Abstract
Rat aortic vascular smooth muscle (VSM) cells synthesize the transmembrane proteoglycan syndecan (Cizmeci-Smith, G., Asundi, V., Stahl, R. C., Teichman, L. J., Chernousov, M., Cowan, K., and Carey, D. J. (1992) J. Biol. Chem. 267, 15729-15736). The present work demonstrated that VSM cells synthesize the related transmembrane proteoglycan fibroglycan and that increased expression of these two proteoglycans is stimulated under different conditions. Fibroglycan synthesis by cultured rat aortic VSM cells was demonstrated by Northern blot analysis with a rat fibroglycan cDNA probe and immunoblot analysis with anti-rat fibroglycan antibodies. Effects of growth factors and vasoactive substances on syndecan and fibroglycan expression were examined by Northern blot analysis. Syndecan mRNA levels increased in response to stimulation of VSM cells with serum, platelet-derived growth factor, or angiotensin II. VSM cells stimulated with platelet-derived growth factor contained more syndecan core protein and processed syndecan than control cells. Fibroglycan mRNA levels either decreased or remained unchanged in response to these agents. Fibroglycan mRNA levels increased following transforming growth factor-beta stimulation, while syndecan mRNA levels decreased. Other agents, including basic fibroblast growth factor, endothelin, and carbacyclin did not alter the expression of either proteoglycan. Syndecan and fibroglycan mRNA levels also varied as a function of cell density. These data demonstrate that syndecan and fibroglycan expression are regulated differently in VSM cells and lend support to the hypothesis that these proteoglycans carry out distinct physiological functions.
Highlights
The probes were labeled by random prime labeling
amplification usingnestedprimers based on the published human fibroblast fibroglycan sequence
The product of this reaction was used as the template
Summary
The probes were labeled by random prime labeling (Prime-a-Gene, Cloning of Rat Fibroglycan cDNA-A neonatal rat aorta Xgtll cDNA library (Clonetech Laboratories, Palo Alto, CA) was used to amplify rat fibroglycan core protein cDNA by polymerase chain reaction (PCR) amplification usingnestedprimers based on the published human fibroblast fibroglycan sequence (Marynen et al., 1989). CCGAATTCGAGGAGGAGAAGGAGGAGGACCCGGGG-3', and the antisense primer was 5'-CCGAATTCCGCATAAAACTCC'M"
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