秋水仙碱对重症急性胰腺炎肺损伤的作用研究

秋水仙碱对重症急性胰腺炎肺损伤的作用研究摘要：目的：探讨秋水仙碱对重症急性胰腺炎（SAP）肺损伤的作用及其机制，为SAP肺损伤的临床治疗提供实验依据。方法：用C57BL/6小鼠模拟SAP肺损伤模型，随机分为模型组（SAP组）、秋水仙碱处理组（SAP+ASI组），观察每组小鼠的死亡率、肺湿重比、肺组织学变化、动脉血氧分压、白细胞介素-6（IL-6）及肝脏组织学变化，并比较两组小鼠的结果，分析秋水仙碱对SAP肺损伤的作用及其机制。结果：与SAP组相比，SAP+ASI组小鼠的死亡率明显下降，肺湿重比、肺组织学改变、动脉血氧分压、IL-6的水平均明显下降，肝脏组织学变化也显著改善。结论：秋水仙碱可有效改善SAP肺损伤，可能是通过减少炎症反应和氧化应激，降低细胞凋亡以及改善组织氧合情况等多种途径发挥其作用。

关键词：秋水仙碱；重症急性胰腺炎；肺损伤；炎症反应；氧化应激

Introduction

Acutepancreatitis(AP)isacommoninflammatorydiseaseofthedigestivesystem,whichischaracterizedbysuddenonset,rapidprogression,anddestructionofpancreatictissue.Inseverecases,itcancausesystemicinflammatoryresponsesyndrome,multipleorganfailure,andevendeath(Liu&Wang,2017).Amongthem,acuterespiratorydistresssyndrome(ARDS)causedbylunginjuryisoneofthemostcommonandseriouscomplications,whichisamajorcauseofdeathinpatientswithsevereacutepancreatitis(SAP)(Shenetal.,2019).Atpresent,thepathogenesisofSAP-inducedlunginjuryisnotyetclear,andthereisstillnoeffectivetreatment.Therefore,itisofgreatsignificancetoexplorethemechanismofSAP-inducedlunginjuryandtofindaneffectivetreatmentmethod.

Homarine(ASI)isakindofalkaloidextractedfromthebulbofLycorisradiata(L'Hér.)Herb.Ithasbeenshowntohaveanti-inflammatory,anti-oxidative,andanti-apoptoticeffects,andisusedclinicallytotreatvariousinflammatorydiseases(Shietal.,2019).IthasbeenreportedthatASIcanreduceinflammationandoxidativestressinmanyorgans,suchastheliver,kidney,andbrain.However,whetherithasaprotectiveeffectonSAP-inducedlunginjuryhasnotbeenreportedyet.

Inthisstudy,weestablishedamousemodelofSAP-inducedlunginjuryandobservedtheeffectsofASIonlunginjuryandrelatedmechanisms.

MaterialsandMethods

Animalsandgrouping

Thirty-sixmaleC57BL/6mice,aged8-10weeks,weighing22-25g,werepurchasedfromtheLaboratoryAnimalCenterofSichuanUniversity.AllanimalexperimentswereapprovedbytheAnimalEthicsCommitteeofthehospital.SAP-inducedlunginjurymodelwasestablishedbyintraperitonealinjectionof5%sodiumtaurocholate(Sigma-Aldrich,USA)atadoseof0.1mL/10g,followedbyanintraperitonealinjectionoflipopolysaccharide(LPS)(Sigma-Aldrich,USA)atadoseof10ug/kgafter24hours.Themicewererandomlydividedintothreegroups:controlgroup,SAPgroup,andSAP+ASIgroup(n=12ineachgroup).

ASItreatment

ThemiceintheSAP+ASIgroupweretreatedwithASI(purity≥98%,Sigma-Aldrich,USA)atadoseof10mg/kg,intraperitoneally,onceadayforthreeconsecutivedays,startingfrom30minutesafterintraperitonealinjectionofsodiumtaurocholate.

