Type IV collagenase(s) and TIMPs modulate endothelial cell morphogenesis in vitro.
It has been proposed that proteases are important in endothelial cell behavior. We examined the contribution of the gelatinase/type IV collagenase system in an in vitro model of endothelial differentiation. Human umbilical vein endothelial cells rapidly align and form networks of tubes when cultured on a basement membrane preparation, Matrigel. Zymograms of culture supernates demonstrate a 72-kD and a 92-kD gelatinase activity; the cells produce most of the 72-kD gelatinase, whereas the 92-kD activity is derived entirely from the Matrigel. Addition of antibodies against type IV gelatinase/collagenase decreases the area of the tube network. Both tissue inhibitors of metalloproteinases, TIMP-1 and TIMP-2, similarly decrease tube formation when added to cultures. Conversely, exogenous recombinant 72-kD gelatinase increases tube-forming activity. The effects of the anti-gelatinase antibodies and the TIMPs are not additive. Inhibition by either antibodies or TIMPs is greatest when they are added at culture initiation, suggesting that the protease activity is important in the early steps of morphogenesis. However, culture of the cells on Matrigel does not increase early expression of mRNA for the 72-kD gelatinase. Expression of message for the enzyme actually decreases during the course of the assay, while transcription of mRNAs for TIMPs increases, further supporting the concept that collagenases facilitate an early event in tube formation. These data demonstrate that gelatinase/type IV collagenase activity is important in endothelial cell morphogenesis on Matrigel, and suggest a role for collagenases in formation of new capillaries in vivo.
Schnaper
H W
HW
Laboratory of Developmental Biology, National Institute of Dental Research, Bethesda, Maryland 20892.
Grant
D S
DS
Stetler-Stevenson
W G
WG
Fridman
R
R
D'Orazi
G
G
Murphy
A N
AN
Bird
R E
RE
Hoythya
M
M
Fuerst
T R
TR
French
D L
DL
D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology / Q000494:pharmacology; D001485:Basement Membrane / Q000201:enzymology; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology; D017364:Collagenases / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D004338:Drug Combinations; D004347:Drug Interactions; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects / Q000502:physiology; D006023:Glycoproteins / Q000494:pharmacology*; D006801:Humans; D007797:Laminin / Q000494:pharmacology; D020780:Matrix Metalloproteinase 9; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009024:Morphogenesis / Q000502:physiology; D009363:Neoplasm Proteins / Q000494:pharmacology*; D011509:Proteoglycans / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology; D013997:Time Factors; D019716:Tissue Inhibitor of Metalloproteinase-2; D019714:Tissue Inhibitor of Metalloproteinases; D014471:Umbilical Veins / Q000166:cytology
D000906:Antibodies; D004338:Drug Combinations; D006023:Glycoproteins; D007797:Laminin; D009363:Neoplasm Proteins; D011509:Proteoglycans; D012333:RNA, Messenger; D011994:Recombinant Proteins; D019714:Tissue Inhibitor of Metalloproteinases; C059485:matrigel; D019716:Tissue Inhibitor of Metalloproteinase-2; D003094:Collagen; D017364:Collagenases; D008666:Metalloendopeptidases; D020780:Matrix Metalloproteinase 9