Elevation difference of Ca^(2+) levels in young leaf cells of bromegrass and induced cold-tolerant enhancement under different controlled chilling temperatures

The changes of Ca2＋ levels in young leaf cells of bromegrass under different controlled chilling temperatures were inves- tigated by an antimonite precipitation cytochemical method. The main results were as follows： under 25/20℃ （day/night） tempera- ture and 14 h photoperiod, electron-dense Ca2＋ antimonite precipitates, indicators of Ca2＋ localization, were mainly localized in the vacuoles, cell walls and intercellular spaces; few Ca2＋ deposits were observed in the cytosol and nuclei. After a 3℃ chilling treatment for 3 h, many Ca2＋ precipitates appeared in the cytosol and nuclei, indicating that Ca2＋ influx had occurred in the cytosol and nuclei. When the 3℃ treatment was prolonged to 8 h, more Ca2＋ deposits appeared in the nuclei and cytosol, but the amount of Ca2＋ deposits in both the cytosol and nuclei decreased markedly after a 24 h treatment and most Ca2＋ deposits were returned to the vacuoles and intercellular spaces after an 8 d treatment. When bromegrass was exposed to 7℃ for 3 h, the Ca2＋ distribution in the cells had no visible changes, compared with that of the 25/20℃ grown control plants. However, when the chilling treatment of 7℃ was prolonged to 8 h, a Ca2＋ influx occurred, where many Ca2＋ deposits were observed in the nuclei and cytosol. More Ca2＋ deposits appeared in the nuclei and cytosol after a 24 h treatment, but the amount of Ca2＋ deposits in the cytosol and nuclei was reduced markedly after an 8 d treatment. After a 14 d treatment, the remaining low level of Ca2＋ was recovered in both the cytosol and nuclei and the Ca2＋ deposits were again located in the vacuoles and the intercellular spaces. The dynamics of subcellular Ca2＋ localization in young leaf cells of bromegrass during a 12℃ chilling treatment were similar to those of the 7℃ treatment. Besides, the results showed that the frost tolerance ofbromegrass exposed to 3℃ for 8 d increased by 6℃, for 7℃ and 8 d by 4℃ and for 12℃ and 14 d by 3℃, compared with the controls. Finally, the relationship between different Ca2＋ dynamics and induced frost tolerance was also explored.

18β-glycyrrhetinic Acid-induced Apoptosis and Relation with Intracellular Ca^2＋ Release in Human Breast Carcinoma Cells

Objective:To study the effects of 18β-glycyrrhetinic acid (GA) on proliferation inhibition, apop totic induction, and the relationship between GA-induced apoptosis and intracellular Ca2+ concentration in human breast carcinoma (MCF-7) cells. Methods: After MCF-7 cells were treated with GA at the concentrations from 50 μmol/L to 250 μmol/L for 24 h, cell viability of proliferation was assessed by MTT assay. After the cells were treated with 100 μmol/L, 150 μmol/L, and 200 μmol/L GA for 24 h, the rates of cell apoptosis were examined by terminal deoxynucleotide transferase mediated dUTP nick-end-labeling method and flow cytometry with Annexin V/propidium iodide fluorescent stain. After the cells treated with 150 μmol/L GA for 24 h, intracellular Ca2+ concentration was measured by Fure-2 fluorescein load method. Results: After the cells were treated with GA at the concentrations from 100 μmol/L to 250 μmol/L, the rates of proliferative inhibition were increased significantly (P<0.05 and P<0.01) in a dose dependent fashion. IC50 of the proliferation inhibition was 234.33 μmol/L. Treated with 100 μmol/L, 150 μmol/L, and 200 μmol/L, the rates of cell apoptosis were increased significantly (P<0.01). Intracellular Ca2+ concentration after treatment with GA was higher evidently than that of control (P<0.05). Conclusion: 18β-glycyrrhetinic acid has the effects of the proliferation inhibition and the apoptotic induction on MCF-7 cells. The rise of intracellular Ca2+ level may be depended on apoptosis induced by GA in MCF-7 cells.

Relationship of Intracellular Free Ca^(2+) Concentration and Calcium-activated Chloride Channels of Pulmonary Artery Smooth Muscle Cells in Rats under Hypoxic Conditions

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.

