| Objective To investigate the inhibitory effect and mechanisms of serine/ threonine kinase Pim-3 gene on hepatocellular apoptosis in acute liver injury.Methods Thirty-two rats were randomly divide into four groups (eight for each group). Group A was normal control. Group B, C and D were pretreated with Ringer's solution, empty vector and recombinant plasmid, repestively, and received intraperitoneal injections of LPS and D-GalN after 1 day. The rats were sacrificed in 8 hour or agonal after injections of LPS/D-GalN, liver tissues were collected. Green fluorescent protein(GFP) expression levels were appraised under fluorescent microscope; Cell apoptosis of liver tissues was detected by the assay of TUNEL; The liver Caspase-3 activity were detected by chromogenic substrate method; Gene expression was detected by RT-PCR and Western blot.Results 1. Overexpression of GFP was found in Group C and D under fluorescence microscope, but Group A and B had no trace of GFP. 2. TUNEL analysis of rats liver cell apoptosis index(AI) respectively were: 3.1%±1.7%, 83.1%±12.6%, 79.9%±13.4% and 10.2%±6.9%,all LPS/D-GalN attacks treated rat liver cell apoptosis index were significantly higher than the normal control group, Group B and C apoptosis index were significantly higher than group D (P<0.01), the difference was not statistically significant in Group B and C (P>0.05). 3. Caspase-3 activity assay were 107.1±75.0 pmol/min.mg, 728.8±185.8 pmol/min.mg, 643.5±242.9pmol/min.mg and 250.1±84.3 pmol/min.mg, repestively, all LPS/D- GalN attacks treated rats Caspase-3 activity were significantly higher than the normal control group, Group B and C were significantly higher than Group D (P<0.01), while, the difference was not statistically significant in Group B and C (P>0.05). 4. Each group rat liver tissue Pim-3 mRNA relative expression levels were 0.29±0.12, 0.10±0.03, 0.16±0.11 and 0.63±0.18 and Pim-3 protein relative expression levels were 1.12±0.56, 0.61±0.48, 0.37±0.29 and 2.52±0.62, mRNA and protein expression of Group D were higher than Group A, B and C (P<0.05), and Group A was higher than Group B and C (P<0.05), while, the difference was not statistically significant in Group B and C (P>0.05). 5. Each group rat liver tissue Bcl-2 mRNA relative expression levels were 0.26±0.06, 0.32±0.11, 0.28±0.09 and 0.86±0.14 and Bcl-2 protein relative expression levels were 0.91±0.22, 0.85±0.19, 0.84±0.23 and 1.93±0.76, mRNA and protein expression of Group D were significantly higher than Group A, B and C (P<0.01), while, the difference was not statistically significant in Group A, B and C (P>0.05). 6. Each group rat liver tissue p53 mRNA relative expression levels were 0.57±0.38, 1.17±0.71, 1.23±0.85 and 0.22±0.16, p53 expression level of Group D was lower than Group A, B and C (P<0.05), and Group A was lower than Group B and C (P<0.05), but the difference was not statistically significant in Group B and C (P> 0.05).Conclusions Pim-3 could inhibit Caspase-3 activity, inhibit p53 expression and promote Bcl-2 expression, inhibit hepatocellular apoptosis. Exogenous Pim-3 gene in vivo transfection could effectively protect rats from LPS/D-GalN induced acute liver injury occurred. Its protective effects produced mainly in the inhibition of hepatocellular apoptosis.
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