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
BACKGROUND： Previous studies have demonstrated that intracellular Ca^2＋ （[Ca^2＋]） overload, excitotoxicity, free radical injury, and nitric oxide toxicity are involved in mechanisms of neuronal death in the ische...BACKGROUND： Previous studies have demonstrated that intracellular Ca^2＋ （[Ca^2＋]） overload, excitotoxicity, free radical injury, and nitric oxide toxicity are involved in mechanisms of neuronal death in the ischemic brain. OBJECTIVE： To investigate the influence of Panax quinquefo/ium saponins （PQS） on multiple factors-induced Ca^2＋ overload in the rat pheochromocytoma （PC12） cell line. DESIGN, TIME AND SETTING： Intergroup comparison, in vitro study. The experiment was performed at the Heilongjiang Key Laboratory of Anti-fibrosis Biotherapy, Mudanjiang Medical University between November 2007 and April 2008. MATERIALS- In vitro cultured PC12 cells in the logarithmic phase were assigned into blank control, model, and drug treatment groups （10 μmol/L nimodipine; 40 μg/L, 100 μg/L, and 250 μg/L PQS）. Nimodipine was purchased from Jiangsu Yangtze River Pharmacy Group Co., China; PQS （purity 〉 95%, HLPC grade） was provided by School of Basic Medical Sciences, Jilin University. Caffeine, Na2S2O4, L-glutamic acid （Glu）, Fura-2/AM, and calcium ionophore A23187 were purchased from Sigma, USA. METHODS： PC12 cells in the model and drug treatment groups were separately incubated in glucose-free Hank＇s buffered saline solution ＋ Na2S2O4 （2 mmol/L） for 6 hours, Glu （200 μmot/L） plus A23187 （0.05 μmol/L） for 6 hours, KCI （50 mmol/L） for 1 hour, and caffeine （5 mmol/L） for 3 hours to establish models of intracellular Ca^2＋ overload induced by oxygen and glucose deprivation, Glu, A23187, high K＋, or caffeine. In addition, control cells were incubated in high-glucose DMEM culture medium. MAIN OUTCOME MEASURES： [Ca^2＋]i changes in PC12 cells exposed to oxygen-glucose deprivation, Glu, A23187, high K^＋, or caffeine were detected using spectrofluorometer. RESULTS： PQS blocked the [Ca^2＋]i increase induced by oxygen-glucose deprivation, Glu, A23187, high K＋, or caffeine. In particular, high-dose PQS was most effective （P 〈 0.01）. PQS significantly inhibited Glu- or caffeine-induced [Ca^2＋]i increases in the absence of extracellular Ca^2＋, but nimodipine did not. CONCLUSION： PQS blocked intracellular Ca^2＋ overload induced by oxygen-glucose deprivation, Glu, A23187, high K^＋, or caffeine. This mechanism might be involved in the attenuation of neuronal apoptosis following ischemic brain injury.展开更多
WT5”BZ] To study the molecular mechanism of the stimulatory effect of low dose radiation(LDR) on T cell activation. [WT5”BX]Methods.[WT5”BZ] Thymocytes from Kunming mice exposed to whole body irradiation(WBI) with ...WT5”BZ] To study the molecular mechanism of the stimulatory effect of low dose radiation(LDR) on T cell activation. [WT5”BX]Methods.[WT5”BZ] Thymocytes from Kunming mice exposed to whole body irradiation(WBI) with different doses of X rays were analyzed for the changes in signal molecules of the phospholipase C phosphatidylinositol biphosphate(PLC IP2) and G protein adenylate cyclase(AC) pathways. [WT5”BX]Results.[WT5”BZ]It was found that[Ca 2+ ] i increased in response to doses within 0 2 Gy which was most marked after 0 075 Gy and the increase was accentuated in the presence of Con A. The changes in CD3 and calcineurin(CN) expression of the thymocytes followed the same pattern as the alterations in [Ca 2+ ] i after LDR. The expression of α,β1 and β2 isoforms of protein kinase C(PKC) was all up regulated after 0 075 Gy with the increase in PKC β1 expression being most marked. The cAMP/cGMP ratio and PKA activity of the thymocytes was lowered after low dose radiation and increased after doses above 0 5 Gy in a dose dependent manner, thus giving rise to J shaped dose response curves. The Ca antagonist TMB 8 and cAMP stimulant cholera toxin suppressed the augmented thymocyte proliferation induced by LDR. [WT5”BX]Conclusion.[WT5”BZ]Data presented in the present paper suggest that activation of the PLC PIP2 signal pathway and suppression of the AC cAMP signal pathway are involved in the stimulation of the thymocytes following WBI with low dose X rays.展开更多
The effect of Epimedium extract （EE） on erythrocytes was investigated by atomic force microscopy （AFM）. The images of the surface structures showed clear concave and progressive increase of surface roughness of er...The effect of Epimedium extract （EE） on erythrocytes was investigated by atomic force microscopy （AFM）. The images of the surface structures showed clear concave and progressive increase of surface roughness of erythrocyte after incubation with EE at concentration of 0.2 or 0.05 g/L, far below its critical hemolytic levels. The AFM results also indicated that the granules of the fine surface structure increased, which caused by aggregation of membrane protein. Further study showed that the change in surface topography of erythrocyte membrane might be connected with the increase of intracellular free Ca^2＋ induced by EE.展开更多
