Abstract
BackgroundConcanavalin A (ConA)-induced hepatitis is an experimental murine model mirroring the pathology of human autoimmune hepatitis.AimTo investigate the effects of intrasplenically transplanted fetal hepatocytes (BNL.CL2) transfected with recombinant adenovirus vector expressing the IL-18 binding protein (IL-18BP) and IL-4 fusion protein on ConA-induced hepatitis in mice.MethodsAd-IL-18BP/IL-4 was used to infect BNL.CL2 cells. IL-4 and IL-18BP fusion protein expression were detected by ELISA and Western blotting. BNL.CL2 cells infected with Ad-IL-18BP/IL-4 were intrasplenically transplanted into mice. After 10 days, mice were injected with ConA (15 mg/kg), and sacrificed 18 hours later. Liver injury was assessed by serum transaminase and liver histology. TNF-α, IL-18, IL-4, IL-10, IL-12p70 and monocyte-chemoattracting protein (MCP)-1 levels in serum and liver homogenates were detected by ELISA. Signaling molecules in liver homogenates were analyzed by Western blotting.ResultsAd-IL-18BP/IL-4 effectively expressed the IL-18BP/IL-4 fusion protein for more than 14 days in BNL.CL12 cells. Treatment of mice with Ad-IL-18BP/IL-4-BNL.CL2 before ConA injection significantly reduced the elevated plasma levels of transaminases compared with ConA control groups. TNF-α, IL-18, IL-12p70 and MCP-1 levels in serum and liver homogenates from mice transplanted with Ad-IL-18BP/IL-4-BNL.CL2 were lower and IL-4 and IL-10 levels were higher than control groups. Phosphorylation levels of NF-κB p65, AKT, p38 and JNK1/2 in liver homogenates were markedly suppressed by Ad-IL-18BP/IL-4.ConclusionsAd-IL-18BP/IL-4 was effectively transfected into mouse BNL.CL2 cells. Intrasplenic transplantation of Ad-IL-18BP/IL-4-BNL.CL12 cells alleviated the severity of inflammation in ConA-induced experimental hepatitis and provides a useful basis for the targeted gene therapy of liver disease.
Highlights
Concanavalin A (ConA) is a lectin originally extracted from the jack-bean, Canavalia ensiformis
Intrasplenic transplantation of Ad-IL18BP/IL-4-BNL.CL12 cells alleviated the severity of inflammation in ConA-induced experimental hepatitis and provides a useful basis for the targeted gene therapy of liver disease
In order to examine the in vitro expression of IL-18 binding protein (IL-18BP) and IL-4 by IL-18BP/IL-4 gene-modified BNL.CL2 cells, culture supernatants were harvested 48 hours after transfection with recombinant adenoviruses at multiplicities of infection (MOI) from 0 to 100, and IL-4 concentrations were measured by Enzyme-linked immunosorbent assay (ELISA)
Summary
Concanavalin A (ConA) is a lectin (carbohydrate-binding protein) originally extracted from the jack-bean, Canavalia ensiformis. ConA is a hemagglutinin and a mitogen which is known for its ability to stimulate mouse T-cell subsets giving rise to four functionally distinct T-cell populations, including precursors to suppressor T-cells. It causes acute inflammation of the liver by the infiltration of activated lymphocytes, resulting in massive necrotic tissue injury of hepatocytes and intrasinusoidal hemostasis [1]. ConA-induced hepatitis in mice is commonly employed as an acute model for human autoimmune hepatitis because it mimics many aspects of this disease, including markedly increased serum levels of alanine transaminase (ALT) and inflammatory responses [2]. Concanavalin A (ConA)-induced hepatitis is an experimental murine model mirroring the pathology of human autoimmune hepatitis
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