Endostatin/collagen XVIII--an inhibitor of angiogenesis--is expressed in cartilage and fibrocartilage.
Aim of the study was to get a deeper insight in the mechanisms regulating avascularity of cartilaginious tissues. In the center of our interest was the expression of the anti-angiogenic fragment of collagen XVIII and its potency to inhibit angiogenesis. We observed a strong endostatin/collagen XVIII production in articular and fibrocartilage and an inhibitory potency concerning the VEGF-signalling pathway.


BACKGROUND
Cartilaginous tissue is mainly avascular and shows a limited intrinsic capacity for healing. Aim of this study was to investigate the expression of the antiangiogenic peptide endostatin/collagen XVIII in cartilage and fibrocartilage.


RESULTS
In fetal epiphyseal cartilage of humans high endostatin/collagen XVIII levels could be detected by ELISA whereas significantly lower levels were found in articular cartilage of adults. In the fibrocartilaginous tissue of the menisci, there was no significant difference in the endostatin/collagen XVIII concentrations between samples of fetuses and adults. But in the menisci of adults, endostatin/collagen XVIII concentrations were higher in the internal avascular two thirds of the meniscus whereas in the fetal menisci higher endostatin/collagen XVIII concentrations were found in the external third. Endostatin/collagen XVIII immunostaining of rat articular cartilage shows that endostatin/collagen XVIII downregulation starts soon after birth. In fetal cartilage and fibrocartilage of rats and humans, endostatin/collagen XVIII could be immunostained in the extracellular matrix and in the pericellular matrix of endothelial cells, fibrochondrocytes and chondrocytes. In adult cells, weak endostatin/collagen XVIII immunostaining was restricted to the pericellular matrix of fibrochondrocytes and chondrocytes. The detection of endostatin/collagen XVIII could be verified by in situ hybridization. Chondrocytes in vitro released measurable amounts of endostatin/collagen XVIII into culture supernatants. Stimulation of chondrocytes with EGF, as an example of a growth factor, or dexamethasone had no influence on endostatin/collagen XVIII expression. Endostatin inhibited VEGF-induced phosphorylation of MAPK in chondrocytes.


CONCLUSIONS
The spatial and temporal expression of endostatin/collagen XVIII in cartilaginous tissue and its potency regarding inactivation of VEGF signalling suggests that this antiangiogenic factor is important not only for the development but also for the maintenance of avascular zones in cartilage and fibrocartilage.


METHODS
We analyzed the spatial and temporal expression of endostatin/collagen XVIII--an endogenous angiogenesis inhibiting factor--in cartilage and fibrocartilage of humans and rats by immunohistochemical and biochemical (ELISA) methods and by in situ hybridization. To elucidate possible factors responsible for the induction or suppression of endostatin/collagen XVIII in cartilaginous tissues, chondrocytes (cell line C28/I2) were exposed to EGF and dexamethason. To study the possible interaction of endostatin/collagen XVIII with angiogenic factors, the immortalized human chondrocytes (C28/I2) have been incubated with VEGF and the phosphorylation of the MAPK Erk 1/2 (extracellular-regulated kinases), a known signal transduction pathway for VEGF has been determined under the influence of endostatin.
Pufe
Thomas
T
Department of Anatomy, Christian-Albrechts-University, Olshausenstrasse 40, D-24098 Kiel, Germany. t.pufe@anat.uni-kiel.de
Petersen
Wolf J
WJ
Miosge
Nicolai
N
Goldring
Mary B
MB
Mentlein
Rolf
R
Varoga
Deike J
DJ
Tillmann
Bernhard N
BN
D000375:Aging / Q000378:metabolism; D020533:Angiogenesis Inhibitors / Q000378:metabolism*; D000818:Animals; D002356:Cartilage / Q000378:metabolism*; D002358:Cartilage, Articular / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism; D043170:Collagen Type XVIII / Q000235:genetics / Q000378:metabolism*; D043169:Endostatins / Q000378:metabolism* / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005333:Fetus; D006801:Humans; D008592:Menisci, Tibial / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis; D009994:Osmolar Concentration; D010766:Phosphorylation / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D020533:Angiogenesis Inhibitors; D043170:Collagen Type XVIII; D043169:Endostatins; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; D009569:Nitric Oxide; D048049:Extracellular Signal-Regulated MAP Kinases