Preliminary Studies On Identification And Differentiation Of Epithelial Stem Cells In The Developing Human Fetal Lungs

As 15own to all,the level of fetal lung develoPment has a eloserelationshiP with the survival ability of the newbom infants.Themorphogenesis of the lung 15 eharacterized by the develoPment of alveoli andthe differentiation of lung ePithelial eells.Retardation of lung alveoli gro11esPeeially the imPaired alveolar ePithelialtyPe 11 eells(AECll)maturationduring develoPment and subsequent sllrietant syntllesis and seeretorydefieieney may lead to diminished lung size and eontnbute to thePathoPhysiogenesis of mally lung disease.Cystie fibrosis(CF),infalltresPiratory distress syndrome and ehronie lung disease(CLD)are severedisorders involving the lungs of the newboms and infants,eharaeterized byePithelialhyPerplasihyPoPlasia and metaplasia that likely eontribute to thedownward sPiral of Physiologie imPairment.All matllre ePithelial eells arederived from the stem eellsdng the embryonie lung develoPment.Elueidation of lung ePithelial stem eells and eell lineage 15 fundamentaltoward understanding meehanisms of lung develoPment and that alterePithelial PhenotyPe.From a theraPeutie standPoint,it 15 also imPortant toidentify stem eells with extensive Progenitorial capacity as Potential targetsfor gene theraPy sueh as CF and lung eareinoma.But Stem eell researeh inthe lung has Progressed rather slowly due to the anatomieal and funetionaleomPlexities associated with mimerous distinct cell tyPes.And there have nooPerative methods to identify the lung ePithelial stem cells till now.Evideneeelearly suggests that multiPotent Progenitors of the eondueting aipayePithelium and gas一exchange alVeolar regions are derived from differentPoPulations of stem eells that are anatomieally seParated in the lung.Inaddition,the existenee in the lung ofa'universal'PluriPotent eell has longbeen sPeeulated uPon and now some initial evidences has emerged with theidentifieation of a sPore一like eell that ean differentiate in vi.0 to bronehiolar 一6一tissue.DesPite many studies directed at understanding the groh ProPertiesof sPeeifie subsets of cells eonstituting the aipay ePithelium,a definitiveProgenitor--Progeny relationshiPs in the normal steady develoPing state oflltlllian fetal lung has not yet been fully elueidated.50 the goals ofthe PresentsdywereattemPttoidentifythePulmonaryePithelialstemeellsandtheirdistribution duringhman fetal lung develoPment through observing themorphogenetie eharaeters of ePithelial eells alld the exPression of diflrentProteins whieh serve an imPortant role in lung growth(TTF一1,SP一A and SP一B)or serve as a marker of stem cells(hTERT,CK19 and nestin).specialattention was foeused on the ehanges in the exPression of these Proteinsduring the develoPment from the Pseudoglandular(PG) and eanalieularstages to Primitive alveolar stages,and on the Possible ilnPlieations of theehanges for the normal fetal lung develoPment and for the signifieanee in theidentifieation of Pulmonary ePithelial stem eells.MsterialS alld MethodS Human lung tissues were available from 41 fetuses of 1034 weeksgestational age with Parental eonsent at indueing abortion with water bag andfrom0 newbom infants who died witllout Pulmonary eauses.Normal adulttissue was obtained from non~afleted Portions of two lung sPecimens takenat the time of lobeetomy.Lung tissues were fixed in4%PhosPhate一buft七redformalin,in most cases within 3 hour after death and the average time 15 6.5hour,dehydrated through graded ethanols,and embedded in Paraffin.Fivem thiek sections were eut and mounted on glass slides.Fortransmission eleetron mieroseoPy(TEM)lung samPles were taken from thePeriPheral Parenchyma and Prefixed in 2.5%glutaral,Posxed inl%osmium tetroxide,dehydrated via a graded aleohol series and then embeddedin fresh ePon.Ultrathin seetions were eut alld mounted on grids,stained withaqueous uranyl aeetate and alkaline lead eite,and examined with TEM at80kV.Immunohistoehemieal analysis was...