[Cloning, expression and antitumor effect of mouse costimulatory molecule 4-1BBL].
OBJECTIVE
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).


CONCLUSIONS
The expression of m4-1BBL by tumor cells is effective in inducing antitumor immune response.
Liu
Cheng-li
CL
Department of Hepatobiliary Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Dou
Ke-feng
KF
Zhu
Bang-fu
BF
Zang
Xiao-xia
XX
Chai
Yu-bo
YB
Du
Ke-jun
KJ
Chen
Su-min
SM
D053330:4-1BB Ligand; D000818:Animals; D019496:Cancer Vaccines / Q000276:immunology; D003001:Cloning, Molecular; D008213:Lymphocyte Activation; D051379:Mice; D008810:Mice, Inbred C57BL; D009374:Neoplasms, Experimental / Q000276:immunology*; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis / Q000235:genetics* / Q000502:physiology
D053330:4-1BB Ligand; D019496:Cancer Vaccines; D012333:RNA, Messenger; C505975:Tnfsf9 protein, mouse; D014409:Tumor Necrosis Factor-alpha