Studies on the calcium antagonistic action of tetrandrine: X VI. Effects of tetrandrine on positive staircase phenomenon and post-rest potentiation of contraction of isolated guinea pig left atrium

Summary Tetrandrine (Tet) 0.32 mmol/1 and verapamil (Ver) 30 μmol/1 inversed positive staircase phenomenon of contraction in left alrium of guinea pig. After the preparation was partially depolarized with K₊ 20 mmol/1, such effect of Tet and Ver became much more pronounced. Propranolol could not inverse the positive staircase phenomenon. In addition, after treatment with Tet 0.32 mmol/1 for 20 min the post -rest potentiation of myocardial contraction in the left atrium was depressed by 2.8±0.3 g (P<0.01). The results suggest that the negative inotropism of Tet is related not only to the inhibition of Ca⁺⁺ influx to the cells, but also to the decrease of the intracellular Ca⁺⁺ release.     

K<Superscript>+</Superscript> channels and their effects on membrane potential in rat bronchial smooth muscle cells

Summary In order to investigate the K⁺ channels and their effects on resting membrane potential (Em) and excitability in rat bronchial smooth muscle cells (BSMCs), the components of outward K⁺ channel currents and the effects of K⁺ channels on Em and tension in rat bronchial smooth muscle were observed by using standard whole-cell recording of patch clamp and isometric tension recording techniques. The results showed that under resting conditions, total outward K⁺ channel currents in freshly isolated BSMCs were unaffected by ATP-sensitive K⁺ channel blocker. There were two types of K⁺ currents: voltage-dependent delayed rectifier K⁺ channel (K_v) and large conductance calcium-activated K⁺ channel (BK_Ca) currents. 1 mmol/L 4-aminopyridine (4-AP, an, inhibitor of K_V) caused a significant depolarization (from −8.7±5.9 mV to −25.4±3.1 mV,n=18,P<0.001). In contrast, 1 mmol/L tetraethylammonium (TEA, an inhibitor of BK_Ca) had no significant effect on Em (from −37.6±4.8 mV to −36.8±4.1 mV,n=12,P>0.05). 4-AP caused a concentration-dependent contraction in resting bronchial strips. TEA had no effect on resting tension, but application of 5 mmol/L TEA resulted in a left shift with bigger pD₂ (the negative logarithm of the drug concentration causing 50% of maximal effect) (from 6.27±0.38 to 6.89±0.54,n=10,P<0.05) in the concentration-effect curve of endothine-1, and a right shift with smaller pD₂ (from 8.10±0.23 to 7.69±0.08,n=10,P<0.05) in the concentration-effect curve of isoprenaline. It was suggested that in rat BSMCs there may be two types of K⁺ channels, K_v and BK_Ca, which serve distinct roles. K_v participates in the control of resting Em and tension. BK_Ca is involved in the regulation of relaxation or contraction associated with excitation. LIU Xiansheng, male, born in 1969, M. D., Ph. D. This project was supported by a grant from the National Natural Sciences Foundation of China (No. 30270583).     

不同糖浓度对颌骨骨髓间充质干细胞成骨分化的影响

目的 研究不同糖浓度对颌骨骨髓间充质干细胞（orofacial bonemesenchymal stem cells, OFMSCs）增殖和成骨分化的影响。方法 体外分离、培养OFMSCs,成骨、成脂、成软骨分化诱导及鉴定,并使用不同含糖量培养基（5.5、11、16.5、25、44 mmol/L）培养OFMSCs,以5.5 mmol/L基准糖浓度为对照组,其余组为实验组。采用CCK-8及流式细胞仪检测各组OFMSCs的增殖活性及增殖指数,OFMSCs成骨诱导后4、7 d检测碱性磷酸酶（ALP）活性,21 d进行茜素红染色及矿化定量分析,同时用RT-PCR检测3、7、14及21 d相关成骨基因Runx2、Osterix的表达。结果 培养的OFMSCs成骨诱导21 d后茜素红染色可见钙结节,成脂诱导14 d 后油红O染色可见红色脂滴,成软骨诱导14 d 后阿利新蓝染色可见蓝色胞浆;糖浓度在5.5～25 mmol/L促进OFMSCs的增殖,但随着糖浓度的继续增加（25～44 mmol/L）,OFMSCs的增殖活性受抑制;成骨诱导时,随着培养液糖浓度升高,ALP活性呈剂量依赖性降低（P Runx2、Osterix mRNA 表达量高于实验组（P Runx2、Osterix mRNA 表达量均出现先上调再下调的趋势。结论 在一定范围的糖浓度内,糖浓度升高可促进OFMSCs的增殖;而糖浓度升高对成骨分化呈抑制效应。     

