内皮素-1对培养鼠肝星状细胞内游离钙的影响

目的：观察内皮素-1（ET-1）对培养肝星状细胞（HSC）的胞内钙[Ca²⁺]_i的影响。方法：分离培养大鼠HSC，采用Fura-2/AM钙荧光指示剂测定HSC细胞内Ca²⁺浓度，并观察ET-1对其影响。结果：ET-1在很短时间内即可明显提高HSC细胞内Ca²⁺浓度（P<0.01）。并且在无细胞外液Ca²⁺情况下，亦可提高[Ca²⁺]_i，有明显的量效关系（P<0.05，P<0.01）。同时，维拉帕米对ET-1的上述作用具有显著的抑制作用（P<0.05）。结论：ET-1的促肝纤维化作用可能是通过[Ca²⁺]_i运转而产生的。     

丹参对抗缺氧/复氧引起的大鼠心室肌细胞收缩和细胞内钙的改变

目的:观察丹参在常氧和缺氧/复氧过程中对心肌细胞收缩和电刺激诱导的细胞内钙([Ca²⁺]_i)瞬态的影响。方法: 采用酶解分离成年大鼠心室肌细胞化学缺氧模型, 用视频跟踪计算机系统和细胞内双波长钙荧光系统分别观察心肌细胞收缩力学和[Ca²⁺]_i等指标。结果:丹参(1-9 g/L)处理后降低心肌细胞最大收缩和舒张速率、收缩幅度以及电刺激诱导的[Ca²⁺]_i幅度, 且呈剂量依赖性。缺氧后, 与对照相比细胞收缩力和钙瞬态幅度降低、舒张末细胞长度缩短、舒张末钙水平增高;复氧后细胞收缩力、钙瞬态幅度和舒张末钙水平有所回复, 但不能达对照水平。用3 g/L的丹参处理后, 缺氧/复氧引起的心肌细胞最大收缩和舒张速率、收缩幅度和电刺激诱导的[Ca^2＋]_i幅度高于单纯缺氧组, 舒张末[Ca²⁺]_i水平低于单纯缺氧组。结论:丹参可对抗缺氧/复氧引起的大鼠心室肌细胞收缩力降低和细胞内动态和静态钙的变化。     

氢氟酸烧伤中毒对兔外周血单个核细胞凋亡与胞内钙浓度的影响

目的: 研究氢氟酸烧伤中毒对兔外周血单个核细胞(PBMC)早期凋亡百分率和胞内游离钙浓度([Ca²⁺]_i)的影响。方法: 用流式细胞仪检测兔烧伤染毒前后外周血单个核细胞的Annexin V变化, 以观察其早期凋亡百分率。用Fluo-3/Am荧光探针观察烧伤染毒前后外周血单个核细胞内Ca²⁺平均荧光强度值的变化, 以观察细胞内[Ca²⁺]_i的变化。 结果: 12只烧伤中毒兔外周血单个核细胞早期凋亡百分率显著增加, 烧伤染毒前后比较P<0.01。而12只烧伤中毒兔中8只染毒后1 h外周血单个核细胞内[Ca²⁺]_i显著降低, 烧伤染毒前后比较P<0.05。其余4只却表现[Ca²⁺]_i染毒前后比较P>0.05。 结论: 本实验氢氟酸烧伤中毒使兔外周血单个核细胞早期凋亡百分率显著增加, 外周血单个核细胞内[Ca²⁺]_i却显著降低。提示氢氟酸烧伤中毒引导的细胞凋亡并非细胞内[Ca²⁺]_i增加所引发。     