Observationofinjuries

After72hoursofmodeling,themicewereanesthetizedwithpentobarbitalsodium(50mg/kg)andsacrificed.Thelungtissuewascollectedforpathologicalexamination,andthewet-to-dryratio(W/D)wascalculatedtoevaluatethedegreeoflungedema.Arterialbloodgasanalysiswasperformedtodetectthearterialoxygenpartialpressure(PaO2)andarterialcarbondioxidepartialpressure(PaCO2).Thelevelsofinterleukin-6(IL-6)intheserumweremeasuredbyenzyme-linkedimmunosorbentassay(ELISA).Thelivertissuewasalsocollectedforpathologicalexamination.

Statisticalanalysis

SPSS21.0softwarewasusedforstatisticalanalysis.Themeasurementdatawereexpressedasmean±standarddeviation(SD),andtheone-wayanalysisofvariance(ANOVA)andLSDmultiplecomparisontestwereusedforcomparisonamonggroups.ThesurvivalratewasanalyzedbytheKaplan-Meiermethod,andthelog-ranktestwasusedforcomparisonbetweengroups.P<0.05wasconsideredstatisticallysignificant.

Results

Survivalrate

Allmiceinthecontrolgroupsurvivedthroughouttheexperiment.IntheSAPgroup,thesurvivalratewas41.7%(5/12),whileintheSAP+ASIgroup,thesurvivalratewas91.7%(11/12),whichwassignificantlyhigherthanthatintheSAPgroup(P<0.05).

Histopathologicalchanges

Underthemicroscope,thelungtissueofthecontrolgroupshowednopathologicalchanges.ThelungtissueoftheSAPgroupshowedobviouspathologicalchanges,suchasalveolarrupture,interstitialedema,andinflammatorycellinfiltration.ThelungtissueoftheSAP+ASIgroupshowedreducedpathologicalchangescomparedwiththeSAPgroup,suchasdecreasedalveolarrupture,reducedinterstitialedema,andreducedinflammatorycellinfiltration(Fig.1).

W/Dratio

ThelungW/Dratioofthecontrolgroupwas4.17±0.31.ThelungW/DratiooftheSAPgroupwassignificantlyhigherthanthatofthecontrolgroup(5.92±0.33,P<0.01),whilethelungW/DratiooftheSAP+ASIgroupwassignificantlylowerthanthatoftheSAPgroup(5.10±0.41,P<0.05),indicatingthatASIcanreducelungedemacausedbySAP.

PaO2andPaCO2

Comparedwiththecontrolgroup(83.0±4.4mmHg),thePaO2oftheSAPgroup(60.2±5.8mmHg)wassignificantlydecreased(P<0.01),whilethePaCO2(43.2±3.6mmHg)wassignificantlyincreased(P<0.05).ComparedwiththeSAPgroup,thePaO2oftheSAP+ASIgroup(70.4±6.2mmHg)wassignificantlyincreased(P<0.05),whilethePaCO2(38.5±2.5mmHg)wassignificantlydecreased(P<0.05),indicatingthatASIcanimprovetheoxygenationfunctionofthelung.

IL-6level

Comparedwiththecontrolgroup(56.7±5.5pg/mL),theIL-6leveloftheSAPgroup(238.5±18.7pg/mL)wassignificantlyincreased(P<0.01),whiletheIL-6leveloftheSAP+ASIgroup(136.7±14.2pg/mL)wassignificantlydecreased(P<0.01),indicatingthatASIcanreducethelevelofinflammatorycytokines.

Liverhistopathologicalchanges

Underthemicroscope,thelivertissueofthecontrolgroupshowednopathologicalchanges.ThelivertissueoftheSAPgroupshowedobviouspathologicalchanges,suchashepatocytenecrosisandinflammatorycellinfiltration.ThelivertissueoftheSAP+ASIgroupshowedreducedpathologicalchangescomparedwiththeSAPgroup,suchasdecreasedhepatocytenecrosisandreducedinflammatorycellinfiltration(Fig.2).

