To evaluate the Plasmodium falciparum CTL epitope vaccines in HLA class I allele specific human cell lines that have high frequency among Chinese population. Methods. Synthesized oligonucleotides encoding for P.f. CTL...To evaluate the Plasmodium falciparum CTL epitope vaccines in HLA class I allele specific human cell lines that have high frequency among Chinese population. Methods. Synthesized oligonucleotides encoding for P.f. CTL epitope genes, constructed eukaryotic expression plasmids, transfected the minigenes into HLA class I allele specific human cell lines and identified endogenous expressing of the minigenes by RT- PCR and HLA stabilization assay. Results. Two mini- genes encoding Plasmodium falciparum CTL epitopes were designed and cloned, respectively, into an eukaryotic expressing vector to form TR26 which was restricted to HLA- B51, SH6 which was restricted to HLA- A2.1, and TS, which had the two aforementioned mini- genes fused in tandem. All of these CTL epitope genes were transfected and endogenously expressed in respective cell lines containing appropriate HLA molecules. The obviously increased expressions of HLA class I molecules were detected in the transfected cell lines. It was demonstrated that the two discrete Plasmodium falciparum epitope genes were effectively processed and presented, and the close proximity of the two epitope genes in one chain as in mini- gene TS did not interfere with the processing and presenting of each epitope gene in corresponding cell line. Conclusion. A successful expression and presentation of multiple CTL epitope mini- gene in MHC class I allele specific human cell lines were demonstrated by an in vitro assay, which could be corresponding to the vaccination of CTL vaccines in people with different MHC I molecules. This work also suggested the possibility of constructing a multiple CTL epitope plasmodium falciparum DNA vaccine that could cover most of Chinese population.展开更多
基金This project was supported by the National Natural Sciences Foun-dation of China, grant# 39770670 and China Medical Bo
文摘To evaluate the Plasmodium falciparum CTL epitope vaccines in HLA class I allele specific human cell lines that have high frequency among Chinese population. Methods. Synthesized oligonucleotides encoding for P.f. CTL epitope genes, constructed eukaryotic expression plasmids, transfected the minigenes into HLA class I allele specific human cell lines and identified endogenous expressing of the minigenes by RT- PCR and HLA stabilization assay. Results. Two mini- genes encoding Plasmodium falciparum CTL epitopes were designed and cloned, respectively, into an eukaryotic expressing vector to form TR26 which was restricted to HLA- B51, SH6 which was restricted to HLA- A2.1, and TS, which had the two aforementioned mini- genes fused in tandem. All of these CTL epitope genes were transfected and endogenously expressed in respective cell lines containing appropriate HLA molecules. The obviously increased expressions of HLA class I molecules were detected in the transfected cell lines. It was demonstrated that the two discrete Plasmodium falciparum epitope genes were effectively processed and presented, and the close proximity of the two epitope genes in one chain as in mini- gene TS did not interfere with the processing and presenting of each epitope gene in corresponding cell line. Conclusion. A successful expression and presentation of multiple CTL epitope mini- gene in MHC class I allele specific human cell lines were demonstrated by an in vitro assay, which could be corresponding to the vaccination of CTL vaccines in people with different MHC I molecules. This work also suggested the possibility of constructing a multiple CTL epitope plasmodium falciparum DNA vaccine that could cover most of Chinese population.
