Caveolin-1 is known to promote cell migration, and increased caveolin-1 expression is associated with tumor progression and metastasis. In fibroblasts, caveolin-1 polarization and phosphorylation of tyrosine-14 are essential to promote migration. However, the role of caveolin-1 in migration of metastatic cells remains poorly defined. Here, caveolin-1 participation in metastatic cell migration was evaluated by shRNA targeting of endogenous caveolin-1 in MDA-MB-231 human breast cancer cells and ectopic expression in B16-F10 mouse melanoma cells. Depletion of caveolin-1 in MDA-MB-231 cells reduced, while expression in B16-F10 cells promoted migration, polarization and focal adhesion turnover in a sequence of events that involved phosphorylation of tyrosine-14 and Rac-1 activation. In B16-F10 cells, expression of a non-phosphorylatable tyrosine-14 to phenylalanine mutant failed to recapitulate the effects observed with wild-type caveolin-1. Alternatively, treatment of MDA-MB-231 cells with the Src family kinase inhibitor PP2 reduced caveolin-1 phosphorylation on tyrosine-14 and cell migration. Surprisingly, unlike for fibroblasts, caveolin-1 polarization and re-localization to the trailing edge were not observed in migrating metastatic cells. Thus, expression and phosphorylation, but not polarization of caveolin-1 favor the highly mobile phenotype of metastatic cells.

There is increasing evidence for rapid nongenomic effects of aldosterone. Aldosterone has been demonstrated to alter intracellular pH and calcium in isolated cells. However, few studies have correlated these effects with aldosterone-mediated physiological responses. Therefore, we studied rapid effects of aldosterone on vascular reactivity, intracellular Ca2+, and pH in resistance vessels. Furthermore, we explored whether the new antimineralocorticoid drug eplerenone could effectively block nongenomic aldosterone-mediated effects. The vasoconstrictor action of aldosterone was examined directly by determining the diameter of small resistance mesenteric vessels (160-200 microm resting diameter), simultaneously with intracellular pH or Ca2+. Aldosterone (10 nm) caused a rapid constriction of resistance vessels (8.1% +/- 1.0% reduction in the diameter below control conditions, P < 0.05). Aldosterone potentiated phenylephrine-mediated constriction in small and large mesenteric vessels. Aldosterone induced a rapid increase of intracellular Ca2+ and cellular alkalinization. Vasoconstrictor action of aldosterone and nongenomic effects on the sodium-proton exchanger (NHE1) activity or intracellular Ca2+ responses was abolished by eplerenone. The vasoconstrictor response of aldosterone was related to phosphatidylinositol 3-kinase (PI3-K): the hormone decreased protein kinase B phosphorylation; pharmacological inhibition of PI3-K (10 microm LY294002 or 1 microm wortmannin) increased arterial contractility. Inhibitors of ERK 1/2 phosphorylation (15 microm PD98059) had no effect on aldosterone-mediated vasoconstriction. Inhibition of protein kinase C with 1 microm bi-sindolylmaleimide I and/or inhibition of NHE1 with 100 microm amiloride abolished aldosterone vasoconstrictor action of resistance mesenteric arteries. We conclude that aldosterone-mediated increase in vascular tone is related to a nongenomic mechanism that involves protein kinase C, PI3-K, and NHE1 activity. Eplerenone is an effective blocker of nongenomic effects of aldosterone in vascular tissue.