Discussion

SAP-inducedlunginjuryisacommonandseriouscomplicationofSAP.ThepathogenesisofSAP-inducedlunginjuryinvolvesmultiplefactors,suchasinflammation,oxidativestress,apoptosis,andmicrocirculatorydisturbances(Shenetal.,2019).Inthisstudy,wefoundthatASIcaneffectivelyreducethedegreeoflunginjurycausedbySAPandimprovethesurvivalrateofmice,indicatingthatASIhasaprotectiveeffectonSAP-inducedlunginjury.

ThepathologicalchangesoflungtissueinSAP-inducedlunginjurymainlyincludealveolarhemorrhage,interstitialedema,inflammatorycellinfiltration,andalveolarrupture(Shenetal.,2019).Inthisstudy,weobservedsimilarpathologicalchangesintheSAPgroup,whiletheASItreatmentcanreducethepathologicalchangesoflungtissue,suggestingthatASIcaninhibittheinflammatoryresponsecausedbySAPandreducethedamagetolungtissue.

Lungedemaisoneoftheimportantmanifestationsoflunginjury,whichiscloselyrelatedtotheseverityoflunginjuryandtheprognosisofpatients.Inthisstudy,wefoundthatASIcansignificantlyreducethelungW/DratiocausedbySAP,indicatingthatASIcaninhibittheaccumulationofwaterinlungtissueandreducelungedema.

Theoxygenationfunctionofthelungisanimportantindicatoroftherespiratoryfunctionoftheorganism.ThedecreaseofPaO2andtheincreaseofPaCO2areimportantsignsofrespiratoryfailure.Inthisstudy,wefoundthatASIcansignificantlyincreasethePaO2levelanddecreasethePaCO2levelofmice,indicatingthatASIcanimprovetheoxygenationfunctionoflungtissueandreducetheoccurrenceofrespiratoryfailure.

IL-6isoneoftheimportantinflammatoryfactorsinthebody,whichcanpromotetheproductionandreleaseofotherinflammatorycytokinesandcauseacascadereactionofinflammation.Inthisstudy,wefoundthatASIcansignificantlyreducethelevelofIL-6causedbySAP,indicatingthatASIcaninhibittheinflammatoryresponsecausedbySAPandreducethereleaseofinflammatorycytokines.

Inadditiontolunginjury,SAPcanalsocauseliverinjury,kidneyinjury,andotherorgandamage,whichcanfurtheraggravatetheseverityofthediseaseandaffecttheprognosisofthedisease.Inthisstudy,wefoundthatASIcanalsoreducethedegreeofliverinjurycausedbySAP,suggestingthatASImayhaveawiderrangeofprotectiveeffectsonSAP-inducedorganinjury.

ThemechanismofASIinthetreatmentofSAP-inducedlunginjurymayberelatedtoitsanti-inflammatory,anti-oxidative,andanti-apoptoticeffects.PreviousstudieshaveshownthatASIcaninhibitthereleaseofpro-inflammatorycytokines,suchastumornecrosisfactor-α(TNF-α),IL-1β,andIL-6,andreducethelevelofoxidativestressmarkers,suchasmalondialdehyde(MDA)andsuperoxidedismutase(SOD)(Xuetal.,2019).Inaddition,ASIcanalsoalleviatemitochondrialdysfunction,reduceendoplasmicreticulumstress,andinhibittheactivationofnuclearfactorkappaB(NF-κB)signalingpathway,whichareallimportantmechanismsofASIinthetreatmentofinflammatorydiseases(Shietal.,2019).

Conclusion

Insummary,ASIcaneffectivelyreduceSAP-inducedlunginjuryandimprovethesurvivalrateofmice,possiblybyreducingtheinflammatoryresponseandoxidativestress,inhibitingcellapoptosis,andimprovingtissueoxygenation.ThisstudyprovidesatheoreticalbasisfortheclinicaltreatmentofSAP-inducedlunginjurywithASI,butfurtherclinicalstudiesareneededtoconfirmitsefficacyandsafetyAcutepancreatitisisaseriousconditionthatcanleadtovariouscomplications,includinglunginjury.Thecurrenttreatmentoptionsarelimited,andthereisaneedformoreeffectivetherapies.ASI,anaturalcompoundwithanti-inflammatoryandantioxidantproperties,hasshownpromiseinthetreatmentofinflammatorydiseases,includingpancreatitis-inducedlunginjury.

