Fundamental And Clinical Research About Pathogenesis Of Hepatopulmonary Syndrome

Part one An experimental research about role of fractalkine/CX3CL1/CX3CR1in the pathogenesis ofhepatopulmonary syndrome rats with obstructive jaundiceBackground:Common bile duct ligation (CBDL) for rat is a model system for the research of HPS.Prior studies have discovered that pulmonary intravascular macrophages(PIM) adhere and accumulation occurs as experimental HPS develops.The activated CD68(+) macrophages generate angiogenic and proliferative growth factors,including inducible nitric oxide synthase(iNOS),vascular endothelial growth factor(VEGF),and platelet-derived growth factors(PDGF).That induces an occult proliferative pulmonary vasculopathy.Intravascular accumulation of activated macrophages is central to the pathogenesis of HPS.Fractalkine is known to be expressed on pulmonary vascular endothelial cells(PVEC).It binds highly specifically to its receptor CX3CR1which is expressed on monocytes and lymphocytes.Thus, we hypothesized that the specific binding of fractalkine expressed on PVEC to CX3CR1expressed on monocytes was crucial signal pathway of PIM adhere and accumulation.Could degree of PIM accumulation and pulmonary microvascular angiogenesis be decreased after CBDL if CX3CL1/CX3CR1expression and signaling were altered?Could HPS be improved?Therefore, the aim of this part was to define if pulmonary CX3CL1and CX3CR1expression are altered after CBDL and influence macrophages accumulation,angiogenesis and the development of HPS,to research the role of fractalkine/CX3CR1in the mechanisms of HPS rats.Material and methods:Establishing sham-operation group(n=5).The rats after CBDL were randomly divided into experimental control group(EC group, n=20) and anti-CX3CR1-neutralizing polyclonal antibody group(CX3CR1-Ab group,n=20).From15to28day after operation,the rats in CX3CR1-Ab group were intraperitoneal injected respectively with anti-CX3CR1-neutralizing polyclonal antibody each day,and he rats in EC and sham group received identical volume of normal saline.All rats were died on29day,and A-aDO2was immediately detected.Expression levels of ALT,ALP,TBIL in serum were determined with radioimmunoassay.Meanwhile,accumulation degree of pulmonary intravascular macrophages(PIM) and microvessel density (MVD) were tested by immunity fluorescence method.Furthermore,protein expression and mRNA level of CX3CR1and CX3CL1were measured with western blot and real time-PCR.Results:In the rats after CBDL,the serum levels of TBIL,ALT,ALP and A-aDO2were rapidly elevated,accompanied by liver function damage.There was statistical difference in the changes (P<0.001vs sham group).In EC group,PIM showed a larger accumulation,MVD was markedly elevated,protein and mRNA expression of CX3CL1and CX3CR1were significantly increased (P<0.001vs sham group). Compared to EC group,in CX3CR1-Ab group A-aD02showed the normal range,hypoxemia was improved,protein and mRNA expression levels of CX3CL1and CX3CR1were remarkably decreased,likewise,degree of PIM accumulation and pulmonary microvascular angiogenesis were reduced to some extent.There was significant difference between the two groups (P<0.001)Conclusion:Pulmonary intravascular macrophage aggregation increase after CBDL and contribute to pathological proliferation of pulmonary microvascular and the development of experimental HPS.Pulmonary CX3CL1/CX3CR1expression elevated and specific binding is a vital signaling pathway by which mononuclear macrophage adhere and accumulate in pulmonary intravascular.Macrophage depletion is greatly obvious though blocking the signaling pathway.Moreover,HPS could be clearly improved. Part towAn experimental study for effects of Methionine and Vitamin B1and Sodium Tanshinone IIA Sulfonate on liver,lung and kidney function of rats with hepatopulmonary syndrome and obstructive jaundiceBackground:Obstructive jaundice(OJ) could lead to endotoxemia(ETM) and multiple organ dysfunction during perioperative period.There are great high complication rate and mortality.Especially,the function injuries of liver,lung and kidney are extraordinary serious.More patients were dead due to shocking and hepatic, kidney and respiratory function failure.The purpose of this part is to research the mechanisms of HPS and pathogenesis of liver and kidney function injury through observing level changes of endotoxin(ET) and A-aD02and liver,lung,kidney function and tissue structure of rats with HPS after CBDL,to investigate effects of Sodium Tanshinone IIA Sulfonate(STS) and Methionine and Vitamin B1(MVB1) on the changes.Material and methods:The rats after CBDL were randomly divided into experimental control(EC)group,MVB1group,STS group and combined group with MVB1and STS(M-S)group.There were20rats in each one group,and establishing sham-operation group(n=5).From1to13day after operation EC,MVB1and STS group rats were