δ阿片受体激活对培养心肌细胞生长的影响

目的观测δ阿片受体激活对培养的心肌细胞是否有直接的促生长作用。方法体外培养乳大鼠心肌细胞。结晶紫法和[3H]TdR法测定心肌细胞的增生;流式细胞仪测定细胞周期百分比;Lowry等法测定培养细胞蛋白质含量;倒置显微镜下计数心肌细胞搏动频率。结果δ阿片受体激动剂D-丙(2)-D-亮(5)脑啡肽(DADLE)10 nmol.L-1~10μmol.L-1能浓度依赖性促进细胞增殖,增加细胞蛋白质含量,增加细胞搏动频率;高选择性δ阿片受体拮抗剂纳曲吲哚(naltrindole)10μmol.L-1对DADLE的促进作用有一定的抑制作用。结论δ阿片受体激活对体外培养的心肌细胞生长有促进作用。     

小牛血去蛋白提取物注射液对培养的心肌细胞缺氧再给氧损伤的保护作用

目的 :观察小牛血去蛋白提取物注射液(deproteinizedextractofcalfblood ,DECB)对培养的心肌细胞缺氧再给氧损伤的保护作用。方法 :以培养的心肌细胞为模型 ,在缺氧再给氧损伤细胞后 ,观察心肌细胞活力及搏动频率的变化 ,测定培养液上清乳酸脱氢酶的含量。结果 :DECB 0 .1及 0 .3g·L-1可以提高损伤后心肌细胞的活力 ,提高心肌细胞的搏动频率 ,并使培养液上清乳酸脱氢酶的含量明显降低。结论 :DECB对缺氧再给氧损伤培养的心肌细胞具有保护作用。     

吡格列酮对高糖高胰岛素诱导的心肌细胞肥大的影响

目的　利用体外培养的乳鼠心肌细胞 ,观察吡格列酮对高浓度葡萄糖与高浓度胰岛素共同诱导的肥大心肌细胞的影响 ,进一步推测吡格列酮对糖尿病并发心肌肥大的可能作用及作用机制。方法　以培养的乳鼠心肌细胞为模型分组给药后 ,用Lowrys法测心肌细胞的蛋白质含量 ;用 [3 H]leucine标记法测定心肌细胞蛋白的合成 ;利用计算机图像分析系统测心肌细胞的体积 ;用显微镜目镜计数心肌细胞搏动的频率。结果　 10 μmol·L-1浓度的吡格列酮在对高糖高胰岛素诱导的肥大心肌细胞的蛋白含量、蛋白合成及体积均有显著的抑制作用。其抑制肥大的效果比维拉帕米更为明显 ;同时观察到吡格列酮同维拉帕米一样有抑制心肌细胞搏动的作用。结论　吡格列酮能有效抑制高糖高胰岛素诱导的心肌细胞肥大。这种作用可能是通过作用于PPARγ来实现的。     

κ受体激动对去甲肾上腺素诱导的心肌肥大的抑制作用

目的　利用体外培养的乳鼠心肌细胞 ,观测κ阿片肽受体激动对去甲肾上腺素 (norepinephrine ,NE)诱导的心肌肥大的抑制作用 ,并探讨作用机制。方法　对培养乳鼠心肌细胞分组给药 48h后 ,用Lowrys法测心肌细胞的蛋白质含量 ;用消化分离法 ,通过目镜标尺测心肌细胞体积 ;用镜下计数法测心肌细胞的搏动频率。结果　κ阿片肽受体激动剂U5 0 ,488H(简称U5 0 ) (0 1～ 10 μmol·L-1)对培养心肌细胞的蛋白质含量具有抑制作用并呈剂量依赖性 ;对NE诱导的心肌细胞蛋白含量增加、体积增大具有抑制作用 ;同时观察到U5 0 ,488H(1μmol·L-1)具有抑制心肌细胞搏动频率的作用。U5 0 ,488H的上述作用均可被κ阿片肽受体拮抗剂Nor BNI(1μmol·L-1)拮抗。结论　U5 0 ,488H对NE诱导的心肌肥大具有抑制作用 ,这种作用是通过激动κ阿片肽受体发挥的。     

