Differential gene expression of vascular smooth muscle cells. Detection byRNA arbitrarily primed polymerase chain reaction

Background: Vascular smooth muscle cells (VSMC) play an important role in t he development of restenotic lesions. However; regulation of proliferation, migration, and matrix synthesis of these cells is still poorly understood The aim of this study was to analyze gene expression of differently stimula ted bovine VSMC. Material and methods: RNA was isolated from stimulated bovine VSMC after di fferent time periods. For stimulation we used growth factors (platelet-deri ved growth factors PDGF-AA, PDGF-BB, basic fibroblast growth factor;) and a nitric oxide donating drug (sodium nitroprusside). Gene expression of stim ulated and control cells was analyzed by non-radioactive RNA fingerprinting (RNA arbitrarily primed polymerase chain reaction, RAP-PCR) and standard g el electrophoresis. Polymorphic fragments were sequenced and further charac terized. Results: By RAP-PCR we detected changes in the RNA fingerprint pattern of s timulated cells compared with unstimulated cells. Sequences of five fragmen ts out of 12 showed high homology to known human genes (serine-methyl-trans ferase, DUTT1, laminin B2, a newly cloned translational regulator (p97), an d a human expressed sequence tag). For laminin B2 we could confirm an upreg ulation after stimulation with growth factors at 1 and 6 hours and after st imulation with SNP at I hour in comparison to controls. For p97 we could sh ow a downregulation after stimulation with SNP: bFGF and PDGF-BB but not PD GF-AA. Conclusion: RAP-PCR is well suited for analysis of VSMC gene expression in vitro. The laminin B2 and p97 gene are differently expressed after growth f actor stimulation in bovine VSMC.