The Study Of Treatment By Artificial Liver Support Systems In Patients With ACLF Based On Intestinal Microecological Metagenomics And NMR Hydrometabonomics

Background and Objective:Acute-on-chronic liver failure(ACLF)is a group of liver failure syndromes manifested by acute jaundice deepening and coagulation dysfunction caused by various triggers on the basis of chronic liver disease.ACLF progresses rapidly,28-day case fatality rates are as high as 30-90%.Systemic hyperinflammatory response plays an important role in the development of ACLF,intestinal microbiome and immune status mainly induce systemic inflammatory responses through pathogen-associated molecular patterns(PAMPs)and virulence factors.Therefore,the generate and progress of ACLF involves multiple factors,multiple stages of intestinal microecology,inflammatory response,immune response and other complex processes.Non-biological ALSS treatment ACLF can improve its biochemical indexes,reduce hepatic encephalopathy,circulatory dysfunction,and immune dysfunction.Artificial liver support system plays an important and irreplaceable role in the treatment of ACLF.The dynamic changes of intestinal flora and metabolites in patients with HBV-ACLF at different stages of disease are still unclear,and the dynamic changes of intestinal flora and metabolites before and after artificial liver treatment are also unclear,at present there are few researches at home and abroad.Finding and confirming serum and body fluid biomarkers for predicting the efficacy of artificial liver treatment,achieving precise treatment with good prognosis for patients,and selecting individual artificial liver treatment schemes are important research directions for ACLF treatment,which have important theoretical significance and high clinical value.there are few studies on the evaluation indicators of the efficacy of artificial liver therapy.This paper will deeply study the related research of artificial liver treatment of ACLF in three parts.The first part:evaluation of the therapeutic efficacy of ACLF patients under different artificial liver treatment modes;The second part:Combining metabolomics and metagenomics to observe the dynamic changes of intestinal flora and metabolites in patients with ACLF at different stages of disease;The third part:Metabolomics based on Hydrogen magnetic resonance spectroscopy(~1HNMR)technology combined with biochemical indicators and metagenomics is used to screen and confirm serum markers of ACLF patients before and after artificial liver treatment.The area under the characteristic curve(AUROC)was used to evaluate the diagnostic value of distinguishing between pre-and post-treatment.PartⅠObjective:Retrospectively analyze the difference in the efficacy of artificial liver support system in ACLF patients under PP mode,PDF mode,PP combined with PDF mode,and evaluation of the efficacy of different artificial liver treatment modes in the treatment of ACLF.Methods:Collect the clinical case data of ACLF patients hospitalized in the Second Affiliated Hospital of Nanchang University from April 2015 to February 2019,and retrospectively analyze the difference in the efficacy of artificial liver support system in ACLF patients under different modes.Results:1.Comparison of clinical indexes before and after treatment in PP mode,PDF mode and PP combined PDF mode of ALSS:TBIL,ALT and AST indexes of the three groups all decreased after treatment,and the difference was statistically significant(P<0.05);MELD scores of the three groups decreased after treatment,and there was no significant difference in PP group,but there was significant difference in PDF group and PP combined PDF group(P<0.05);The PTA value of the three groups of patients increased after treatment,and there was no statistical difference between PP group and PP combined PDF group,but there was statistical difference in PDF group(P<0.05);The INR of PP group was prolonged after treatment,and the difference was not statistically significant.The INR of PDF group and PP combined with PDF group was decreased after treatment,and the difference was statistically significant(P<0.05);The albumin level of PP group,PP combined PDF group decreased after treatment,and the difference was not statistically significant.The albumin level of PDF group increased after treatment,and the difference was statistically significant(P<0.05).2.Comparison of clinical indexes between the three groups before and after treatment:TBIL value,PT and MELD score of the three groups decreased,while PTA value increased,with no statistical significance(P>0.05);After treatment,the white blood cell count increased,but the platelet decreased,the difference was no statistically significant(P>0.05);the albumin level of PDF group increased,while that of PP group and PP combined PDF group decreased.,there was no significant difference among the three groups(P>0.05).3.Secrection of cytokines before and after artificial liver treatment:cytokines after artificial liver treatment decreased to varying degrees compared with that before artificial liver treatment,among which IL-10 and IL-6 showed significant differences before and after artificial liver treatment(P<0.05).4.Comparison of prognosis of patients with different artificial liver treatment modes:the survival rate of patients in PP group after treatment was 70.83%,that in PDF group after treatment was 75.64%,and that in PP combined with PDF group after treatment was 76.66%,there was no significant difference among the three groups(P>0.05).5.The prognosis of patients with liver failure at different stages