Objective:To explore the protective role of recombinant plasmid pEGFP-N2/Pim-3 transfecting in rat pancreatic acinar cells AR4-2J damaged by lipopolysaccharide (LPS). Methods: AR4-2J cells were divided into 4 groups: Group A, as the normal control group; Group B, treated with 5 μg/mL of LPS; Group C, transfected with blank plasmid pEGFP-N2 and treated with 5 -滋g/mL of LPS; Group D, transfected with recombinant plasmid pEGFP-N2 /Pim-3 and treated with 5 μg/mL of LPS. Flow cytometry examined the apoptosis of AR4-2J cells in four groups after 24 h. RT-PCR was performed to detect the mRNA expression of Pim-3, intercellular cell adhesion molecule-1 (ICAM-1), and Occludin in AR4-2J cells of four groups treated in 0, 6, 12, 24 h, and Western blot was performed to detect the protein expression of Pim-3, ICAM-1, and Occludin in each group. Results: The apoptotic rates of Groups B and C were (53.13 ± 5.73)% and (51.76 ± 5.17)%, respectively, both significantly higher than that of Group D (21.13 ± 4.15)%(both P < 0.05). The expressions of Pim-3 mRNA and protein in Groups A, B and C remained a low level. The Pim-3 expression of Group D was significantly higher than those in Groups A, B and C (both P < 0.05). The ICAM-1 mRNA and protein expression levels of Groups B, C and D were constantly upregulated 6 h later, all significantly higher than those of Group A (all P < 0.01), and the ICAM-1 expressions of Groups B and C were significantly higher than Group D (both P < 0.05). The Occludin mRNA and protein expression levels of Groups B, C and D began to increase 6 h later, and peaked 12 h later, all significantly higher than those of Group A (all P < 0.01), and the Occludin expressions of Groups B and C were significantly lower than that of Group D (both P < 0.05). Conclusion: Pim-3 gene can inhibit the apoptosis of pancreatic acinar cell damaged by LPS, and also can upregulate the expression of Occludin and downregulate the expression of ICAM-1 to inhibit the pancreatic inflammatory reaction induced by LPS.