Cardiac adrenoceptors: physiological and pathophysiological relevance

At present, nine adrenoceptor (AR) subtypes have been identified: alpha(1A)-, alpha(1B)-, alpha(1D)-, alpha(2A)-, alpha(2B)-, alpha(2C)-, beta(1)-, beta(2)-, and beta(3)AR. In the human heart, beta(1)- and beta(2)AR are the most powerful physiologic mechanism to acutely increase cardiac performance. Changes in betaAR play an important role in chronic heart failure (CHF). Thus, due to increased sympathetic activity in CHF, betaAR are chronically (over)stimulated, and that results in beta(1)AR desensitization and alterations of down-stream mechanisms. However, several questions remain open: What is the role of beta(2)AR in CHF? What is the role of increases in cardiac G(i)-protein in CHF? Do increases in G-protein-coupled receptor kinase (GRK)s play a role in CHF? Does betaAR-blocker treatment cause its beneficial effects in CHF, at least partly, by reducing GRK-activity? In this review these aspects of cardiac AR pharmacology in CHF are discussed. In addition, new insights into the functional importance of beta(1)- and beta(2)AR gene polymorphisms are discussed. At present it seems that for cardiovascular diseases, betaAR polymorphisms do not play a role as disease-causing genes; however, they might be risk factors, might modify disease, and/or might influence progression of disease. Furthermore, betaAR polymorphisms might influence drug responses. Thus, evidence has accumulated that a beta(1)AR polymorphism (the Arg389Gly beta(1)AR) may affect the response to betaAR-blocker treatment.     

Possible therapeutic targets in cardiac myocyte apoptosis

Since Kerr described programmed cell death (apoptosis) as a process distinct from necrosis, there have been many studies of apoptosis in disease, especially of immunological origin. Because cardiac myocytes are terminally differentiated cells, they have typically been assumed to die exclusively by necrosis. However, during the last decade this view has been challenged by several studies demonstrating that a significant number of cardiac myocytes undergo apoptosis in myocardial infarction, heart failure, myocarditis, arrhythmogenic right ventricular dysplasia, and immune rejection after cardiac transplantation, as well as in other conditions of stress. These are potentially relevant observations, because apoptosis--unlike necrosis--can be blocked or reversed at early stages. Specific inhibition of this process may confer a considerable degree of cardioprotection, but requires a thorough understanding of the underlying mechanisms. Recent progress includes a better understanding of the importance of mitochondria-initiated events in cardiac myocyte apoptosis, of factors inducing apoptosis in heart failure and during hypoxia, and of the dual pro-apoptotic and anti-apoptotic effects of hypertrophic stimuli such as beta-adrenoceptor agonists, angiotensin converting enzyme inhibitors, nitric oxide and calcineurin. The investigation of cytoprotective and apoptotic signal transduction pathways has revealed important new insights into the roles of the mitogen-activated protein kinases p38, extracellular signal regulated kinase and c-Jun N-terminal kinase in cardiac cell fate. Our present review focuses on the intracellular signal transduction pathways of cardiac myocyte apoptosis and the possibility of specific inhibition of the process.     

Molecular mechanisms of apoptosis in the cardiac myocyte.

Cardiac myocytes can undergo programmed cell death in response to a variety of insults and apoptotic elimination of myocytes from the adult myocardium can lead directly to cardiomyopathy and death. Although it remains to be shown that therapy specifically targeting apoptosis will improve the prognosis of ischemic heart disease or heart failure, a number of studies in the past year have shed light on potential ways to intervene in the process. Progress in the past year includes a better understanding of the importance of mitochondria-initiated events in cardiac myocyte apoptosis, of factors inducing apoptosis during hypoxia, and of the dual pro-apoptotic and anti-apoptotic effects of hypertrophic stimuli such as beta-adrenoceptor agonists, nitric oxide and calcineurin. Further evidence supports the pathophysiologic relevance of apoptosis in human heart disease. The tracking of cytoprotective and apoptotic signal transduction pathways has revealed important new insights into the roles of the mitogen-activated protein (MAP) kinases p38, extracellular signal regulated kinase (ERK) and c-Jun N-terminus kinase (JNK) in cardiac cell fate.     

