紫菀乙醇提取物对豚鼠离体气管平滑肌收缩功能的影响

本试验旨在研究紫菀乙醇提物对豚鼠离体气管平滑肌收缩功能的影响。采用离体气管恒温灌流的方法,将豚鼠气管制成气管螺旋条,通过 BL-420F 生物机能实验系统测定其张力的变化,观察紫菀乙醇提物对离体气管平滑肌静息状态下的舒张作用,以及在氯化乙酰胆碱(Ach)、磷酸组胺(His)、CaCl₂条件下和无钙下Ach诱导细胞内钙释放和外钙内流所致两种收缩条件下对离体气管平滑肌张力变化的影响。试验结果显示,低浓度紫菀乙醇提取物(0.002~0.008 g/mL)对静息状态下豚鼠离体气管平滑肌具有一定的收缩作用,高浓度(0.008~0.196 g/mL)时具有舒张作用;低、中、高3个剂量组均可抑制Ach、His和CaCl₂引起的气管平滑肌收缩强度,使各致痉剂的量效曲线非平行右移并降低最大效应。紫菀乙醇提取物对His的抑制作用强度最大,其次为Ach,最后是CaCl₂,且均呈剂量依赖性。以上结果表明紫菀乙醇提物具有舒张气管平滑肌的作用,其机制可能与抑制豚鼠气管平滑肌M受体、H₁受体和阻断Ca²⁺通道从而抑制细胞Ca²⁺内流有关。     

MCU在低氧诱导肉鸡心肌细胞线粒体损伤中的作用

旨在探讨线粒体钙单向转运体（mitochondrial calcium uniporter,MCU）介导线粒体Ca²⁺转运是否参与低氧诱导的肉鸡心肌细胞线粒体损伤。试验通过分离白羽肉鸡鸡胚原代心肌细胞,在低氧条件下（3% O₂,5% CO₂,92% N₂）培养24、48、72 h,同时使用RU360抑制MCU的表达并在低氧条件下处理72 h后,使用流式细胞术检测细胞内Ca²⁺浓度、线粒体Ca²⁺浓度、线粒体活性氧水平和线粒体膜电位,检测MCU及其调节因子的mRNA和蛋白表达。结果显示,通过差速贴壁方法培养的心肌细胞纯度可达90%以上;低氧培养24 h,诱导心肌细胞MCU、MICU1 mRNA表达增加（P<0.05）,胞浆和线粒体内Ca²⁺浓度显著上升（P<0.05）,线粒体膜电位增加（P<0.05）;低氧培养48 h,诱导MCUR1和MICU1 mRNA表达减少（P<0.05）,细胞Ca²⁺浓度上升（P<0.05）;低氧培养72 h,诱导MCU mRNA表达增加（P<0.01）,细胞和线粒体内Ca²⁺增加（P<0.01）,线粒体膜电位下降（P<0.01）,活性氧增加（P<0.01）。低氧处理72 h后,与低氧组相比,RU360预处理组细胞和线粒体内Ca²⁺减少,线粒体膜电位上升,活性氧生成减少（P<0.01）,MCU mRNA表达减少（P<0.01）。结果表明：低氧诱导MCU上调导致线粒体钙超载,促使线粒体功能降低并发生损伤,进而心肌细胞发生损伤;抑制MCU表达可以减轻低氧诱导的心肌细胞线粒体钙超载,保护线粒体。     

