The Roles And Mechanism Of CD147 Glycosylation Mediated By Glycosyltransferase ?3GnT2 And ?3GnT8 In The Invasion And Metastasis Of Colon Cancer Cell Lines SW620

Objective:To ascertain the roles and mechanism of CD147 glycosylation mediated by ?3GnT2and ?3GnT8 in the metastasis of colon cancer cells SW620.These results will provide theoretical basis for targeting therapy of colon cancer.In summary,our project is to find a new mechanism for colon cancer targeting therapy from the glycobiology point of view.Methods:(1)Clone the full-length gene of ?3GnT2,?3GnT8 and establish the ?3GnT2,?3GnT8sense vector pEX-?3GnT2,pEX-?3GnT8.(2)Select the effective si RNA fragment of ?3GnT2 and ?3GnT8 with Real time PCR.(3)Exogenous ?3GnT2,?3GnT8 sense plasmid vector was transfected into SW620 cells and ?3GnT2,?3GnT8 shRNA interference vector was transfected into SW620 cells by lipofectamine.(4)The mRNA and protein expression of ?3GnT2,?3GnT8,CD147,MMP2 and MMP14 were detected by Real time PCR and Western blot.(5)Flow cytometor(FCM)was used to detect the level of polylactosamine.(6)Cell scratch repair assessment was used to detect the migration ability of the cells.Results:(1)?3GnT2 and ?3GnT8 sequence did not appear in variation after inserting into the plasmid pEX-2,which was detected by Blast and NCBI.(2)The effective siRNA fragments(si?3GnT2-1336 and si?3GnT8-505)were got by Real time PCR.(3)The expression of HG-CD147,MMP2 and MMP14 were upregulated with the overexpression of?3GnT2,?3GnT8.The expression of HG-CD147,MMP2 and MMP14 were downregulated with the knockdown of?3GnT2,?3GnT8.(4)Flow cytometor(FCM)results showed the level of polylactosamine in cell surface was increased in SW620 cells transfected ?3GnT2??3GnT8 sense vector and decreased while ?3GnT2??3GnT8 were interferenced compared with control groups.(5)The potential of migration and invasion of SW620 were enhanced when ?3GnT2??3GnT8 was up regulated.Migration and invasion abilities were declined following the interference expression of ?3GnT2??3GnT8.Conclusion:?3GnT2 and ?3GnT8 may both play an essential role in the invasion and migration of colon cancer cells SW620 by synthesis the formation of polylactosamine and changing level of glycosylation of CD147.Overexpression of ?3GnT2 and ?3GnT8 can enhance the potential of invasion and migration of SW620 cell lines.