在急性呼吸窘迫综合征中分泌的磷脂酶A2与肺表面活性剂的相互作用及其病理生理关系

Phospholipases A2 (PLA2s) belong to a family of enzymes that hydrolyze phospholipids at the sn-2 position leading to the liberation of fatty acids and lysophospholipids (Fig 1). Mammalian PLA2s are divided into two major classes according to their location: high molecular mass intracellular PLA2 and low     

Force-frequency relationship in intact mammalian ventricular myocardium: physiological and pathophysiological relevance

The force–frequency relationship (FFR) is an important intrinsic regulatory mechanism of cardiac contractility. The FFR in most mammalian ventricular myocardium is positive; that is, an increase in contractile force in association with an increase in the amplitude of Ca²⁺ transients is induced by elevation of the stimulation frequency, which reflects the cardiac contractile reserve. The relationship is different depending on the range of frequency and species of animal. In some species, including rat and mouse, a ‘primary-phase’ negative FFR is induced over the low-frequency range up to approximately 0.5–1 Hz (rat) and 1–2 Hz (mouse). Even in these species, the FFR over the frequency range close to the physiological heart rate is positive and qualitatively similar to that in larger mammalian species, although the positive FFR is less prominent. The integrated dynamic balance of the intracellular Ca²⁺ concentration ([Ca²⁺]_i) is the primary cellular mechanism responsible for the FFR and is determined by sarcoplasmic reticulum (SR) Ca²⁺ load and Ca²⁺ flux through the sarcolemma via L-type Ca²⁺ channels and the Na⁺-Ca²⁺ exchanger. Intracellular Na⁺ concentration is also an important factor in [Ca²⁺]_i regulation. In isolated rabbit papillary muscle, over a lower frequency range (<0.5 Hz), an increase in duration rather than amplitude of Ca²⁺ transients appears to be responsible for the increase in contractile force, while over an intermediate frequency range (0.5–2.0 Hz), the amplitude of Ca²⁺ transients correlates well with the increase in contractile force. Over a higher frequency range (>2.5 Hz), the contractile force is dissociated from the amplitude of Ca²⁺ transients probably due to complex cellular mechanisms, including oxygen limitation in the central fibers of isolated muscle preparations, while the amplitude of Ca²⁺ transients increases further with increasing frequency (‘secondary-phase’ negative FFR). Calmodulin (CaM) may contribute to a positive FFR and the frequency-dependent acceleration of relaxation, although the role of calmodulin has not yet been established unequivocally. In failing ventricular myocardium, the positive FFR disappears or is inverted and becomes negative. The activation and overexpression of cardiac sarcoplasmic reticulum Ca²⁺ ATPase (SERCA2a) is able to reverse these abnormalities. Frequency-dependent alterations of systolic and diastolic force in association with those of Ca²⁺ transients and diastolic [Ca²⁺]_i levels are excellent indicators for analysis of cardiac excitation-contraction coupling, and for evaluating the severity of cardiac contractile dysfunction, cardiac reserve capacity and the effectiveness of therapeutic agents in congestive heart failure.     

Proteinases and signalling: pathophysiological and therapeutic implications via PARs and more

Proteinases like thrombin, trypsin and tissue kallikreins are now known to regulate cell signaling by cleaving and activating a novel family of G-protein-coupled proteinase-activated receptors (PARs 1-4) via exposure of a tethered receptor-triggering ligand. On their own, short synthetic PAR-selective PAR-activating peptides (PAR-APs) mimicking the tethered ligand sequences can activate PARs 1, 2 and 4 and cause physiological responses both in vitro and in vivo. Using the PAR-APs as sentinel probes in vivo, it has been found that PAR activation can affect the vascular, renal, respiratory, gastrointestinal, musculoskeletal and nervous systems (both central and peripheral nervous system) and can promote cancer metastasis and invasion. In general, responses triggered by PARs 1, 2 and 4 are in keeping with an innate immune inflammatory response, ranging from vasodilatation to intestinal inflammation, increased cytokine production and increased or decreased nociception. Further, PARs have been implicated in a number of disease states, including cancer and inflammation of the cardiovascular, respiratory, musculoskeletal, gastrointestinal and nervous systems. In addition to activating PARs, proteinases can cause hormone-like effects by other signalling mechanisms, like growth factor receptor activation, that may be as important as the activation of PARs. We, therefore, propose that the PARs themselves, their activating serine proteinases and their associated signalling pathways can be considered as attractive targets for therapeutic drug development. Thus, proteinases in general must now be considered as 'hormone-like' messengers that can signal either via PARs or other mechanisms.     