OCX-32基因表达载体构建及其对鸡子宫上皮细胞Ca2+浓度和脂质指标的影响

为探究卵黄蛋白原-32（ovocalyxin-32,OCX-32）基因过表达与鸡子宫上皮细胞Ca²⁺含量和脂质指标之间的关联性,本研究采用Ⅳ型胶原酶法分离培养长顺绿壳蛋鸡子宫上皮细胞,构建pcDNA3.1（+）+OCX-32+Flag表达载体并转染子宫上皮细胞,测定OCX-32基因过表达量、Ca²⁺浓度和脂质指标并进行相关性分析。结果显示,试验组的总胆固醇（TCHO）浓度显著高于对照组（P<0.05）,对照组的Ca²⁺浓度显著高于试验组（P<0.05）。OCX-32基因过表达量与Ca²⁺浓度呈显著负相关关系（P<0.05）,与甘油三酯（TG）浓度呈显著正相关关系（P<0.05）,与TCHO浓度呈极显著负相关关系（P<0.01）,TCHO浓度与TG浓度呈极显著负相关关系（P<0.01）,研究结果揭示了OCX-32基因过表达能够调控鸡子宫上皮细胞的Ca²⁺含量和脂质的合成转运,为深入阐释鸡子宫上皮细胞中脂质和钙分泌的调控机制提供试验基础和数据支持。     

松萝酸对兔离体肠平滑肌运动的舒张作用

以兔离体肠平滑肌为研究对象,考察松萝酸对兔离体肠平滑肌运动的影响,并探讨其作用机制。利用BL-420E生物机能实验系统,观察松萝酸对兔离体小肠平滑肌自主收缩的影响;采用工具药乙酰胆碱、组胺、氯化钡诱导小肠平滑肌收缩,并记录小肠平滑肌收缩频率和振幅。采用IP3受体阻断剂肝素、肌浆网ryanodine受体阻断剂钌红(RR)和一氧化氮合酶抑制剂左旋硝基精氨酸甲脂(L-NAME),探明松萝酸对兔离体肠平滑肌作用的机制。结果发现,松萝酸可抑制兔离体小肠平滑肌收缩,具有浓度依赖性,且对乙酰胆碱、组胺、氯化钡诱导的肠肌收缩具有显著的抑制作用。IP3受体阻断剂肝素可增强松萝酸舒张肠平滑肌运动的频率(P<0.01)和振幅(P<0.05),肌浆网ryanodine受体阻断剂钌红和一氧化氮合酶抑制剂左旋硝基精氨酸甲酯可显著增强松萝酸舒张兔离体肠平滑肌运动的频率,对振幅影响不显著。说明松萝酸可显著抑制兔离体小肠平滑肌收缩的频率和振幅,其机制可能与增加一氧化氮浓度,抑制IP3受体和肌浆网ryanodine受体介导的内钙释放有关。     

青木香提取物对兔离体肠段运动的影响

为了探讨青木香对腹痛腹泻的治疗作用机制,采用离体肠肌试验,应用BL-420E生物机能实验系统,比较青木香提取物对正常肠段及工具药酚妥拉明、氯化钡作用下离体肠平滑肌活动的影响,记录肠段的收缩频率和振幅。结果显示,青木香提取物可浓度依赖性地抑制兔小肠段自发性收缩的频率和振幅;2．5mg／mL青木香提取物对由氯化钡引起的离体肠段的兴奋性收缩有明显的拮抗作用,而对酚妥拉明阻断肾上腺素能受体后,仍能显著降低离体肠段收缩的振幅。提示青木香提取物对小肠蠕动有一定的抑制作用。     