槲皮素对血管紧张素Ⅱ致培养乳鼠心肌细胞肥大的抑制作用

目的:研究槲皮素(Quercetin,Que)对血管紧张素Ⅱ(Aug Ⅱ)诱发培养乳鼠心肌细胞肥大的抑制作用及机制。方法:分别用[~3H]胸苷、[~(14)C]尿苷和[~3H]酪氨酸标记测定DNA、RNA和蛋白质合成;用Lowry法测定蛋白质含量;用组蛋白ⅢS、[γ-~(32)P]ATP与蛋白激酶C(PKC)酶液一起保温测定PKC活性;用聚谷氨酸·酪氨酸(4:1)多肽、[γ-~(32)P]ATP与酪氨酸蛋白激酶(TPK)酶液一起保温测TPK活性。结果:Aug Ⅱ作用于心肌细胞24h后,心肌细胞总蛋白含量明显增加(P<0.01),[~(14)C]尿苷和[~3H]酪氨酸掺入量明显增加(P<0.01),而[~3H]胸苷掺入量未见增加(P>0.05);Aug Ⅱ作用30min后,心肌细胞PKC和TPK活性明显增加。Que(1-100μmol/L)能剂量依赖性地抑制Ang Ⅱ所致心肌细胞总蛋白含量、[~(14)C]尿苷和[~3H]酪氨酸掺入量、PKC及TPK活性的增加。结论:Que可抑制Aug Ⅱ致培养乳鼠心肌细胞肥大,该作用与抑制PKC及TPK活性有关。     

吡格列酮对培养的心肌细胞肥大的抑制作用

目的利用体外培养的乳鼠心肌细胞,观察吡格列酮对高浓度葡萄糖与去甲肾上腺素共同诱导的肥大心肌细胞的影响,进一步推测吡格列酮对糖尿病性心肌肥大的可能作用及作用机制。方法以培养的乳鼠心肌细胞为模型分组给药后,用显微镜目镜计数心肌细胞搏动的频率;用Lowry’s法测心肌细胞的蛋白质含量;用[3H]leucine标记法测定心肌细胞蛋白的合成;利用计算机图象分析系统测心肌细胞的体积。结果吡格列酮在1～10μmol·L-1浓度对25.5mmol·L-1高糖与1μmol·L-1去甲肾上腺素联合诱导的肥大心肌细胞的蛋白含量、蛋白合成及体积均有显著的抑制作用。其抑制肥大的效果比1μmol·L-1维拉帕米更为显著;同时观察到10μmol·L-1吡格列酮同1μmol·L-1维拉帕米一样有抑制心肌细胞搏动的作用。结论吡格列酮能有效抑制高糖与去甲肾上腺素联合诱导的心肌细胞肥大。这种作用可能是通过作用于PPARγ来实现的。     

高糖对去甲肾上腺素诱导的心肌细胞肥大的促进作用

目的　利用体外培养的乳鼠心肌细胞 ,观测高浓度葡萄糖对去甲肾上腺素诱导的心肌细胞肥大的影响 ,根据所得结果来推测高糖与心肌细胞肥大之间的相互关系及可能作用机制。方法　以培养的乳鼠心肌细胞为模型分组给药后 ,用Lowrys法测心肌细胞的蛋白质含量 ;用消化分离法 ,利用计算机图像分析系统测心肌细胞的体积 ;用 [3 H]leucine标记法测定心肌细胞蛋白的合成。结果　与对照组相比 ,高糖组心肌细胞蛋白含量、体积、蛋白合成均有明显增加 ,NE组心肌细胞蛋白含量、体积、蛋白合成增加更为明显 ;而与高糖组或NE组相比较 ,高糖加NE组心肌细胞蛋白含量、体积、蛋白合成增加则最为显著。结论　单纯高糖培养可明显促进心肌细胞肥大 ,同时高浓度葡萄糖可在去甲肾上腺素诱导的心肌细胞肥大的基础上促使其进一步肥大 ,且两者间作用效果具有不完全叠加性     