K~+ Channels and Their Effects on Membrane Potential in Rat Bronchial Smooth Muscle Cells

In order to investigate the K~+ channels and their effects on resting membrane potential(Em) and excitability in rat bronchial smooth muscle cells (BSMCs), the components of outward K~+channel currents and the effects of K~+ channels on Em and tension in rat bronchial smooth musclewere observed by using standard whole-cell recording of patch clamp and isometric tension recordingtechniques. The results showed that under resting conditions, total outward K~+ channel currents infreshly isolated BSMCs were unaffected by ATP-sensitive K~+ channel blocker. There were two typesof K~+ currents: voltage-dependent delayed rectifier K~+ channel (Kv) and large conductance calcium-activated K~+ channel (BK_(Ca)) currents. 1 mmol/L 4-aminopyridine (4-AP, an inhibitor of Kv)caused a significant depolarization (from — 8.7±5.9mV to —25.4±3.1mV, n=18, P<0.001).In contrast, 1 mmol/L tetraethylammonium (TEA, an inhibitor of BK_(Ca)) had no significant effect onEm (from —37.6±4.8 mV to —36.8±4.1 mV, n=12, P>0.05). 4     

吡哆胺和替米沙坦对人肾小管上皮细胞活性氧生成的影响

 目的  通过体外培养人肾小管上皮细胞株（HK-2）细胞,探讨吡哆胺和替米沙坦对肾小管上皮细胞活性氧生成影响。方法  HK-2细胞培养并按干预方式不同分为HK-2正常对照组,血管紧张素Ⅱ组（10^-6 mol/L,AngⅡ组）,替米沙坦组［替米沙坦（10 ^-6 mol/L）+ AngⅡ（10^-6 mol/L）,T组］,吡哆胺1组［吡哆胺（1 mmol/L）+ AngⅡ（10^-6 mol/L）,P1 组）,吡哆胺2 组［吡哆胺（10 mmol/L）+ AngⅡ（10^-6 mol/L）,P2］,替米沙坦+吡哆胺联合组［替米沙坦（10^-6 mol/L）垣吡哆胺（10 mmol/L）+ AngⅡ（10^-6 mol/L）,TP 组］。流式细胞仪检测细胞上清液中的活性氧（ROS）浓度,荧光实时定量PCR方法检测HK-2 细胞所表达的转化生长因子β1（TGF-β1 ）和晚期糖基化终产物受体（RAGE ）mRNATGF-β1 表达量。结果  检测细胞上清液中的ROS 浓度,与AngⅡ组比较,P1 组和P2 组均显著降低ROS 浓度（P < 0.01）,TP 组的ROS 生成也明显减少（ P< 0.01）,但T组与AngⅡ组间比较差异无统计学意义（ P>0.05）;与AngⅡ组比较,T组、P2 组和TP 组的HK-2 细胞的, mRNA表达量明显降低（ P< 0.05）;与T 组和P2 比较,TP组HK-2 细胞的mRNA表达进一步降低（ P< 0.01）。结论  替米沙坦和吡哆胺可协同下调HK-2 细胞转化生长因子TGF-β1 和RAGE 的表达;在改善氧化应激功能上,吡哆胺作用强于替米沙坦。     

