Intracellular distributing and interferon-γ secretion of human interleukin-18 in BxPC-3 cells.
OBJECTIVE
To investigate the characteristics of interleukin-18 (IL-18) in vitro, explore IL-18, interferon-γ (IFN-γ) and interleukin-2 (IL-2) secretive activity in BxPC-3 line cells with interleukin-18 mutants.


METHODS
Human IL-18 full-length gene (hIL-18-F) and the hIL-18 presumed mature protein gene (hIL-18-M) were inserted into the expression vector pEGFP-N1, to construct recombinant plasmids as Mu0, Mu1, Mu2, Mu3, and Mu4, and the recombinant plasmids were then transferred into BxPC-3 line cells. There are significant differences between Mu1, Mu2 and the pEGFP-C1 control group (P<0.05) by 3-(4,5-dimethiazol- 2-yl)- 2,5-diphenyltetrazolium bromide (MTT) for a proliferation assay, and the fluorescence of the Mu1 and Mu 2 appeared targeted to the membranous region in the BxPC-3 cells after transfected 24h by confocal laser scanning microscope (OLSM).To characterize the intracellular distribution of hIL-18, recombinant IL-18 were each fused to the enhanced green fluorescent protein gene, and expressed in BxPC-3 cells.


RESULTS
Results showed that the Mu1 tended to the membranous region in BxPC-3 cells, this indicates that the N-terminal former amino acid peptide helped ChIL-18 target to BxPC-3 cellS membranes. ELISA results demonstrated that IFN-γ and IL-18 secreted levels of BxPC-3 cells transfecting with recombinant plasmid showed an significant difference (P<0.01); refers to IL-2 expression, the two BxPC-3 cells groups transfecting with recombinant plasmid have no significant function (P>0.05).


CONCLUSIONS
The results showed that hIL-18 and hIL-18 presumed mature protein can induce the secretion of IFN-γ in BxPC-3 cells, and increase the expression of IL-18, but they have no effects on IL-2.
1. Department of General Surgery, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou 310016, China.
Jin
J
Yang
1. Department of General Surgery, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou 310016, China.
Linlin
L
Chen
1. Department of General Surgery, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou 310016, China.
Bin
B
Xu
2. Life Science College, Zhejiang Chinese Medical University, Hangzhou 310053, China.
Jian
J
Xu
2. Life Science College, Zhejiang Chinese Medical University, Hangzhou 310053, China.
Jinquan
J
Sun
2. Life Science College, Zhejiang Chinese Medical University, Hangzhou 310053, China.
Wen
W
Shen
2. Life Science College, Zhejiang Chinese Medical University, Hangzhou 310053, China.
Ting
T
Zhang
D002460:Cell Line; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D007371:Interferon-gamma / Q000235:genetics / Q000378:metabolism*; D020382:Interleukin-18 / Q000096:biosynthesis / Q000378:metabolism*; D007376:Interleukin-2 / Q000096:biosynthesis / Q000378:metabolism*; D014162:Transfection
D020382:Interleukin-18; D007376:Interleukin-2; D007371:Interferon-gamma
BxPC-3 cells; IFN-γ; hIL-18 mutants; intracellular distribution