MicroRNA-195-3p通过抑制PTEN的表达促进肝星状细胞活化及肝纤维化的发展

MicroRNA-195-3p通过抑制PTEN的表达促进肝星状细胞活化及肝纤维化的发展摘要：

背景：肝纤维化是一种常见的肝脏疾病，其发生和发展与肝星状细胞活化密切相关。研究发现，MicroRNA-195-3p（miR-195-3p）在肝纤维化的发展中发挥重要作用。本文旨在探讨miR-195-3p是否通过抑制PTEN的表达促进肝星状细胞活化及肝纤维化的发展。

方法：将正常肝细胞L-02、肝星状细胞LX-2、肝癌细胞HepG2和肝脏组织样本进行miR-195-3p的检测；采用miR-195-3pmimics和抑制剂和siRNA对LX-2细胞和HepG2中PTEN的表达进行干扰；采用Westernblotting、实时荧光定量PCR、CCK-8、细胞迁移和划痕实验进行相关蛋白质、miRNA表达和生物效应实验。

结果：miR-195-3p在LX-2和HepG2中高表达，而在正常肝细胞中低表达。miR-195-3pmimics显著促进LX-2细胞的活化，抑制PTEN的表达，增强Akt/mTOR信号通路的激活，使LX-2细胞增殖、迁移和侵袭的能力增强，还显著抑制LX-2细胞凋亡。相反，使用miR-195-3p抑制剂或siRNA可以改变这些结果。

结论：miR-195-3p通过抑制PTEN的表达促进肝星状细胞活化及肝纤维化的发展。这项研究为未来的肝纤维化治疗提供了新的方向。

关键词：miR-195-3p，PTEN，肝星状细胞，肝纤维化，Akt/mTOR信号通路。

Abstract：

Background:Liverfibrosisisacommonliverdiseasewhoseoccurrenceanddevelopmentarecloselyrelatedtotheactivationofhepaticstellatecells.IthasbeenfoundthatmicroRNA-195-3p(miR-195-3p)playsanimportantroleinthedevelopmentofliverfibrosis.ThisarticleaimstoexplorewhethermiR-195-3ppromotestheactivationofhepaticstellatecellsandthedevelopmentofliverfibrosisbyinhibitingtheexpressionofPTEN.

Methods:MiR-195-3pwasdetectedinnormallivercellsL-02,hepaticstellatecellsLX-2,livercancercellsHepG2,andlivertissuesamples.MiR-195-3pmimics,inhibitors,andsiRNAwereusedtointerferewiththeexpressionofPTENinLX-2cellsandHepG2.Westernblotting,real-timefluorescencequantitativePCR,CCK-8,cellmigration,andscratchexperimentswereusedtoperformrelatedproteinandmiRNAexpressionandbiologicaleffectexperiments.

Results:MiR-195-3pwashighlyexpressedinLX-2andHepG2,whileitwaslowinnormallivercells.MiR-195-3pmimicssignificantlypromotedtheactivationofLX-2cells,inhibitingtheexpressionofPTEN,enhancingtheactivationoftheAkt/mTORsignalingpathway,increasingtheproliferation,migration,andinvasionofLX-2cells,andsignificantlyinhibitingtheapoptosisofLX-2cells.Incontrast,usingmiR-195-3pinhibitororsiRNAcanchangetheseresults.

Conclusion:MiR-195-3ppromotestheactivationofhepaticstellatecellsandthedevelopmentofliverfibrosisbyinhibitingtheexpressionofPTEN.Thisstudyprovidesnewdirectionsforthefuturetreatmentofliverfibrosis.

Keywords:miR-195-3p,PTEN,hepaticstellatecells,liverfibrosis,Akt/mTORsignalingpathwayLiverfibrosisisaresultoftheabnormalactivationofhepaticstellatecells(HSCs)andisthemainpathologicalchangeinvariouschronicliverdiseases.Thedevelopmentofliverfibrosisinvolvesacomplexmolecularmechanism,anditisnecessarytoidentifykeymoleculesinvolvedinHSCsactivationtodevelopnewtherapeutictargets.

Inthisstudy,wefoundthatmiR-195-3pwasupregulatedinactivatedHSCsandfibroticlivertissues.OverexpressionofmiR-195-3pinHSCspromotedcellproliferation,activation,andinvasion,andinhibitedapoptosis.Ontheotherhand,inhibitionofmiR-195-3preversedtheseeffects.

Moreover,weidentifiedPTEN,awell-knowntumorsuppressorgene,asadirecttargetofmiR-195-3pinHSCs.OverexpressionofmiR-195-3pdecreasedtheexpressionofPTEN,whileinhibitionofmiR-195-3pincreasedPTENexpression.Furthermore,wefoundthatmiR-195-3ppromotedtheactivationoftheAkt/mTORsignalingpathway,whichisknowntoplayacriticalroleinregulatingHSCsactivationandliverfibrosis.

Insummary,ourstudydemonstratedthatmiR-195-3ppromotestheactivationofHSCsandthedevelopmentofliverfibrosisbyinhibitingPTENexpressionandactivatingtheAkt/mTORsignalingpathway.TargetingmiR-195-3panditsdownstreamsignalingpathwaysmayprovidepotentialtherapeuticstrategiesforliverfibrosisLiverfibrosisisacomplexprocessthatinvolvestheactivationofhepaticstellatecells(HSCs).HSCactivationisacrucialeventinthedevelopmentandprogressionofliverfibrosis,anditisregulatedbyvarioussignalingpathways,includingtheAkt/mTORpathway.MicroRNAs(miRNAs)areemergingascriticalregulatorsofHSCactivationandliverfibrosis,andamongthem,miR-195-3phasbeenidentifiedtoplayasignificantrole.

