Preliminary Study On The Mechanism Of Antiplatelet Intervention In LPS/D-GalN Induced Acute Liver Failure In Mice

Background Acute liver failure(ALF)is caused by severe hepatocyte damage which induces coagulation dysfunction and different degrees of hepatic encephalopathy,and eventually with fatal results.Rapid decline in platelets is a common clinical phenomenon in patients with liver failure.In acute liver failure,the rapid decline in platelet is closely related to systemic inflammatory response,multiple organ failure,and short-term mortality.D-galactosamine(D-GalN)is a special hepatic uridine nucleotide blocker.Toll-like receptors(TLRs)constitute a large class of innate immune recognition receptors that recognize microbial components and spread through a well-defined signal cascade.LPS(lipopolysaccharide)/D-GaIN is a commonly used animal model of ALF,which is similar to clinical ALF.When administered to LPS/D-GaIN model mice,Agkisacucetin and tirofiban which block platelet adhesion and aggregation,respectively,can improve the survival of LPS/D-GalN induced ALF mice.In addition to LPS/D-GalN,acetaminophen(APAP),concanavalin A(ConA),polyinosinic-polycytidylic acid[poly(I:C)]/D-GalN and CpG(cytosine-phosphate-guanine)DNA/D-GalN are also common ALF models.Methods Establish mouse ALF models LPS/APAP,LPS/ConA,poly(I:C)/D-GalN,and CpG DNA/D-GalN,then intervene with agkisacucetin and(or)tirofiban to observe whether they have a therapeutic effect.Quantitative proteomics analysis of the liver in these three D-GalN-related ALF mice were performed to reveal the possible mechanisms of TLR agonists and D-GalN induced ALF.Liver tissue immunofluorescence was used to verify the expression of relevant protein markers.Results We successfully constructed four ALF models:LPS/APAP,LPS/ConA,poly(1:C)/D-GalN,and CpG DNA/D-GalN.The experimental results showed that and agkisacucetin and(or)tirofiban had no therapeutic effect on the four ALF mouse models except LPS/D-GalN.Among them,poly(I:C)/D-GalN and CpG DNA/D-GalN were similar to the LPS/D-GalN model in mouse serum alanine aminotransferase(ALT)levels,liver tissue damage and death time.Proteomic analysis of LPS/D-GalN,poly(I:C)/D-GalN and CpG DNA/D-GalN showed that,compared to healthy controls,208,89,and 121 differentially expressed proteins were identified at 2 hours after treatments in LPS/D-GalN,poly(I:C)/D-GalN,and CpG DNA/D-GalN groups,respectively.Among them,24 proteins were common expressed proteins,153,36,and 51 were model-specific proteins.At 4 hours of treatments,321,227,and 114 differentially expressed proteins were identified,of which 52 were common expressed proteins,80,195,and 23 were model-specific proteins.The data showed that although the three D-GalN related ALF models had similar phenotypes,the annotation and classify of identified proteins and the function enrichment such as platelet and coagulation dysfunctions were very different.Platelets and coagulation related proteins Alpha-1-antitrypsin,Alpha-2-antiplasmin,Mannose-binding protein A,and High affinity immunoglobulin epsilon receptor subunit gamma were specifically expressed in the LPS/D-GalN model;immue-related proteins Lactotransferrin,Protein S100-A9,Neutrophil gelatinase-associated lipocalin,Integrin alpha-M,Annexin A1,and Myeloperoxidase were significantly expressed in the LPS/D-GalN model.Conclusion The antiplatelet drugs agkisacucetin and(or)tirofiban are model specific for the treatment ofALF in mice.Although the phenotypes were similar in LPS/D-GalN,poly(I:C)/D-GalN and CpG DNA/D-GalN induced ALF in mice,the pathogenesis were very different,including differences in coagulation and inflammation levels,further research on related proteins will help to understand the mechanism of antiplatelet therapy in LPS/D-GalN induced ALF mice.