Calcium ions are important in many vital neuron processes, including spontaneous neurotransmitter release. Extracellular calcium has long been known to be related to spontaneous neurotransmitter release, but the detai...Calcium ions are important in many vital neuron processes, including spontaneous neurotransmitter release. Extracellular calcium has long been known to be related to spontaneous neurotransmitter release, but the detailed mechanism for the effect of intracellular Ca^2＋ on synaptic release has not yet been understood. In this research, 1,2-bis-（o-aminophenoxy）-ethane-N, N, N′, N′-tetraacetic acid tetraacetoxymethyl ester （BAPTA-AM） was used to combine with cytosolic free Ca^2＋ in a calcium free medium of cultured Xenopus neuromuscular junctions （NMJ）, The spontaneous synaptic current （SSC） frequency was obviously reduced. Then, drugs were applied to interrupt and activate the Ca2＋ release channels in the endoplasmic reticulum （ER） membrane, but the SSC frequency was not affected. The results show that spontaneous neurotransmitter release depends on intracellular rather than ER calcium in cultured Xenopus NMJ without extracellular calcium.展开更多
Spontaneous secretions occur in both neurons and non-neuronal cells, and calcium is important for these secretion processes. However, the detailed roles of calcium on the secretions have not yet been identified. In th...Spontaneous secretions occur in both neurons and non-neuronal cells, and calcium is important for these secretion processes. However, the detailed roles of calcium on the secretions have not yet been identified. In the present study, cultured Xenopus myocytes loaded with exogenous acetylcholine （ACh） into the cytoplasm in the absence of extracellular Ca^2＋ undergo spontaneous quantal ACh secretion as detected by the appearance of pulsatile miniature endplate currents. Analysis of the frequencies, amplitudes, and time courses of these currents suggests that similar cellular mechanisms are involved in the secretions of ACh in normal medium and Ca^2＋-free solution. Various doses of ryanodine were used to regulate the intra- cellular Ca^2＋ to different levels. The spontaneous ACh secretion from myocytes in Ca^2＋-free medium was decreased by reducing intracellular Ca^2＋ levels and enhanced by increasing cytosolic Ca^2＋ levels. These observations demonstrate that the spontaneous secretion from isolated myocytes and the effect of ryanodine on ACh-loaded cells are both independent of extracellular Ca^2＋ while Ca^2＋ in the sarcoplasmic reticulum plays a crucial role in the secretions.展开更多
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.
基金Supported by:the Natural Science Foundation of Heilongjiang Province,No ZA2006-07
文摘BACKGROUND： Previous studies have demonstrated that intracellular Ca^2＋ （[Ca^2＋]） overload, excitotoxicity, free radical injury, and nitric oxide toxicity are involved in mechanisms of neuronal death in the ischemic brain. OBJECTIVE： To investigate the influence of Panax quinquefo/ium saponins （PQS） on multiple factors-induced Ca^2＋ overload in the rat pheochromocytoma （PC12） cell line. DESIGN, TIME AND SETTING： Intergroup comparison, in vitro study. The experiment was performed at the Heilongjiang Key Laboratory of Anti-fibrosis Biotherapy, Mudanjiang Medical University between November 2007 and April 2008. MATERIALS- In vitro cultured PC12 cells in the logarithmic phase were assigned into blank control, model, and drug treatment groups （10 μmol/L nimodipine; 40 μg/L, 100 μg/L, and 250 μg/L PQS）. Nimodipine was purchased from Jiangsu Yangtze River Pharmacy Group Co., China; PQS （purity 〉 95%, HLPC grade） was provided by School of Basic Medical Sciences, Jilin University. Caffeine, Na2S2O4, L-glutamic acid （Glu）, Fura-2/AM, and calcium ionophore A23187 were purchased from Sigma, USA. METHODS： PC12 cells in the model and drug treatment groups were separately incubated in glucose-free Hank＇s buffered saline solution ＋ Na2S2O4 （2 mmol/L） for 6 hours, Glu （200 μmot/L） plus A23187 （0.05 μmol/L） for 6 hours, KCI （50 mmol/L） for 1 hour, and caffeine （5 mmol/L） for 3 hours to establish models of intracellular Ca^2＋ overload induced by oxygen and glucose deprivation, Glu, A23187, high K＋, or caffeine. In addition, control cells were incubated in high-glucose DMEM culture medium. MAIN OUTCOME MEASURES： [Ca^2＋]i changes in PC12 cells exposed to oxygen-glucose deprivation, Glu, A23187, high K^＋, or caffeine were detected using spectrofluorometer. RESULTS： PQS blocked the [Ca^2＋]i increase induced by oxygen-glucose deprivation, Glu, A23187, high K＋, or caffeine. In particular, high-dose PQS was most effective （P 〈 0.01）. PQS significantly inhibited Glu- or caffeine-induced [Ca^2＋]i increases in the absence of extracellular Ca^2＋, but nimodipine did not. CONCLUSION： PQS blocked intracellular Ca^2＋ overload induced by oxygen-glucose deprivation, Glu, A23187, high K^＋, or caffeine. This mechanism might be involved in the attenuation of neuronal apoptosis following ischemic brain injury.