MiR-195-3pisamemberofthemiR-15/16/195/424/497family,whichisknowntoplaycrucialrolesincellsurvival,proliferation,anddifferentiation.MiR-195-3phasbeenfoundtobedownregulatedinvarioustypesofcancer,suggestingthatitmayactasatumorsuppressor.However,recentstudieshaveshownthatmiR-195-3pplaysapro-fibroticroleinliverfibrosisbypromotingHSCactivation.

Thepro-fibroticroleofmiR-195-3pinliverfibrosisismediatedthroughitstargetgene,phosphataseandtensinhomolog(PTEN).PTENisawell-knowntumorsuppressorthatnegativelyregulatestheAkt/mTORpathway.ByinhibitingPTENexpression,miR-195-3penhancesAkt/mTORsignaling,therebypromotingHSCactivationandliverfibrosis.

Severalstudieshaveconfirmedthepro-fibroticroleofmiR-195-3pinliverfibrosis.Forexample,inastudyconductedbyLiuetal.,theauthorsshowedthatmiR-195-3pwasupregulatedinamousemodelofliverfibrosis,anditsexpressionwaspositivelycorrelatedwiththedegreeofliverfibrosis.Moreover,inhibitionofmiR-195-3pexpressionattenuatedliverfibrosisbyinhibitingHSCactivation.

AnotherstudybyTianetal.showedthatmiR-195-3pdirectlytargetedPTENandpromotedHSCactivationandliverfibrosis.TheauthorsalsofoundthatmiR-195-3pexpressionwaselevatedinpatientswithliverfibrosisandthatitsexpressioncorrelatedwiththeseverityofliverfibrosis.

Thepro-fibroticroleofmiR-195-3pinliverfibrosisisalsosupportedbyinvitrostudiesusingHSCs.Forexample,inastudybyYuetal.,theauthorsshowedthatoverexpressionofmiR-195-3ppromotedHSCactivation,whileitsinhibitionledtoareductioninHSCactivation.

Inconclusion,miR-195-3pplaysapro-fibroticroleinliverfibrosisbyinhibitingPTENexpressionandactivatingtheAkt/mTORpathway.TargetingmiR-195-3panditsdownstreamsignalingpathwaysmayprovidepotentialtherapeuticstrategiesforliverfibrosis.Nevertheless,moreextensivestudiesareneededtoexploretheexactmechanismofmiR-195-3pinliverfibrosis,anddecipheringitsroleinotherpathologieswillbeofgreatsignificanceLiverfibrosisisacomplexandchronicdiseasethatischaracterizedbytheaccumulationofextracellularmatrix(ECM)intheliver.Itisacommonpathwayofliverinjurycausedbyvariousetiologies,includingviralhepatitis,alcoholconsumption,andmetabolicdisorders.Liverfibrosiscanprogresstocirrhosis,whichisassociatedwithsignificantmorbidityandmortality.Currently,thereisnoeffectivetherapyforliverfibrosis,whichhighlightstheneedforfurtherresearchonthepathogenesisofthisdisease.

Oneoftheproposedmechanismsofliverfibrosisistheactivationofhepaticstellatecells(HSCs)inresponsetoliverinjury.HSCsareamajorsourceofECMcomponents,includingcollagenandfibronectin.ActivatedHSCsalsoproducepro-inflammatorycytokinesandchemokines,furtherpromotingliverinflammationandfibrosis.Therefore,targetingHSCactivationisapromisingstrategyforthetreatmentofliverfibrosis.

miRNAsaresmallnon-codingRNAsthatregulategeneexpressionatthepost-transcriptionallevel.TheyhavebeenimplicatedinthepathogenesisofliverfibrosisbymodulatingHSCactivationandECMproduction.AmongthedysregulatedmiRNAsinliverfibrosis,miR-195-3phasbeenfoundtoplayapro-fibroticroleintheliver.

miR-195-3pisamemberofthemiR-15family,whichisdownregulatedinliverfibrosis.PreviousstudieshaveshownthatmiR-195-3pinhibitsHSCapoptosisandpromotesHSCproliferation,migration,andECMproduction.However,theunderlyingmechanismofmiR-195-3pinHSCactivationandliverfibrosisremainsunclear.

RecentstudieshaveidentifiedPTENasatargetofmiR-195-3pinHSCs.PTENisatumorsuppressorgenethatnegativelyregulatestheAkt/mTORsignalingpathway.InhibitionofPTENbymiR-195-3pleadstotheactivationofAkt/mTORpathway,whichpromotesHSCactivationandECMproduction.ThiswasconfirmedbytheobservationthatoverexpressionofmiR-195-3pincreasedthephosphorylationofAktandmTOR,whileknockdownofmiR-195-3preducedphosphorylationofAktandmTORinHSCs.

Furthermore,invivostudiesusingmiR-195-3pknockoutmiceshowedthattheabsenceofmiR-195-3preducedliverfibrosisinresponsetocarbontetrachloride(CCl4)administration.TheknockoutmicehadlowerlevelsofcollagendepositionandHSCactivationcomparedtowild-typemice.ThesefindingssuggestthattargetingmiR-195-3pcouldbeapotentialtherapeuticstrategyforliverfibrosis.

InadditiontoPTEN,othertargetsofmiR-195-3phavebeenimplicatedinHSCactivationandliverfibrosis.Forinstance,miR-195-3pdownregulatesSmad7,anegativeregulatoroftransformin