Rapid identification of differentially expressed endothelial cell genes by RNA display.
Endothelial cells line the inside of all blood vessels forming a structurally and functionally highly heterogenous population of cells. Here we describe the application of the differential RNA display technique to the analysis of the heterogeneity of endothelial cells. Multiple fragment cDNAs from quiescent resting and from activated migrating endothelial cells were amplified by RT-PCR using random 10mer 5' primers and T11XY 3' primers. Labelled amplification products were displayed on a sequencing gel. Expression patterns of more than 5000 bands of the two cell populations were approximately 98% identical. Of the differentially expressed bands, 26 fragment cDNAs were reamplified, sequenced, and used as probes for Northern blots. Approximately 50% of the analyzed fragment cDNAs could be confirmed as being differentially expressed by Northern blot analysis. Among the differentially expressed cDNAs was follistatin, which was exclusively expressed by migrating and not by quiescent arrested endothelial cells. Stimulation by exogenous bFGF, however, induced follistatin expression in arrested endothelial cells. These experiments support the use of the differential RNA display technique as a rapid cloning strategy for the identification of differentially expressed genes and suggest a role of the follistatin/activin system in the autocrine control of endothelial cell growth and differentiation.
Kozian
D H
DH
Dept. of Gynecology and Obstetrics, University of Göttingen Medical School, Germany.
Augustin
H G
HG
D028341:Activins; D000595:Amino Acid Sequence; D000818:Animals; D001013:Aorta, Thoracic; D015152:Blotting, Northern; D002417:Cattle; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D017931:DNA Primers; D018076:DNA, Complementary / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D038681:Follistatin; D015870:Gene Expression; D006023:Glycoproteins / Q000096:biosynthesis / Q000737:chemistry; D006133:Growth Substances / Q000096:biosynthesis; D006801:Humans; D007265:Inhibins / Q000096:biosynthesis / Q000737:chemistry; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D013552:Swine
D017931:DNA Primers; D018076:DNA, Complementary; D038681:Follistatin; D006023:Glycoproteins; D006133:Growth Substances; D012333:RNA, Messenger; D016222:Fibroblast Growth Factor 2; D028341:Activins; D007265:Inhibins