G1 phase arrest of human smooth muscle cells by heparin, IL-4 and cAMP is linked to repression of cyclin D1 and cdk2.

Smooth muscle cell proliferation is a key event in the development of atherosclerosis. Inhibition of this proliferation may lead to better prevention and treatment of the disease. While a number of agents have been found to inhibit SMC proliferation, their mechanisms of action are not fully understood. We wanted to determine the effects of three physiologically relevant anti-mitogenic agents on two classes of proteins which have major roles in cellular proliferation, namely cyclins and cyclin-dependent kinases (cdks). Following stimulation with fetal calf serum (FCS), quiescent human umbilical artery smooth muscle cells (HUASMC) synthesised cyclin D1 mRNA and protein and cdk2 mRNA in the G1 phase, whereas cdc2 protein was expressed after the onset of the S phase. Heparin, a strong inhibitor of HUASMC proliferation, strongly down-modulated the levels of cyclin D1 mRNA and protein, cdk2 mRNA and cdc2 protein. Interleukin-4 (IL-4) or 8-bromo-adenosine 3',5'-cyclic monophosphate (cAMP) also lowered the levels of these cell cycle regulatory proteins, although their effects were relatively weak, reflecting their only partial inhibition of HUASMC DNA synthesis. There was specificity in the cell cycle targets of the agents since none appeared to affect the levels of cdk4 protein.

Interleukin-4 inhibits human smooth muscle cell proliferation.

Although proliferation of smooth muscle cells is a key event in the pathogenesis of atherosclerosis, the signals which regulate this proliferation are not fully understood. It is likely that proliferation is regulated by cytokines released by cells found in the plaque, such as T cells. In this study we report that the T cell-derived cytokine, interleukin-4 (IL-4), can inhibit proliferation of cultured human umbilical artery smooth muscle cells. Maximum inhibitory effect was achieved at IL-4 concentrations of 20 U/ml or greater. In addition, the data showed that IL-4 acted early in the G1 phase of the cell cycle, thereby preventing cells from entering S phase. The mechanism of IL-4 inhibition did not appear to involve stimulation of prostanoid synthesis since similar data were obtained when experiments were performed in the presence of a cyclooxygenase inhibitor. We propose that IL-4 may act as a protective factor released by T-cells in an atherosclerotic lesion in order to minimise the size of the plaque.

Human arterial smooth muscle cells synthesize granulocyte colony-stimulating factor in response to interleukin-1 alpha and tumor necrosis factor-alpha.

Vascular smooth muscle cells (SMC) are a major cell type comprising the walls of blood vessels. We report the synthesis of granulocyte colony-stimulating factor (G-CSF) by cultured human SMC obtained from the internal mammary artery and thoracic aorta. Interleukin-1 alpha (IL-1 alpha) greatly increased in a dose-dependent manner the amount of this cytokine produced by the SMC, with tumor necrosis factor-alpha (TNF-alpha) being less effective. Newly formed G-CSF could be detected in culture supernatants within 6 hours after IL-1 alpha or TNF-alpha treatment. Northern blot analysis of SMC stimulated with IL-1 alpha and TNF-alpha showed an increase in the amount of mRNA for G-CSF as compared with control cells. Enhanced G-CSF mRNA levels were observed when SMC were treated with cycloheximide in the absence or presence of added cytokine. In vasculitis, the walls of blood vessels become inflamed as evidenced by a leucocytic infiltrate usually dominated by polymorphonuclear neutrophil leukocytes (PMNs). G-CSF is known to stimulate PMNs, and our findings raise the possibility that G-CSF made by SMC contributes to the development of vasculitis lesions.