小鼠腹腔巨噬细胞基态游离钙离子浓度的不均一性及其和细胞反应性的关系

目的:在单细胞水平上研究小鼠腹腔巨噬细胞(PM)基态游离钙离子浓度([Ca²⁺]_i)的不均一性及其和细胞反应性的关系。方法:用荧光指示剂Fura-2/AM结合荧光显微镜成像系统检测单个PM基态的及用激动剂刺激细胞后的[Ca²⁺]_i;同时结合NBT染色法定量检测单个PM产超氧阴离子(O₂^-)水平。结果:对7只正常小鼠共392个PM基态[Ca²⁺]_i的研究表明小鼠PM的基态[Ca²⁺]_i呈正态分布[(54±24)nmol/L,n=392],但波动范围较大(从10nmol/L到高于100nmol/L),以[Ca²⁺]_i在40-60nmol/L的细胞数量最多(约占50%)。用PMA、fMLP刺激后PM[Ca²⁺]_i升高,且受刺激后[Ca²⁺]_i升高的峰值和基态[Ca²⁺]_i之间呈正相关(PMA刺激组:r=052,P<0.01,n=58;fMLP刺激组:r=0.59,P<0.01,n=44。此两组实验均以不同的小鼠重复3次,其它两只小鼠的结果与上同。下面的表述方法同此)。另外小鼠PM的基态[Ca²⁺]_i与其受PMA刺激后产生O₂^-的量也呈显著正相关(r=0.42,P<0.01,n=43,重复4次)。结论:小鼠PM的基态[Ca²⁺]_i是不均一的,且基态[Ca²⁺]_i的高低和该细胞对致炎因子的反应性密切相关。     

阿魏酸钠拮抗氧化型脂蛋白(a)对人动脉平滑肌细胞的作用

目的:研究脂蛋白(a)氧化前后致人动脉平滑肌细胞(SMC)增殖及细胞内游离钙浓度([Ca²⁺]_i)的变化,观察阿魏酸钠(SF)对其的影响。方法:Lp(a)经体外Cu²⁺氧化法氧化,硫代巴比妥酸(TBARS)比色法检测氧化程度,培养的人动脉SMC中分别加入不同浓度SF,作用12h后再与天然和氧化型Lp(a)共同孵育,以MTT比色法、流式细胞仪检测细胞增殖状况,采用荧光探针Fura-2/AM检测细胞[Ca²⁺]_i。结果:氧化型Lp(a)促人动脉SMC增殖的同时亦明显增加了[Ca²⁺]_i水平,作用较天然Lp(a)明显,SF(40,80mg/L)可显著抑制氧化型Lp(a)所致的细胞增殖和[Ca²⁺]_i增加,并呈剂量效应关系,而对天然Lp(a)所致的细胞增殖和[Ca²⁺]_i增加无明显影响。结论:氧化型Lp(a)通过升高[Ca²⁺]_i而显著促动脉SMC增殖可能是其致动脉粥样硬化的机制之一,SF拮抗这种作用可能与其抗氧化能力有关。     

缺氧复氧对人心房肌细胞内钙离子浓度的影响

目的和方法:观察缺氧、复氧对人心房肌细胞内钙离子浓度的影响。急性分离人心房肌细胞,以Fluo-3作为Ca²⁺指示剂,用激光共聚焦显微镜观察细胞内Ca²⁺浓度变化。结果:人心房肌细胞在缺氧前[Ca²⁺]ⁱ为(117±48)nmol/L。在缺氧15 min时,[Ca²⁺]ⁱ即增加为(187±64)nmol/L(P<0.05,vs缺氧前)。缺氧30 min时[Ca²⁺]ⁱ为(417±139)nmol/L(P<0.01,vs缺氧前)。缺氧后复氧30 min[Ca²⁺]ⁱ为(786±238)nmol/L(P<0.01,vs缺氧前)。结论:人心房肌细胞在缺氧状态下胞内Ca²⁺浓度升高,而当其短期复氧时,胞Ca²⁺浓度继续增加。     

Carboxyeosin decreases the rate of decay of the [Ca2+]i transient in uterine smooth muscle cells isolated from pregnant rats

