Down-regulation Of MRPL35 Or RasGRF2 On The Effect Of Growth And Metastasis Of Colorectal Cancer Xenograft In Nude Mice

Objective:To explore the effect of RNA interference-mediated Mitochondrial ribosomal protein L35(MRPL35)or Ras protein-specific guanine nucleotide-releasing factor 2(RasGRF2)downregulation on the growth and metastasis of colorectal cancer xenograft in nude mice.Methods:1.Lentiviral vector containing MRPL35 shRNA was constructed and infected into HCT116 cells.The cells with stable MRPL35 knockdown were selected and inoculated subcutaneously to BALB/c nude mice.The growth of tumors were observed and the growth curve was plotted as well.The mRNA level of MRPL35 was detected by real-time polymerase chain reaction(PCR).The expression of Ki-67 and Cleaved caspase-3 in tumor tissues were detected by immunohistochemistry(IHC).The difference of tumor volume was analyzed by t-test method.2.Lentiviral vector containing RasGRF2 shRNA was constructed and infected into HCT116 cells.The cells with stable RasGRF2 knockdown were selected and injected into BALB/c nude mice through tail vein.The breathing,weight,activity and mental status of nude mice were observed.The lungs were removed,and visible tumors on the lung surface were counted.The expression of Ras GRF2 and matrix metallo-proteinase(MMP)-9 in tumor tissues were detected by immunohistochemistry(IHC).The survival rate was analyzed by Kaplan-Meier method.Results:1.The HCT116 xenograft model was established.The gross tumor volume of MRPL35 shRNA was significantly smaller than those in control shRNA groups(P<0.05).The mRNA level of MRPL35 was significantly decreased in MRPL35 shRNA group as compared to control shRNA group(P<0.05).The expression of Ki-67 dropped about 72%and Cleaved caspase-3 increased about 61%,respectively.2.In pulmonary metastasis model,the nude mice appeared obvious weight loss,bow back,apathetic phenomenon after 8 weeks.The numbers of lung metastasis tumor were lower in RasGRF2 shRNA group than those in the control group(P<0.05).Knockdown of RasGRF2 reduced the number of metastatic nodules compared with the control group.IHC staining showed the lung tumors expressed Ras GRF2 and MMP-9.Kaplan-Meier analysis indicated that the survival rate in RasGRF2 shRNA group was higher than that in the control group(P<0.05).Conclusion:Downregualtion of MRPL35 can inhibit the growth of tumor.Downregulation of RasGRF2 can suppress the lung metastasis of colorectal cancer cell.