PLCγ1基因对绵羊早期胚胎体外发育的影响

旨在探究磷脂酶Cγ1（phospholipase Cgamma 1,PLCγ1）基因对绵羊早期胚胎体外发育的影响,为揭示PLCγ1基因在早期胚胎发育过程中的作用机制奠定基础。本研究选取包裹3层以上颗粒细胞的绵羊卵母细胞为试验材料,体外培养成熟后将其分为两组,利用显微注射技术将PLCγ1基因导入MⅡ期（减数第二次分裂中期）卵母细胞（n=127）中为试验组,以显微注射空载体pcDNA3.1-EGFP作为对照组（n=120）,经48 h后统计其卵裂率;并利用Ca²⁺探针Rhod 2-AM监测MⅡ卵母细胞和显微注射后16、48、72、96、120 h时各时期卵母细胞以及早期胚胎内Ca²⁺波动。结果显示,PLCγ1基因显微注射后卵母细胞被激活,卵裂率为（（19.67±0.2）%,P<0.01）,桑椹胚发育率为（（9.00±0.17）%,P<0.05）;PLCγ1基因注射后,卵母细胞和早期胚胎不同发育时期有Ca²⁺浓度变化,可观察发现Ca²⁺主要分布在胞质中且注射48 h时可达到峰值（P<0.01）。结果表明,PLCγ1基因可以引起绵羊卵母细胞发育过程中发生Ca²⁺振荡,启动其卵裂,并促使早期胚胎的继续发育。     

槐花对兔离体小肠运动性能的影响

为了观察不同浓度槐花水煎液对家兔离体小肠运动性能的影响,笔者采用离体器官实验法,将兔离体肠管放置于恒温(38.0±0.5℃)平滑肌槽中,依次加入0.2 g/L、0.4 g/L、0.6 g/L、0.8 g/L浓度的槐花水煎液,记录给药前后十二指肠、空肠平滑肌运动性能的变化。结果表明,与用药前比较,水煎液浓度为0.2 g/L时,离体十二指肠的收缩张力明显下降(P0.01),收缩振幅、收缩频率无显著变化(P0.05);离体空肠的收缩张力、收缩振幅明显下降(P0.01),收缩频率不显著(P0.05)。随着浓度增加到0.4 g/L、0.6 g/L、0.8 g/L时,离体十二指肠和空肠的收缩张力、收缩振幅、收缩频率极显著下降(P0.01)。说明槐花水煎液能显著降低离体十二指肠和空肠的收缩张力、收缩振幅和收缩频率,且呈现量效关系。     

小裂芽梅衣对兔离体肠平滑肌的舒张作用及其机制

采用兔离体肠肌试验,应用BL-420E生物机能实验系统比较小裂芽梅衣对正常肠肌及乙酰胆碱、氯化钡、组胺所致肠平滑肌运动的影响。应用IP3受体阻断剂肝素、肌浆网ryanodine受体阻断剂钌红、一氧化氮合酶抑制剂左旋硝基精氨酸甲酯,探讨小裂芽梅衣对兔离体肠平滑肌的作用机制。结果表明,小裂芽梅衣可浓度依赖性抑制兔小肠平滑肌的收缩,且对乙酰胆碱、氯化钡、组胺诱导的肠平滑肌收缩有明显抑制作用(P0.05)。肝素能显著增强小裂芽梅衣对兔小肠平滑肌运动的抑制作用(P0.05),左旋硝基精氨酸甲酯和钌红能部分增强小裂芽梅衣对兔小肠平滑肌运动的抑制作用(P0.01)。小裂芽梅衣乙醇提取物对兔小肠平滑肌收缩的抑制作用机制可能与增加NO浓度、抑制IP3受体和肌浆网ryanodine受体介导的内钙释放有关。     

菊花水提物对家兔离体小肠平滑肌收缩性的影响

研究菊花水提物在有乙酰胆碱、组胺和阿托品的条件下对家兔离体小肠平滑肌收缩性的影响。以家兔离体小肠平滑肌条为模型,考察在正常克氏液条件下分别加入乙酰胆碱、组胺和阿托品后家兔平滑肌条的收缩状况及在该条件下分别加入0.5g/mL菊花水提液物后家兔小肠平滑肌条的收缩状况。结果:菊花水提物对家兔离体小肠平滑肌的收缩具有一定的促进作用,菊花水提物能协同促进乙酰胆碱对家兔离体小肠平滑肌的收缩作用,拮抗阿托品的作用,对组胺的小肠收缩振幅影响不显著。菊花水提物对家兔小肠平滑肌收缩活动具有明显的兴奋作用,这种兴奋作用可由M受体介导。     

