Contribution of synthetic phenotype on the enhanced angiotensin II-generating system in vascular smooth muscle cells from spontaneously hypertensive rats

Objective We have demonstrated that cultured vascular smooth muscle cells ( VSMC) from spontaneously hypertensive rats (SHR), but not from normotensive Wistar-Kyoto (WKY) rats, produce angiotensin II (Ang II) in a homogeneous culture with increased levels of angiotensinogen, cathepsin D and angiotens in converting enzyme (ACE) at early passages. In the current study, we inve stigated how changes in the cell phenotype affect the Ang II-generating sys tem and the growth of VSMC from SHR. Design and methods We evaluated basal DNA synthesis by [H-3]thymidine incor poration, immunofluorescence of alpha-smooth muscle (SM) actin, mRNA expres sion of phenotype markers such as SM22 alpha appeared by contractile phenot ype, Ang II-generating system components and growth factors by reverse tran scription and polymerase chain reaction analysis, and Ang II levels by radi oimmunoassay in quiescent VSMC from WKY/lzumo rats and SHR/lzumo at passage s 4, 8 and 12. Results nasal DNA synthesis in VSMC from WKY rats increased with increasing passage number, whereas in cells from SHR it was markedly higher at early passages and was not affected by the passages. At early passage numbers, im munofluorescence of alpha-SM actin was stronger in VSMC from WKY rats than in cells from SHR, but decreased after several passages. Expression of SM22 alpha mRNA was higher in VSMC from WKY rats than in cells from SHR at earl y passages, and decreased after several passages in cells from both rat str ains. Expression of matrix Gla mRNA was higher in VSMC from SHR than in cel ls from WKY rats at early passage, and increased after several passages in cells from both rat strains. Ang II was not detected at early passages but increased in VSMC from WKY rats with increasing passage, whereas it was det ected in VSMC from SHR at early passages and did not change with the passag es. Expression of angiotensinogen mRNA was higher in VSMC from SHR than in cells from WKY rats, and was not affected by the passages. Expressions of c athepsin D and ACE mRNA were higher in VSMC from SHR than in cells from WKY rats at early passage, and were increased by the passages in VSMC from WKY rats. Expressions of transforming growth factor-beta 1, platelet-derived g rowth factor A-chain, and basic fibroblast growth factor mRNA were signific antly higher in VSMC from SHR than in cells from WKY rats, and were increas ed by the passages. Conclusion These data indicate that early in culture VSMC from SHR have the synthetic phenotype, whereas VSMC from WKY rats have the contractile pheno type which then changes to the synthetic phenotype after increased passage numbers, with increased expression of cathepsin D and ACE, which produce An g II, and increased expression of Ang II-related growth factors, which indu ce the exaggerated growth observed in VSMC from SHR. I Hypertens 1999, 17:1 099-1107 (C) Lippincott Williams & Wilkins.