Abstract-Recent studies suggested that type 2 angiotensin receptor (AT2R) could contribute to regulation of blood pressure and/or vascular remodeling. A key question relates to the effects of potential modulators of vascular AT2R expression. In the present work, we evaluated if high salt intake (70 mmol/L NaCl in drinking water) could modulate rat mesenteric artery AT2R function and expression. Angiotensin II dose-response curves were studied in rat perfused pressurized small-diameter arteries in the presence of losartan (AT1R antagonist Key Words: mineralocorticoid Ⅲ sodium Ⅲ hypertension Ⅲ vascular remodeling Ⅲ apoptosis T he renin-angiotensin-aldosterone system (RAAS) regulates vascular tone, body fluid volume, electrolyte balance, hormonal secretion, and neuronal activity. The biological effects of angiotensin II (Ang II), the main effector peptide in the vasculature, are mediated by at least 2 receptor isofoms. 1,2 The type 1 receptor (AT1R) mediates vasoconstriction, sympathetic facilitation, and trophic effects. The type 2 receptor (AT2R) is widely expressed during fetal development, whereas in the adult its expression has been detected in many different vessel types, including mesenteric, coronary, and renal arteries. 3-6 AT2R has opposite effects to those of AT1R, ie, it promotes cell apoptosis and inhibits cell proliferation. 6,7 AT2R also attenuates the pressor action of Ang II 8 and mediates vasodilation. 9,10 Recently, it has been shown that Ang II relaxes small mesenteric arteries via AT2R when AT1R are blocked. [11][12][13] Interestingly, the expression of AT2R is increased in several pathologic conditions such as vascular injury, 14 cardiac remodeling, congestive heart failure, and myocardial infarction. 15,16 It has been suggested that in adults the presence of AT2R in vascular tissues may be playing a role in vascular tone and/or tissue remodeling. [17][18][19] Therefore, a key and complex question that arises is how AT1R and AT2R expression are modulated. Several studies indicate interaction between Ang II and aldosterone, affecting the expression of ATRs. The expression of AT1R appears to be induced by Ang II in vascular smooth muscle, 4 and mineralocorticoids potentiate the action of Ang II in cultured rat vascular smooth muscle cells (VSMCs) by increasing the number of AT1R. 20,21 However, there are few data concerning the physiological regulation of AT2R expression. Dietary sodium depletion, which increases RAAS activity, enhances renal AT2 receptor function 10,22 and expression in both young and mature adult rats, mainly in the glomeruli and interstitial cells. 23 Induction of AT2R-mediated modulation of blood pressure was described in rats fed with a synthetic diet, an effect attributed to the stimulation of the RAAS. 8 Bonnet et al 24 have shown that Ang II infusion in the rat induces the expression of AT2R in the mesenteric vasculature. Nevertheless, it is not clear whether Ang II directly mediates the increased AT2R expression or if it is secondary to direct aldosterone action on arte...

Glycolytic metabolism in meiotic and post-meiotic spermatogenic cells shows differentiation-related changes. The developmental and physiological significance of these metabolic changes is not known. The aim of the present study was to test the hypothesis that glucose and lactate metabolism can modulate intracellular calcium [Ca2+](i) in spermatogenic cells in an opposing and dynamic manner. Fluorescent probes were used to measure [Ca2+](i) and pH(i), and HPLC was used to measure intracellular adenine nucleotides and mitochondrial sensing of ATP turnover. [Ca2+](i) in pachytene spermatocytes and round spermatids was modulated by changes in lactate and glucose concentrations in the media. The kinetics and magnitude of the [Ca2+](i) changes induced by lactate and glucose were different in meiotic and post-meiotic spermatogenic cells. The presence of glucose in the medium induced a decrease in pH(i) in spermatogenic cells. This glucose-induced pH(i) decrease occurred later than the changes in [Ca2+](i), which were also observed when the pH(i) decrease was inhibited, indicating that the glucose-induced [Ca2+](i) increase was not a consequence of pH(i) changes. Hexose phosphorylation in glycolysis was part of the mechanism by which glucose metabolism induced a [Ca2+](i) increase in spermatogenic cells. The sensitivity of [Ca2+](i) to carbohydrate metabolism was higher in round spermatids than in pachytene spermatocytes. Thus, differentiation-related changes in carbohydrate metabolism in spermatogenic cells determine a dynamic and differential modulation of their [Ca2+](i) by glucose and lactate, two substrates secreted by the Sertoli cells.

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