Endothelin receptor subtypes and their functional relevance in human small coronary arteries

1. The potent constrictor peptide endothelin (ET) has been implicated in various cardiovascular disorders including myocardial infarction and atherosclerosis. We have investigated the nature of ET receptor subtypes present on human small coronary arteries.
2. Small coronary arteries were mounted in a wire-myograph for in vitro pharmacology. To investigate the ET receptor subtypes present in different segments of the coronary vascular tree, arteries were grouped according to internal diameter. Responses in arteries with small internal diameters (mean 316.7±7.9 μm; Group B) were compared to those in larger arteries (mean 586.2±23.1 μm; Group A).
3. ET-1 consistently and potently contracted arteries from Group A and B, with EC₅₀ values of 1.7 (0.9–3.2) nM (n=15) and 2.3 (1.4–4.2) nM (n=14), respectively. No correlation was observed between ET-1 potency and internal diameter. The response to ET-1 was potently antagonized by the selective ET_A receptor antagonist PD156707 in both Group A and Group B, yielding pA₂ values of 8.60±0.12 (n=4–6) and 8.38±0.17 (n=4–6), respectively. Slopes from Schild regression were not significantly different from unity.
4. In contrast to ET-1, individual responses to ET-3 were variable. While all arteries from Group A responded to ET-3 (EC₅₀∼69 (23–210) nM) (n=12), no response was obtained in 5 of the 14 tested in Group B. Of those responding, many failed to reach a maximum at concentrations up to 1 μM. ET-1 was more potent than ET-3 in all arteries tested. A biphasic ET-3 response was observed in 8 arteries suggesting that a small ET_B population was also present in some patients. The selective ET_B receptor agonist sarafotoxin S6c had little or no effect up to 10 nM (n=4–6).
5. Responses to ET-1 and ET-3 were unaffected by removal of the endothelium in arteries from both groups suggesting a lack of functional, relaxant ET_B receptors on endothelial cells (n=5).
6. Using autoradiography, specific high density binding of the non-selective, ET_A/ET_B ligand [¹²⁵I]-ET-1 and selective ET_A ligand [¹²⁵I]-PD151242 was detected on the vascular smooth muscle layer of small intramyocardial coronary arteries (n=5). In contrast, little or no binding of the selective ET_B receptor ligand [¹²⁵I]-BQ3020 was observed (n=5). Similarly, [¹²⁵I]-ET-1 binding to vascular smooth muscle was absent in the presence of the selective ET_A receptor antagonist PD156707.
7. We conclude that human small epi- and intramyocardial coronary arteries express predominantly ET_A receptors and it is these receptors which mediate ET-induced contractions. A constrictor ET_B receptor population may exist in some patients. However, these receptors may have a limited role as contractions to ET-1 can be blocked fully by the selective ET_A receptor antagonist PD156707.

     

缺血/再灌注过程中心肌细胞自噬研究进展

自噬是一种广泛存在于真核细胞中的生命现象,心肌细胞营养缺乏、缺血/再灌注损伤、心衰等均可诱发细胞自噬。缺血/再灌注过程中的心肌细胞自噬可以维持心肌细胞稳态、减少细胞缺失,但是自噬作用也可导致心肌细胞死亡。     

Endothelin levels in experimental diabetes combined with cardiac hypertrophy

Diabetes mellitus is associated with endothelial and cardiac dysfunction, and endothelin has been suggested to alter cardiac function by being a positive inotropic agent, modulating the Frank-Starling response, contracting the coronary arteries and inducing tissue proliferation. We investigated endothelin levels in diabetic and in healthy dogs, 1 and 3 days after placing arteriovenous shunts (8 weeks after diabetes induction) in the femoral regions. Right and left ventricular weight/body weight ratios and Nterminal- atrial natriuretic peptide were increased in shunted animals (P < 0.05). Plasma endothelin levels were comparable in healthy and diabetic dogs. Shunted circulation did not change systemic endothelin levels in healthy dogs but reduced endothelin levels in diabetic dogs. The functional significance of altered endothelin responses to acute hemodynamic burden in experimental diabetes needs further investigation.     

Chymase: its pathophysiological roles and inhibitors

Chymase is a chymotrypsin-type serine protease mainly localized in mast cells (MCs). Human, primate, and dog chymase generate angiotensin II (Ang II) from Ang I, while mouse and rat chymases degrade Ang II. It is suggested that chymase generating Ang II might be an alternative Ang II-forming enzyme to angiotensin-converting enzyme (ACE) in the renin-angiotensin system in tissues, but not in blood, and cause hypertrophy and remodeling of cardiovascular tissues. Chymase also degrades extracellular matrix, and processes procollagenase, inflammatory cytokines and other bioactive peptides. As a result, chymase plays important roles in inflammatory tissues through its proteolytic activities to cause tissue remodeling, that is, a chymase inhibitor may have the ability to prevent diseases caused by the above inflammatory reactions. The investigation of chymase inhibitors by pharmaceutical companies has yielded peptide and peptide mimetic inhibitors. We also found potent non-peptide low molecular inhibitors. However, the in vivo functions of chymase have not been verified so far by applying a chymase inhibitor to in vivo pathological models. In this article, we overview the pathophysiological roles of chymase and chymase inhibitors proposed to date, and discuss the structure-activity relationships of substituted 3-phenylsulfonyl-1-phenylimidazolidine-2,4-dione derivatives.     