Differential activation of mitogen-activated protein kinases by γ-irradi-ation in IEC-6 cells: Role of intracellular Ca^(2+)

Objective: To explore the effects of γ-irradiation on mitogen-activated protein kinases (MAPKs) and role of intracellular calcium in this event in intestinal epithelial cell line 6 (IEC-6 cells). Methods: After cultured rat IIEC-6 cells with or without the pretreatment of intracellular Ca2+ chelator were exposed to Y-ir-radiation of 6 Gy, the total and phosphorylated MAPKs in the cells were determined with Western blotting and apoptosis was examined with flow cytometry. Activities of Extracellular signal-regulated protein kinase (ERK) and p38 MAPK were determined by using immuoprecipitation followed by Western blotting. Results: In response to γ-irradiation, phosphorylation of ERK was not significantly observed, while the levels of phosphorylated c-Jun NH2-terminal kinase (JNK) and p38 MAPK were increased in 30 min and reached the peak 2 h after exposure to 6 Gy γ-irradiation, though the cell viability was significantly lowered 12 h. On the other hand, no obvious changes were seen in the total protein levels of ERK, JNK and p38 MAPK. Chelation of intracellular Ca2+ almost completely suppressed the JNK and p38 MAPK phosphorylation induced by γ-irradia-tion, but removal of external Ca2+ had no such effect. Activation of p38 MAPK, but not of ERK, was seen to have a correlation with γ-irradiation induced apoptosis. Conclusion: The results suggest that γ-irradiation is a potent activator for JNK and p38 MAPK, and Ca2+ mobilized from intracellular stores plays an important role in the activation of MAPKs and the induction of apoptosis in IEC-6 cells.

Influence of Panax quinquefolium saponins on increased intracellular Ca^(2+) in PC12 cells

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.

Ca^2＋信号参与铝诱导黑麦根系分泌有机酸的调控

【目的】揭示铝诱导根系分泌有机酸的机制,探讨胞质钙信号对铝诱导黑麦根系分泌有机酸的调控作用。【方法】采用药理学研究方法和激光共聚焦扫描显微技术探讨铝胁迫下根尖胞质游离钙离子浓度([Ca2+]cyt)及其与有机酸分泌的关系。【结果】铝不仅诱导黑麦根系分泌柠檬酸和苹果酸,而且使根尖细胞Ca2+的荧光强度增强、波动加剧。铝引起的根尖细胞Ca2+荧光强度的变化在受Ca2+通道抑制剂异搏定、胞内Ca2+通道阻断剂辽红干扰后,显著减少了铝诱导的有机酸分泌。铝诱导的根尖[Ca2+]cyt的升高及有机酸分泌还受CaM阻断剂三氟拉嗪的干扰和抑制。钙螯合剂EGTA处理不仅减弱了铝诱导的Ca2+荧光信号、加大了Ca2+信号的波动幅度,而且显著抑制铝诱导的柠檬酸分泌。此外,在铝胁迫下,根系有机酸分泌受阴离子通道抑制剂尼氟灭酸的抑制。【结论】根尖[Ca2+]c可能是介导铝诱导黑麦根系分泌有机酸的胞内信号因子,而质膜上的阴离子通道可能是铝诱导的钙信号传导途径中的下游效应器。

广枣总黄酮对大鼠心室肌细胞IC_a、I_(to)和细胞[Ca^(2+)]_i的影响

目的 观察广枣总黄酮对大鼠心室肌细胞L 型钙通道电流 (ICa)和瞬时外向钾通道电流 (Ito)以及对心肌细胞内游离钙浓度 ([Ca2 + ]i)的影响 ,探讨其抗心律失常作用机制。方法 全细胞膜片钳记录大鼠心室肌细胞ICa、Ito,激光共聚焦显微镜观察细胞 [Ca2 + ]i 的变化。结果 在钳制电压- 4 0mV ,实验电压 - 4 0～ +5 0mV时 ,广枣总黄酮 10 0mg·L-1对心室肌细胞ICa无显著影响 ;在钳制电压 - 6 0mV ,实验电压 - 4 0～ +5 0mV时显著抑制瞬时外向钾通道Ito(P <0 0 5 ) ;而激光共聚焦显微镜结果显示广枣总黄酮在 5 0、10 0、2 0 0mg·L-1却降低缺氧复氧心肌细胞收缩期和静息期[Ca2 + ]i 的浓度。结论 广枣总黄酮对心肌细胞ICa无显著影响 ,可显著抑制瞬时外向钾通道Ito,并可明显降低心肌细胞收缩期和静息期细胞 [Ca2 + ]i 浓度。这可能是其抗心律失常和保护缺血心肌的主要作用机制。