叶下珠生物碱对兔离体肠肌的舒张作用

目的:以兔离体小肠平滑肌为研究对象,研究叶下珠生物碱对兔离体肠肌收缩的影响,并探讨其作用机制。方法:利用BL-420E生物机能系统,观察叶下珠生物碱对兔离体肠肌自发性收缩和工具药乙酰胆碱、阿托品、新斯的明、组胺作用下离体肠肌活动的影响,观察并记录收缩频率和振幅。结果:叶下珠生物生物碱可显著抑制兔离体肠肌自发性收缩的频率和振幅(P0.05);可显著或极显著抑制由乙酰胆碱、新斯的明和组胺引起的离体肠肌挛缩,加强阿托品对离体肠肌的舒张作用。结论:叶下珠抗腹泻作用可能与抑制小肠蠕动有关,叶下珠生物碱抑制肠肌运动可能与促进肠上皮细胞释放钙离子、阻断H1受体促进胞外钙离子内流相关。     

Effects of Alcohol Extract of Aster tataricus L.f.on the Contraction of Guinea Pig Tracheal Smooth Muscle in vitro

The aim of the experiment was to study the effect of alcohol extract of Aster tataricus L.f.on the contraction of isolated guinea pig tracheal smooth muscle.The guinea pig tracheal smooth muscle was prepared by isolated tracheal thermostatic perfusion method.The effects of Aster tataricus L.f.on tracheal smooth muscle under the resting state and contraction stimulated by acetylcholine (Ach),histamine (His),CaCl₂,Ca²⁺ release and influx in cells without calcium were measured by BL-420F biological function experiment systems.The results showed that Aster tataricus L.f.could induce the contraction of isolated trachea at resting state at a low concentrations of 0.002 to 0.008 g/mL and exerted a relaxation on the isolated trachea when at the higher concentrations of 0.008 to 0.196 g/mL.All of the three concentrations of Aster tataricus L.f.could inhibit the tension stimulated by Ach,His and CaCl₂ which were antispasmodic agents,led to the non-parallelled rightward moving of cumulative concentration-response curve and the decline of maximal responses.The suppressive effect on His was the strongest,followed by Ach and CaCl₂,and the suppression could counteract the contraction induced by extracellular Ca²⁺influx significantly,and they were all expressed concentration-dependent manner.These results indicated the alcohol extract of Aster tataricus L.f.could relax tracheal smooth muscle by acting on M-receptor and histamine receptor and blocking Ca²⁺ passage,thus inhibiting extracellular Ca²⁺influx.     

Effects of bethanechol on canine urinary bladder smooth muscle function

We studied whether the effects of bethanechol are mediated via a muscarinic receptor, the role of extracellular calcium on bladder contraction, and down-regulation of bladder contraction by bethanechol after activation with potassium chloride (KCl) and acetylcholine (Ach). Smooth muscle strips of normal urinary bladder were studied with standard methods to measure isometric force. Bethanechol caused a dose-dependent increase in bladder contraction. The potency of bethanechol is higher than Ach, as shown by higher peak active isometric stress (P(max)) and lower half-maximal contraction (ED(50)) (P< 0.01). The contractile responses to bethanechol were diminished in the presence of atropine, nifedipine and in calcium-free medium as shown by P(max) decreased by 58%, 87% and 65% and ED(50) increased by 314-, 24- and 16-fold, respectively. When bladder strips were stimulated with KCl and Ach, pre-treatment with bethanechol reduced the responses to KCl by 116-242% (P<0.05), while the contractile responses to Ach were unaltered. Thus, bethanechol induces bladder contraction via muscarinic receptor activation while both intracellular and extracellular calcium play a crucial role on bladder smooth muscle contraction. The mechanisms of down-regulation by bethanechol may be related to interference with calcium influx into the smooth muscle cells, rather than the desensitisation of muscarinic receptors or post-receptor steps of signal transduction following bethanechol binding to the receptor.     

