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Published on: January 14, 2011
[Cloning, expression and antitumor effect of mouse costimulatory molecule 4-1BBL]
Cheng-li Liu1, Ke-feng Dou, Bang-fu Zhu
1Department of Hepatobiliary Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Aim:
To clone mouse 4-1BBL gene, construct its eukaryotic expression vector, and evaluate antitumor activity of the expression product.
Methods:
RT-PCR was used to amplify mouse 4-1BBL gene from total RNA of C57BL/6 splenocytes stimulated by PHA. Then m4-1BBL cDNA was subcloned into eukaryotic expression vector pcDNA3.1(+) and transfected into mouse hepatocellular carcinoma cell line Hepa1-6. The expression of m4-1BBL in transfected cells was detected by RT-PCR, indirect immunofluorescence staining, and flow cytometry. Non-adherent splenocytes from non-immunized C57BL/6 mice were incubated with mitomycin-treated non-transfected Hepa1-6(Hepa1-6-wt) or transfected Hepa1-6 cells (Hepal-6-m4-1BBL), respectively. Then the lymphocytes were tested for cytotoxic activity to Hepa1-6-wt cells.
Results:
The Hepa1-6 cells transfected by pcDNA3.1(+)-m4-1BBL could efficiently express m4-1BBL. As compared with Hepa1-6-wt cells,Hepa1-6-m4-1BBL cells could induce more efficiently cytotoxic activity of lymphocytes to Hepa1-6-wt cells (P<0.01).
Conclusion:
The expression of m4-1BBL by tumor cells is effective in inducing antitumor immune response.