小檗碱对培养大鼠神经细胞内游离Ca^(2+)的影响

以Ｆｕｒａ２／ＡＭ为细胞内钙离子的荧光指示剂，用ＡＲＣＭＭＩＣ阳离子测定系统，直接测定了体外培养的新生大鼠神经细胞内游离钙（［Ｃａ２＋］ｉ）值，并观察了小檗碱（Ｂｅｒ）的影响。结果表明，Ｂｅｒ对神经细胞静息［Ｃａ２＋］ｉ无明显影响，Ｂｅｒ１～１００μｍｏｌ·Ｌ－１能剂量依赖地抑制去甲肾上腺素和Ｈ２Ｏ２引起的［Ｃａ２＋］ｉ升高，其ＩＣ５０分别为３９．９和１７．９μｍｏｌ·Ｌ－１。高剂量Ｂｅｒ（１０～１００μｍｏｌ·Ｌ－１）能抑制高Ｋ＋引起的［Ｃａ２＋］ｉ升高。姐果提示，Ｂｅｒ对去甲肾上腺素，高Ｋ＋及Ｈ２Ｏ２引起的［Ｃａ２＋］ｉ升高的抑制作用可能是其抗脑缺血作用机制之一。

18β-甘草次酸诱导人乳腺癌细胞凋亡及其细胞内Ca^(2+)水平的变化

背景与目的:18-甘草次酸是甘草的重要成分,近年来的研究发现18-甘草次酸具有抑制人淋巴细胞性白血病、肝癌和肺癌的细胞增殖。本研究探讨18-甘草次酸对人乳腺癌MCF-7细胞诱导凋亡的作用。目前研究已证实细胞内游离Ca2+浓度([Ca2+]i)的动态变化在诱发细胞凋亡过程的多个环节中起重要作用,因此本研究也探讨由18-甘草次酸诱导的MCF-7细胞凋亡发生与[Ca2+]i变化的关系。方法:用50~250μmol/L浓度梯度的18β-甘草次酸处理MCF-7细胞24h,用MTT比色法测定细胞增殖能力。100μmol/L和150μmol/L 18β-甘草次酸处理细胞24 h,用末端脱氧核苷酸转移酶介导dUTP末端标记法、Annexin V流式细胞仪法和单细胞凝胶电泳法检测凋亡细胞。150μmol/L 18β-甘草次酸处理细胞24 h,用Fura-2荧光负载方法测定[Ca2+]i的变化。分别用100μmol/L BAPTA-AM和0.5 mmol/L EGTA与150μmol/L 18β-甘草次酸联合处理MCF-7细胞24 h,用单细胞凝胶电泳法检测凋亡细胞。结果:从100μmol/L 18β-甘草次酸浓度起对MCF-7细胞的增殖抑制率显著升高(P<0.01和P<0.05),呈剂量依赖性,半增殖抑制浓度(IC 50)为234.33μmol/L。100μmol/L和150μmol/L 18β-甘草次酸使细胞凋亡率显著升高(P<0.01和P<0.05);18β-甘草次酸处理组的[Ca2+]i也明显高于对照组(P<0.05)。18β-甘草次酸与BAPTA-AM联合处理组和18β-甘草次酸与EGTA联合处理组的凋亡率均明显低于单纯的150μmol/L18β-甘草次酸处理组(P<0.05和P<0.01)。结论:18-甘草次酸具有抑制MCF-7细胞增殖和诱导其凋亡的作用,而18-甘草次酸诱导MCF-7细胞凋亡的作用在抑制该细胞增殖方面起主要作用。18-甘草次酸诱导MCF-7细胞凋亡依赖于细胞内Ca2+水平上调,而细胞外Ca2+内流是导致细胞内Ca2+水平上调的原因之一。

慢性染铅对大鼠海马区神经细胞Ca^(2+)浓度及Ca^(2+)-ATP酶活性的影响

目的：观察慢性染铅对大鼠海马区神经细胞Ｃａ２＋浓度及Ｃａ２＋－ＡＴＰ酶活性的影响。方法：用０．１５％醋酸铅饲养大鼠建立慢性染铅动物模型，参照Ｄｉｌｄｙ法和徐友涵法测定海马神经细胞Ｃａ２＋浓度及Ｃａ２＋－ＡＴＰ酶活性。结果发现：细胞内Ｃａ２＋浓度，染铅组（２０３．８３±３０．５０）ｎｍｏｌ／Ｌ，对照组（９７．６２±１９．８３）ｎｍｏｌ／Ｌ，ｔ＝８．３１Ｐ＜０．００５；Ｃａ２＋－ＡＴＰ酶活性，染铅组（３２６．４２±４０．０６）ｎｍｏｌ（Ｐｉ）·ｍｇ－１·ｍｉｎ－１，对照组（２５３．０７±２５．４０）ｎｍｏｌ（Ｐｉ）·ｍｇ－１·ｍｉｎ－１，ｔ＝３．５４，Ｐ＜０．０１。结论：慢性染铅可使大鼠海马区神经细胞内Ｃａ２＋浓度升高。

