il1 ifnγ对机械损伤后肝星状细胞迁移及生物学活性的影响-effects of il1 ifn γ on the migration and biological activity of hepatic stellate cells after mechanical injury.docx

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2018-05-29 发布于上海
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il1 ifnγ对机械损伤后肝星状细胞迁移及生物学活性的影响-effects of il1 ifn γ on the migration and biological activity of hepatic stellate cells after mechanical injury.docx

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il1 ifnγ对机械损伤后肝星状细胞迁移及生物学活性的影响-effects of il1 ifn γ on the migration and biological activity of hepatic stellate cells after mechanical injury

ABSTRACTLiverfibrosisisacommonprocessinthedevelopmentofchronicliverdiseasetocirrhosis,suchasviralhepatitis,schistosomiasis,andalcoholicliverdisease.Inliverfibrosis,theextracellularmatrix(especiallytypeIcollage)isexcessdepositionwhichbreakthedynamicblanceofmatrixproductionanddegradation.Thenthematrixstiffnesshasbeenchanged.However,liverfibrosiscanbereversed,soagreatdealofresearchhasbeenperformedtounderstandthemolecularmechanismsofliverfibrosis.Followingafibrogenicstimulus,Hepaticstellatecells(HSCs)changefromaquiescenttoanactivatedandbecamethemostimportantECM-producingcell.Thus,themechanismsofactivationandproliferationinHSCareattractingconsiderableattention.Thetissueinjuryiscellinjuryatall,itwouldresultintwokindsofpathologicalphenomenon:①functionalfailureinorgans②thetissue3-Dstructurealterationinducedbythespaceaftercelldeath.Thespacedefectsinducedbythecelldeathcanalsodestructmechanicalmicroenvironmentofcells.Thecellswillresponsetothissignal.Howthedifferencecellwillresponsetoinjuresignalwithdifferentcytokinestimuli.Inthisstudy,inordertomimicthespaceinvivo,weestablishedmechanicalscratchbythetipsinculturedmonolayerHSCs.Weusebio-mechanics,cellbiology,molecularbiology,immunohistochemistryandcomputerimageprocessingtoobservethedifferenceofHSC’sbioactivity.Theabbreviatedetailsareasfollow:WeimprovedtheHSCsisolationprotocolbasingonreferences.HSCswerepurifiedfrommaleSDratsbyoriginallyperfusion,sequentialdigestiontheliverwithpronaseandcollagenase,followedbydensitygradientcentrifugationwithOptiPrep.Thepurityofcultured-activatedHSCwasassessedbyimmunohistochemistryusingα-smoothmuscleactin;WeestablishedmechanicalscratchinculturedmonolayerHSCs.ThentheHSCsshoweddirectionalmigrationtothewoundarea.Themigrationdistancewithin12hafterscratchwasmuchhigherthan12-24hand24-36h.Thisindicatedthatthecapacityinproliferationandrepairwithin12hhadrelativelystrong.TheratioofSphaseinHSCswassignificantlyincreased,andtheexpressionofTGF-β1andtypeⅠcollagenmRNAwasup-regulated,indicatingthatmechanicaldamageef