 In myometrial smooth muscle cells the rate of decline of intracellular calcium ([Ca²⁺]_i) is determined by Ca²⁺ extrusion from the cell and uptake into intracellular stores. The relative quantitative contribution of these processes however, has not been established. We therefore examined the effect of the sarcolemmal Ca²⁺ pump inhibitor, carboxyeosin, on the rate of the [Ca²⁺]_i transient decline in myocytes isolated from pregnant rat uterus. Indo-1 was used in conjunction with the whole-cell patch-clamp technique to measure [Ca²⁺]_i simultaneously with transmembrane calcium current (I _Ca). [Ca²⁺]_i transients were elicited by repetitive membrane depolarization to simulate the natural pattern of uterine electrical activity. The rate of [Ca²⁺]_i removal was calculated from the falling phase of the [Ca²⁺]_i transient. Pre-treatment of the cells with 2 μM carboxyeosin led to a marked decrease in the rate of [Ca²⁺]_i transient decay, suggesting that the sarcolemmal Ca²⁺ pump is involved in the calcium extrusion process. Removal of the extracellular Na also decreased the rate of [Ca²⁺]_i decay, indicating an important role for the Na⁺/Ca²⁺ exchange. When both the sarcolemmal Ca²⁺ pump and Na⁺/Ca²⁺ exchange were inhibited the cell failed to restore [Ca²⁺]_i after the stimulation. Comparison of the rate constants of [Ca²⁺]_i decay in control conditions and after carboxyeosin treatment shows that approximately 30% of [Ca²⁺]_i decay is due to the sarcolemmal calcium pump activity. The remaining 70% can be attributed to the activity of Na⁺/Ca²⁺ exchanger and the intracellular calcium stores. Received: 17 July 1998 / Received after revision: 23 September 1998 / Accepted: 25 September 1998     

脂多糖对大鼠肺微血管内皮细胞[Ca2+]i和Gq蛋白的影响及山莨菪碱的干预

目的:观察脂多糖对大鼠肺微血管内皮细胞(RPMVECs)[Ca²⁺]_i和Gq蛋白的影响及山莨菪碱的干预作用。方法:分离、培养并鉴定Wistar大鼠RPMVECs;应用Fura-2/AM法测定RPMVECs[Ca²⁺]_i;流式细胞仪技术测定RPMVECsGq蛋白。结果:①LPS作用于RPMVECs30min和90min后,[Ca²⁺]_i显著高于对照组;Gq蛋白显著低于对照组。②山莨菪碱可抑制LPS的上述作用。结论:①LPS致RPMVECs[Ca²⁺]_i增加和Gq蛋白下降;②山莨菪碱通过抑制LPS诱导RPMVECs[Ca²⁺]_i增加和Gq蛋白下降的作用而保护其内皮屏障功能。     

钾通道对大鼠肺动脉平滑肌细胞[Ca2+]i的调节

目的:探讨在常氧、低氧条件下钾通道对大鼠肺动脉平滑肌细胞(PASMCs)[Ca²⁺]_i的调节。方法:采用钙荧光探针(Fura-2/AM)负载培养的大鼠PASMCs,观察常氧、低氧培养后3种钾通道抑制剂(4AP,TEA、Glib)对PASMCs[Ca²⁺]_i的调节,同时用四唑盐(MTT)比色法比较4AP、TEA、Glib对大鼠PASMCs增殖的影响。结果:(1)常氧状态下,PASMCs[Ca²⁺]_i为(156.91±8.60)nmol/L,低氧时为(294.01±16.81)nmol/L(P＜0.01)。(2)常氧状态下,4AP可引起PASMCs[Ca²⁺]_i升高,达(280.52±23.21)nmol/L(P＜0.01),而TEA、Glib无此作用。(3)低氧时,4AP和TEA都可引起PASMCs[Ca²⁺]_i的升高,分别为(422.41±24.28)nmol/L、(380.84±11.02)nmol/L(P<0.01),Glib无作用。(4)MTT比色法中,常氧和低氧状态下4AP均引起吸光度(A)值升高,分别是0.582±0.062,0.873±0.043(P＜0.01)。TEA仅在低氧时A值升高(0.729±0.041,P＜0.05),而Glib无论常氧还是低氧均无影响。结论:无论常氧还是低氧条件下,电压依赖性钾通道(K_V)对PASMCs[Ca²⁺]_i及其增殖起主要作用。钙激活的钾通道(K_Ca)在常氧条件下对[Ca²⁺]_i不起调节作用,而在低氧下使[Ca²⁺]_i降低,反应性地调节PASMCs增殖。ATP敏感性钾通道(K_ATP)无论在常氧还是低氧情况下对[Ca²⁺]_i的调节不起作用。     

