兰尼碱受体2参与重大心脏疾病发生发展机制的研究进展

心血管疾病是全球范围内首要的致死原因,严重威胁人类的生命健康。兰尼碱2受体(RyR2)是心肌细胞肌浆网上重要的钙释放通道,其调控机制复杂,一些相关研究仍存在争议。RyR2的基因突变、结构改变、功能障碍或与相关蛋白的相互作用异常均可造成RyR2受体的钙释放阈值降低,引起肌浆网钙异常释放,诱发或加重多种心脏疾病。以RyR2为靶点的药物对心脏疾病很可能具有治疗作用。一种针对RyR2靶点的新药已进入Ⅱ期临床试验,随着RyR2结构、功能及作用机制研究的进一步深入,必将为心血管疾病的早期诊断、鉴别、药物研发等提供更加准确可靠的信息。本文就RyR2参与重大心脏疾病发生与发展机制的研究进行综述。     

内皮素1对胞浆Ca~(2+)升高调控作用的研究进展

由21个氨基酸残基组成的内皮素1(ET-1)不仅是已知最强的缩血管活性肽,还对血管形成与重构、细胞增殖、细胞外基质合成和感觉神经活化等诸多生理活动具有调控作用。其生理效应多通过升高胞浆Ca2+继而促发连锁反应来实现。增强细胞外Ca2+内流和促进胞内Ca2+释放是ET-1升高胞浆Ca2+浓度的两条基本途径,同时,通过抑制胞内的Ca2+外排与重摄取及对细胞核等非典型Ca2+库内的Ca2+信号的调控同样可以达到升高胞浆Ca2+浓度的目的,本文主要就ET-1升高胞浆Ca2+的调控途径作一综述。     

Liguzinediol甲基替代衍生物的合成及其正性肌力活性研究

目的设计合成liguzinediol甲基替代衍生物,并分析其对正常大鼠离体心脏的正性肌力作用。方法分别以2,5-二甲基吡嗪和吡嗪为原料,通过与醛进行自由基取代得烷基酰化产物,经还原、氧化、BOEKELHEIDE重排等反应,硅胶柱层析分离纯化得到目标化合物;采用1H-NMR、ESI-MS对其结构进行了表征;采用Langendorff离体灌流装置观察化合物对大鼠离体心脏心功能的影响。结果合成了五个liguzinediol甲基替代衍生物,正性肌力活性实验结果显示,2,5-二羟甲基-3,6-二乙基吡嗪、2,5-二羟甲基-3,6-二丙基吡嗪、2,5-二羟甲基-3,6-二丁基吡嗪对正常大鼠离体心脏产生正性肌力作用;2,5-二羟甲基吡嗪、2,5-二羟甲基-3,6-二异丁基吡嗪对正常大鼠离体心脏无正性肌力作用。结论当liguzinediol的二个甲基被H和异丁基取代时,无正性肌力活性,而被乙基、丙基及丁基取代时表现出一定的正性肌力活性,且随着碳链的延长,正性肌力活性显著下降。     

正性肌力药物作用靶点的研究进展

正性肌力药物能影响心肌收缩力,是治疗心力衰竭的主要药物之一,用于治疗心力衰竭的药物疗效及用药患者终期生存率取决于作用靶点。与心肌收缩力有关的靶点包括:β肾上腺素受体、磷酸二酯酶、L-型钙通道、蛋白激酶A、蛋白激酶C、蛋白磷酸酶、Na+-Ca2+交换体、Na+-K+-ATP酶、肌浆网钙泵、受磷蛋白、兰尼碱受体、三磷酸肌醇受体、与钙增敏剂相关的心肌收缩蛋白等。文中对心肌收缩力相关的作用靶点进行综述,分析临床现今使用的和未来的正性肌力药物应具备的靶点特征。     

小檗碱的降糖作用及安全性评价

小檗碱是中药黄连的主要活性成分，具有降血糖的功能。作者综述了近年来小檗碱的降糖机制、临床疗效及安全性评价。     

MCI-154对心肌肥厚大鼠离体心脏的正性变力作用及其机制(英文)