玻璃化冷冻对哺乳动物卵母细胞Ca2+的影响机制

卵母细胞冷冻保存是胚胎生物技术(如体外受精、胞浆内单精子注射、体细胞克隆)的重要组成部分,对优良种畜和濒危动物种质资源保存,加速家畜品种改良进程都具有重要意义。与常规冷冻法相比,玻璃化冷冻具有操作简单、降温速率快、耗时短、冷冻效率高等优点,被越来越广泛地应用于家畜卵母细胞的冷冻保存。然而,与新鲜卵母细胞相比,玻璃化冷冻卵母细胞的受精率及发育能力仍不理想,这严重影响了玻璃化冷冻卵母细胞的应用潜力。玻璃化冷冻会引起卵母细胞Ca²⁺浓度升高及钙振荡模式异常,导致其透明带硬化、受精信号紊乱等问题。综合前人研究进展,作者分析了玻璃化冷冻对胞内Ca²⁺浓度、钙振荡模式的影响和作用机制,并指出胞外Ca²⁺内流和胞内钙库Ca²⁺释放是导致冷冻卵母细胞胞内Ca²⁺浓度升高的主要原因,1,4,5-三磷酸肌醇(IP3)Ⅰ型受体分布异常和线粒体损伤可能是导致冷冻卵母细胞钙振荡模式异常的重要原因,以期为正向调控冷冻卵母细胞Ca²⁺浓度及钙振荡模式提供技术参考,从而进一步提高玻璃化冷冻卵母细胞的受精和后续的发育能力。     

番石榴叶止泻机制的初步研究

用80%酒精提取番石榴叶中的有效成分, 观察提取物对细菌和离体小肠的作用。结果表明: 提取物能抑制大肠杆菌等6种病原菌生长; 降低家兔十二指肠、空肠、回肠等三肠段收缩张力。     

Characterization of muscarinic receptor subtype mediating contraction and relaxation in equine coronary artery in vitro

In coronary arterial rings isolated from horses, 10-^-8-10^-6 mol/l acetylcholine (ACh) induced concentration-dependent contractions which were potentiated by the removal of endothelium and by pretreatment with I,-nitro-arginine (LNAG) or methylene blue (MB). Relatively lower concentrations of Ach 10-¹⁴-10-⁸ mol/l) induced relaxation when the coronary rings were contracted by phenylephrine (PE). ACh-induced contractions in the coronary rings without endothelium were competitively inhibited by each muscarinic subtype selective antagonist in the following order of potency: 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) > pirenzepine ≥ parafluoro-hexahydrosiladiphenidol (pFHHSiD) > methoctramine. ACh-induced relaxation in the rings with endothelium was inhibited by LNAG or MB, and by each selective antagonist in the following order of potency: 4-DAMP < pFHHSiD ≥ pirenzepine ≥ methoctramine. These results suggest that the ACh-induced contraction and relaxation in equine coronary arteries are mediated mainly by an M₃-receptor located on the smooth muscle cells and endothelial cells, respectively, and that the stimulation of the M₃-receptor on the endothelial cells liberates nitric oxide.     

Effects of muscarinic receptor antagonists on acetylcholine-induced contractions of jejunal smooth muscle in horses

