One of the most critical cellular events in disease states such as vascular restenosis is the cytokine-induced activation of vascular smooth muscle cells (VSMCs) resulting in intimal thickening. Identification of the molecular mechanisms of VSMC activation is crucial in understanding the initiation of vascular restenosis. In this report, we show that one 14-3-3 protein family isoform, gamma, is transcriptionally up-regulated in rat carotid arteries after balloon angioplasty. 14-3-3 gamma protein induction localizes to both the media and neointima in such injured vessels. Because it has been shown that some members of the 14-3-3 family may play an important role in cellular proliferation by binding to and activating the protein kinase Raf-1 and VSMCs constitute the major cellular component of the restenotic lesion, we investigated the expression of this message in serum- and cytokine-stimulated human VSMCs. Both serum and selected cytokines induce 14-3-3 gamma mRNA and protein, the magnitude of which correlates with the degree of cellular stimulation. 14-3-3 gamma mRNA, however, does not increase when other cell types are stimulated with specific growth factors. Human tissue distribution of 14-3-3 gamma mRNA indicates that in contrast to other 14-3-3 proteins, the gamma isoform is highly expressed in VSMCs and skeletal and heart muscle, suggesting an important role for the gamma isoform in muscle tissue as well. These results indicate that 14-3-3 gamma expression increases in response to vessel damage and proliferative signals and may implicate a role for the gamma isoform of 14-3-3 in VSMC activation and metabolism.