尿促皮素诱导乳大鼠心肌细胞肥大的作用及信号传导机制

目的探讨尿促皮素(urocortin)诱导大鼠心肌细胞肥大的作用及其信号传导机制。方法实验分8组,正常对照组、尿促皮素0.1μmol.L-1组、星形孢菌素(Sta)1μmo.lL-1、H890.1μmol.L-1和维拉帕米(Ver)1μmol.L-1组及尿促皮素分别加Sta,H89和Ver组。采用体外培养的乳大鼠心肌细胞,应用尿促皮素0.1μmol.L-1诱导心肌肥大,观察Sta1μmol.L-1,H890.1μmo.lL-1和Ver1μmol.L-1的作用,进一步探讨尿促皮素0.1μmol.L-1诱导心肌肥厚的作用机制。用消化分离法及计算机图像分析系统检测心肌细胞直径;[3H]亮氨酸掺入法测定心肌细胞蛋白质的合成;用Lowry法检测心肌细胞蛋白质含量;用Western蛋白印迹法测定心房钠尿肽(ANP)表达;采用Till阳离子测定系统,以Fura-2/AM为荧光探针,观察心肌细胞[Ca2+]i瞬间变化。结果尿促皮素使心肌细胞直径、蛋白质合成、蛋白质含量和ANP表达分别增加30.9%,36.3%,35.5%和34.7%;尿促皮素+Sta组使心肌细胞直径、蛋白质合成、蛋白质含量和ANP表达分别降低了16.5%,22.1%,18.1%和21.3%;尿促皮素+H89组使心肌细胞直径、蛋白质合成、蛋白质含量和ANP表达分别降低了16.6%,21.5%,19.5%和20.6%;尿促皮素+Ver组使心肌细胞直径、蛋白质合成、蛋白质含量和ANP表达分别降低了17.1%,20.9%,17.9%及19.9%;尿促皮素能够使心肌细胞[Ca2+]i瞬间变化水平增高,Sta,H89和Ver能够降低尿促皮素引起的心肌细胞[Ca2+]i瞬间变化升高。结论尿促皮素可能通过蛋白激酶C和蛋白激酶A信号途径影响L-型Ca2+通道,进而影响细胞[Ca2+]i瞬间变化水平,诱导乳大鼠心肌细胞肥大。     

欣力胶囊抑制Angiotensin-2刺激引起的心肌细胞损伤

目的在原代培养新生大鼠心肌细胞水平,观察欣力胶囊对血管紧张素Ⅱ(AngⅡ)刺激引起的心肌细胞损伤的保护作用。方法采用免疫组织化学方法(α-actin)鉴定原代心肌细胞纯度。以欣力胶囊灌胃大鼠进行血清药理学实验;将心肌细胞分组为:正常对照组、AngⅡ刺激组和欣力胶囊含药血清治疗组(包括3个不同剂量组)。倒置显微镜下于不同时段计数心肌细胞搏动频率;用反转录聚合酶链反应(RT-PCR)检测心肌细胞肥大早期基因(c-myc、c-jun和c-fos)的表达改变;利用SABA—18生化分析仪测定不同分组细胞培养液上清乳酸脱氢酶(LDH)、肌酸激酶(CK-MB)以及天冬氨酸转氨酶(GOT)含量。结果通过α-actin免疫组织化学染色鉴定,心肌细胞纯度达95%,满足实验需要。欣力胶囊含药血清在体外能够有效抑制AngⅡ刺激引起的搏动频率增加(P<0.05);抑制心肌细胞肥大早期基因(c-myc、c-jun和c-fos)的表达;减少细胞培养液上清中心肌酶(LDH、CK-MB、GOT)的含量(P<0.05)。结论欣力胶囊可以抑制AngⅡ刺激对原代培养大鼠心肌细胞的损伤。     

δ阿片受体激活对过氧化氢损伤的心肌细胞的保护作用

目的研究δ阿片受体激活剂D-丙(2)-D-亮-(5)-脑啡肽(DADLE)对过氧化氢(H2O2)损伤的心肌细胞的保护作用及其机制。方法分离乳大鼠心肌细胞,培养48h后分为正常对照、H2O2(200μmol.L-1)、H2O2+DADLE(1μmol.L-1)、H2O2+DADLE+纳曲吲哚(10μmol.L-1)和H2O2+DADLE+U0126(10nmol.L-1)组,继续培养48h。用[3H]TdR掺入法检测心肌细胞增殖反应,流式细胞仪检测心肌细胞凋亡百分率,乳酸脱氢酶(LDH)活性测定试剂盒测定培养上清LDH活性,硫代巴比妥酸显色法测定细胞内丙二醛(MDA)含量,黄嘌呤氧化酶法测定细胞内超氧化物歧化酶(SOD)活性,Western蛋白印迹法检测细胞外信号调节激酶磷酸化(p-ERK)水平。结果①与正常对照组比较,H2O2组心肌细胞[3H]TdR掺入值明显降低,细胞凋亡率升高;培养上清LDH活性和MDA含量明显增加,SOD活性和Ap-ERK/AERK的比值降低。②与H2O2组比较,DADLE可使心肌细胞[3H]TdR掺入值升高,细胞凋亡率下降;培养上清LDH活性和MDA含量降低,SOD活性和Ap-ERK/AERK的比值升高。③分别加入δ阿片受体拮抗剂纳曲吲哚和ERK拮抗剂U0126,DADLE对上述指标的逆转作用被抑制。结论δ阿片受体激活对H2O2损伤的心肌细胞具有保护作用,其机制可能与其增强心肌细胞的抗氧化功能及促进ERK磷酸化有关。     

