Specific Antitumor-immunity Induced By MIL-12 Gene Modified Dendritic Cells Transfected With Murine Colon Carcinoma RNA

Partâ… SpecificactivityofcytotoxicTlymphocyteinducedbymIL-12genemodifieddendriticcellstransfectedwiththeRNAofmurinecarcinomaofcolonObjective : To investigate specific activity of cytotoxic Tlymphocyte (CTL) induced by mIL-12 gene modified dendritic cells(DC)transfected with the total RNAof CT-26(a cell line of murine carcinomaofcolon).Methods:In vitro proliferation of DC from the culture of murinebone marrow, was stimulated by rmGM-CSF and rmIL-4, the purity ofwhich was detected by flow-cytometry. The generated DC were thenmodified byadenovirus with mIL-12 genewhich wereproliferatedin293cells. The total RNA of CT-26 was obtained by Trizol's process, withmIL-12 gene modified DC transfected by TransMessenger in vitro. Thelevel of mIL-12 both in vitro and in vivo and the activity of CTL in vivowere estimated by ELISA assay and modified LDH release assay expectively.Results:Plenty of DC were obtained from the culture of murinebone marrow, with over 90 percent of CD11c+ cells.DC modified bymIL-12 gene could induce high level of mIL-12 both in vitro and invivo,and the difference compaired with control groups was significant(P<0.01).Vaccination with mIL-12 gene modified DC transfected withthetotalRNAcouldinducehighlevelofspecificCTLactivityinvivo,andthe Ad-LacZ modified DC and DC transfected with the total RNAinduced moderate level of specific CTLactivity in vivo, meanwhile quitelow level of specific CTL activity was estimated in the mice immunizedwithDCandPBS.Conclusion:mIL-12 gene modified DC transfected with the totalRNAof tumor could improve the level of mIL-12 in mice,and could alsoactivatespecificCTLeffectively. Partâ…¡Specificantitumor-immunityinducedbymIL-12genemodifieddendriticcellstransfectedwithmurinecoloncarcinomaRNAObjective: To investigate the specific antitumor effect of mIL-12gene modified dendritic cells(DC) transfected with the total RNA ofCT-26(murinecoloncarcinomacells).Methods: DC were cultured from murine bone marrow in thepresence of rmGM-CSF and rmIL-4. The purity was detected byflow-cytometry. The generated DC were then modified by adenoviruswith mIL-12 gene which was proliferated in 293 cells. The total RNAofCT-26 was extracted by Trizol reagent into mIL-12 gene modified DCand by TransMessenger in vitro. Mice were immunized with thesemodified DC. The level of mIL-12 both in vitro and in vivo and thespecificactivityofcytotoxicTlymphocyte(CTL)invivowereestimatedby ELISAassay and modified LDH release assay expectively.The tumorgrowth and animal survival time of immunized mice after chanllengewithparentaltumorcellswereestimated.Results: DC were obtained from the culture of murine bonemarrow.TheCD11c+cellswereover90percent.Vaccinationwith mIL-12 genemodifiedDCtransfectedwiththetotalRNAcouldinducehighlevelof specific CTL activity in vitro , 100 percent of the immunized micecould survive. Vaccination with Ad-LacZ modified DC and DCtransfected with the total RNA induced moderate level of specific CTLactivity in vitro(P<0.01)and 60 percent of the immunized micesurvived,low level specific CTL activity was estimated in the miceimmunizedwithDCandPBS,andthemicealldied.Conclusions: mIL-12 gene modified DC transfected with the totalRNA of tumor could present endogenetic tumor antigen,and could alsoactivate CTL, thus inducing specific anti-tumor immune responseeffectively.