In this study, a novel approach to measure the absolute cytoplasmic Caconcentration([Ca]cyt) using the Caindicator fluo-3 AM was established. The parameters associated with the probe fluo-3 AM were optimized to accura...In this study, a novel approach to measure the absolute cytoplasmic Caconcentration([Ca]cyt) using the Caindicator fluo-3 AM was established. The parameters associated with the probe fluo-3 AM were optimized to accurately determine fluorescence intensity from the Ca-bound probe. Using three optimized parameters(final concentration of 6 m M probe,incubation time of 135 min, loading probe before plasma treatment), the maximum fluorescence intensity(F?=?527.8 a.u.) and the minimum fluorescence intensity(F?=?63.8 a.u.) were obtained in a saturated Casolution or a solution of lacking Ca. Correspondingly, the maximum [Ca]cytinduced by cold plasma was 1232.5 n M. Therefore, the Caindicator fluo-3 AM was successfully applied to measure the absolute [Ca]cytin Saccharomyces cerevisiae stimulated by cold plasma at atmospheric air pressure.展开更多
Using the method of dual-wavelength measurement of platelet [Ca2+]i, and Fura-2 as the Ca2+ fluorophore probe, we measured the effect of acidic Mu-copolysaccharide from Sticopus Japonicus Selenka (SJAMP) on platelet [...Using the method of dual-wavelength measurement of platelet [Ca2+]i, and Fura-2 as the Ca2+ fluorophore probe, we measured the effect of acidic Mu-copolysaccharide from Sticopus Japonicus Selenka (SJAMP) on platelet [Ca2+]i. The results showed that the most significant increase in platelets [Ca2+]i was seen when the concentration of SJAMP was 100μg/ml and the elevation of normal platelet [Ca2+]i was 93. 96±10. 24 nmol/L (n = 10). In the presence of extracellular Ca2+(1 mmol/L), the magnitude of platelet [Ca2+]i response to SJAMP was increased and the [Ca2+]i, could reach 116. 72 + 10. 66 nmol/L (n= 10). On the other hand, the magnitude of increased platelet [Ca2+]i; induced by SJAMP was smaller and the duration of [Ca2+]i reaching the highest level was longer when compared with other platelet aggregation agents. In the mean time, if platelets were first incubated with cyclooxygenase inhibitor, the rise of [Ca2+]i evoked by SJAMP was inhibited. The results indicated that the mechanism of the rise of [Ca2+]i induced by SJAMP might be dependent upon the generation of prostaglandin endoperoxides and (or) TXA?.展开更多
Object. The effects of ATP-introduced a rise in cytosolic free Ca2+ concentration and inhibition of nitric oxide were investigated. Method. Measurement of free Ca2+([Ca2+] i)of cultured rat tail arterial smooth muscle...Object. The effects of ATP-introduced a rise in cytosolic free Ca2+ concentration and inhibition of nitric oxide were investigated. Method. Measurement of free Ca2+([Ca2+] i)of cultured rat tail arterial smooth muscle cells using Fura-2/AM dual excitation wavelength spectrofluorometer. Results. There are two components of [Ca2+] i can be evoked by ATP. One part is Ca2+ entry from Ca2+ channel and formed a plateau. The another part is a peak that released from Ca2+ store. Both of them can be inhibited by NO. Conclusion. The ATP induced [Ca2+] i rise that release Ca2+ from both Insp 3 and ryanochine receptors and Ca2+ entry through calcium channels. The inhibition of NO on ATP induced [Ca2+] i rise that was mediated by cGMP.展开更多
To investigate the relationship between intracellular free Ca^2＋ concentration （[Ca^2＋ ]i ） and calcium-activated chloride （Clca） channels of pulmonary artery smooth muscle cells （PASMCs） in rats under acute a...To investigate the relationship between intracellular free Ca^2＋ concentration （[Ca^2＋ ]i ） and calcium-activated chloride （Clca） channels of pulmonary artery smooth muscle cells （PASMCs） in rats under acute and chronic hypoxic conditions, acute hypoxia-induced contraction was observed in rat pulmonary artery by using routine blood vascular perfusion in vitro. The fluorescence Ca^2＋ indicator Fura-2/AM was used to observe [Ca^2＋ ]i of rat PASMCs under normal and chronic hypoxic condition. The effect of Clca channels on PASMCs proliferation was assessed by MTT assay. The Clca channel blockers niflumic acid （NFA） and indaryloxyacetic acid （IAA-94） exerted inhibitory effects on acute hypoxia-evoked contractions in the pulmonary artery. Under chronic hypoxic condition, [Ca^2＋ ]i was increased. Under normoxic condition, [Ca^2＋ If was （123.634-18.98） nmol/ L, and in hypoxic condition, [Ca^2＋]i wag （281. 754-16.48） nmol/L （P〈0. 01）. Under normoxic condition, [Ca^2＋ ]i showed no significant change and no effect on Clca channels was observed （P〉 0. 05）. Chronic hypoxia increased [Ca^2＋ ]i which opened Clca channels. The NFA and IAA-94 blocked the channels and decreased [Ca^2＋ ]i from （281.75± 16.48） nmot/L to （117.66 ±15.36） nmol/L （P〈0.01）. MTT assay showed that under chronic hypoxic condition NFA and IAA-94 decreased the value of absorbency （A value） from 0. 459±0. 058 to 0. 224±0. 025 （P〈0. 01）. Hypoxia increased [Ca^2＋ ]i which opened Cl~ channels and had a positive-feedback in [Ca^2＋ ]i. This may play an important role in hypoxic pulmonary hypertension. Under chronic hypoxic condition, Clca channel may play a part in the regulation of proliferation of PASMCs.展开更多
基金supported by National Natural Science Foundation of China (Grant Nos. 21246012, 21306015 and 21476032)
文摘In this study, a novel approach to measure the absolute cytoplasmic Caconcentration([Ca]cyt) using the Caindicator fluo-3 AM was established. The parameters associated with the probe fluo-3 AM were optimized to accurately determine fluorescence intensity from the Ca-bound probe. Using three optimized parameters(final concentration of 6 m M probe,incubation time of 135 min, loading probe before plasma treatment), the maximum fluorescence intensity(F?=?527.8 a.u.) and the minimum fluorescence intensity(F?=?63.8 a.u.) were obtained in a saturated Casolution or a solution of lacking Ca. Correspondingly, the maximum [Ca]cytinduced by cold plasma was 1232.5 n M. Therefore, the Caindicator fluo-3 AM was successfully applied to measure the absolute [Ca]cytin Saccharomyces cerevisiae stimulated by cold plasma at atmospheric air pressure.