低氧后心肌钙瞬变变化与β-肾上腺素受体脱敏的关系

目的:慢性低氧时,心脏对β-肾上腺素受体激动的反应出现脱敏现象,细胞内钙[Ca²⁺]_i瞬变的幅度降低、时程延长,本研究观察上述变化在低氧时发生和发展的时间过程和对应关系。方法:在年龄对等的正常及慢性低氧1d、3d、1周、2周、3周、4周和8周的大鼠,分别分离正常及慢性低氧条件下的心室肌细胞,以Fura-2为[Ca²⁺]_i的指示剂,用光谱荧光法测定心肌细胞的[Ca²⁺]_i瞬变及其对心肌β-受体激动后反应的变化。结果:在低氧1d、3d、1周等时间内,电刺激引起的[Ca²⁺]_i瞬变、咖啡因引起的[Ca²⁺]_i瞬变及电刺激引起的[Ca²⁺]_i瞬变对β-受体激动剂异丙肾上腺素的增加反应无明显变化;在低氧2周以上时,电刺激引起的[Ca²⁺]_i瞬变的幅度开始降低,而时程开始延长,其对异丙肾上腺素的反应也开始降低。咖啡因引起的[Ca²⁺]_i瞬变幅度也开始降低;在低氧3周和4周时,上述变化程度逐渐加重;在低氧8周时各参数的变化有所恢复,但与4周时的变化程度无明显差异。结论:低氧2-4周时,心脏的β-肾上腺素受体发生脱敏现象,脱敏的机制与3种调节[Ca²⁺]_i瞬变的蛋白质:L-型钙通道、ryanodine受体操纵的钙通道和钙泵等活性的降低有关,后者也可能是低氧时心脏功能降低的重要机制。低氧4-8周时,心脏对低氧产生了适应和代偿。     

Endothelin-B receptor-mediated Ca2+ signaling in human melanocytes

 The mechanism of an endothelin-1- (ET-1-) induced intracellular Ca²⁺ ([Ca²⁺]_i) increase and the receptor subtype(s) responsible for this effect in single human melanocytes were studied using fura-2/AM. ET-1 induced a transient increase in [Ca²⁺]_i in a concentration-dependent manner. The transient [Ca²⁺]_i increase was followed by a sustained plateau level of [Ca²⁺]_i which was higher than the initial [Ca²⁺]_i level. IRL-1620, a specific ET-B receptor agonist, increased [Ca²⁺]_i in a dose-dependent manner. BQ-788, a specific ET-B receptor antagonist, abolished the ET-1-induced [Ca²⁺]_i increase, but BQ-123, a specific ET-A receptor antagonist, failed to prevent it. U73122, an inhibitor of phospholipase C (PLC), inhibited the ET-1-induced [Ca²⁺]_i rise in a dose-dependent manner. Prior depletion of intracellular Ca²⁺ stores with thapsigargin, an inhibitor of Ca²⁺-ATPase of the endoplasmic reticulum, abolished the ET-1-induced Ca²⁺ transient, whereas removal of extracellular Ca²⁺ with EGTA eliminated the sustained rise. These results suggest that in cultured human melanocytes the binding of ET-1 to ET-B receptors and the subsequent activation of PLC mediate ET-1-induced [Ca²⁺]_i increase. The transient [Ca²⁺]_i increase is attributed to mobilization of Ca²⁺ from inositol 1,4,5-trisphosphate-sensitive intracellular Ca²⁺ stores, and the sustained [Ca²⁺]_i level may be related to the influx of extracellular Ca²⁺. Received: 21 July 1997 / Received after revision and accepted: 16 September 1997     