基金This work was supported by a grant from NSFC (No.39570188)
文摘WT5”BZ] To study the molecular mechanism of the stimulatory effect of low dose radiation(LDR) on T cell activation. [WT5”BX]Methods.[WT5”BZ] Thymocytes from Kunming mice exposed to whole body irradiation(WBI) with different doses of X rays were analyzed for the changes in signal molecules of the phospholipase C phosphatidylinositol biphosphate(PLC IP2) and G protein adenylate cyclase(AC) pathways. [WT5”BX]Results.[WT5”BZ]It was found that[Ca 2+ ] i increased in response to doses within 0 2 Gy which was most marked after 0 075 Gy and the increase was accentuated in the presence of Con A. The changes in CD3 and calcineurin(CN) expression of the thymocytes followed the same pattern as the alterations in [Ca 2+ ] i after LDR. The expression of α,β1 and β2 isoforms of protein kinase C(PKC) was all up regulated after 0 075 Gy with the increase in PKC β1 expression being most marked. The cAMP/cGMP ratio and PKA activity of the thymocytes was lowered after low dose radiation and increased after doses above 0 5 Gy in a dose dependent manner, thus giving rise to J shaped dose response curves. The Ca antagonist TMB 8 and cAMP stimulant cholera toxin suppressed the augmented thymocyte proliferation induced by LDR. [WT5”BX]Conclusion.[WT5”BZ]Data presented in the present paper suggest that activation of the PLC PIP2 signal pathway and suppression of the AC cAMP signal pathway are involved in the stimulation of the thymocytes following WBI with low dose X rays.
文摘The effect of Epimedium extract （EE） on erythrocytes was investigated by atomic force microscopy （AFM）. The images of the surface structures showed clear concave and progressive increase of surface roughness of erythrocyte after incubation with EE at concentration of 0.2 or 0.05 g/L, far below its critical hemolytic levels. The AFM results also indicated that the granules of the fine surface structure increased, which caused by aggregation of membrane protein. Further study showed that the change in surface topography of erythrocyte membrane might be connected with the increase of intracellular free Ca^2＋ induced by EE.
基金Supported by the Natural Science Foundation of Beijing (No. 5052015)
文摘Calcium ions are important in many vital neuron processes, including spontaneous neurotransmitter release. Extracellular calcium has long been known to be related to spontaneous neurotransmitter release, but the detailed mechanism for the effect of intracellular Ca^2＋ on synaptic release has not yet been understood. In this research, 1,2-bis-（o-aminophenoxy）-ethane-N, N, N′, N′-tetraacetic acid tetraacetoxymethyl ester （BAPTA-AM） was used to combine with cytosolic free Ca^2＋ in a calcium free medium of cultured Xenopus neuromuscular junctions （NMJ）, The spontaneous synaptic current （SSC） frequency was obviously reduced. Then, drugs were applied to interrupt and activate the Ca2＋ release channels in the endoplasmic reticulum （ER） membrane, but the SSC frequency was not affected. The results show that spontaneous neurotransmitter release depends on intracellular rather than ER calcium in cultured Xenopus NMJ without extracellular calcium.
基金Supported by the National Natural Science Foundation of China (No. 30370468)
文摘Spontaneous secretions occur in both neurons and non-neuronal cells, and calcium is important for these secretion processes. However, the detailed roles of calcium on the secretions have not yet been identified. In the present study, cultured Xenopus myocytes loaded with exogenous acetylcholine （ACh） into the cytoplasm in the absence of extracellular Ca^2＋ undergo spontaneous quantal ACh secretion as detected by the appearance of pulsatile miniature endplate currents. Analysis of the frequencies, amplitudes, and time courses of these currents suggests that similar cellular mechanisms are involved in the secretions of ACh in normal medium and Ca^2＋-free solution. Various doses of ryanodine were used to regulate the intra- cellular Ca^2＋ to different levels. The spontaneous ACh secretion from myocytes in Ca^2＋-free medium was decreased by reducing intracellular Ca^2＋ levels and enhanced by increasing cytosolic Ca^2＋ levels. These observations demonstrate that the spontaneous secretion from isolated myocytes and the effect of ryanodine on ACh-loaded cells are both independent of extracellular Ca^2＋ while Ca^2＋ in the sarcoplasmic reticulum plays a crucial role in the secretions.