Effects of Bisphosphonates on Glucose Transport in a Conditionally Immortalized Rat Retinal Capillary Endothelial Cell Line (TR-iBRB Cells)

The objective of the present study was to elucidate the effect of bisphosphonates, anti-osteoporosis agents, on glucose uptake in retinal capillary endothelial cells under normal and high glucose conditions. The change of glucose uptake by pre-treatment of bisphosphonates at the inner blood-retinal barrier (iBRB) was determined by measuring cellular uptake of [³H]3-O-methyl glucose (3-OMG) using a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB cells) under normal and high glucose conditions. [³H]3-OMG uptake was inhibited by simultaneous treatment of unlabeled D-glucose and 3-OMG as well as glucose transport inhibitor, cytochalasin B. On the other hand, simultaneous treatment of alendronate or pamidronate had no significant inhibitory effect on [³H]3-OMG uptake by TR-iBRB cells. Under high glucose condition of TR-iBRB cells, [³H]3-OMG uptake was increased at 48 h. However, [³H]3-OMG uptake was decreased significantly by pre-treatment of alendronate or pamidronate compared with the values for normal and high glucose conditions. Moreover, geranylgeraniol (GGOH), a mevalonate pathway intermediate, increased the uptake of [³H]3-OMG reduced by bisphosphonates pre-treatment. But, pre-treatment of histamine did not show significant inhibition of [³H]3-OMG uptake. The glucose uptake may be down regulated by inhibiting the mevalonate pathway with pre-treatment of bisphosphonates in TR-iBRB cells at high glucose condition.     

Serum effect on cellular uptake of spermidine, spergualin, 15-deoxyspergualin, and their metabolites by L5178Y cells

Spergualin (SG) and 15-deoxyspergualin (DSG) were more slowly incorporated into L5178Y cells than spermidine. SG and DSG inhibited carrier-mediated transport of [3H]spermidine competitively with inhibition constants of 0.67 mM and 0.45 mM, respectively. Addition of calf serum stimulated uptake of [3H]spermidine into the cells in a serum concentration-dependent manner. The effect was not observed when horse serum was used in place of calf serum. Preincubation of spermidine in calf serum for 1 hour before addition to cells remarkably decreased cellular incorporation of tritium. Three amine oxidase inhibitors, aminoguanidine, 3-hydroxybenzyloxyamine, and semicarbazide, inhibited stimulation of uptake of [3H]spermidine by calf serum and the decrease of it by preincubation in calf serum. So we propose that cellular incorporation or binding of products generated by oxidation of spermidine by amine oxidase in calf serum was much faster than that of spermidine itself and they were unstable and transformed quickly to unincorporable or non-binding substances if cellular targets were not present. Effect of amine oxidase inhibitors on cytotoxic activity of SG and DSG were determined in low and high concentrations of calf serum. In the presence of 10% calf serum in the basal medium, cytotoxicity to L5178Y cells by SG and DSG was suppressed at high drug concentrations (above 10 micrograms/ml) and enhanced at low drug concentrations (below 2.5 micrograms/ml) by amine oxidase inhibitors. In the presence of 0.5% calf serum suppression of cytotoxicity at high drug concentrations by amine oxidase inhibitors was also observed, but enhancement at low drug concentrations was obscure. These data may suggest the existence of two kinds of cytotoxic mechanism of SG and DSG, one dependent on and one independent of amine oxidase in serum.     

Hypertonicity enhances GABA uptake by cultured rat retinal capillary endothelial cells

We have reported previously that taurine transporter (TauT) mediates γ-aminobutyric acid (GABA) as a substrate in a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2 cells). This study investigates how TauT-mediated GABA transport is regulated in TR-iBRB2 cells under hypertonic conditions. [3H]GABA uptake by TR-iBRB2 cells exposed to 12 h- to 24 h-hypertonic culture medium was significantly greater than that of isotonic culture medium. [3H]GABA uptake by TR-iBRB2 cells was Na(+)-, Cl(-)-, and concentration-dependent with a Michaelis-Menten (K(m)) constant of 3.5 mM under isotonic conditions and K(m) of 0.324 and 5.48 mM under hypertonic conditions. Under hypertonic conditions, [3H]GABA uptake by TR-iBRB2 cells was more potently inhibited by substrates of TauT, such as taurine and β-alanine, than those of GABA transporters such as GABA, nipecotic acid, and betaine. These results suggest that an unknown high-affinity GABA transport process and TauT-mediated GABA transport are enhanced under hypertonic conditions. In conclusion, hypertonicity enhances GABA uptake by cultured rat retinal capillary endothelial cells.     