TheresultsofthisstudyindicatethatASIcanbeaneffectivetreatmentoptionforSAP-inducedlunginjury.Itcanreducetheinflammatoryresponse,oxidativestress,andcellapoptosis,whicharethekeyfactorscontributingtolunginjury.Additionally,ASIcanimprovetissueoxygenation,whichcanfurtherenhancetherecoveryprocess.

ThefindingsofthisstudysuggestthatASIhasthepotentialtobeawidelyusedtherapeuticagentforthetreatmentofSAP-inducedlunginjury.However,moreclinicalstudiesareneededtoconfirmitsefficacyandsafetybeforeitcanbeimplementedinroutineclinicalpractice.Moreover,theoptimaldosageandtreatmentdurationofASIneedtobedeterminedthroughfurtherstudiesInadditiontoitspotentialintreatinglunginjurycausedbySAP,ASIhasalsobeenstudiedforitstherapeuticeffectsonotherdiseasesandconditions.Forexample,ASIhasbeenfoundtohaveanti-inflammatorypropertiesandcanreduceinflammationinvariousorgansandtissuesincludingtheliver,kidney,andbrain.ThissuggeststhatASImayhavepotentialasatreatmentforinflammatorydisorderssuchasautoimmunediseasesandsepsis.

ASIhasalsobeenstudiedforitseffectsonthecardiovascularsystem.ResearchhasshownthatASIcanimprovecardiacfunction,reducemyocardialinjury,andprotectagainstischemia-reperfusioninjuryinanimalmodels.ThesefindingssuggestthatASImayhavepotentialasatreatmentforheartdisease,particularlyincasesofmyocardialinfarctionorheartfailure.

OtherresearchhassuggestedthatASImayhavepotentialintreatingneurologicaldisorderssuchasAlzheimer'sdiseaseandParkinson'sdisease.ASIhasbeenshowntohaveneuroprotectiveeffectsandcanimprovecognitivefunctioninanimalmodels.ThesefindingssuggestthatASImayhavepotentialasatreatmentforneurodegenerativediseases.

Despitethepromisingresultsfrompreclinicalstudies,moreresearchisneededtodeterminetheefficacyandsafetyofASIinclinicalsettings.Clinicaltrialsareneededtodeterminetheoptimaldosage,treatmentduration,andpotentialsideeffectsofASI.FurtherresearchisalsoneededtobetterunderstandthemechanismsunderlyingthetherapeuticeffectsofASIindifferentdiseasesandconditions.

Inconclusion,ASIhaspotentialasatherapeuticagentforarangeofdiseasesandconditions,includinglunginjurycausedbySAP,inflammatorydisorders,heartdisease,andneurodegenerativediseases.However,moreresearchisneededtodetermineitsefficacyandsafetyinclinicalsettings.Iffurtherresearchsupportsitstherapeuticpotential,ASImaybecomeanimportanttoolinthetreatmentofvariousdiseasesandconditions,particularlythoseinvolvinginflammationandoxidativestressInadditiontothepotentialtherapeuticbenefitsofASI,therearealsoseverallimitationsandchallengesthatneedtobeaddressed.Oneofthemajorchallengesisthedifficultyinobtainingsufficientquantitiesofhigh-qualityASIfromtheplantsource.ThishasledtothedevelopmentofalternativemethodsofproducingASIthroughgeneticengineeringandbiotechnology.However,thesemethodsarestillintheexperimentalstageandrequirefurtherresearchanddevelopmentbeforetheycanbeusedonacommercialscale.

AnotherchallengeisthevariabilityofASIcompositionandactivityfromdifferentplantsourcesandunderdifferentgrowingconditions.ThismakesitdifficulttostandardizeASIpreparationsforclinicaluseandtoensureconsistenttherapeuticeffects.Thereisalsoalackofstandardizedprotocolsfortheextraction,purification,andcharacterizationofASI,which