生后至8个月龄大鼠肝贮脂细胞的电镜观察

用透射电镜观察了生后至8个月龄大鼠肝(贝宁)脂细胞的超微结构并讨论了其功能。新生鼠的(贝宁)脂细胞呈幼稚状态,核仁不明显;生后1—2个月的大鼠(贝宁)脂细胞核切迹深而多;成鼠(贝宁)脂细胞核质比率减小,核仁明显,粗面内质网发达。(贝宁)脂细胞含较多脂滴和致密小体,在脂滴和致密小体周围有糖元颗粒包绕,这支持了糖元可由吞饮摄取的葡萄糖合成的观点;并认为碳水化合物参与脂肪的合成。在狄氏隙内有大量胶原原纤维,提出(贝宁)脂细胞有合成胶原原纤维的作用。     

Inhibitory effects of chlorpheniramine and astemizolum on contraction of isolated rat and rabbit uteri induced by oxytocin and PGF2α

Summary The contraction of isolated rat and rabbit uteri induced by oxytocin and PGF_2α was markedly inhibited by chlorpheniramine (Chl) and astemizolum (Ast), both of which also decreased the resting tension of uteri, and their spontaneous contraction. The inhibitory effects of both drugs were dose-dependent. At high concentrations, Chl 7.4× 10^-4 mol/L and Ast 10^-4 mol/L could counteract the contraction of the uteri induced by Oxy and PGF_2α, and their spontaneous contraction as well. They decreased the resting tension to the lower level. The mechanism of their non-special relaxed action on uteri could not be completely explained only by their H₁-receptor blocking action. Whether they act by blocking calcium channel or by inhibiting calmodulin (CaM) remains to be further explored.     

二氧化硫衍生物舒张大鼠离体主动脉血管环的效应及其机制研究

目的:探讨二氧化硫(sulfur dioxide, SO2)及其衍生物的舒张血管作用及其机制.方法:离体大鼠主动脉环灌流,应用去甲肾上腺素(noradrenaline, NE)预收缩主动脉环后,观察其对SO2供体--亚硫酸钠/亚硫酸氢钠混合液(Na2SO3/NaHSO3, 3:1物质的量比)的舒张反应;观察应用KATP通道阻断剂格列本脲和钙通道阻断剂尼卡地平对Na2SO3/NaHSO3血管效应的影响;观察应用内源性SO2生成酶抑制剂天冬氨酸异羟肟酸(hydroxamate,HDX)和Na2SO3/NaHSO3预孵育血管组织后NE缩血管效应的变化.结果:大鼠离体主动脉环对Na2SO3/NaHSO3呈浓度(0～12 mmol/L)依赖性的舒张反应,IC50值为(7.28±0.12) mmol/L,最大舒张率(Emax)为78.79%±3.24%.格列本脲(1×10^-6 mol/L)抑制低浓度Na2SO3/NaHSO3(≤4 mmol/L)的舒血管效应,而对高浓度(＞6 mmol/L)的舒血管效应无明显影响.经尼卡地平(1×10^-9 mol/L)预孵育的血管环对NE的收缩反应明显减弱,Na2SO3/NaHSO3则不能舒张该血管.反之,预先用HDX(1×10^-4 mol/L)孵育阻断内源性SO2生成后,血管环对NE的收缩反应增强[EC50从(6.48±0.84)×10^-7 mol/L降至(3.97±1.63)×10^-7 mol/L,P＜0.01];而用Na2SO3/NaHSO3预先孵育的血管对NE的收缩反应曲线右移[EC50从(6.48±0.84)×10^-7 mol/L升至(4.93±0.81)×10^-5 mol/L,P＜0.01].结论:SO2具有明显的舒张血管平滑肌作用,其作用机制与钙离子通道及KATP通道有关,推测机体内源性SO2具有血管功能调节意义.     