目的　探讨钙增敏剂MCI 15 4〔6 [4 (4 吡啶氨基 )苯基 ] 4 ,5 二氢 3(2H)哒嗪酮〕对心肌肥厚心脏与对正常心脏的作用是否不同及有关的机制。方法　利用离体心脏灌流技术观察MCI 15 4对心肌肥厚大鼠心功能的影响 ;应用离子影像学分析系统同步测定心肌细胞钙浓度瞬变和细胞长度。结果　①MCI 15 410 0～ 4 0 0 μmol·L- 1范围内浓度依赖性地提高了心肌肥厚大鼠心功能的各项指标。在 4 0 0μmol·L- 1时 ,左室主动收缩压 (左室收缩峰压与左室舒张末压之差 )及左室压最大上升速率 (+dp/dtmax)与对照值相比显著增加 ,左室压最大下降速率(-dp/dtmax)有增高趋势 ,但无统计学意义 ;②MCI 15 410～ 10 0 μmol·L- 1在钙瞬变无明显改变情况下 ,呈浓度依赖性增加肥厚心肌细胞的缩短程度和钙敏感性 ;③MCI 15 4对肥厚心肌细胞钙瞬变的 5 0 %和90 %恢复时间影响不大。结论　在心肌肥厚大鼠心脏上 ,和在正常大鼠心脏上一样 ,MCI 15 4主要通过钙增敏作用发挥其正性变力作用。     

心肌α_1肾上腺素受体研究进展──正性肌力作用机制及受体亚型

心肌α_１肾上腺素受体研究进展──正性肌力作用机制及受体亚型杨黄恬，杨毓麟（南通医学院药理教研室，江苏２２６００１）自１９４８年Ａｈｌｑｕｉｓｔ将肾上腺素受体分为α和β受体直至１９６６年Ｗｅｎｚｅｌ等报道α受体亦参与心肌功能调节为止，β受体一直被认为...     

乌头碱与钾离子通道激动剂合用对离体大鼠心脏的正性肌力作用研究

目的　研究钾离子通道激动剂吡那地尔与乌头碱合用时乌头碱对心功能不全大鼠离体心脏的正性肌力作用。方法　本实验利用Langendorff离体心脏灌流装置 ,以左室收缩压 (LVSP)、左室压上升最大速率 ( +dp/dtmax)、左室压下降最大速率 ( -dp/dtmax)、左室舒张末压 (LVEDP)为指标 ,在①单纯使用乌头碱②乌头碱与吡那地尔 (KATP通道激动剂 )合用两种情况下 ,观察了乌头碱对腹主动脉狭窄法造成的心功能减退大鼠离体心脏的正性肌力作用。结果　实验结果表明①乌头碱对心功能不全大鼠离体心脏具有一定正性肌力作用 ,②乌头碱在激动钾通道的情况下 ,与单纯使用乌头碱相比 ,强心的有效浓度范围增大 ,强心效果明显增强 (P <0 0 5 )。结论　乌头碱对心功能减退的心脏具有强心作用 ,激动心肌钾通道可显著增强乌头碱的强心作用和有效浓度范围。     

中文关键词索引(2012年第26卷)

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吴茱萸次碱的心脏保护作用涉及辣椒素敏感的感觉神经

目的 观察吴茱萸次碱对大鼠心肌缺血再灌注损伤的保护作用是否与激活辣椒素敏感的感觉神经有关。方法 大鼠在实验前10 min静脉注射吴茱萸次碱,然后结扎冠状动脉左前降支60 min,再灌注3 h。测定心肌梗死面积、血清肌酸激酶(CK)活性和血浆降钙素基因相关肽(CGRP)浓度。结果 吴茱萸次碱(100、300μg·kg-1,iv)能显著缩小心肌梗死面积,降低血清CK水平,升高血浆CGRP浓度。这些作用被预先给予竞争性辣椒素受体拮抗药Capsazepine(38 mg·kg-1,s.c.)或选择性感觉神经耗竭剂辣椒素(50 mg·kg-1,s.c.)所完全取消。结论 吴茱萸次碱对大鼠心肌缺血再灌注损伤有保护作用,其保护作用与通过激动辣椒素受体而激活辣椒素敏感的感觉神经有关。     

Ryanodine receptors as pharmacological targets for heart disease

Calcium release from intracellular stores plays an important role in the regulation of muscle contraction and electrical signals that determine the heart rhythm. The ryanodine receptor （RyR） is the major calcium （Ca^2＋） release channel required for excitation-contraction coupling in the heart. Recent studies have demonstrated that RyR are macromolecular complexes comprising of 4 pore-forming channel subunits, each of which is associated with regulatory subunits. Clinical and experimental studies over the past 5 years have provided compelling evidence that intracellular Ca^2＋ release channels play a pivotal role in the development of cardiac arrhythmias and heart failure. Changes in the channel regulation and subunit composition are believed to cause diastolic calcium leakage from the sarcoplasmic reticulum, which could trigger arrhythmias and weaken cardiac contractility. Therefore, cardiac RyR have emerged as potential therapeutic targets for the treatment of heart disease. Consequently, there is a strong desire to identify and/or develop novel pharmacological agents that may target these Ca^2＋ signaling pathways. Pharmacological agents known to modulate RyR in the heart, and their potential application towards the treatment of heart disease are discussed in this review.     

