Comprehensive Evaluation Of Urinary Trypsin Inhibitor Against Sepsis

BackgroundSepsis is a common fatal disease, the most threatening in critically ill patients and thecommonest cause of death. Integrated results of published epidemiological and economicdata shows extremely severe disease burden, even topped as the1stone of all diseases.Information summary from existing pathogenesis and relevant research of sepsis displaysno uniform model but some common pattern between the models, including coexistingexcessive inflammation and immune suppression both as the causes of death. Data analysisof current clinical trials of some clinical strategies indicates some living hope ofanti-inflammatory strategies’ success and long way to go for immune-stimulation.Conclusion from historical development and failed trials clues a potential breakthroughfrom the disease polymorphism, pathogenesis complexity, time-point uncertainty, commonmechanism of self-injury and death channel.Urinary trypsin inhibitor (UTI) was selected as adjunct therapy against sepsis basedon the above analysis and its following effects: anti-inflammation against excessiveinflammatory response upstream in the sepsis pathogenesis; anti-oxidation againstoxidative stress injury downstream in the sepsis, enzyme-inhibition against enzymaticactivation of the inflammatory cascade upstream and enzymatic hydrolysis injury causedby inflammatory effector cells downstream.Part One. META Analysis of UTI against SepsisObjective: to investigate the clinical effect of UTI against sepsis.Methods: Cochrane protocol of systematic review was prepared ahead. Following theprotocol, literature search, screening, quality assessment, data extraction and pooling wereconducted in turn. GRADE system was used to evaluate the recommendation level andevidence power. Rank correlation and meta-regression was performed to further validatespecific program of UTI using and to screen the confounding factors may affectingmortalityResults:①Meta analysis showed that UTI treatment for sepsis could significantlyreduce the overall mortality [RR=0.58(0.44,0.76), P <0.0001(Z=3.93)] and28-daymortality [RR=0.51(0.37,0.71), P <0.0001,(Z=4.05)], and could alleviate the diseaseseverity of sepsis [standardized mean difference (SMD)=-0.77(-1.07,0.47), P <0.0001 (Z=5.0)];②GRADE recommended "very low" Level to the pooled effects;③rankcorrelation analysis showed a positive correlation (coefficient=-0.8660, P=0.0012)between the number of usage times (frequency) of UTI per day and28-day mortality;④Meta regression clued the constitution of primary disease may affect the efficacy of UTIwhen used for treating sepsis.[Data not shown fully here due to word limit]Conclusions:①All the pooled effects was graded as "very low" by GRADEsystem, indicating uncertain results with no value to be believed or denied. It could not bedetermined as the effective or ineffective medicine.②Increased frequency of UTI usingmay be more useful for treating sepsis.③UTI may be of better efficacy when applied tothe sepsis with trauma or pancreatitis as the primary disease.Part Two. Multi-Center Retrospective Analysis of UTI forExtensive Burns and Burn Sepsis TreatmentObjective: Based on conclusion of Part One "UTI may be of better efficacy whenapplied to the sepsis with trauma or pancreatitis as the primary disease", this study wasconducted to investigate UTI’s effect for the treatment of extensive burns and burn sepsis.Methods: A retrospective collection of multi-center data of extensive burns wasanalyzed with UTI use as "exposure" factor according retrospective cohort analysis.Univariate model was used to screen the related outcomes and potential confoundingfactors, followed by multivariate regression model to fulfill the secondary screening andremove the real confounding factors. Stratification analysis was used to determine thespecific confounding effects of these confounding factors.Results:①Univariate model showed UTI applications could improve overallsurvival of extensive burns [OR=3.168(2.010,4.992), P <0.001] and the subgroupsurvival of burn sepsis [OR=3.722(1.627,8.515), P=0.001], reduce MODS incidence[OR=0.505(0.305,0.836), P=0.007], complication incidence in cardiovascular system[OR=0.501(0.322,0.779), P=0.002], in respiratory system [OR=0.582(0.385,0.881),P=0.010] and in urinary system [OR=0.538(0.296,0.977), P=0.039]. But sevenpotential confounding factors (anti-oxidative drugs, insulin, albumin, liver protectivemedicine, immunoglobulin, Traditional Chinese medicine, thymosin) were meanwhileshown significant relation with UTI.②UTI could improve overall survival in patientswith extensive burns [OR=2.097(1.077,4.086), P=0.029] and in non-sepsis subgroup [OR=3.147(1.192,8.304), P=0.021], reduce complication risk in respiratory system[OR=0.146(0.043,0.488), P=0.002], cardiovascular system [OR=0.460(0.249,0.848),P=0.013]. No significant effect was tested on the survival rate of burn sepsis.③Stratification analysis showed a statistical benefit of UTI ’[OR=4.77(1.39,16.41), P=0.017] when liver protective medicine was not used in burn sepsis subgroup, but no value[OR=2.02(0.59,6.96), P=0.260] when liver protective medicine used.