Effects and its possible mechanism of Radix Saposhnikoviae on rat colonic smooth muscle in vitro

Objective: To determine the effect of different concentrations of Radix Saposhnikoviae (RS) on the contraction of smooth muscle strips and the Ca2+ mobilization of cultured smooth muscle cells of rat colon and its possible mechanism of action. Methods: Strips of rat colon longitudinal muscle were prepared and smooth muscle cells from rat colon were isolated and cultured. In the experiments, in vitro muscle strips were suspended in an organ bath and the contraction of the strips was recorded. In the cell-experiments, intracellular Ca2+ was assessed using fluorescent intensity (FI) of smooth muscle cells loaded with Fluo-4/AM, measured with a laser scanning confocal microscope and related software. Results: In the in vitro experiment, RS (0.02, 0.2, 2, 20 g/L) inhibited contraction of muscle strips in a concentration-dependent manner, and this inhibition was significant for the three higher RS concentrations (P < 0.01) for both Peak (the maximal contraction amplitude) and Area (the area under curves). Similarly, RS inhibited Ach-induced contraction. In these experiments the inhibition of the Peak values in the RS 2 and 20 g/L groups was significant (P < 0.01), as was the inhibition of the Area values in all RS groups (P < 0.05). Naloxone and propranolol did not significantly affect the inhibitory effect of RS on smooth muscle contractility, while phentolamine significantly reduced the inhibitory effect (P < 0.01). In experiments using primary smooth muscle cell cultures in Ca2+ - containing buffer, the post-treatment fluorescence of cells in the RS 0.2, 2 and 20 g/L groups differed significantly from pre-treatment values (P < 0.05), and the percent inhibition of fluorescence in the RS 2 g/L and 20 g/L groups was significant (P < 0.01). However, in Ca2+-free buffer, FS had no significant effect on cell fluorescence. Conclusion: RS inhibited both the spontaneous and Ach-stimulated contraction of rat colonic smooth muscle strips. This RS effect appeared to involve α-adrenoceptors, but not β-adrenoceptors or opioid receptors. In cultured primary smooth muscle cells, RS reduced the mobilization of Ca2+ from extracellular sources, but may had no effect on the release of Ca2+ from sarcoplasmic reticulum and endoplasmic reticulum.     

高糖经内质网应激途径诱导胰岛内皮细胞凋亡

目的:探讨高糖诱导胰岛微血管内皮细胞株MS-1凋亡及其可能机制?方法:体外培养MS-1细胞,用含有不同浓度葡萄糖(5.6?25.0和33.6 mmol/L)的培养液分组培养12?24 h?流式细胞术分析各组细胞凋亡率变化;四甲基偶氮唑盐(MTT)法检测各组细胞增殖率变化;RT-PCR分析GRP78?CHOP?caspase-3?caspase-12 mRNA表达变化情况;Western blot分析GRP78?CHOP蛋白表达变化情况?结果:与5.6 mmol/L组相比,培养12 h及24 h后,25.0和33.6 mmol/L组MS-1细胞凋亡率显著增加(P < 0.05);而细胞增殖率显著下降(P < 0.05),且呈浓度和时间依赖性?进一步研究表明,在高糖刺激12 h及24 h后,caspase-3?caspase-12 mRNA表达显著上调(P < 0.05),CHOP mRNA和蛋白表达水平均显著上调(P < 0.05);而GRP78 mRNA和蛋白表达水平在刺激12 h后显著上调,24 h后却显著下降(P < 0.05)?结论:高糖可以促进胰岛内皮细胞凋亡增加,并呈浓度和时间依赖性升高,其机制可能与启动胰岛内皮细胞内质网应激有关?     

高磷诱导大鼠血管平滑肌细胞钙化及其电镜表现

目的观察高磷条件体外培养的原代大鼠血管平滑肌细胞(vascular smooth muscle cells,VSMCs)是否发生钙含量的变化及其电镜下的微观表现。方法应用大鼠原代血管平滑肌细胞进行体外实验,将培养细胞分为对照组和高磷组。在培养的10d内观察细胞的形态学变化,并应用原子分光光度计检测细胞钙含量,应用电镜观察细胞的微观变化。结果对照组细胞VSMCs钙含量在10d内无明显增加,高磷组VSMCs钙含量在培养第8天后明显增加,与换液当天比较差异有统计学意义(P<0.01)。2组细胞电镜下第10天VSMCs细胞外基质中均可见大量胶原纤维,高磷组还可见高密度电子致密物呈颗粒状沿胶原纤维沉积。结论高磷条件可诱导体外培养的大鼠VSMCs细胞钙含量增加。电镜下高密度电子致密物呈颗粒状并沿胶原纤维沉积。     