after artificial liver treatment:the survival rate of patients with early liver failure was 83.56%,the survival rate of patients with intermediate liver failure was 68.75%,and the survival rate of patients with late liver failure was 27.27%.Conclusion:The application of different models of artificial liver can remove harmful toxins and inflammatory mediators in the body,improve the internal environment of the body,and promote the regeneration of hepatocytes;the artificial liver support system is effective for early stage of ACLF,while is not effective for advanced ACLF.And there was no difference between different modes of treatment.PartⅡObjective:The objective is to investigate the relationship between HBV-ACLF progression and intestinal flora,especially to determine the specific microflora of ACLF and its relationship with serum biochemical indices of patients.Methods:From November 2017 to April 2018,109 patients with HBV-ACLF were included in the study.Utilizing 16S rDNA high-throughput sequencing and metabolomics Research(among them,feces of 9 ACLF patients were collected all three different stages of disease development(liver failure ascending stage,plateau stage and descending stage),observe the dynamic changes of microflora.Results:1.With the progress of ACLF,the intestinal flora is gradually disordered,and the diversity and abundance of intestinal flora are significantly reduced.2.Compared with the healthy control group,the relative abundance of Bacteroidetes in ACLF patients was significantly reduced,and the potential pathogenic bacteria veronicus,Streptococcus,Enterococcus and Klebsiella were highly enriched.3.The level of AFP was negatively correlated with the abundance of Bacteroidetes,and TBIL was positively correlated with the abundance of Veronica.The abundance of faecalis was negatively correlated with tbil-inr and positively correlated with PTA.Conclusion:The gut microbiota plays an important role in the development of HBV-ACLF.The diversity and abundance of the gut microbiome significantly decreased with the progression of HBV-ACLF,The relative abundance of the Bacteroidetes phylum in the microbiome was significantly reduced,whereas the abundance of potentially pathogenic bacteria,such as Veilonella,Streptococcus,Enterococcus,and Klebsiella,was highly enriched in the HBV-ACLF group.The abundance of Bacteroidetes was negatively correlated with the level of serum alpha fetoprotein,and the abundance of Veilonella was positively correlated with serum total bilirubin(TBIL).Furthermore,the abundance of Coprococcus was significantly negatively correlated with the level of serum TBIL and the international normalized ratio and positively correlated with prothrombin time activity.Improving the intestinal microbial composition and function of patients of ACLF is helpful for the outcome of disease.PartⅢObjective:Metabolomics using hydrogen magnetic resonance spectroscopy(1HNMR)technology was used to identify the differential metabolites in serum samples of ACLF patients before and after artificial liver treatment,and the area under the characteristic curve(AUROC)was used to evaluate the biomarkers of the efficacy of artificial liver treatment.Methods:Serum samples were obtained from 57 early to middle stage ACLF patients before and after ALSSs treatment and analyzed by metabonomics.The diagnostic values were evaluated by the area under receiver operating characteristic curve(AUROC).A retrospective cohort analysis was further employed.Results:1.Using metabolomics technology based on nuclear magnetic resonance,it was found that artificial liver treatment mainly changed amino acid and energy metabolism,Metabonomic study showed that serum ratios of lactate:creatinine in ACLF patients is significantly al-tered and then restored to normal levels after ALSSs treatment.2.A retrospective cohort analysis validated that the lactate:creatinine ratio of ACLF patients in the one-month death group remained unchanged after ALSSs treatment,but fell markedly in the survival group with AUROC of 0.682 for diagnosis of survival group from death group.3.ACLF patients with PTA between 20.0 and 40.0 before artificial liver treatment were screened to design a prospective experimental study.The results show,The AUROC(95%confidence interval)of the lactate-creatinine ratio after ALSSs treatment for diagnosis ofthe survival group from the death group was0.682(0.502-0.862),the sensitivity was 44.8%,and the specificity was 82.1%.Whereas,The AUROC of PTA after ALSSs treatment for diagnosis of those was0.594(0.40-0.783),the sensitivity was 41.7%,and the specificity was 81.8%serum lactic acid/creatinine is more sensitive measure than measures of prothrombin time activity(PTA)to evaluate the therapeutic effect of ALSSs treatment.Conclusion:Through the use of 1HNMR metabonomics in this study,the lactate:creatinine ratio in serum has been found to be a good indicator to distinguish between ACLF patients before and after ALSSs treatment,the greater the decline in the serum lactate:creatinine ratio with better effective treatments of ALSSs in the ACLF patients with early to middle stage.Further a prospective analysis validation that the lactate:creatinine ratio can be used as a novel marker to evaluate the likely prognosis for early to middle stage ACLF patients who have undertaken ALSSs treatment.This marker may offer superior results to those offeredby measurement of PTA.Application of the marker inredicting the outcomes of ALSSs treatment in early to middle stage ACLF patients may help avoid unnecessary ALSSs and perform liver transplantation.