced mammary tumorigenesis and protects from lung metastasis[J].Nat Genet,2007,39:338-346
[8]Sun T,Aceto N,Meerbrey KL,et al.Activation of multiple proto-oncogenic tyrosine kinases in breast cancer via loss of the PTPN12phosphatase[J].Cell,2011,144:703-718
[9]Han T,Xiang DM,Sun W,et al.PTPN11/Shp2 overexpression enhances liver cancer progression and predicts poor prognosis of patients[J].J Hepatol,2015,63:651-660
[10]Yuan T,Wang Y,Zhao ZJ,et al.Protein-tyrosine phosphatase PTPN9 negatively regulates Erb B2 and epidermal growth factor receptor signaling in breast cancer cells[J].J Biol Chem,2010,285:14861-14870
[11]Du WW,Fang L,Li M,et al.MicroRNA miR-24 enhances tumor invasion and metastasis by targeting PTPN9 and PTPRF to promote EGF signaling[J].J Cell Sci,2013,126:1440-1453
[12]Ahmad KA,Iskandar KB,Hirpara JL,et al.Hydrogen peroxide-mediated cytosolic acidification is a signal for mitochondrial translocation of Bax during drug-induced apoptosis of tumor cells[J].Cancer Res,2004,64:7867-7878
[13]Zhao LC,Li J,Liao K,et al.Evodiamine Induces Apoptosis and Inhibits Migration of HCT-116 Human Colorectal Cancer Cells[J].Int JMol Sci,2015,16:27411-27421
[14]Bu Y,Su F,Wang X,et al.Protein tyrosine phosphatase PTPN9 regulates erythroid cell development through STAT3 dephosphorylation in zebrafish[J].J Cell Sci,2014,127:2761-2770
[15]Tonks NK.Protein tyrosine phosphatases:from genes,to function,to disease[J].Nat Rev Mol Cell Biol,2006,7:833-846
[16]Den Hertog J,Ostman A,Bohmer FD.Protein tyrosine phosphatases:regulatory mechanisms[J].FEBS J,2008,275:831-847
[17]Hu B,Yan X,Liu F,et al.Downregulated Expression of PTPN9 Contributes to Human Hepatocellular Carcinoma Growth and Progression[J].Pathol Oncol Res,2016,22:555-565
[18]Ray S,Zhao Y,Jamaluddin M,et al.Inducible STAT3 NH2 terminal mono-ubiquitination promotes BRD4 complex formation to regulate apoptosis[J].Cell Signal,2014,26:1445-1455
[19]Wang P,Xue Y,Han Y,et al.The STAT3-binding long noncoding RNA lnc-DC controls human dendritic cell differentiation[J].Science,2014,344:310-313
[20]Inghirami G,Chiarle R,Simmons WJ,et al.New and old functions of STAT3:a pivotal target for individualized treatment of cancer[J].Cell Cycle,2005,4:1131-1133