The power of single channel recording and analysis: its application to ryanodine receptors in lipid bilayers

1. Since the inception of the patch-clamp technique, single-channel recording has made an enormous impact on our understanding of ion channel function and its role in membrane transport and cell physiology. 2. However, the impact of single-channel recording methods on our understanding of intracellular Ca2+ regulation by internal stores is not as broadly recognized. There are several possible reasons for this. 3. First, ion channels in the membranes of intracellular organelles are not directly accessible to patch pipettes, requiring other methods that are not as widely known as the patch-clamp techniques. 4. Second, bulk assays for channel activity have proved successful in advancing our knowledge of Ca2+ handling by intracellular stores. These assays include Ca2+ imaging, ryanodine binding assays and measurements of muscle tension and Ca2+ release and uptake by vesicles that have been isolated from internal stores. 5. The present review describes methods used for single- channel recording and analysis, as applied to the calcium release channels in striated muscle, and details some of the unique contributions that single-channel recording and analysis have made to our current understanding of the release of Ca2+ from the internal stores of muscle. 6. With this in mind, the review focuses on three aspects of channel function and shows how single-channel investigations have led to an improved understanding of physiological processes in muscle. 7. Finally, the review describes some of the latest improvements in membrane technology that will underpin future advances in single-channel recording.     

Proteins within the intracellular calcium store determine cardiac RyR channel activity and cardiac output

SUMMARY: The contractile function of the heart requires the release of Ca(2+) from intracellular Ca(2+) stores in the sarcoplasmic reticulum (SR) of cardiac muscle cells. The efficacy of Ca(2+) release depends on the amount of Ca(2+) loaded into the Ca(2+) store and the way in which this 'Ca(2+) load' influences the activity of the cardiac ryanodine receptor Ca(2+) release channel (RyR2). The effects of the Ca(2+) load on Ca(2+) release through RyR2 are facilitated by: (i) the sensitivity of RyR2 itself to luminal Ca(2+) concentrations; and (ii) interactions between the cardiac Ca(2+) -binding protein calsequestrin (CSQ) 2 and RyR2, transmitted through the 'anchoring' proteins junctin and/or triadin. Mutations in RyR2 are linked to catecholaminergic polymorphic ventricular tachycardia (CPVT) and sudden cardiac death. The tachycardia is associated with changes in the sensitivity of RyR2 to luminal Ca(2+) . Triadin-, junctin- or CSQ-null animals survive, but their longevity and ability to tolerate stress is compromised. These studies reveal the importance of the proteins in normal muscle function, but do not reveal the molecular nature of their functional interactions, which must be defined before changes in the proteins leading to CPVT and heart disease can be understood. Herein, we discuss known interactions between the RyR, triadin, junctin and CSQ with emphasis on the cardiac isoforms of the proteins. Where there is little known about the cardiac isoforms, we discuss evidence from skeletal isoforms.     

Ca2+ stores regulate ryanodine receptor Ca2+ release channels via luminal and cytosolic Ca2+ sites