Effect of acetylcholine on membrane potential in toad dorsal root ganglion neurons and its underlying ionic basis

Intracellular recordings were made to investigate the responses of membrane potential to acetylcholine (ACh) on neurons in isolated toad dorsal root ganglion (DRG). In the 73 neurons examined, 67 were of type A, and the remaining 6 of type C cell. The resting membrane potential of these two types of cells was −67.5±1.3 mV (× ±SE). During the application of ACh (4 × 10^-4–6 × 10^-4 mol/L), the changes in membrane potential were as follows: 1) hyperpolarization, with amplitude of 9.1±3.0 mV (X ± SE; n = 23); 2) depolarization, with amplitude of 12.9 ±2.2 mV (X ±SE; n = 20); 3) biphasic response, i.e., hyperpolarization with amplitude of 8.0±2.4 mV (X±SE) followed by depolarization with amplitude of 10.9±2.1 mV (X±SE) (n=24); no effect (n=6). The hyperpolarization induced by ACh was blocked by superfusion with atropine (1.3 × 10^-5 mol/L; n = 23), while ACh depolarization was blocked by the mixture of d-tubocurarine (1.4 × 10^-5 mol/L) and hexamethonium (1.4 ×10^-5 mol/L) (n = 18). When ACh caused hyperpolarization, the membrane conductance wascin reased by 13.8% and the reversal potential was about -96 mV (n=3). TEA (20 mmol/L) superfusion enhanced ACh depolarization amplitude by 48.2 ±3.2 % (× ± SE;n = 6), and depressed ACh hyperpolarization amplitude by 79.4 ±4.3 % (× ± SE; n= 8).MnCl₂ (4 mmol/l) superfusion decreased the amplitudes of ACh depolarization and hyperpolarization by 54.2 ±7.2 % (X ±SE; n= 5) and by 69.2±6.4 % (X±SE; n = 14) respectively. These results suggest that the depolarization and hyperpolarization induced by ACh are mediated by nicotinic and muscarinic receptors on the soma of toad DRG neurons separately, and it seems that ACh hyperpolarization involves activation of calcium-activated potassium conductance.     

硫酸脱氢表雄酮对MIN6细胞葡萄糖刺激的胰岛素分泌的影响

目的探讨硫酸脱氢表雄酮(DHEAS)对胰岛β细胞株MIN6葡萄糖刺激的胰岛素分泌的影响。方法以葡萄糖刺激浓度为2.8mmol/L和16.7mmol/L的对数生长期MIN6细胞作为实验对象,分别以1、5、10μmol/LDHEAS干预10min和24h(不同浓度DHEAS组),以5μmol/LDHEAS或与10μmol/L糖皮质激素受体阻断剂RU486联合干预24h(DHEAS和DHEAS+RU486组),设立空白对照组。ELISA法测定细胞培养上清液中胰岛素分泌量,Real-TimePCR检测细胞胰岛素mRNA表达。结果干预后10min和24h时点,5μmol/L和10μmol/LDHEAS组MIN6细胞培养上清液中的胰岛素分泌量均显著高于空白对照组(P<0.05);干预后24h时点,DHEAS+RU486组与DHEAS组MIN6细胞培养上清液中胰岛素分泌量比较差异无统计学意义(P>0.05),但均显著高于空白对照组(P<0.05)。干预后24h时点,5μmol/L和10μmol/LDHEAS组MIN6细胞胰岛素mRNA表达均显著高于空白对照组(P<0.05)。结论 DHEAS对MIN6细胞葡萄糖刺激的胰岛素...     