基金This project was supported by grant from the National Nature Science Foundation of China (No.39370322)
文摘Using the method of dual-wavelength measurement of platelet [Ca2+]i, and Fura-2 as the Ca2+ fluorophore probe, we measured the effect of acidic Mu-copolysaccharide from Sticopus Japonicus Selenka (SJAMP) on platelet [Ca2+]i. The results showed that the most significant increase in platelets [Ca2+]i was seen when the concentration of SJAMP was 100μg/ml and the elevation of normal platelet [Ca2+]i was 93. 96±10. 24 nmol/L (n = 10). In the presence of extracellular Ca2+(1 mmol/L), the magnitude of platelet [Ca2+]i response to SJAMP was increased and the [Ca2+]i, could reach 116. 72 + 10. 66 nmol/L (n= 10). On the other hand, the magnitude of increased platelet [Ca2+]i; induced by SJAMP was smaller and the duration of [Ca2+]i reaching the highest level was longer when compared with other platelet aggregation agents. In the mean time, if platelets were first incubated with cyclooxygenase inhibitor, the rise of [Ca2+]i evoked by SJAMP was inhibited. The results indicated that the mechanism of the rise of [Ca2+]i induced by SJAMP might be dependent upon the generation of prostaglandin endoperoxides and (or) TXA?.
文摘Object. The effects of ATP-introduced a rise in cytosolic free Ca2+ concentration and inhibition of nitric oxide were investigated. Method. Measurement of free Ca2+([Ca2+] i)of cultured rat tail arterial smooth muscle cells using Fura-2/AM dual excitation wavelength spectrofluorometer. Results. There are two components of [Ca2+] i can be evoked by ATP. One part is Ca2+ entry from Ca2+ channel and formed a plateau. The another part is a peak that released from Ca2+ store. Both of them can be inhibited by NO. Conclusion. The ATP induced [Ca2+] i rise that release Ca2+ from both Insp 3 and ryanochine receptors and Ca2+ entry through calcium channels. The inhibition of NO on ATP induced [Ca2+] i rise that was mediated by cGMP.
文摘To investigate the relationship between intracellular free Ca^2＋ concentration （[Ca^2＋ ]i ） and calcium-activated chloride （Clca） channels of pulmonary artery smooth muscle cells （PASMCs） in rats under acute and chronic hypoxic conditions, acute hypoxia-induced contraction was observed in rat pulmonary artery by using routine blood vascular perfusion in vitro. The fluorescence Ca^2＋ indicator Fura-2/AM was used to observe [Ca^2＋ ]i of rat PASMCs under normal and chronic hypoxic condition. The effect of Clca channels on PASMCs proliferation was assessed by MTT assay. The Clca channel blockers niflumic acid （NFA） and indaryloxyacetic acid （IAA-94） exerted inhibitory effects on acute hypoxia-evoked contractions in the pulmonary artery. Under chronic hypoxic condition, [Ca^2＋ ]i was increased. Under normoxic condition, [Ca^2＋ If was （123.634-18.98） nmol/ L, and in hypoxic condition, [Ca^2＋]i wag （281. 754-16.48） nmol/L （P〈0. 01）. Under normoxic condition, [Ca^2＋ ]i showed no significant change and no effect on Clca channels was observed （P〉 0. 05）. Chronic hypoxia increased [Ca^2＋ ]i which opened Clca channels. The NFA and IAA-94 blocked the channels and decreased [Ca^2＋ ]i from （281.75± 16.48） nmot/L to （117.66 ±15.36） nmol/L （P〈0.01）. MTT assay showed that under chronic hypoxic condition NFA and IAA-94 decreased the value of absorbency （A value） from 0. 459±0. 058 to 0. 224±0. 025 （P〈0. 01）. Hypoxia increased [Ca^2＋ ]i which opened Cl~ channels and had a positive-feedback in [Ca^2＋ ]i. This may play an important role in hypoxic pulmonary hypertension. Under chronic hypoxic condition, Clca channel may play a part in the regulation of proliferation of PASMCs.
