Cell Biology International Reports, Vol. 14, Abstracts Supplement 7990 233

P617 THE EXPRESSION OF THE VITRONECTIN RECEPTOR IS DOUN REGIHATED BY TNF-a AND IFN-s IN

HUMrAN ENDOTHELIAL CELLS . Pa0la Derllippl, Gabrlella Trufia, otlgl 1eho Stefanuto, Fiorella Altruda, Lorenzo Silengo, Guide rar0ne. Dipartimento dl Genetlca, Riolopia e C’limlca, Universita’ di Tcrino, Torino, Italy.

I i3 this woa k \<e show that two l,llpol-tant mediators of inflammation, TNF-a and IFN-T, alter the e~.pre5e,o,, of extracellular matrix I ecep tars (integrins) in cultured human endothelial cell5 (HEC). leadl,,g to a different Interactlow of HEC with extracellular matrix. HEC grown in standard co,-ldi tlons express at their sur-face t eceptol-e for flbrcnectin, collagen, laminin and vitronectin. After proloiiged exposul-e to THF-a and IFH-I (24-72 hours), the level of the vitrcnectin receptor (alohaV:beta3! at the cell sur-face wacj decreased by 70%, while the amounts of the beta1 integrin subunit I emained uncha,>ged. This effect a-equii.es the combined treatment with TNF-a and IFN-7 since each c,tukli>e used alone did Cl0 t induce s*gl,lficant alterations. The decreased expression of the

alphaV:beta3 complex at the i-e11 surface 15 dile to a selective effect of TNF-a and IFN-T on the svnthesis and processing of the beta3 protein. In fact, while tee levels of the mRNAs for both alphaV and beta3 are comparable 14) contl-ol and t#-eated cell5 , the svnthesis of the beta3 subunit was decreased by a

factor of xxi. The ,-ate of pa-ocessing of the beta3

was also specifically slowered and cytokine treated

ccl 1s accumulate an BGkDa precursor forul. T I re

svlthes;s and processing of the alphaV subunlt were lwchanged :i, i,toklhe treated cells. I id agreemen e with the decreased expressloi, of the receptor. c,toklne-treat& IiEC showed deem-eased abllitr tu ad?ere to vitronectln but adhered notmallv to fibzonectln #II collage,>.

P619 INTERACTION BETWEEN bFGF AND AN INTEGRIN- LIKE MOLECULE IN ENDOTHELIAL CELLS (EC) Marco Rusnati. Chiara Urbinati, Mirella

Belleri, Marco Presta. Giovanni Ragnotti. Gen. Pathol., Dept. Biomed. Sci. & Biotech., School of Medicine, Univ. of Brescia, 25123 Brescia, Italy

Basic fibroblast growth factor (bFGF), a 18,000 daltons mitogen, induces cell proliferation in EC. bFGF is known to interact with the heparan-sulfate of the extracellular matrix and with a well- characterized cell membrane receptor in EC. Here we report the first evidence that bFGF interacts also with an integrin-like molecule, this interaction being necessary for bFGF to exert its mitogenic activity in EC. bFGF contains two DGR sequences that are the inverse of the RGD sequence, a well known recognition site of many adhesive proteins. DGR and RGD-related peptides inhibit mitogenic activity and cell-adhesive capacity @S bFGF without affecting the binding of I-bFGF to its cell membrane receptor in EC. The data suggest that bFGF interacts in a RGD- dependent manner with an integrin-like molecule. The validity of the distinction between bFGF interaction with its canonic receptor and with an integrin-like molecule is substantiated by the fact that EGTA-induced calcium deprivation inhibits ~1~d4~~~~ve2s~P~~~~yt~fi~~G~e~~t no&,,‘;:

receptor. The characterization of the bFGF-integrin interaction will allow a better underst;z;ing of the mechanism(s) responsible bFGF-induced cell proliferation and motility.

P618 STUDY OF ExPI~E~YIOB OFe6$4 IBTBDRIN IN HUMAN LEE3 CARCIIIOMAS

Rite Faloioni, Rena.to Mariani Costantini, Pasquale Battista, Gabriella Zupi, Irene Ventura, Ada Sacohi Lab. Onoogeneai loleoolam, Regina Elena Cancer Institlkte, Rome - Italy

The expression of thea6b4 integrin complex ~86 analyzed in human lung osroinomas using two monoclg nal antibodies which recognise the integrin subunit a6 (mAb 135-l 3C) and P4 (mAb 439-9B). Immunoprecipit~ tion pattern obtained from human luw carcinoma cell lines demostrated that the a6 and theP4subunits were differentially expressed in carcinomas of different type. The Quantitative analysis on biopsies from primary lung tumor and nonal lung confirmed that the oomplex was differentially expressed in non-small cell versus small cell lung cancer and that it was also deteotable in normal lung at low levels. The highest l.evels ofa6S4 were found in moderately differentiated squsmoua cell oarcinomaa. By immunohistochamistry the (34 subunit was detectable in all the equemo.~ cell carcinoma and bdenocaroinomae tested but not in the small cell oancer. The pattern of immunoreactivity was con+ stent with the expected distribution of membrane glycoprotein and in squamous cell carcinomas was suggestive of the localization displayed by molec;

les involved in carcinom~t3troma interuotion. Partsally supported by AIRC and CBR-PF ‘Oncology’

P620 mAh SD9 REACIS WITH a,-INTEGIUN AND STAINS THE STRATUM SPINOSUM OF ADULT SKIN.

BartDcStrooper,SytvieVekemangGecrtCarmelict,MartineJaspers, BemadetteVanderSchue~ RoagRoug Wu,Jean-Jaque.sCaAian. Center for Human Genetic+ UaiversIty of Leuvea, Leuveu, Belgium.

The in&pins are a famiIy of celkell and eeII-extraceIIuIar matrix receptors. Each integrin is composed of au a- aad a O-subunit. Meg&s may ecmtaiu aa ideutieaI subunit, for iuatauas the “Very Late Antigens” ail share the same B+buit. We have used mAb DHl2, wbIc.b resets speeitIeaBy wItIt this Ii,-subunit, for the afBuity pmifieatkm of &grIas from placenta. I&ted material was evakmted on 7% poIya+mide geIs with siIver stabdug before and after reductmu, At least 5 proteiac compatible with a-subunits were idea&d. yields were between 100 aad 500 pg B,-complexes per placenta. mAbs were raised agaiust this material. Screening by Immuuoblottiag yielded 2 mAbs reactive with the same a-subunit and one mAb recogui+ the g,-subuait. One of the a-speeitic mAbs was subclouul aud used for tke af6aity pmifscatIon of the recognized antigen. Its N-terminus was identical Ia 10 out of 10 amino acids with tbat of the as-subunit of the humau fibronectia receptor. The mAb rcackd partIeuIarIy well ia ImmuuoblottIug with reduced and uuredueed as-subunit and a wide eeII aad tissue distribution of this protem was demonstrated. ‘Ike mAb reacted aho ia immuuocyto- chemistry with focal adhesion plaques iu skia fibrobhsts seeded on tibrouedia. Howcvcr, ia adult humaa skia the mAb stained strongiy the stratum sphtosum but not the basal layers. ‘Ibis coutrasts with previous redt.s obtahud with the &qedfic mAb DHI2 and with mAb 16 agaiust tke a+hmit( kindly provided by Dr. K. Yamada). It is possible that the stew qmAb cross reacts with a moleetde speeificaIIy induced during keratiacqte dIIfereutiatioa or that as-subunit is present iu the stratum spiuosum ia aa aberraut eonformation.