RESUMO
Hydrogen peroxide at concentrations below cytotoxic ones causes an increase in the cytoplasmic calcium concentration in human umbilical vein endothelial cells as a result of calcium release from intracellular stores. Two-pore calcium channel blocker trans-NED19 partially suppresses the increase in the level of calcium ions in the cells in response to the addition of hydrogen peroxide. The staining of endothelial cells with the fluorescent stereoisomer cis-NED19 and LysoTracker confirmed the localization of two-pore calcium channels in lysosomes and endolysosomal vesicles.
Assuntos
Canais de Cálcio/metabolismo , Cálcio/metabolismo , Citoplasma/efeitos dos fármacos , Citoplasma/metabolismo , Células Endoteliais da Veia Umbilical Humana/citologia , Peróxido de Hidrogênio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Carbolinas/farmacologia , Humanos , Piperazinas/farmacologia , PorosidadeRESUMO
The blocker of two-pore channels trans-NED 19 and hydrogen peroxide were found to inhibit histamine-induced relaxation of rat-aorta. The degree of inhibition depended on histamine concentration. The relaxation in response to I µM histamine of rat aorta preconstricted with 30 mM KCI, serotonin, or endothelin- 1, was completely abolished by 30 µM trans-NED 19. On the other hand, trans-NED 19 decreased the relaxation of the aorta in the presence of 10 µM histamine only by 2.1-fold to 2.4-fold, and there was almost no inhibition by trans-NED 19 of the relaxationinduced by 100 ptM histamine.) Relaxation of precontracted with serotonin aorta in response to 10 and 100 µM histamine was reduced by hydrogen peroxide (200 M) by 10- and 2.5-fold, respectively. Suppression of aorta relaxation by trans-NED 19 and H202 correlated with their inhibitory effect on the histamine-induced increase in the cytoplasmic free calcium concentration in human umbilical vein endothelial cells. With the use of a fluorescent probe LysoTracker, the cis-NED19 binding sites were demonstrated to be localized in endolysosomes of the endothelial cells. These data indicate that two-pore calcium channels participate in the histamine-induced endothelium-dependent relaxation of rat aorta. Furthermore, their functional role is exhibited much more clearly at low histamine concentrations. We suggest that hydrogen peroxide evokes depletion of intracellular calcium depots thereby suppressing the response to histamine.
Assuntos
Aorta/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Carbolinas/farmacologia , Células Endoteliais/metabolismo , Peróxido de Hidrogênio/farmacologia , Piperazinas/farmacologia , Vasodilatação/efeitos dos fármacos , Animais , Aorta/fisiopatologia , Histamina/farmacologia , Masculino , Ratos , Ratos WistarRESUMO
The ratio of early apoptosis and late apoptosis (necrosis) in the cultured human umbilical vein endothelial cells was estimated after exposure to hydrogen peroxide (H2O2) in vitro trying to keep them close to the physiological conditions (high cell density, high serum content, H2O2 concentration not over 500 µM). Cell viability was assessed using flow cytometry and simultaneous staining with fluorescent dyes PO-PRO-1 to detect early apoptotic cells, and DRAQ7 to detect late apoptotic and necrotic cells. The data obtained suggest that the primary mechanism of cytotoxic response is apoptosis. The critical concentration of H2O2 causing the death of the cell population in a dense monolayer is 250 µM. Lower concentrations of H2O2 (up to 200 µM) cause death of individual cells; however, viability of endothelial cell population is retained, and response to calcium activating agonists does not change compared with control cells.
Assuntos
Apoptose/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Necrose/induzido quimicamente , Antraciclinas , Antígenos CD/genética , Antígenos CD/metabolismo , Apoptose/genética , Benzoxazóis , Biomarcadores/metabolismo , Cálcio/metabolismo , Agonistas dos Canais de Cálcio/farmacologia , Contagem de Células , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Fator VIII/genética , Fator VIII/metabolismo , Citometria de Fluxo , Corantes Fluorescentes , Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Indóis/farmacologia , Necrose/genética , Necrose/metabolismo , Necrose/patologia , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Receptores de Serotonina/genética , Receptores de Serotonina/metabolismo , Tiofenos/farmacologiaRESUMO
Serotonin in cardiovascular system plays an important role in blood coagulation, allergy, and inflammation, as well as in blood vessel tone regulation. In this review, the mechanisms of serotonin effects upon the cells of blood vessels are considered and the list of main agonists and antagonists is presented. The signaling pathways activated by serotonin and their interaction in normal and pathological states are described.