环维黄杨星D对分离大鼠心室肌细胞内Ca^(2+)和L型钙电流的影响

目的 研究环维黄杨星D(CD)对大鼠心室肌细胞内Ca2 + 动员和L型钙电流 (ICa L)的影响。方法 采用全细胞膜片钳和激光扫描共聚焦显微术研究CD对心肌细胞ICa L以及氯化钾、咖啡因诱发心肌细胞内Ca2 + 动员的影响。结果 CD浓度依赖性抑制ICa L。指令电压为 10mV时 ,1和 10 μmol·L- 1 CD分别使ICa L 电流密度从 (- 9 9±1 8)pA pF降至 (- 6 4± 1 4 )pA pF和 (- 4 2± 0 6 )pA pF。共聚焦实验显示 1和 10 μmol·L- 1 CD不影响静息心肌细胞 [Ca2 + ]i,对氯化钾诱发 [Ca2 + ]i升高水平无明显抑制作用 ;咖啡因引起的细胞内Ca2 + 动员可被CD进一步增强。结论 CD浓度依赖性抑制大鼠心室肌细胞ICa L,并有促进咖啡因诱发心肌细胞内Ca2 +

大黄酸对IL-2诱发T淋巴细胞增殖和细胞[Ca^(2＋)]_i变化的作用

研究大黄酸对IL-2引起大鼠T淋巴细胞增殖和细胞内[Ca^(2+)]_i变化的影响.采用MTT法检测细胞增殖,用单细胞钙成像系统记录胞内游离Ca^(2+)浓度([Ca^(2+)]_i).结果表明,大黄酸对IL-2引起的细胞增殖有显著抑制作用,且抑制效果与剂量有关;IL-2引起[Ca^(2+)]_i升高,并持续维持高[Ca^(2+)]_i水平,[Ca^(2+)]_i升高的机制包括胞内Ca^(2+)释放和胞外Ca^(2+)内流;大黄酸显著抑制IL-2引起的[Ca^(2+)]_i升高.结果提示大黄酸可抑制T淋巴细胞增殖,作用机制可能与抑制细胞内[Ca^(2+)]_i升高相关.

绞股蓝总黄酮对缺氧心肌细胞损伤标记物及细胞内游离Ca^2＋浓度的影响

探讨绞股蓝总黄酮(TFG)对缺氧心肌细胞损伤标记物及细胞内游离Ca2+浓度的影响.在离体乳鼠心肌细胞原代培养基础上制备心肌细胞缺氧模型,随机分为正常对照组、缺氧损伤组和绞股监总黄酮干预缺氧损伤组.分别测定各组心肌培养基中乳酸脱氢酶(LDH)、肌钙蛋白I(cTn I)的含量,心肌细胞内游离Ca2+的浓度(荧光探针法).缺氧损伤组心肌培养基中LDH和cTnI的含量及心肌细胞内游离Ca2+浓度较正常对照组明显增加(P<0.05或0.01);经TFG干预后细胞培养基中LDH利cTnI的含量及心肌细胞内游离Ca2+浓度较缺氧损伤组明显降低(P<0.05或0.01).TFG对缺氧心肌细胞具有保护作用,其机理可能与减轻心肌细胞内钙超负荷有关.