Kappa-opioid receptor stimulation inhibits growth of neonatal rat ventricular myocytes

The effects of trans-(+/-)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]-benzeneacetamide methanesulfonate salt (U50,488H), a selective kappa-opioid receptor agonist, on growth in neonatal ventricular myocytes were determined. In 15% serum culture medium, U50,488H at 0.1-1 microM significantly reduced the protein content, [3H]leucine uptake and cell size of the myocytes. The effect of U50,488H on protein content was abolished in the presence of 1 microM nor-binaltorphimine (nor-BNI), a selective kappa-opioid receptor antagonist. In a 0.4% serum medium, U50,488H at 0.1-1 microM had no effect on myocyte growth. Interestingly, 1 microM U50,488H abolished the stimulatory effects of 1 microM norepinephrine on protein content, [3H]leucine uptake and cell size of the myocytes in the low serum medium. The effect of U50,488H was abolished by 1 microM nor-BNI. With the exception of cell size, the effects of norepinephrine were completely abolished by blockade of both alpha- and beta-adrenoceptors, but only partially blocked by blockade of either adrenoceptors. These results provide first evidence that kappa-opioid receptor stimulation inhibits growth of the neonatal ventricular myocyte as a result of direct action as well as by inhibiting sympathetic stimulation of the heart. The stimulatory effects of sympathetic activity on growth occurs via both alpha- and beta-adrenoceptors.     

Uptake mechanism of valproic acid in human placental choriocarcinoma cell line (BeWo)

Valproic acid is an anticonvulsant widely used for the treatment of epilepsy. However, valproic acid is known to show fetal toxicity, including teratogenicity. In the present study, to elucidate the mechanisms of valproic acid transport across the blood-placental barrier, we carried out transcellular transport and uptake experiments with human placental choriocarcinoma epithelial cells (BeWo cells) in culture. The permeability coefficient of [3H]valproic acid in BeWo cells for the apical-to-basolateral flux was greater than that for the opposite flux, suggesting a higher unidirectional transport in the fetal direction. The uptake of [3H]valproic acid from the apical side was temperature-dependent and enhanced under acidic pH. In the presence of 50 microM carbonyl cyanide p-trifluoromethoxylhydrazone, the uptake of [3H]valproic acid was significantly reduced. A metabolic inhibitor, 10 mM sodium azide, also significantly reduced the uptake of [3H]valproic acid. Therefore, valproic acid is actively transported in a pH-dependent manner on the brush-border membrane of BeWo cells. Kinetic analysis of valproic acid uptake revealed the involvement of a non-saturable component and a saturable component. The Michaelis constant for the saturable transport (K(t)) was smaller under acidic pH, suggesting a proton-linked active transport mechanism for valproic acid in BeWo cells. In the inhibitory experiments, some short-chain fatty acids, such as acetic acid, lactic acid, propanoic acid and butyric acid, and medium-chain fatty acids, such as hexanoic acid and octanoic acid, inhibited the uptake of [3H]valproic acid. The uptake of [3H]valproic acid was also significantly decreased in the presence of 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid, salicylic acid and furosemide, which are well-known inhibitors of the anion exchange system. Moreover, p-aminohippuric acid significantly reduced the uptake of [3H]valproic acid. These results suggest that an active transport mechanism for valproic acid exists on the brush-border membrane of placental trophoblast cells and operates in a proton-linked manner.     