[Data not shownfully here due to word limit]Conclusions:①UTI could improve overall survival in patients with extensiveburns and non-sepsis subgroup.②No significant benefit was found when UTI wasapplied for treating burn sepsis. Liver protective medicine was the cofounding factor,covering part effect of UTI, thus resulting in a difference between the survival rates ofsepsis and non-sepsis burns. And the covering effect of liver protective medicine on UTIwas more obvious in sepsis subgroup than that in non-sepsis subgroup.③UTI couldreduce the risk of complications in the respiratory system, cardiovascular system,especially in respiratory c system (reduced incidence risk by7times). Basic condition ofburns (including the burn size, depth, concurrent injuries, especially inhalation injury) andprehospital fluid resuscitation were still important factors in determining prognosticoutcome of extensive burns. Detailedly, burn shock at admission was a powerful indicatorpredicting poor prognosis (increased risk of death by8times). Antioxidant treatment wasthe most commonly used adjuvant therapy in the treatment of extensive burns; respiratorycomplication was the commonest one in all the complications. Liver protective medicineimproved survival rate significantly in sepsis or non-sepsis subgroup and showed a greatefficacy in decreasing the incidence of MODS.Part Three. UTI on Sepsis and Sepsis-Related Lung Injury inLow Acute Phase Reaction MiceObjective: Based on conclusion of Part Two "Liver protective medicine was thecofounding factor, covering part effect of UTI ", to validate the relationship between UTIand hepatic function, we did the present study.Methods: Establishing sepsis model in Smad3knock-out mice using cecal ligationand puncture procedure and validating the low level of acute phase reaction were thefoundation of all the next experiment and finished first. UTI treatment was used in knock-out mice suffering cecal ligation and puncture and the mortality change, serum levelof some indicators and sepsis-induced lung injury was observed24hours after modelsetting. Dose-response effect of UTI and comparison between UTI andalpha-1-antitrypsin (A1AT as representative complementarities of acute phase protein) wasconducted to further explore the therapeutic efficacy of UTI and its interaction with hepaticacute phase reaction.Results: cecal ligation and puncture at the one third distal cecum was a proper modelfor Smad3knockout mice to obtain the predestined mortality rate and biological effects ofexcessive inflammation, oxidative stress and enzyme activation. Digital expressionprofiling of liver acute phase protein showed that compared to wild-type mice, Smad3knockout mice had a less number and a lower degree in mRNA changing of all the acutephase proteins, proving that Smad3knockout mice were low acute phase reaction model.UTI could greatly improve the survival rate of septic mice with low acute phase reaction(61.5%vs.23.1%, Log-rank chi-square test shows P=0.020), with lower serum level ofTNF-alpha, IL-6, MDA, SOD and elastase activity, but with no significant difference inwild type survival. In lung tissue, UTI could reduce the transcriptional level of IL-1betaand IL-6and the translational level of TNF-alpha and IL-6, decrease lung tissue SOD andelastase activity thus causing significant decrement of pathological score of lung injury.Enzymatic activity testing in vitro confirmed the quantitative relationship betweenA1AT and UTI (1unit of UTI=2.5micrograms A1AT).50kU/kg UTI and0.125g/kgA1AT treatment after sepsis modeling showed similar results between the two survivalcurves (P==0.784). Doses-response effect of UTI (0kU/kg,10kU/kg,50kU/kg,100kU/kg) displayed a statistical linear trend although with no significant difference inLog-rank test.[Data not shown fully here due to word limit]Conclusions:①UTI could improve sepsis survival in mice with low hepatic acutephase reaction by a dose-response model and roof effect, maybe involving themechanisms of its anti-inflammatory, anti-oxidant, enzyme-inhibition property.②UTIcould alleviate sepsis-related lung injury occurring after CLP in low acute phase reactionmice, by mechanisms of anti-inflammatory, anti-oxidant and enzyme inhibition effects.③There are similarities and common mechanisms against sepsis pathogenesis between acutephase reaction and UTI treatment, so that when in the same application simultaneously, theacute phase reaction would undermine some efficacy of UTI treatment against sepsis.④Hepatic acute phase protein A1AT could also improve the sepsis survival in low acute phase reaction mice. The same inhibition of enzyme activity may predict the sameimprovement in survival rate. So UTI could be considered as an alternative to A1AT.⑤Hepatic acute phase reaction is the mechanism by which Smad3gene protects the body,and maintains the resistance against sepsis and Smad3knockout mice could be used as alow hepatic acute phase reaction model.