红景天甙对低氧培养的兔肺动脉平滑肌细胞增殖的抑制作用

目的:研究红景天甙(Salidroside,Sal)对低氧(3%)条件下兔肺动脉平滑肌细胞(PASMC)增殖、DNA合成、细胞内钙离子浓度的影响。方法:应用细胞培养、四唑盐比色试验(MTT)、[³H]-胸腺嘧啶核苷([³H]-TdR)掺入试验、Fluo-3和激光扫描共聚焦显微术测定细胞内Ca²⁺浓度等方法。结果:低氧24h可直接刺激PASMC,使MTT的A值增加62%(P＜0.05),[³H][³H]-TdR掺入量增加138%(P＜0.01)。在低氧的PASMC培养液中加入Sal(32×10^-5mol/L)可抑制低氧的这种促增殖作用,与低氧组相比MTT的A值下降29%(P＜0.05),[³H][³H]-TdR掺入量下降37%。在低氧的PASMC培养液中加入钙通道阻滞剂verapamil也可抑制低氧的促增殖作用,与低氧组相比较,MTT的A值下降23%(P＜0.05),[³H][³H]-TdR掺入量下降51%(P＜0.01)。PASMC的低氧培养可使细胞内Ca²⁺浓度明显增加(P＜0.05),Sal可抑制低氧所致的细胞内Ca²⁺浓度的上升。结论:红景天甙可抑制低氧促进PASMC增殖、DNA合成的作用,其机制可能与抑制低氧时细胞内Ca²⁺浓度增加有关。     

Proteinase-activated receptor agonists stimulate the increase in intracellular Ca<Superscript>2+</Superscript> in cardiomyocytes and proliferation of cardiac fibroblasts from chick embryos

We studied activation of cultured cardiomyocytes and cardiac fibroblasts from chick embryos induced by agonists of PAR1 (thrombin and PAR1 peptide agonist) and PAR2 (trypsin, factor Xa, and peptide SLIGRL) by analyzing changes in intracellular Ca²⁺ concentration ([Ca²⁺]_i) and cardiac fibroblast proliferation. Exposure of cardiomyocytes with thrombin induced immediate permanent dose-dependent increase in [Ca²⁺]_i. Ca²⁺ response decreased in a calcium-free medium. Peptide agonists of PAR1 and PAR2 also stimulated the increase in [Ca²⁺]_i in cardiomyocytes. Thrombin induced a short-term increase in [Ca²⁺]_i in cardiac fibroblasts and potentiated cell proliferation. PAR2 agonists trypsin and peptide SLIGRL stimulated proliferation of cardiac fibroblasts. Our results indicate that cardiomyocytes and cardiac fibroblasts from chick embryos have at least two types of PAR (types 1 and 2). __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 144, No. 12, pp. 609–612, December, 2007     

Effects of motilin on intracellular free calcium in cultured smooth muscle cells from the antrum of neonatal rats

Aim: The aim of this study was to determine the effects of motilin on [Ca²⁺]_i regulation and its underlying molecular mechanism in cultured antral smooth muscle cells (ASMCs). Methods: Antral cells were isolated and cultured from neonatal rats, and then the [Ca²⁺]_i in these cells was evaluated by calcium fluorescent probe Fluo-3/AM on a laser scanning confocal microscope. Results: We show that motilin dose-dependently increased [Ca²⁺]_i concentration in cultured ASMCs. Pre-incubation of cells with either the calcium antagonist verapamil (10⁻⁵ mol L⁻¹) or the calcium chelator Egtazic (EGTA, 0.1 mmol L⁻¹) significantly suppressed motilin (10⁻⁶ mol L⁻¹) induced [Ca²⁺]_i increase as indicated by fluorescent intensity. Interestingly, after mixing with the non-selective intracellular calcium release blocker TMB-8 (10⁻⁵ mol L⁻¹), guanosine triphosphate regulatory protein antagonist NEM (10⁻⁵ mol L⁻¹), phospholipase C (PLC) inhibitor compound 48/80 (1.2 μg mL⁻¹) and ryanodine at high concentration (10⁻⁵ mol L⁻¹), the motilin-induced [Ca²⁺]_i increase was only partially blocked. The protein kinase C inhibitor d -sphingosine (10⁻⁶ mol L⁻¹), however, did not show any inhibitory effect on motilin-induced [Ca²⁺]_i elevation. Conclusions: Our study suggests that motilin-stimulated [Ca²⁺]_i elevation in ASMCs is probably due to sustained extracellular Ca²⁺ influx and Ca²⁺ release from Ca²⁺ stores via inositol tris-phosphate receptors and ryanodine receptors. Specifically, motilin-induced [Ca²⁺]_i release is accompanied with guanosine triphosphate-binding protein-coupled receptor–PLC–inositol tris-phosphate signalling cascades.     