高钾离子引起PC12细胞内游离Ca^(2+)浓度升高的机制

目的：分析高钾离子（Ｋ＋）引起ＰＣ１２细胞内游离钙离子浓度（［Ｃａ２＋］ｉ）升高的可能机制。方法：利用ＭｉｒａＣａｌＩｍａｇｅＳｙｓｔｅｍ检测［Ｃａ２＋］ｉ，Ｃａ２＋荧光探针为Ｆｕｒａ－２／ＡＭ。结果：（１）ＫＣｌ浓度为３０～１００ｍｍｏｌ／Ｌ时，可剂量依赖性地诱导ＰＣ１２细胞［Ｃａ２＋］ｉ升高；（２）在细胞外无Ｃａ２＋时，高Ｋ＋对ＰＣ１２细胞［Ｃａ２＋］ｉ无影响；（３）Ｌ－型电压门控钙通道阻滞剂维拉帕米、地尔硫和硝苯地平的浓度分别为５×１０－５，１×１０－４和５×１０－３ｍｏｌ／Ｌ时，可完全阻断高Ｋ＋诱导的ＰＣ１２细胞［Ｃａ２＋］ｉ升高，但Ｎ－型电压门控钙通道阻滞剂ω－ｃｏｎｏｔｏｘｉｎＧＶＩＡ在浓度为１×１０－６ｍｏｌ／Ｌ时，对高Ｋ＋诱导的ＰＣ１２细胞［Ｃａ２＋］ｉ升高没有影响；（４）５×１０－５ｍｏｌ／Ｌ维拉帕米对细胞外由无Ｃａ２＋到有Ｃａ２＋过程引起的［Ｃａ２＋］ｉ升高无抑制作用。结论：高Ｋ＋引起ＰＣ１２细胞［Ｃａ２＋］ｉ升高以细胞质膜上Ｌ－型电压门控Ｃａ２＋通道开放为基础。

单细胞内Ca^（2＋）时空变化的激光共聚焦显微测定

应用激光扫描共聚焦显微系统（ＬＳＣＭ）和Ｆｌｕｏ－３／ＡＭ荧光探剂标记技术，测定了单个活细胞胞内游离Ｃａ２＋的动态变化与立体分布影像．结果显示，在３７℃，Ｆｌｕｏ－３／ＡＭ终浓度为６μｍｏｌ／Ｌ的条件下，Ｃ５７ＢＬ／６Ｊ小鼠巨噬细胞负载１ｈ左右即可获得良好的标记效果．相反，若探剂浓度太高或负载时间太长，胞内荧光强度太强，影响在共聚焦显微镜镜下分辨细胞内结构．因此用ＬＳＣＭ研究细胞内游离Ｃａ２＋变化时，荧光探剂的负载应以获得最适荧光信号而不是以最大荧光强度为标准．上述方法在其他如平滑肌细胞、卵母细胞中的测定亦获得满意的结果，这对进一步研究各种生理和病理条件下细胞内Ｃａ２＋信号的动态变化、与跨膜Ｃａ２＋梯差的关系及对活细胞功能活动的调节提供了一种可行的、直观的研究手段。

神经肽Y经Ca^2＋/CaM依赖的钙调神经磷酸酶途径诱导心肌细胞肥大

目的观察神经肽Y(NPY)诱导心肌细胞肥大效应,及Ca2+/CaM依赖的钙调神经磷酸酶(CaN)途径在其中起的作用。方法用NPY(10、100nmol)刺激心肌细胞,用环胞素A(CsA)特异性抑制CaN活性。测定心肌细胞蛋白合成速率(3H-Leu掺入量)、CaN-蛋白表达、c-junmRNA表达、CaN酶活性、细胞内游离钙含量。结果(1)在100nmolNPY作用下,心肌细胞3H-Leu掺入量及c-junmRNA表达量均较对照组显著增高(P<0.05,P<0.001)。NPY+CsA组和对照组相比无显著差别。(2)在100nmolNPY作用下,NPY组心肌细胞内CaN酶比活力、CaN-α表达、胞浆及核内钙含量较对照组显著增高(P<0.05,P<0.05,P<0.001,P<0.001)。结论NPY可刺激心肌细胞肥大,CaN信号途径在其中起重要作用。

羟墓自由基引起神经细胞［Ca^（2＋）］i增高的机制和Ebselen的抑制作用

用Ｆｕｒａ－２显微荧光测量技术研究了羟基自由基对单个皮层神经细胞内游离钙离子浓度［Ｃａ２＋］ｉ影响和硒化合物Ｅｂｅｌｅｎ对［Ｃａ２＋］ｉ的抑制作用。结果表明羟基自由基的作用首先引起胞内［Ｃａ２＋］ｉ以时间常数τ＝３８９５．４±５０７．２Ｓ速度缓慢增加，然后加入了以τ＝４２０．６±１２２．０Ｓ的外钙大量涌入。钙通道阻断剂、疏基还原剂、疏基还原制和自由基清除剂对羟基自由基损伤作用的影响提示外钙的大量涌入部分与通道的开放有关，疏基损伤在羟基自由基引起的［Ｃａ２＋］ｉ升高中起着重要的作用。具有类谷胱甘肽过氧化酶活性的小分子硒化合物Ｅｂｓｅｌｅｎ（１０－５ｍｏｌ／Ｌ和１０－６ｍｏｌ／Ｌ）抑制羟基自由基引起的［Ｃａ２＋］ｉ升高，推测它可以抑制钙库的释放或促进内钙的外排以及抑制外钙的流入。