1. In muscle, intracellular calcium concentration, hence skeletal muscle force and cardiac output, is regulated by uptake and release of calcium from the sarcoplasmic reticulum. The ryanodine receptor (RyR) forms the calcium release channel in the sarcoplasmic reticulum. 2. The free [Ca2+] in the sarcoplasmic reticulum regulates the excitability of this store by stimulating the Ca2+ release channels in its membrane. This process involves Ca2+-sensing mechanisms on both the luminal and cytoplasmic sides of the RyR. In the cardiac RyR, these have been shown to be a luminal Ca2+ activation site (L-site; 60 micromol/L affinity), a cytoplasmic activation site (A-site; 0.9 micromol/L affinity) and a cytoplasmic Ca2+ inactivation site (I2-site; 1.2 micromol/L affinity). 3. Cardiac RyR activation by luminal Ca2+ occurs by a multistep process dubbed 'luminal-triggered Ca2+ feed-through'. Binding of Ca2+ to the L-site initiates brief (1 msec) openings at a rate of up to 10/s. Once the pore is open, luminal Ca2+ has access to the A-site (producing up to 30-fold prolongation of openings) and to the I2-site (causing inactivation at high levels of Ca2+ feed-through). 4. The present paper reviews the evidence for the principal aspects of the 'luminal-triggered Ca2+ feed-through' model, the properties of the various Ca2+-dependent gating mechanisms and their likely role in controlling sarcoplasmic reticulum (SR) Ca2+ release in cardiac muscle. 5. The model makes the following important predictions: (i) there will be a close link between luminal and cytoplasmic regulation of RyRs and any cofactor that prolongs channel openings triggered by cytoplasmic Ca2+ will also promote RyR activation by luminal Ca2+; (ii) luminal Mg2+ (1 mmol/L) is essential for the control of SR excitability in cardiac muscle by luminal Ca2+; and (iii) the different RyR isoforms in skeletal and cardiac muscle will be controlled quite differently by the luminal milieu. For example, Mg2+ in the SR lumen (approximately 1 mmol/L) can strongly inhibit RyR2 by competing with Ca2+ for the L-site, whereas RyR1 is not affected by luminal Mg2+.     

右兰索拉唑缓释胶囊健康人体生物等效性及安全性评价

目的:评价右兰索拉唑缓释胶囊在健康受试者空腹、餐后状态下的人体生物等效性及安全性.方法:采取随机、开放、四周期、两交叉单次给药设计,空腹组及餐后组各有30例健康受试者入选,采用高效液相色谱,串联质谱法测定受试者每周期口服右兰索拉唑缓释胶囊受试制剂或参比制剂(Dexilant(R))后15个不同时间点的右兰索拉唑浓度.采...     

CAR-T治疗诱导细胞因子释放综合征的机制及临床前安全性评价

嵌合抗原受体T细胞（CAR-T）疗法极大地改变了癌症的治疗前景,细胞因子释放综合征（CRS）是病理性免疫系统激活的严重全身炎症反应,也是CAR-T治疗过程中最常见且致命的毒性反应。目前CRS机制尚未完全阐明,临床前研究的实验模型主要包括异种移植免疫缺陷小鼠模型、同源小鼠模型、转基因小鼠模型、人源化小鼠模型和非人灵长类动物模型。由于种属差异且缺乏体外细胞模型,临床前预测CAR-T细胞引起CRS的风险仍是亟待解决的问题。从CRS的相关指导原则及法规、机制及临床前安全性评价方法和模型3个方面对CAR-T诱发的CRS进行综述,以期为CAR-T细胞治疗产品的临床前安全性评价策略提供新的思路。     

药物心脏安全性评估与全面QT/QTc研究

许多非抗心律失常药物可以导致心电图QT间期延长,甚至引发尖端扭转型室性心动过速.因此在新药上市前,进行的心脏安全性评估,应该包括药物对QT间期影响的特点.全面QT/QTC研究旨在通过测量QT间期,明确药物是否具有延长QT间期的作用,判断其引发恶性心律失常的风险,并为决定药物是否进入下一步研发提供数据支持.     

A gain of function ryanodine receptor 2 mutation (R1760W-RyR2) in catecholaminergic polymorphic ventricular tachycardia

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited arrhythmia syndrome associated with Ca²⁺ leak predominantly caused by ryanodine receptor 2 (RyR2) mutations. We identified a R1760W-RyR2 mutation located between the N-terminal domain and the central domain of RyR2 in a CPVT patient by DNA sequencing. Recombinant mutant RyR2_{−2801mcherry} plasmid generated by the overlap extension polymerase chain reaction and seamless cloning was transfected in HEK293 cells for the cell model. Single-cell luminal and cytosolic Ca²⁺ imaging was measured by endoplasmic reticulum (ER) luminal Ca²⁺-sensitive protein D1ER and Fura-2 AM on a confocal laser scanning microscope, respectively. We found that in RyR2 mutant cells, the propensity for store-overload-induced Ca²⁺ release (SOICR) was enhanced representing increased Ca²⁺ oscillations, reduced activation and termination thresholds of spontaneous Ca²⁺ release; and the sensitivity to cytosolic Ca²⁺ activation was increased manifesting reduced steady state ER Ca²⁺ levels. Our results indicated that R1760W-RyR2 mutation induced calcium leak, representing a gain of function. Further, antiarrhythmic drugs propafenone and flecainide significantly suppressed SOICR caused by the R1760W-RyR2 mutation at a concentration of 20 μM, which was lower than the concentration at which carvedilol suppressed SOICR.