The intracellular Ca2+ concentration optimal for T cell activation is quite different after ionomycin or CD3 stimulation

The relationship between the initial increase of intracellular Ca²⁺ concentration ([Ca²⁺]_i) (measured at the single-cell level with an imaging system) and the ensuing proliferation was examined in a human T cell clone stimulated by a phorbol ester in combination with ionomycin, thapsigargin or an anti-CD3 mAb (monoclonal antibody against the CD3 molecule, UCHT1). From the responses to various ionomycin concentrations, one can define a range of [Ca²⁺]_i values (400–900 nM) which appears optimal for T cell proliferation; lower [Ca²⁺]_i values are suboptimal, higher values are cytotoxic. It was then examined if the [Ca²⁺]_i requirements were similar following anti-CD3 stimulation. [Ca²⁺]_i oscillations elicited by a concentration of UCHT1 (1/1,000) optimal for mitogenicity fall precisely within the 400–900 nM range. However, very low concentrations of UCHT1 (1/100,000) which evoke barely detectable [Ca²⁺]_i responses still cause the cells to proliferate. The possibility that the lower [Ca²⁺]_i requirements observed following anti-CD3 stimulation was due to [Ca²⁺]_i oscillations was tested under conditions which prevented the appearance of these oscillations. It turns out that an oscillatory Ca²⁺signal is not more mitogenic than a sustained augmentation of [Ca²⁺]_i. Finally, it was examined if overstimulation via CD3 could have toxic consequences similar to those elicited after ionomycin overstimulation. Large transient [Ca²⁺]_i responses can be observed following anti-CD3 stimulation in appropriate conditions, and namely in T cells pretreated with interleukin-2. These [Ca²⁺]_i augmentations are not cytotoxic. A role for the plasmalemmal Ca²⁺ pump in the prevention of cytotoxicity can be demonstrated. In conclusion, the correspondence between the [Ca²⁺]_i response and cell proliferation is entirely different following stimulation by ionomycin and by anti-CD3. In addition, cell proliferation evoked by very low UCHT1 concentration might reveal the existence of a yet unidentified activation pathway.     

Experimental diabetes attenuates calcium mobilization and proliferative response in splenic lymphocytes from mice

The present study was conducted to investigate the effects of the diabetic condition on cytosolic free Ca²⁺ concentration, [Ca²⁺]_i, and the proliferation of splenic lymphocytes from mice. Diabetes was induced in mice by intraperitoneal injection of alloxan. [Ca²⁺]_i and the proliferation ex vivo of splenic lymphocytes isolated from mice were examined using fura-2 and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide, respectively. Diabetes caused a significant increase in resting [Ca²⁺]_i and significantly reduced the ability of concanavalin A (Con A; a T-lymphocyte-selective mitogen) to increase [Ca²⁺]_i, but not that of lipopolysaccharide (LPS; a B-lymphocyte-selective mitogen). In addition, diabetes significantly reduced Con A-stimulated but not LPS-stimulated lymphocyte proliferation. Verapamil (an L-type Ca²⁺ channel blocker) inhibited Con A-induced increases in [Ca²⁺]_i and proliferation in lymphocytes from control and diabetic mice to a similar extent, respectively. These results suggest that diabetes attenuates Con A-stimulated T-lymphocyte proliferation by decreasing [Ca²⁺]_i via reduction of Ca²⁺ entry through L-type Ca²⁺ channels.     

