The Effects Of Extracellular-regulated Kinase Signaling Pathways On Functions Of Rat Hepatic Stellate Cells Stimulated By Acetaldehyde

Background and aim:In recent years,how the signal transduction inhepatiC stellate cellS(HSC)regulate has been still remained inradical debate．More researchers are paying close attentions toextracellular-regulated kinase(ERK)signaling pathways,as aconverge or common signal passageway,among others,who regulate theHSC proliferation and differentiation．It has been confirmed that theactivation of HSC triggered by direct action of acetaldehyde,as akey factor, resulting in extracelluar mattiX oversecretioncontributes to alcoholic hepatic fibrosiS in preyiOUS researchesHowever,the molecular mechanism of acetaldehyde-induced hepaticfibrosis has not hitherto been well known．We sought to investigatedacetaldehyde-induced change of ERK and the effect,raring withdifferent activated ERK,on proliferation,type I collagensynthesis,transforming growth factorβ_l(TGF-β₁)and Na⁺／Ca²+exchanger gene expresSion in rat HSC, which to show what a role ERKsignaling transduction pathway plays in rat HSC activated byacetaldehyde and to redound to clarify the pathogenesiS of hepaticfibrosiS and open the possibility to assess the future potentialantifibrotic treatment．Materials and Methods:HSC were isolated from rats liver by the methodof density gradient centrifugation after the liver tissue wasdigested in pronase E and IV collagenase．Pre-disposed in differentdose PD98059(20,50,100 μ mol／L),then st imulated by acetaldehyde,HSC were collected for detection of the ERK,proliferation,type Icollagen synthesis,transforming growth factor beta l and Na⁺／Ca2+exchanger mRNA expression,by means of Immunohistochemistry,MTT,cell-count,Insitu-hybridization and RT-PCR respectively．Results:(1)HSC were isolated successfully in our research,with thehigh cells' obtain and purity rate compared with traditionalmethod．(2)The ERK began to expressed in nuclei migrated fromcytoplasm,since HSC had been exposed in acetaldehyde for no more than5 minutes．The expression of ERK in nuclei grew to increase graduallyand got up to the maximum 20 minutes later whi le that in cytoplasmdescended step by step．The course as above could be blocked by PD98059with dosage dependence:PD98059 with a concentration of 20umol／hcould partiallY inhibit the ERK migration from cytoplasm to nuclei,and a apparently inhibition were shown at a concentration of 50umoI／L．When the PD98059 concentration got to lOOumol／L,the process was all but completely inhibited．(3)The cells proliferation,the I collagen secretion and the expression of α 1(I)collagen mRNA,TGFβ-lmRNA and Na⁺／Ca²+ exchanger mRNA were enhanced apparently after HSC were stimulated by acetaldehyde．After acetaldehyde stimulation,the measured value of MTT,cell counting,ELISA,in situ hybridization, RT-PCR and gel electrophoresiS assay increased apparently,the difference has statistical significant(P(0．01)comDared with pro-stimulation． (4)PD98059 can suppressed the HSC Droliferation,I collagen secretion and the expression of α 1(I)co1lagen mRNA and TGF β₁ mRNA in HSC stimulated by acetaldehyde．Theinhibition of PD98059 with a concentration of 20 μ mol／L was not rathersignificant (P>O．05),the inhibition of PD98059 with theconcentration of 50 μ mol／L and 100 u mol／L is significant(P+／Ca²+exchanger mRNA was not apparently affected byPD98059．Conclusions:(1)ERK has been activated significantly after HSC werestimulated by acetaldehyde．(2)Acetaldehyde can activate quiescentHSC and promote HSC proliferation,I collagen synthesiS,and theexpression of TGF β₁ mRNA and Na⁺／Ca²+exchanger mRNA．(3)ERK signal ingpathways in HSC stimulated by acetaldehyde play an important roleon r...