关键词: 电穿孔, RNA

Abstract: Abstract:Objective To investigate the protein expression in mature dendritic cells（mDCs） after transfection with total RNA from carcinoembryonic antigen(CEA)-expressing colon adenocarcinoma SW480 by electroporation and test whether electroporation can influence on the function and phenotype of DCs in order to lay a foundation for the preparation and clinical application of DCs nucleic acid tumor vaccine.Methods DCs were generated in vitro from human peripheral blood mononuclear cells(PBMC) in the presence of recombinaht human granulocyte monocyte colony stimulating factor(rhGM-CSF) and recombinant human interleukin-4 (rhIL-4),and mDCs were induced with 0tumor necrosis factor-α (TNF-α) after treated for 24 h.There were two groups in the experiment:mDCs group and immature DCs(imDCs) group.The total tumor RNA was extracted from SW480 by Trizol and was transfected into mDCs by electroporation.The phenotype of DCs and expression of CEA protein before and after electroporation were assessed by flow cytometry.The secretion of IL-12 in each group was investigated by ELISA.Results DCs with characteristic of morphological features were generated in vitro from PBMC by rhGM-CSF,rhIL-4 and TNF-α.After cultivated for 5 d，the cells gained the phenotype of imDCs,showed strong expression of CD1a,weak expression of CD80 and no expression of CD83.After maturation by TNF-α， the expressions of CD1α,CD80 and CD83 were positive and they were also positive in RNA transfection DCs group.The expression of CEA protein was meaured at 4 h after electroporation.The secretion levels of IL-12 in mDCs and RNA transfection DCs groups were significantly higher than that in imDCs group (P<0.01),but there was no significant difference between RNA transfection DCs group and mDCs group(P>0.05). Conclusion The total RNA from colon adenocarcinoma cells can be transfected i nto mDCs by electroporation and electroporation doesn’[KG-*3]t alter the phenotyp e and function of mDCs．

Key words: electroporation , RNA