Objective: To provide evidence that blocking the receptor-interacting protein 2 (Rip2) expression can decrease inflammatory cytokine production by macrophage and protect mice from endotoxin lethality.
Methods: Murine Rip2 small interfering RNA (siRNA) plamids were constructed and transfected into macrophages and Rip2 expression was assessed with RT-PCR and Western blot. Cell proliferation was assayed with MTT; TNFalpha concentration was assayed with ELISA and high-mobility group box 1 protein (HMGB1) level with semi quantitative Western blot after lipopolysaccharide (LPS) stimulation. LPS challenge was given after the plasmids were injected into mice and the survival rate was calculated. Rip2 and HMGB1 expression in liver was assessed with Western blot and serum TNFalpha level with ELISA.
Results: Rip2 siRNA plasmids could block the mRNA and protein expression of Rip2 and promote cell proliferation. Blocking of Rip2 could attenuate LPS-induced TNFalpha and HMGB1 production. HMGB1 expression in liver were decreased to (40.21 +/- 11.03) pg/g and serum TNFalpha level was decreased to (300.43 +/- 59.26) ng/L (P < 0.05). The survival rate of endotoxemic mice was also improved (P < 0.05).
Conclusion: The results demonstrate that Rip2 siRNA plasmids can block the expression of Rip2, and decrease the production of TNFalpha and HMGB1, thus protect mice from lethal endotoxemia.