Expression and Function of Osteopontin in Vascular Adventitial Fibroblasts and Pathological Vascular Remodeling

Osteopontin is known to play important roles in various diseases including vascular disorders. However, little is known about its expression and function in vascular adventitial fibroblasts. Adventitial fibroblasts have been shown to play a key role in pathological vascular remodeling associating with various vascular disorders. In this study, we measured activation of Osteopontin and its biological functions in cultured adventitial fibroblasts and injured rat carotid injury arteries induced by balloon angioplasty. Our results showed that angiotensin II and aldosterone increased Osteopontin expression in adventitial fibroblasts in a time- and concentration-dependent manner. MAPKs and AP-1 pathways were involved in Osteopontin upregulation. In addition, Adventitial fibroblast migration stimulated by Angiotensin II and aldosterone required OPN expression. Perivascular delivery of antisense oligonucleotide for Osteopontin suppressed neointimal formation post-injury. We concluded that upregulation of Osteopontin expression in adventitial fibroblasts might be important in the pathogenesis of vascular remodeling after arterial injury

Ion channel clustering enhances weak electric field detection by neutrophils: apparent roles of SKF96365-sensitive cation channels and myeloperoxidase trafficking in cellular responses

We have tested Galvanovskis and Sandblom’s prediction that ion channel clustering enhances weak electric field detection by cells as well as how the elicited signals couple to metabolic alterations. Electric field application was timed to coincide with certain known intracellular chemical oscillators (phase-matched conditions). Polarized, but not spherical, neutrophils labeled with anti-K v 1.3, FL-DHP, and anti-TRP1, but not anti-T-type Ca 2+ channels, displayed clusters at the lamellipodium. Resonance energy transfer experiments showed that these channel pairs were in close proximity. Dose-field sensitivity studies of channel blockers suggested that K + and Ca 2+ channels participate in field detection, as judged by enhanced oscillatory NAD(P)H amplitudes. Further studies suggested that K + channel blockers act by reducing the neutrophil’s membrane potential. Mibefradil and SKF93635, which block T-type Ca 2+ channels and SOCs, respectively, affected field detection at appropriate doses. Microfluorometry and high-speed imaging of indo-1-labeled neutrophils was used to examine Ca 2+ signaling. Electric fields enhanced Ca 2+ spike amplitude and triggered formation of a second traveling Ca 2+ wave. Mibefradil blocked Ca 2+ spikes and waves. Although 10 μM SKF96365 mimicked mibefradil, 7 μM SKF96365 specifically inhibited electric field-induced Ca 2+ signals, suggesting that one SKF96365-senstive site is influenced by electric fields. Although cells remained morphologically polarized, ion channel clusters at the lamellipodium and electric field sensitivity were inhibited by methyl-β-cyclodextrin. As a result of phase-matched electric field application in the presence of ion channel clusters, myeloperoxidase (MPO) was found to traffic to the cell surface. As MPO participates in high amplitude metabolic oscillations, this suggests a link between the signaling apparatus and metabolic changes. Furthermore, electric field effects could be blocked by MPO inhibition or removal while certain electric field effects were mimicked by the addition of MPO to untreated cells. Therefore, channel clustering plays an important role in electric field detection and downstream responses of morphologically polarized neutrophils. In addition to providing new mechanistic insights concerning electric field interactions with cells, our work suggests novel methods to remotely manipulate physiological pathways.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/46726/1/249_2005_Article_1.pd

Aldosterone Enhances IGF-I-Mediated Signaling and Biological Function in Vascular Smooth Muscle Cells

The IGF-I pathway and renin-angiotensin-aldosterone axis are both involved in the pathogenesis of hypertension and atherosclerosis, but no information is available about IGF-I and aldosterone interaction or their potential synergistic effects in vascular smooth muscle cells (VSMCs). The aims of this study were to investigate whether aldosterone influences IGF-I signaling and to determine the mechanism(s) by which aldosterone affects IGF-I function. Aldosterone resulted in significant increases in the Akt (1.87 ± 0.24, P < 0.001), MAPK (1.78 ± 0.13, P < 0.001), p70S6kinase (1.92 ± 0.15, P < 0.001), IGF-I receptor (1.69 ± 0.05, P < 0.01), and insulin receptor substrate-1 (1.7 ± 0.04, P < 0.01) (fold increase, mean ± SEM, n = 3) phosphorylation responses to IGF-I compared with IGF-I treatment alone. There were also significant increases in VSMC proliferation, migration, and protein synthesis (1.63 ± 0.03-, 1.56 ± 0.08-, and 1.51 ± 0.04-fold increases compared with IGF-I alone, respectively, n = 3, P < 0.001). Aldosterone induced osteopontin (OPN) mRNA expression and activation of αVβ3-integrin as well as an increase in the synthesis of IGF-I receptor. The enhancing effects of aldosterone were inhibited by eplerenone (10 μmol/liter), actinomycin-D (20 nmol/liter), and an anti-αVβ3-integrin antibody that blocks OPN binding. The antioxidant N-acetylcysteine (2 mmol/liter) completely inhibited the ability of aldosterone to induce any of these changes. In conclusion, our results show that aldosterone enhances IGF-I signaling and biological actions in VSMCs through induction of OPN followed by its subsequent activation of the αVβ3-integrin and by increasing IGF-I receptor. These changes are mediated in part through increased oxidative stress. The findings suggest a new mechanism by which aldosterone could accelerate the development of atherosclerosis