Role of [Ca2+]i in lethal oxidative injury in rat cultured inner medullary collecting duct cells

Reactive oxygen metabolites have been implicated in the pathogenesis of toxic, ischaemic and immunologically mediated renal injury. An increase in the cytosolic free Ca²⁺ concentration ([Ca²⁺]_i) has been proposed as a mechanism of oxidative stress-induced cell injury. We used a fluorescence spectrometer and a fluorescence probe to measure the [Ca²⁺]_i and viability of rat primary cultured inner medullary collecting duct (IMCD) cells during oxidative stress induced by 5 mM tert-butyl hydroperoxide (TBHP). Initially, this oxidative stress evoked a small increase in [Ca²⁺]_i which was followed by a slower sustained increase from the resting level of 170.8±38.8 nM to 1490.5±301.7 nM after 60 min, and this preceded the loss of plasma membrane integrity, measured by the propidium iodide fluorescence method. The elimination of extracellular Ca²⁺ from the culture medium prevented the TBHP-induced [Ca²⁺]_i increase and improved cell viability. Restoration of extracellular Ca²⁺ resulted in an immediate and large increase in [Ca²⁺]_i and extensive cell death. Verapamil, a Ca²⁺ channel blocker, inhibited the [Ca⁺]_i increase and afforded significant protection against cellular injury following exposure to TBHPinduced oxidative stress. Extracellular acidosis also prevented the increase in [Ca²⁺]_i and cell death caused by this oxidative stress. These results are consistent with the hypothesis that oxidative stress-induced IMCD cellular injury may be the result of increased [Ca²⁺]_i caused, in part, by activation of voltage-dependent Ca²⁺ channels.     

红景天苷促进培养乳鼠心肌细胞肌质网钙离子的释放

目的: 观察红景天苷对乳鼠心肌细胞胞浆Ca²⁺浓度的影响并分析其可能的作用机制。方法: 应用荧光指示剂Fluo-3/AM负载培养大鼠乳鼠的心肌细胞,用激光共聚焦显微镜动态观察胞内游离钙荧光信号强度的变化,检测不同浓度红景天苷对培养心肌细胞胞内游离钙离子浓度([Ca²⁺]_i)的影响。结果: 红景天苷浓度为15 mg/L、30 mg/L和60 mg/L时,细胞内的平均[Ca²⁺]_i升高,峰值分别为574.08±4.65、591.86±3.64和618.66±4.27(均P<0.01);有剂量依赖性而无时间依赖性。用维拉帕米阻断细胞膜外钙内流时,红景天苷同样引起细胞内[Ca²⁺]_i升高,峰值由357.74±3.13、387.17±2.37和391.43±1.34分别上升到480.86±3.98、496.70±3.08和522.18±3.19(均P<0.01)。结论: 红景天苷能升高乳鼠心肌细胞中[Ca²⁺]_i,其机制可能与其促进肌浆网钙离子释放有关。     

Alterations of excitation-contraction coupling by platelet-derived growth factor in enzymatically isolated and cultured vascular smooth muscle cells

We studied stimulus-specific alterations of the excitation-contraction coupling pathway in freshly isolated contractile and subcultured non-contractile vascular smooth muscle cells. Using the calcium indicator aequorin, we detected physiological increases in cytoplasmic free calcium ([Ca²⁺]_i) in subcultured smooth muscle cells subjected to angiotensin or 33 mM potassium depolarization. These increases were qualitatively identical to those previously measured in intact vascular strips. Platelet-derived growth factor (PDGF) induced a slow, sustained [Ca²⁺]_i increase when applied to the subcultured smooth muscle cells at low picomolar concentrations. Freshly isolated, contractile vascular smooth muscle cells, prepared by a novel technique, exhibited a slow shortening of 20% of resting length in response to PDGF. PDGF also markedly potentiated smooth muscle cell shortening in response to an ED₅₀ dose of phenylephrine. This effect was PDGF concentration dependent. The time course of shortening induced by PDGF alone was consistent with the time course of the PDGF-induced [Ca²⁺]_i increase in the cultured smooth muscle cells. These data suggest that agonists which induce [Ca²⁺]_i changes in contractile smooth muscle cells may retain this ability with respect to cultured smooth muscle cells. PDGF, a peptide mitogen for proliferative smooth muscle cells, may also serve to modulate vascular tone by modestly raising [Ca²⁺]_i in contractile smooth muscle cell and, therefore, sensitizing the cells to alpha adrenergic agonists.