染氟成纤维细胞内Ca^(2+)水平的变化及其意义

目的研究氟对成纤维细胞Ca2+浓度的影响并探讨其意义。方法应用扫描共聚焦显微技术测定不同染氟条件下(0.0001、1、10mg/LF-)成纤维细胞Ca2+水平的变化。结果染氟1mg/L组成纤维细胞在染氟500s左右时[Ca2+]i水平开始升高,约1500s达到高峰,然后开始下降,3000s左右回到起点位置。结论成纤维细胞染氟1mg/LF-组[Ca2+]i有一过性升高。认为这种[Ca2+]i浓度的变化在影响成纤维细胞中cbfa1和某些生长因子之间的相互关系、促进细胞增殖、分化及成骨表型转换中可能起重要作用。

水蛭对Ang-Ⅱ刺激鼠肝星状细胞活化Ca^(2+)效应的抑制作用

目的研究水蛭药物血清是否具有抑制肝星状细胞(HSCs)胞浆游离钙([Ca^(2+)]i)增高的作用。方法将32只健康SD大鼠按随机数字表随机分为4组肝纤维化模型水蛭组(8只)皮下注射用精制橄榄油配制的40%CCl4溶液9周,然后灌服水蛭6d;肝纤维化模型对照组(8只)皮下注射用精制橄榄油配制的40%CCl4溶液9周,然后灌服0.9%氯化钠溶液6d;正常大鼠水蛭组(8只)仅皮下注射等量的精制橄榄油9周,然后灌服水蛭6d;正常对照组(8只)仅皮下注射等量的精制橄榄油9周,然后灌服0.9%氯化钠溶液6d。结束后,经下腔静脉取血并分离血清。采用盲法用上述10%血清培养HSCs24h并负载好Fluo-3/AM后使用激光扫描共聚焦显微镜(LSCM)检测HSCs[Ca2+]i。结果(1)经正常大鼠水蛭组及肝纤维化模型水蛭组大鼠血清预处理的HSCs,其[Ca^(2+)]i荧光强度相对值均明显低于肝纤维化模型对照组(P<0.05);且两者与正常对照组间差别也均有显著性意义(P<0.05)。(2)经正常大鼠水蛭组及肝纤维化模型水蛭组大鼠血清预处理HSCs后加入Ang-Ⅱ(1.1×10-7mol/L),HSCs[Ca^(2+)]i荧光强度变化百分数显著低于肝纤维化模型对照组(P<0.01)。结论水蛭能抑制肝星状细胞活化[Ca^(2+)]i的升高,此可能是其发挥抗肝纤维化作用的重要途径之一。

aFGF对肺腺癌AGZY-83A细胞株TPK、PKC活性及Ca^(2+)浓度的影响

目的：观察ａＦＧＦ与细胞膜上特异受体结合后引起的细胞内信号转导途径，探讨ａＦＧＦ导致细胞增殖的机理。方法：以不同浓度的ａＦＧＦ处理ＡＧＺＹ－８３Ａ细胞，利用［γ－３２Ｐ］ＡＴＰ掺入外源性底物的方法测定受体的酪氨酸蛋白激酶活性（ＴＰＫ）及蛋白激酶Ｃ（ＰＫＣ）活性；用Ｆｕｒａ－２／ＡＭ为荧光指示剂测定［Ｃａ２＋］ｉ。结果：随着ａＦＧＦ浓度增加，ＴＰＫ及ＰＫＣ活性随之升高。当ａＦＧＦ浓度为１．１２μｇ／ｍｌ时ａＦＧＦ处理组的ＴＰＫ是对照组的４倍；膜ＰＫＣ活性也是对照组的４倍，胞浆ＰＫＣ活性是对照组的１．７５倍。［Ｃａ２＋］ｉ是对照组的３倍。结论：该细胞株中ａＦＧＦ受体具有ＴＰＫ活性。ＴＰＫ激活后进一步促进蛋白质和酶磷酸化，而使ＰＫＣ活性及［Ｃａ２＋］ｉ升高，即ＰＫＣ和Ｃａ２＋是ＴＰＫ的下游信号分子，进一步促进ｃ－ｆｏｓ、ｊｕｎ基因表达增加。