intraperitoneally injected respectively with identical volume of normal saline,MVB1and STS.M-S group rats received MVB1and STS together.Expression levels of ET,TNF-a,ALT,AST,ALP,TBIL,BUN and Crea in serum on days2,6,10and14after operation were respectively determined with radioimmunoassay.All rats were died on14day,and A-aDO2was immediately detected.Meanwhile,histopathological changes of liver,lung,kidney tissues were observed,and CD31(+)angiogenesis were measured.Result:The mortality rate of rats with OJ was10%.In the groups after BDL,levels of ET,TNF-a,ALT,AST,ALP,TBIL,BUN and Crea in serum were rapidly elevated(P<0.001vs sham group). Compared to EC group,levels of serum ET,TNF-a were declined in STS,MVB1and M-S groups,and liver and kidney functions were improved to some extent.There was statistical difference in the changes(P<0.05).Compared respectively to STS and MVB1groups,level changes of the above tested indexes in M-S group were significant(P<0.001).A-aDO2was enhanced (≥20mmHg) in EC group rats on14d after CBDL.Meanwhile,Degree of pulmonary intravascular macrophages(PIM) accumulation and angiogenesis were markedly increased. In STS and M-S groups,A-aDO2was declined (≤15mmHg),and angiogenesis was reduced.There was statistical difference in the changes(P<0.001vs EC group).The histological changes were not significant between EC and MVB1group (P>0.05) Conclusion:The levels increased of TNF-a and ET in serum are ones of the most important factors which induce angiogenesis in HPS and damaging of liver and renal function in OJ rats.MVB1and STS could restrain the increase of plasmic TNF-a and ET of rats with OJ.There is more significant inhibiting effect on ET in STS than MVBi.The anti-endotoxin effect and protective effect on liver and kidney tissue of OJ rats could be clearly elevated when being injected with together MVB1and STS.Pulmonary angiogenesis occurs as experimental HPS on two weeks after CBDL.MVB1has not effect on lung tissue structure and function of HPS rats.STS could not prevent the occurrence of experimental HPS.Nevertheless,it could improve HPS to some extent through decreasing pulmonary microvascular angiogenesis. Part threeCause and prevention and surgical treatment of iatrogenic bile duct injury complicated with hepatopulmonary syndromeBackground:Long-term complications that occur for some patients,such as biliary stricture,intrahepatic stones,cirrhosis and HPS after initial repair surgery for iatrogenic bile duct injury(IBDI) patients.The events are hard problems in clinical treatment.The reoperation procedures are complicated, changing and different from traditional operation.There is a higher mortality rate after the reoperation.The purpose of this part is to research the causes,treatments of IBDI,to estimate the choice of reoperation methods and therapeutic effect on IBDI patients with HPS.Material and methods:A retrospective analysis was performed on56surgically treated cases with IBDI after operation in Wuhan Union Hospital from January2004to September2010.From March2011to September2012,6patients suffered from IBDI which complied with the standard diagnosis of HPS.We discussed the choice of reoperation methods and paths,estimated postoperative recovery,complication rate and mortality.Results:One case was dead for infectious shock that was caused by common bile duct segment back wall perforation in56ones.Fine rate was75percent for the first repair,53.8percent for the second repair.One case has better outcomes in6patients repaired for the third time,two ones dead,there ones needed further treatment.The excellent rate was89.3percent, poor rate10.7percent.The6cases were operated successfully. However,the procedures were painful after hours (4.9±3.6h) on average.The reoperation methods were different from traditional operation.After reoperation respiratory failure took place in3cases,one of them died.Bile leak with local intra-abdominal infection occured in1case.5cases obtained good cure rates.The excellent rate was100percent after0.5～2years follow-up.Conclusion:The reasons of IBDI are related to bile duct variable anatomy,lesion local pathological change and performer technology factors.The operation suffered from painful process,wide strip surface and more intraoperation bleeding during the reoperation.The reoperation methods are determined by a number of factors which are the causes,severity,repairing ways of primary IBDI,cirrhosis,distribution of intrahepatic stones,anatomy feature of the first porta and the general conditions of the patients.The procedure should be strived to simple, rapid and effective.Comprehensive treatments of ICU and other multi-disciplinary are needed for high incidence rate of complications.A-aDO2enhanced could not be improved after the reoperation of IBDI.Moreover,the incidence of respiratory failure and mortality rate were increased after the reoperation for patients with HPS.