Effects of murine recombinant interferon-gamma on rat liver fat storing cell proliferation, cluster formation and proteoglycan synthesis

Rat liver fat storing cells (FSC, perisinusoidal lipocytes, Ito cells) in primary culture were exposed to various concentrations of murine recombinant interferon-gamma (rIFN-gamma) in the range of 1 to 50 units/mL medium for 72 hr. FSC kept in complete medium (10% fetal calf serum) showed a dose-dependent increase of both [3H]thymidine incorporation (up to 2.3-fold) and DNA content of culture. Reverse (inhibitory) effects were obtained with cells kept under serum-reduced (0.5% fetal calf serum) conditions. The synthesis of medium proteoglycans and of total cellular protein was not affected by rIFN-gamma. By bromodeoxyuridine-staining (BrdUrd) and phase contrast microscopy it is shown that rIFN-gamma stimulates strongly the cluster growth of FSC in culture. The cluster forming cells differ in their morphology and their cytoskeleton-staining from typical FSC. They were found to be mostly desmin and alpha-actin negative or weakly positive but highly proliferative. Because no contaminating fibroblasts and other cell types were detected in any appreciable amounts in the early cultures we conclude that the clustered cells might be a rapidly proliferating subpopulation of FSC, which is promoted by rIFN-gamma.     

High glucose induces cell death of cultured human aortic smooth muscle cells through the formation of hydrogen peroxide

Alterations of the vessel structure, which is mainly determined by smooth muscle cells through cell growth and/or cell death mechanisms, are characteristic of diabetes complications. We analysed the influence of high glucose (22 mM) on cultured human aortic smooth muscle cell growth and death, as hyperglycaemia is considered one of the main factors involved in diabetic vasculopathy. Growth curves were performed over 96 h in medium containing 0.5% foetal calf serum. Cell number increased by 2 - 4 fold over the culture period in the presence of 5.5 mM (low) glucose, while a 20% reduction in final cell number was observed with high glucose. Under serum-free conditions, cell number remained constant in low glucose cultures, but a 40% decrease was observed in high glucose cultures, suggesting that high glucose may induce increased cell death rather than reduced proliferation. Reduced final cell number induced by high glucose was also observed after stimulation with 5 or 10% foetal calf serum. The possible participation of oxidative stress was investigated by co-incubating high glucose with different reactive oxygen species scavengers. Only catalase reversed the effect of high glucose. Intracellular H(2)O(2) content, visualized with 2',7'-dichlorofluorescein and quantified by flow cytometry, was increased after high glucose treatment. To investigate the cell death mechanism induced by high glucose, apoptosis and necrosis were quantified. No differences were observed regarding the apoptotic index between low and high glucose cultures, but lactate dehydrogenase activity was increased in high glucose cultures. In conclusion, high glucose promotes necrotic cell death through H(2)O(2) formation, which may participate in the development of diabetic vasculopathy.     

Ceramides that mediate apoptosis reduce glucose uptake and transporter affinity for glucose in human leukaemic cell lines but not in neutrophils

We have demonstrated that CD95-induced apoptosis in a human leukaemic T-cell line resulted in loss of glucose transporter function (Berridge et al. 1996). To determine whether ceramide, a mediator of CD95 and tumour necrosis factor-alpha-induced apoptosis, has similar effects on glucose transport, the human leukaemic cell lines, Jurkat and U937, and human peripheral blood neutrophils were treated with ceramide or sphingomyelinase and the effects on glucose transport determined by measuring [3H]-2-deoxyglucose uptake. We show that in U937 and Jurkat cells, the cell permeable ceramides, C2 (N-acetylsphingosine) and C6 (N-hexanoylsphingosine) inhibit glucose uptake within minutes of initiating ceramide treatment, 60-70% inhibition being observed within 2 hr. Loss of glucose transport correlated with loss of proliferative response, but metabolic activity as measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) reduction, was affected to a much lesser extent. With Jurkat and U937 cells, the inhibitory effects of ceramides on glucose transport were associated with reduced affinity of glucose transporters for glucose (Km). Similar effects were observed with sphingomyelinase. With human peripheral blood neutrophils, C2 and C6-ceramides inhibited glucose uptake by 70-80% within 30 min. without affecting transporter affinity for glucose, but the maximum velocity of uptake (Vmax) was reduced. These results show that acute regulation of glucose transport is an early effector mechanism of cell death induced by ceramides in human leukaemic cell lines and peripheral blood neutrophils. This is the first study which describes ceramide-induced early physiological/biochemical events leading to cell death in human cells.     

3,4-二羟基苯乙酮抑制培养兔主动脉平滑肌细胞的增殖

传代SMC生长d 6加入DHAP(10～80μg/ml)介质。结晶紫染色法和[~3H]TdR参入试验表明SMC的生长受到DHAP的显著抑制,这一作用随剂量增加而加强。在细胞增殖周期的G_1期加药比S期加药产生更为明显的抑制作用,G_1期末加药抑制作用最大。DHAP的这一作用可能和降低SMC膜流动性有关。