Let-7g对BEL7402/5-FU细胞CRD-BP表达及5-FU敏感性的影响
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摘要
目的 :探讨微小RNA(miRNA)对肝癌耐药细胞株BEL7402/5-Fu细胞CRD-BP表达及5-FU敏感性的影响。方法 :构建let-7g表达载体,将Let-7g质粒表达载体转染至BEL7402/5-FU细胞;MTT试验及流式细胞术检测细胞生长变化;逆转录PCR(RT-PCR)及免疫印迹试验(Western-Blot)检测CRD-BP mRNA水平和相关蛋白表达水平的变化。结果 :以浓度梯度5-氟尿嘧啶(5-Fu)处理肝癌耐药细胞株BEL7402/5-FU后,与未转染组和空载体转染组相比,let-7g转染组的细胞生长抑制率增高,而CRD-BP、P-gp蛋白表达水平均降低。结论 :let-7g可能通过下调BEL7402/5-FU细胞CRD-BP、c-myc和P-gp蛋白的表达,逆转BEL7402/5-FU细胞对5-Fu的耐药性。
Objective To investigate the effect of miRNA on CRD-BP expression and 5-FU sensitivity in BEL7402/5-Fu cells of hepatocellular carcinoma. Methods Let-7 g expression vector was constructed and Let-7 g plasmid expression vector was transfected into BEL7402/5-FU cells; MTT test and flow cytometry were used to detect the cell growth. Reverse transcription PCR(RT-PCR) and immunoblotting test(Western-Blot) were used to detect the changes of the level of CRD-BP mRNA and the expression of related proteins. Results The cell growth inhibition rate of let-7 g transfected group was higher than that of the untransfected group and the empty carrier transfected group, but the expression level of CRD-BP and P-gp protein decreased after the concentration gradient 5-fluorouracil(5-Fu) was used to treat the BEL7402/5-FU of the liver cancer resistant cell line. Conclusion let-7 g may reverse the resistance of BEL7402/5-FU cells to 5-Fu by down regulating the expression of CRD-BP, c-myc and P-gp proteins in BEL7402/5-FU cells.
Objective To investigate the effect of miRNA on CRD-BP expression and 5-FU sensitivity in BEL7402/5-Fu cells of hepatocellular carcinoma. Methods Let-7 g expression vector was constructed and Let-7 g plasmid expression vector was transfected into BEL7402/5-FU cells; MTT test and flow cytometry were used to detect the cell growth. Reverse transcription PCR(RT-PCR) and immunoblotting test(Western-Blot) were used to detect the changes of the level of CRD-BP mRNA and the expression of related proteins. Results The cell growth inhibition rate of let-7 g transfected group was higher than that of the untransfected group and the empty carrier transfected group, but the expression level of CRD-BP and P-gp protein decreased after the concentration gradient 5-fluorouracil(5-Fu) was used to treat the BEL7402/5-FU of the liver cancer resistant cell line. Conclusion let-7 g may reverse the resistance of BEL7402/5-FU cells to 5-Fu by down regulating the expression of CRD-BP, c-myc and P-gp proteins in BEL7402/5-FU cells.
引文
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[15]Hosseinahli N,Aghapour M,Duijf PHG,et al.Treating cancer with microRNA replacement therapy:A literature review[J].J Cell Physiol.2018,doi:10.1002/jcp.26514.[Epub ahead of print]
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[17]Singleton KR,Crawford L,Tsui E et al.Melanoma Therapeutic Strategies that Select against Resistance by Exploiting MYC-Driven Evolutionary Convergence[J].Cell Rep.2017,21(10):2796-2812.
[18]Benjamin Boyerinas,Sun-Mi Park,Noam Shomron,et al.Identification of Let-7-Regulated Oncofetal Genes[J].Cancer Res,Apr 2008;68:2587-2591.
[2]Ladep NG,Khan SA,Crossey MM,et al.Incidence and mortality of primary liver cancer in England and Wales:changing patterns and ethnic variations[J].World J Gastroenterol,2014,20(6):1544-1553.
[3]Marin JJG,Briz O,Herraez E et al.Molecular bases of the poor response of liver cancer to chemotherapy[J].Clin Res Hepatol Gastroenterol,2018.
[4]Meltzer P S.Cancer genomics:small RNAs with big impacts[J].Nature,2005,435(7043):745-746.
[5]Mens MMJ,Ghanbari M.Cell Cycle Regulation of Stem Cells by Micro RNAs[J].Stem Cell Rev,2018.
[6]Hou N,Guo Z,Zhao G,et al.Inhibition of microRNA-21-3p suppresses proliferation as well as invasion and induces apoptosis by targeting RBPMS through Smad4/ERK signaling pathway in human colorectal cancer HCT116 cells[J].Clin Exp Pharmacol Physiol,2018.
[7]Mayr C,Hemann MT,Bartel DP.Disrupting the pairing between Let-7and hmga2 enhances oncogenic transformation[J].Science,2007,315(5818):1576-1579.
[8]Boyerinas B,Park SM,Shom ron N,et al.Identification of Let-7-regulated oncofetal genes[J].Cancer Res,2008,68(8):2587-2591.
[9]Sparanese D,Lee HC.CRD-BP shields c-myc and MDR-1 RNA from endonucleolytic attack by a mammalian endoribonuclease[J].Nucleic Acids Res,2007,35(4):1209-1221.
[10]Boyerinas B,Park SM,Shomron,N,et al.Identification of Let-7-Regulated Oncofetal Genes[J].Cancer Res,2008,68(8):2587-2591.
[11]Levchenko A,Mehta BM,Niu X,et al.Intercellular transfer of P-glycop rotein mediates acquired multidrug resistance in tumor cells[J].Proc Natl Acad Sci USA,2005,102(6):1933-1938.
[12]N B,K R C.Tetrandrine and cancer-An overview on the molecular approach[J]Biomed Pharmacother,2018,97:624-632.
[13]Dilger K,Schwab M,Fromm MF.Identification of budesonide and prednisone as substrates of the intestinal drug efflux pump P-glycoprotein[J].Inflamm Bowel Dis,2004,10(5):578-583.
[14]Rosenberg MF,Callaghan R,Modok S et al.Three dimensional structure of P-glycoprotein:the transmembrane regions adopt an asymmetric configuration in the nucleotide 2 bound state[J].J Biol Chem,2005,280(4):2857-2862.
[15]Hosseinahli N,Aghapour M,Duijf PHG,et al.Treating cancer with microRNA replacement therapy:A literature review[J].J Cell Physiol.2018,doi:10.1002/jcp.26514.[Epub ahead of print]
[16]Tvingsholm SA,Hansen MB,Clemmensen KKB,et al.Let-7 microR-NA controls invasion-promoting lysosomal changes via the oncogenic transcription factor myeloid zinc finger-1[J].Oncogenesis,2018.
[17]Singleton KR,Crawford L,Tsui E et al.Melanoma Therapeutic Strategies that Select against Resistance by Exploiting MYC-Driven Evolutionary Convergence[J].Cell Rep.2017,21(10):2796-2812.
[18]Benjamin Boyerinas,Sun-Mi Park,Noam Shomron,et al.Identification of Let-7-Regulated Oncofetal Genes[J].Cancer Res,Apr 2008;68:2587-2591.