Experimental Study on A549 Cell Death Mediated by Xenoantigen α-gal 
in Human Serum

Abstract

Background and objective The absence of α-gal in humans is caused by the inactivity of α-1,3GT gene. However, humans have pre-existing and abundant anti-gal antibodies. Xenotransplantation procedures have indicated the high potential of introducing α-1,3GT gene to synthesize α-gal for cancer gene therapy by mimicking hyper-acute rejection. The aim of this study is to construct a lung cancer A549 cell line that expressed α-gal, and to observe the antitumor mechanisms mediated by human serum. Methods A549 cells were transfected with pEGFP-N1-GT plasmids constructed in a previous study. A stable transgenic cell line, A549-GT, was then selected and cultivated. The biological characteristics of A549-GT cells, including morphology and proliferation, were examined. α-1,3GT mRNA expression was detected by RT-PCR. Direct immunofluorescence staining and flow cytometry (FCM) were used to analyze the synthesis of α-gal in A549-GT. The binding of human serum IgM and C3 with A549-GT were also detected. Results α-1,3GT mRNA was expressed in A549-GT. Direct immunofluorescence staining and FCM indicated a high and stable α-gal expression rate in A549-GT. Compared with parental A549 cells, the biological characteristics of A549-GT were unaltered. α-Gal expression was not detected in the human fetal lung fibroblast cell line MRC-5 even though A549-GT and its culture medium were cultivated with the enzyme. Immunofluorescence staining and FCM also indicated abundant binding between A549-GT treated with human serum and IgM/C3. Conclusion α-Gal expression in tumor cells by gene transduction can induce complement-dependent cytototic antitumor effects.

Keywords

• α-1,3-Galactosyltransferase
• α-Galactosyl
• Xenoantigen
• Human adenocarcinoma cell A549