血管紧张素II对肾小管上皮细胞凋亡的影响及冬虫夏草提取液对其的干预作用

目的:研究冬虫夏草提取液(Cordyceps sinensis, C. sinensis)对血管紧张素II( AngII )诱导的肾小管上皮细胞凋亡的保护机制。方法:C. sinensis(0, 5, 10, 20, 40 mg/L)与10^-8 mol/L AngII共同孵育NRK-52E 24, 48, 72 h, 了解C. sinensis对细胞增殖的影响, 确定最佳干预浓度; 取AngII(0, 10^-12, 10^-10, 10^-8, 10^-6 mol/L)与NRK-52E共同培育24 h及10^-8 mol/L AngII与NRK-52E共同培养24, 48, 72 h, 观察AngII对NRK-52E凋亡的影响, 确定AngII最佳干预浓度、时间后设立对照组, AngII(10^-8 mol/L)组, AngII(10^-8 mol/L)+ C. sinensis(40 mg/L)组, AngII(10^-8 mol/L)+ 福辛普利(10^-5 mol/L)组和AngII(10^-8 mol/L)+C. sinensis(40 mg/L)+ 福辛普利(10^-5 mol/L)组, 培养24 h后进行实验。利用MTT法检测不同浓度(0,5,10,20,40 mg/L)C. sinensis对NRK-52E作用24,48,72 h后的增殖情况,流式细胞仪Annexin V/PI双染检测凋亡率,比色法测定caspase-3酶活性。结果:C. sinensis(10~40 mg/L)在一定范围内可呈浓度依赖地促进培养的肾小管上皮细胞增殖(P<0.05); AngII呈浓度(10^-10~10^-6 mol/L)和时间(12~24 h)依赖地诱导NRK-52E凋亡率增加, caspase-3酶活性增加(P<0.05)。C. sinensis(10~40 mg/L)能部分地抑制AngII诱导的NRK-52E凋亡和降低caspase-3酶活性(P<0.01), 与福辛普利对细胞凋亡的抑制无明显区别, 二者联合用药效果与单用效果差异无统计学意义(P>0.05)。结论:C. sinensis对AngII诱导的肾小管上皮细胞凋亡具有一定抑制作用, 可能与抑制NRK-52E中caspase-3的激活有关。     

组织胺对肠粘膜屏障功能的调节作用研究

The model of CaCo2 cell monolayer system has been generally accepted as a standard method for in vitro study of the relationship between the epithelium and microbial invasion. In this study, we set up the model with a simplified method and used it to observe if histamine, which is an important inflammatory factor in gastroenteric tract, can offer some barrier protection from E. coli invasion. After two weeks' routine incubation, the established CaCo2 cells monolayer systems were co-cultured with 1 x 10(-8) mmol/L, 1 x 10(-7) mmol/L and 1 x 10(-6) mmol/L histamine DMEM for 2 hours, and with frank DMEM as control for the same length of the time. For observation on time course effect of histamine, the co-cultures with 1 x 10(-6) mmol/L histamine DMEM were kept up for 0.5, 1.0, 2.0, 4.0 and 18.0 hours. Bacterial invasion ws assessed by quantitating the number of E. coli within the coultured epithelial cells. The results showed that histamine significantly inhibited the invasion of E. coli to epithelial cells(P < 0.05). The colony counts in co-cultures with 1 x 10(-7) mmol/L and 1 x 10(-8) mmol/L histamine DMEM were 52.5, 30.3 and 91.3 respectively, compared with 563 in control group. In the study of the time course effect of histamine, the colony conuts of co-culture with histamine DMEM for 0.5, 1.0, 2.0, 4.0 and 18.0 hours were 135.5, 64.0, 29.5, 36.0 and 22.5 respectively. It was concluded that histamine can enhance the protective barrier function of intestinal epithelium against E. coli invasion.     