This study investigated the effects of a muscarinic type 1 (M(1)), 2 (M(2)), and 3 (M(3)) antagonists (4-DAMP, pirenzepine, and methoctramine, respectively) on acetylcholine (Ach)-induced contractions of longitudinal jejunal muscle strips of horses. Strips were irrigated with Krebs-Henseleit solution gassed with 95% O(2) and 5% CO(2), and the developed tension in response to Ach was recorded before and after incubation with increasing concentrations of 4-DAMP (10(-8)-10(-6) M), pirenzepine (10(-6)-10(-4) M), and methoctramine (10(-5)-10(-3) M). When competitive antagonism was characterized, the affinity constant (pA(2)) was calculated by Schild plots. A parallel rightward shift in the concentration-response curves was observed after 4-DAMP and pirenzepine. Methoctramine presented a dual effect on the concentration-response curves: lower concentrations induced a parallel rightward shift without altering the maximum intensity of contraction (E(max)), while the highest concentration increased slope of the concentration-response curve and increased E(max). The pA(2) for 4-DAMP and pirenzepine was 9.18 and 7.13, respectively. Acetylcholine-induced contractions of longitudinal jejunal smooth muscle are mediated mainly via M(3) receptors. The complex role of M(2) receptors in jejunal smooth muscle contractions was evident because methoctramine potentiated the contractile response to higher doses of Ach.     

谷氨酸棒状杆菌全细胞催化合成Levan果聚糖条件优化分析

试验旨在探究微生物合成Levan果聚糖的新途径并优化其合成条件。利用转入果聚糖蔗糖转移酶基因的重组谷氨酸棒状杆菌进行全细胞催化合成Levan果聚糖,并采用响应面法以Levan果聚糖产量为响应值优化催化合成条件。首先设计Plackett-Burman试验从7个相关因素（蔗糖浓度、菌体浓度、转化时间、转化pH、PBS中Ca²⁺和Mg²⁺浓度、Levan果聚糖提取时的乙醇终浓度）中筛选出影响Levan果聚糖产量的3个主要因素（蔗糖浓度、乙醇终浓度和Ca²⁺浓度）;再用筛选出的3个最重要因素进行最陡爬坡试验逼近最大响应值区域;最后运用BBD响应面法优化确定3个主要因素之间的交互作用和最佳合成条件。结果显示,初始试验中Levan果聚糖平均产量为0.069 g/mL。经过响应面试验优化对利用重组谷氨酸棒状杆菌进行全细胞催化合成Levan果聚糖产量影响最大的因素为蔗糖浓度,其次为提取时的乙醇终浓度,最后是转化液中的Ca²⁺浓度。当蔗糖浓度为52.71%、乙醇终浓度为85.31%、Ca²⁺浓度为0.04 g/L时,利用重组谷氨酸棒状杆菌进行全细胞催化合成的Levan果聚糖产量平均值为0.238 g/mL,与模型预测最大值0.233 g/mL相近,转化率约为44%,转化率较初始试验提高了约1.9倍,产量亦较初始试验提高了约3.4倍。研究结果为Levan果聚糖工业化生产提供了依据,并为其在动物医学和营养方面的研究奠定基础。     

Inhibition of spontaneous smooth muscle contractions in rat and rabbit intestine by blockers of the thromboxane A2 pathway

The effect of inhibitors of the thromboxane A2 pathway on spontaneous contractions of intestinal smooth muscle preparations was studied. The thromboxane A2 antagonists Bay u3405, SK and F 88046 and KW-3635 concentration-dependently inhibited both the amplitude and the frequency of spontaneous contractions of the longitudinal muscle from the rat proximal colon. A concentration-dependent inhibition of the myogenic contractions was also observed with the thromboxane A2 synthase inhibitor U-51605, and with the combined cyclooxygenase/lipoxygenase inhibitor nordihydroguaiaretic acid, whereas indomethacin, a pure cyclooxygenase inhibitor, was ineffective. None of these inhibitors affected the contractile response evoked by the cholinergic agonist carbachol, excluding non-specific actions on intestinal motility. A similar response was observed for the rabbit jejunum, which, in contrast to the rat colon, exhibits more regular, high-frequency spontaneous contractions, which were inhibited by Bay u3405, SK and F 88046 and KW-3635 in a concentration-dependent manner, whereas the response to carbachol remained unaffected. These results suggest a role for thromboxane A2 in the generation and/or facilitation of spontaneous smooth muscle contractions in the gut.