Pathogenesis of abdominal aortic aneurysms: possible role of differential production of proteoglycans by smooth muscle cells.
OBJECTIVE
In vivo and in vitro observations strongly suggest that marked differences exist in the phenotype, growth, and matrix-producing capabilities of distinct smooth muscle cell subpopulations. An earlier study from our laboratory showed differences in matrix metalloproteinase expression patterns in cultures of medial smooth muscle cells from tissue affected by abdominal aortic aneurysm (AAA) or atherosclerotic occlusive disease and from normal arterial tissue. In this study we were interested in ascertaining whether smooth muscle cells from the same sample groups also synthesized different proteoglycan profiles that correlated with vascular disease.


METHODS
Proteoglycans from smooth muscle cell monolayer cultures from tissue affected by AAA or atherosclerotic occlusive disease and from normal arterial tissue were examined by means of immunoblotting and affinity-blotting composite agarose polyacrylamide gel electrophoresis (CAPAGE) and sodium dodecyl sulphate PAGE. Enzyme-linked immunosorbent assay (ELISA) was used to quantitate perlecan levels in smooth muscle cell monolayer media samples.


RESULTS
Versican, perlecan, and biglycan levels were significantly elevated in AAA smooth muscle cell cultures. Two populations of smooth muscle cell versican were identified by means of CAPAGE-immunoblotting and by means of a novel affinity-blotting technique with biotinylated hyaluronan. A small keratan sulfate-substituted proteoglycan was present in similar levels in all smooth muscle cell cultures. This proteoglycan had a free core protein of about 55 kd after keratanase digestion and had a relatively high charge-to-mass ratio, as was evident from its electrophoretic mobility in CAPAGE; this proteoglycan was tentatively identified as keratocan. Immunoblotting with monoclonal antibodies 3-G-10 (anti-delta heparan sulfate, heparan sulfate stubs generated by heparitinase treatment) and 10-E-4 (anti-native heparan sulfate chains) helped identify several smooth muscle cell heparan sulfate-substituted proteoglycans. Elevated levels of intact and processed perlecan core protein were identified in AAA cultures by means of immunoblotting with a monoclonal antibody to perlecan core protein (A76). ELISA measurements confirmed that perlecan levels were significantly higher in AAA smooth muscle cell cultures compared with the normal arterial tissue and tissue affected by atherosclerotic occlusive disease.


CONCLUSIONS
Because heparan sulfate proteoglycans can bind growth factors, their elevated synthesis by AAA smooth muscle cells in combination with an increased expression of matrix metalloproteinases may at least partly explain the differential proliferative capacity of the AAA smooth muscle cells examined and may govern the pattern of abnormal cellular proliferation and matrix protein synthesis observed in the pathogenesis of vascular disease.
Melrose
J
J
Department of Surgery, The University of Sydney at The Royal North Shore Hospital, St Leonards, New South Wales, Australia.
Whitelock
J
J
Xu
Q
Q
Ghosh
P
P
D000368:Aged; D000369:Aged, 80 and over; D001012:Aorta, Abdominal / Q000378:metabolism; D017544:Aortic Aneurysm, Abdominal / Q000378:metabolism*; D001161:Arteriosclerosis / Q000378:metabolism; D058578:Biglycan; D002478:Cells, Cultured; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D016326:Extracellular Matrix Proteins; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000032:analysis; D006801:Humans; D015151:Immunoblotting; D037181:Lectins, C-Type; D008875:Middle Aged; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D011509:Proteoglycans / Q000032:analysis / Q000096:biosynthesis*; D053675:Versicans
D058578:Biglycan; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D037181:Lectins, C-Type; D011509:Proteoglycans; D053675:Versicans; C546884:BGN protein, human; C508322:VCAN protein, human; C071927:perlecan