阿司匹林诱导内皮细胞铁蛋白表达与抗氧化损伤作用的研究

目的探讨阿司匹林(aspirin,As)在抗H2O2损伤内皮细胞过程中对铁蛋白(ferritin,Fn)表达的作用.方法体外培养人血管内皮细胞,酶联免疫吸附法(ELISA)测定As在不同浓度(0.1～3mmol/L)、不同处理时间(4～24h)对细胞Fn表达的影响,以及消炎痛和水杨酸钠对Fn表达的作用;并观察经As预处理后的细胞加入H2O2(0.5mmol/L)后乳酸脱氢酶(lactatedehydrogenase,LDH)释放率、丙二醛(malondialdehyde,MDA)和细胞存活率的改变.用单因素方差分析对上述指标进行统计学处理.结果小剂量As(0.1mmol/L)即可明显诱导内皮细胞Fn表达(5.8ng/106细胞数±0.3ng/106细胞数),与正常对照组比较P<0.05;且As与Fn之间表现出剂量和时间依赖关系,0.5mmol/L、1mmol/L、2mmol/L、3mmol/L组Fn浓度分别为(6.4±0.4)ng/106细胞数、(7.0±0.7)ng/106细胞数、(7.4±0.4)ng/106细胞数和(7.7±0.5)ng/106细胞数;4h组Fn尚未明显增加(5.8ng/106细胞数±1.0ng/106细胞数,P>0.05),但8h组Fn浓度(6.5±1.0)ng/106细胞数与正常对照组比较P<0.05,24h组Fn浓度达最大值(7.8±0.8)ng/106细胞数.As诱导Fn表达后能明显增强细胞抗氧化的能力,0.1mmol/L的As可减少细胞50%的LDH释放率,保护74.4%的细胞避免H2O2的损伤,同时使氧自由基指标MDA明显下降.并随As剂量的增大保护作用逐渐增强,其结果与未饱和铁蛋白组相似.而水杨酸钠及消炎痛则无诱导Fn表达的作用.结论As在较小剂量(0.1mmol/L)时即可明显诱导内皮细胞Fn的表达,增强其抗氧化损伤的能力;但其他非甾体类抗炎药不具有这个作用.     

外源性硫化氢通过ATP敏感钾通道诱导人子宫肌瘤细胞凋亡

目的探讨ATP敏感钾通道（K_ATP）在外源性硫化氢（H₂S）诱导人子宫肌瘤细胞凋亡中的作用。方法用硫氢化钠(NaHS)作为H₂S的供体处理原代培养的人子宫肌瘤细胞,K_ATP通道抑制剂格列本脲(Gly)预处理细胞。实验分为对照组、不同浓度NaHS组、ATP敏感性钾通道（K_ATP）抑制剂组和Gly + NaHS组。用流式细胞术检测细胞凋亡率,采用实时定量PCR和Western blot分别检测p53和bcl-2 mRNA和蛋白的表达。结果与对照组比较,NaHS（10^-4 和10^-3 mol/L）处理人子宫肌瘤细胞24和48 h后以剂量和时间依赖的方式增加了细胞的凋亡率（均P<0.05）。与对照组比较,NaHS（10^-4 mol/L）处理人子宫肌瘤细胞24 h后p53的表达显著增加（P＜0.05）,而bcl-2表达显著降低（P＜0.05）。Gly没有影响人子宫肌瘤细胞的凋亡率、p53和bcl-2的表达,但部分地拮抗NaHS的致凋亡作用以及NaHS诱导的子宫肌瘤细胞中p53表达的上调和bcl-2表达的下调（均P<0.05）。结论外源性H₂S诱导子宫肌瘤细胞凋亡,其机制与H₂S开放K_ATP通道有关。     

Suppressive effects of genomic imprinted gene PEG10 on hydrogen peroxide-induced apoptosis in L0<Subscript>2</Subscript> cells

