Therapeutic Effects Of Anti-rage Antibodies On Cholestasis-induced Liver Injury In Rats

The change of dietary pattern leads to the increase of population of cholestasis-induced patients,especially due to cholelithiasis,in China and effectively affects our healthy life.In this project,it is proposed to explore and investigate new recombinant proteins for the treatment of cholestasis-induced liver injury and two recombinant proteins,recombinant mouse G-CSF protein and recombinant rat sRAGE protein,were selected for candidate protein materials.Then rabbit anti-RAGE antibodies were obtained using recombinant rat sRAGE protein,named anti-RAGE antibodies.Therapeutic effect of anti-RAGE antibodies on cholestasis-induced liver injury was tested based on rat bile duct ligand(BDL)model.1.Two candidate recombinant proteins,mouse G-CSF and rat sRAGE,have been succefully obtained.We used Escherichia coli expression system to produce a sufficient amount of recombinant mG-CSF(rmG-CSF)with super quality and bioactivity,and fulfilled the purpose of homologous administration of mG-CSF in mice.rmG-CSF was demonstrated to make no therapeutic effect on liver injury/fibrosis.2.High-efficiency production of bioactive recombinant rat sRAGE(rrsRAGE)was obtained in a eukaryotic system using Pichia pastoris strain X-33 as the host strain.Treatment with rrsRAGE resulted in significantly lower liver fibrosis and lower serum level of ALT,suggesting that sRAGE prevent liver from injury and fibrosis.3.We investigated whether blocking the RAGE receptor with polyclonal anti-RAGE antibodies could regulate acute liver injury and fibrosis in a rat bile duct ligation(BDL)model.Anti-RAGE antibodies improved the gross appearance of the liver and the rat survival rate.Data of liver tissue histology and serum levels of relevant biochemistry indicated that anti-RAGE antibodies attenuated liver necrosis,inflammation,liver fibrosis,and duct proliferation in the BDL model.4.The further role of anti-RAGE antibodies in BDL-induced injury were studied.Anti-RAGE antibodies could decrease liver inflammation/necrosis through the regulation of the level of interleukin-1 beta(IL-1?)that was secreted from Kupffer cells.Anti-RAGE antibodies could decrease liver fibrosis through the regulation of ratio of metalloproteinases-1(TIMP-1)to matrix metalloproteinase(MMPs)that were secreted from hepatic stellate cells(HSCs).In addition,anti-RAGE antibodies could decrease duct proliferation through regulation of transcriptional level of cholesterol 7?-hydroxylase(Cy7?1).Anti-RAGE antibodies could also make an effective effect on the decrease of collegen and cell viability in LX-2 HSCs.Above all,anti-RAGE antibodies can exert a protective effect on cholestasis-induced liver injury,which supported a potential target for such disease.