Najia Jin

H2O2-induced egr-1, fra-1, and c-jun gene expression is mediated by tyrosine kinase in aortic smooth muscle cells

Abstract Hydrogen peroxide (H2O2) has recently been shown to have a dual effect on cell growth by stimulating proliferation and triggering apoptosis. Apoptosis induced by H2O2 is a direct consequence of oxidant injury, while the proliferative response to H2O2 is thought to be a protective mechanism against oxidant injury. Signaling of the H2O2-induced proliferative effect has been proposed to occur via the activation of mitogen-activated protein kinase (MAPK) and increase in expression of transcription factors. In the present study, H2O2-induced mitogenic signaling in aortic smooth muscle cells (ASMC) was investigated with a specific focus on the roles of tyrosine kinase and tyrosine phosphatase in the regulation of the H2O2-stimulated egr-1, fra-1, and c-jun transcription. The results show that H2O2-induced increases in egr-1, fra-1, and c-jun mRNA levels, as measured by Northern blot analysis, are time and dose dependent with the peak of the response within 2 h. Tyrosine kinase inhibitors (genistein, amino-genistein, and tyrphostin 51) significantly attenuated H2O2-induced expression of these genes and a tyrosine phosphatase inhibitor (perox-vanadate) stimulated their expression. H2O2 stimulated tyrosine kinase activities and caused protein tyrosine phosphorylation, which was blocked by tyrphostin 51. H2O2 also caused tyrosine phosphorylation of platelet derived growth factor (PDGF) receptor. These data show that H2O2 increases egr-1, fra-1, and c-jun mRNA levels in vascular smooth muscle cells, and the increase in expression of these genes is mediated by activation of tyrosine kinase. Our data also provide evidence that the H2O2-induced mitogenic response is, in part, mediated through the receptor tyrosine kinase, PDGF receptor.

Influence of microtubules on vascular smooth muscle contraction

Microtubules are ubiquitous in eukaryotic cells and play key roles in many cellular activities. The purpose of this study was to investigate the influence of microtubules on vascular smooth muscle contraction. Quantitative immunocytochemical analysis of rat aortic tissue revealed that, relative to the control group, colchicine (15 μM, 90 min) and nocodazole (15 μM, 90 min) decreased the microtubule density by 40–50% while taxol (10 μM, 90 min) increased the microtubule density by 33%. Isometric contraction studies demonstrated that both colchicine and nocodazole caused an upward shift in the phenylephrine (10−8 to 10−5 M) dose–response curve while taxol caused no significant change when compared to the control group. Potassium chloride (30 mM) induced 55 ± 5% P0 contraction in DMSO treated vessel rings. The active tension increased to 73 ± 5% P0 and 71 ± 6% P0 after pretreatment of the aortic rings with colchicine or nocodazole, respectively. Taxol did not cause a significant change in the active tension (56 ± 7% P0). These results indicate that microtubule depolymerization enhances isometric contraction of vascular smooth muscle and this enhanced contraction is not receptor dependent. Pretreatment of the aortic rings with an inhibitor of nitric oxide synthase (NOS) (Nω-nitro-L-arginine) did not change the increased contractile response to phenylephrine due to microtubule depolymerization suggesting that this phenomenon is not mediated by endothelium dependent relaxation.

Microtubule disruption modulates the Rho-kinase pathway in vascular smooth muscle

Microtubules constitute one of the main cytoskeletal components in eukaryotic cells. Recent studies have shown that microtubule disruption induced significant vasoconstriction or enhanced agonist-induced contraction in vascular smooth muscle. However, the underlying mechanisms are not clear. We hypothesize that microtubule disruption may affect contractile signaling in vascular smooth muscle and lead to the enhanced contraction. The present study demonstrates that both colchicine and nocodazole induced a small but sustained contraction (4–6% P0) in rat aortic rings. This microtubule disruption-induced contraction was abolished by co-treatment with either HA 1077 or Y-27632, both of which are relatively specific Rho-kinase inhibitors. However, co-treatment with ML-9, an inhibitor of myosin light chain kinase, (MLCK) did not have a significant effect on the colchicine-induced contraction. The enhanced KCl-induced contraction due to treatment with colchicine was also blocked by inhibition of Rho-kinase, but not by inhibition of MLCK. These results indicate that microtubule disruption modulates contractile signaling in vascular smooth muscle, mainly through the Rho-kinase pathway, but not MLCK. Interestingly, the colchicine-enhanced, phenylephrine-induced contraction was not completely blocked by inhibition of Rho-kinase suggesting that other signaling pathways might also be involved.

Platelet Activating Factor Causes Relaxation of Isolated Pulmonary Artery and Aorta

Platelet activating factor (PAF) is a lipid chemical mediator produced by a variety of activated cells, including basophils, eosinophils, platelets, monocytes, endothelial cells, macrophages, and neutrophils (Tamura, 1987; Vercellotti, 1988). A number of pathophysiological responses, such as inflammatory and allergic reactions, bronchoconstriction, platelet aggregation, pulmonary hypertension, anaphylactic shock, endotoxic shock, and systemic hypotension are mediated by PAF (Benveniste and Vargaftig, 1983; Heffner, 1983a; 1983b; Terashita, 1985; 1987; Hanahan, 1986; Barnes, 1988). These pathological conditions are linked to changes in smooth muscle tone. For example, bronchoconstriction is due to the increase in tone of airway smooth muscle and hypotension is due to the decrease in tone of vascular smooth muscle.

Norepinephrine Stimulates Inositol Trisphosphate Formation in Rat Pulmonary Arteries

Although the discovery of the “phosphoinositide effect” occurred over 35 years ago, its mechanisms were explored only over the last decade. It now is clear that hydrolysis of phosphoinositides generates second messengers for multiple cellular functions when receptors are activated by a wide array of hormones and agonists in a variety of cell types (for review see Rana and Hokin, 1990). Early studies focused on the role of phosphoinositides on regulation of secretion or control of release of secretory cell contents (Hokin and Hokin, 1953, 1960; Freinkel, 1957; Hokin et al., 1958, 1963; Axen et al., 1983). In recent years, studies of the physiological effects of phosphoinositide hydrolysis have extended to such cell types and tissues as cerebral cortical slices (Kendall and Nahorski, 1984), sympathetic ganglia (Bone et al., 1984), adrenal glomerulosa cells (Kojima et al., 1986), rod outer segments (Brown et al., 1987), epithelium (Anderson and Welsh, 1990), leukocytes (Bradford and Rubin, 1986), skeletal muscle (Volpe et al., 1985), and smooth muscle (Akhtar and Abdel-Latif, 1980; Bielkiewicz-vollrath et al., 1987).