The effects of PEG10 on hydrogen peroxide （H2O2）-induced apoptosis in human normal liver cell line L02 were investigated. The PEG10 gene was transfected into L02 cells by lipofectamine, the positive clone was screened by G418 and defined as L02/PEG10, while the cell transfected with empty expression vector （pEGFP-N1） was defined as L02/vector. L02/vector and parental L02 cells served as control. RT-PCR and Western blotting were employed to detect the expression of target genes. H2O2 （50–400 mmol/L） was administered to induce the apoptosis of L02 cells. Cells viability was measured by MTT and the morphological changes of apoptotic cells were determined by fluorescence microscopy using hoechst33342 nuclei staining. DNA fragmentation was observed by agarose gel electrophoresis. PEG10 mRNA and protein levels in L02/PEG10 cells were significantly increased as compared with those in the control cells. After treatment with 400 mmol/L H2O2 for 24 h, the cellular growth inhibition rate of L02/PEG10 cells was significantly lower （58.2%） than that of L02 （92.5%） and L02/vector （88%）. Distinct morphological changes characteristic of cell apoptosis such as karyopyknosis and conglomeration were not observed in L02/PEG10. Ladder-like DNA fragmentation in a dose-dependent manner was observed in both L02 and L02/vector cell lines, but not in L02/PEG10. PEG10 over-expression significantly inhibited cytotoxicity induced by H2O2 on human normal liver cell line L02 by antagonizing H2O2-induced apoptosis.     

Monoclonal anti-human T cell antibody and PAP-s conjugate —Preparation and selective cytotoxic properties on leukemic cell

Pokeweed antiviral protein (PAP-s) was prepared from seeds of Phytolacca americana. Monoclonal antibody against human pan-T lymphocyte Wu71 was linked to PAP-s by a disulfide bond. The results of SDS-PAGE, double immunodiffusion of active monoclonal antibody and PAP-s showed that the conjugate was highly cytotoxic to the human T-leukemic cell line CEM, but not to antigen-negative cell line SP₂/O. At a concentration of 10⁻⁹mol/L, 76.4 % of the target cells were killed, as compared with 10.1 % 10⁻⁹mol/L of free PAP-s. Treatment of the CEM cells with conjugate at 10⁻⁹mol/L reduced their rate of protein synthesis by 72.4 %, as determined with¹⁴C-leucine incorporation. The immunotoxin may be useful for the in-vitro eradication of leukemic cells in autologous bone marrow transplantation to leukemia patients.     

细胞外液低钾对小鼠心室肌细胞跨膜电位影响的定量分析

目的 研究不同程度细胞外低钾对心肌细胞跨膜电位的效应,阐明低钾对心肌细胞电生理特性的详细影响。方法 分离C57BL/6J小鼠的左心室乳头肌,采用标准玻璃微电极胞内记录技术记录心室肌细胞的跨膜电位,观察细胞外液K⁺浓度由正常的5.4mmol/L分别降为3、2、1和0mmol/L时,心室肌细胞跨膜电位各参数的变化。结果 低钾对心肌细胞的静息电位(RP)有双向影响:细胞外K⁺浓度降为3mmol/L时,RP显著增大(超极化)(P=0.000),而细胞外K⁺浓度降为2、1和0mmol/L时,细胞RP先显著增大后显著减小(P=0.000),这些结果异于传统观点。当细胞外液K⁺浓度为3mmol/L时,动作电位振幅(APA)和0期最大除极速度(Vmax)均增大,动作电位时程APD10、APD20、APD50和APD90均显著缩短(P<0.05),而动作电位复极到APD90后,复极速度减慢,即复极化有拖尾现象。当细胞外K⁺浓度为2mmol/L时,APA极度减小,Vmax明显减慢,AP呈侏儒型,而当细胞外K⁺浓度为1和0mmol/L时,细胞兴奋性丧失,电刺激不能诱发动作电位。此外,低钾可诱发早期后除极以及连串的触发活动,且后者两种形式,也表现为量-效和时-效的特点。结论 细胞外液低钾对心室肌细胞的RP、APA和Vmax有双重影响:中度低钾(K⁺ 3mmol/L)使这3个参数均增大;重度低钾(K⁺ 2mmol/L及以下)使这3个参数均减小;低钾使动作电位早期复极加快,晚期复极减慢;极度低钾(K⁺ 1mmol/L及以下)会导致心室肌细胞的兴奋性丧失;中、重度低钾可导致心室肌细胞发生早期后除极及连串的触发活动,后者相当于细胞水平的心动过速,但不是工作细胞获得了自律性。本研究在一定程度上澄清了以往对低钾的心肌电生理效应的模糊认识。