Browsing by Author "Figueroa, XF"

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    Clonidine-induced nitric oxide-dependent vasorelaxation mediated by endothelial α2-adrenoceptor activation
    (2001) Figueroa, XF; Poblete, MI; Boric, MP; Mendizábal, VE; Adler-Graschinsky, E; Huidobro-Toro, JP
    1 To assess the involvement of endothelial alpha (2)-adrenoceptors in the clonidine-induced vasodilatation, the mesenteric artery of Sprague Dawley rats was cannulated and perfused with Tyrode solution (2 ml min(-1)). We measured perfusion pressure, nitric oxide (NO) in the perfusate using chemiluminescence, and tissue cyclic GMP by RIA.
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    Histamine reduces gap junctional communication of human tonsil high endothelial cells in culture
    (2004) Figueroa, XF; Alviña, K; Martínez, AD; Garcés, G; Rosemblatt, M; Boric, MP; Sáez, JC
    The regulation of gap junctional communication by histamine was studied in primary cultures of human tonsil high endothelial cells (HUTECs). We evaluated intercellular communication, levels, state of phosphorylation, and cellular distribution of gap junction protein subunits, mainly connexin (Cx)43. Histamine induced a time-dependent reduction in dye coupling (Lucifer yellow) associated with reduction in connexin43 localized at cell-cell appositions (immunofluorescence), without changes in levels and phosphorylation state of connexin43 (immunoblots). These effects were prevented with chlorpheniramine, an H-1 receptor blocker; indomethacin, a cyclooxygenase blocker; or GF109203X, a protein kinase C inhibitor. Treatment with phorbol myristate acetate, a protein kinase C activator, and 4bromo (4Br)-A23187, a calcium ionophore, mimicked the histamine-induced effects on dye coupling. 8Bromo-cAMP doubled the dye coupling extent and prevented the histamine-induced reduction in incidence of dye coupling. After 24-h histamine treatment, known to desensitize H, receptors, reapplication of histamine increased cell coupling in a way prevented by ranitidine, an H-2 receptor blocker. Thus, activation of H-1 and H-2 receptors, which increase intracellular levels of free Ca2+ and cAMP, respectively, may affect gap junctional communication in opposite ways. Stabilization of actin filaments with phalloidine diminished but did not totally prevent histamine-induced cell shape changes and reduction in dye coupling. Hence, the histamine-induced reduction in gap junctional communication between HUTEC is mediated by cytoskeleton-dependent and -independent mechanisms and might contribute to modulate endothelial function in lymphoid tissue. (C) 2004 Elsevier Inc. All fights reserved.
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    Stimulation of NO production and of eNOS phosphorylation in the microcirculation in vivo
    (2000) Durán, WN; Seyama, A; Yoshimura, K; González, DR; Jara, PI; Figueroa, XF; Boric, MP
    The role of nitric oxide (NO) in microvascular permeability is controversial, in part because the regulation of its endothelial constitutive synthase, eNOS, has been studied in vitro but not in vivo. Our study was designed to detect the morphologic and functional presence of eNOS and to test whether eNOS could be phosphorylated by platelet-activating factor (PAF), an agent that induces hyperpermeability. Immunocytochemistry was applied using human anti-eNOS antibodies in the hamster cheek pouch (hcp). The hcp microvessels demonstrated positive reaction products in the endothelium. The functional presence of eNOS in hcp was investigated by topical application of 10(-7) M PAF to the hcp and by measuring NO production by chemiluminescence. The mean baseline value of NO release was 63.3 +/- 6.9 pmol/ml (mean +/- SE), Application of PAF led to an increase in mean NO release to 120.8 +/- 31.2 pmol/ml (P < 0.05). In another series of experiments, 10(-7) M PAF was applied topically to hcp preincubated with [P-32]orthophosphoric acid. Immunoprecipitation and Western blots detected P-32-labeled bands that migrated with the mobility of positive eNOS indicating phosphorylated eNOS protein. The intensity of the radioactive bands was evaluated by computer;assisted image analysis. Comparison of the net band intensities yielded a mean PAF-treated/control ratio of 1.6 +/- 0.1. Our data demonstrate the morphologic and functional presence of eNOS in the microcirculation. The data also provide evidence that the function of microvascular eNOS is subject to regulation by phosphorylation. (C) 2000 Academic Press.