Pathophysiological relevance of the cardiac β2-adrenergic receptor and its potential as a therapeutic target to improve cardiac function

β-adrenoceptors are members of the G protein-coupled receptor superfamily which play a key role in the regulation of myocardial function. Their activation increases cardiac performance but can also induce deleterious effects such as cardiac arrhythmias or myocardial apoptosis. In fact, inhibition of β-adrenoceptors exerts a protective effect in patients with sympathetic over-stimulation during heart failure. Although β₂-adrenoceptor is not the predominant subtype in the heart, it seems to importantly contribute to the cardiac effects of adrenergic stimulation; however, the mechanism by which this occurs is not fully understood. This review summarizes the current knowledge on the role of β₂-adrenoceptors in the regulation of cardiac contractility, metabolism, cardiomyocyte survival and cardiac arrhythmias. In addition, therapeutic considerations relating to stimulation of the β₂-adrenoceptor such as an increase in cardiac contractility with low arrythmogenic effect, protection of the myocardium again apoptosis or positive regulation of heart metabolism are discussed.     

α_1受体对不同状态心肌细胞的作用

目的　通过激活和阻断不同状态心肌的α1 受体 ,探讨α1 受体的活化状态与不同状态心肌的关系。方法　采用Langedorff灌流技术复制动物模型。第 1个实验检验不同浓度的α1 受体激动剂 phenylephrine和阻断剂 prazosin(0 0 1、0 1、1、1 0、1 0 0 μmol·L- 1 )在缺血前 1 0min、缺血期和再灌注时的效应。第 2个实验研究 phenylephrine和 prazosin的时间依赖性。每组实验的末期 ,测量磷酸肌酸激酶、细胞凋亡和坏死情况。结果　 0 1、1 μmol·L- 1 的 phenylephrine可发挥最大效益 ,但prazosin浓度超过 1 0 μmol·L- 1 可产生有害作用。缺血前激活、缺血期间阻断α1 受体可保护心肌。再灌注时激活α1 受体效应不明确 ,但阻断α1 受体有害。结论　激动或阻断α1 受体所产生的效应与心肌的状态有关     

Inhibition of matrix metalloproteinases prevents peroxynitrite-induced contractile dysfunction in the isolated cardiac myocyte

BACKGROUND AND PURPOSE: The potent oxidant peroxynitrite (ONOO(-)) induces mechanical dysfunction in the intact heart in part through activation of matrix metalloproteinase-2 (MMP-2). This effect may be independent of the proteolytic actions of MMPs on extracellular matrix proteins. The purpose of this study was to examine the effects of ONOO(-) on contractile function at the level of the single cardiac myocyte and whether this includes the action of MMPs.EXPERIMENTAL APPROACH: Freshly isolated ventricular myocytes from adult rats were superfused with Krebs-Henseleit buffer at 21 degrees C and paced at 0.5 Hz. Contractility was measured using a video edge-detector. ONOO(-) or decomposed ONOO(-) (vehicle control) were co-infused over 40 min to evaluate the contraction cease time (CCT). The effects of ONOO(-) on intracellular [Ca(2+)] were determined in myocytes loaded with calcium green-1 AM. MMP-2 activity was measured by gelatin zymography.KEY RESULTS: ONOO(-) (30-600 microM) caused a concentration-dependent reduction in CCT. Myocytes subjected to 300 microM ONOO(-) had a shorter CCT than decomposed ONOO(-) (14.9+1.5 vs 32.2+3.5 min, n=7-8; P<0.05) and showed increased MMP-2 activity. The MMP inhibitors doxycycline (100 microM) or PD 166793 (2 microM) reduced the decline in CCT induced by 300 microM ONOO(-). ONOO(-) caused shorter calcium transient cease time and significant alterations in intracellular [Ca(2+)] homoeostasis which were partially prevented by doxycycline.CONCLUSIONS AND IMPLICATIONS: This is the first demonstration that inhibition of MMPs protects the cardiac myocyte from ONOO(-)-induced contractile failure via an action unrelated to proteolysis of extracellular matrix proteins.     

次氯酸介导的氧化应激及其病理生理学意义

机体内次氯酸(HClO)主要由髓过氧化物酶和血管过氧化物酶催化生成,HClO氧化性极强,基础水平的HClO在机体防御系统中起重要作用。然而,高浓度的HClO则给机体组织细胞造成氧化损伤。研究表明,HClO介导的氧化应激参与了动脉粥样硬化、细胞凋亡和衰老等多种病理生理过程。HClO可通过损害血管内皮功能、氧化修饰脂蛋白和增加斑块的不稳定性等多条途径促进动脉粥样硬化形成;通过影响线粒体渗透性转运孔和凋亡信号分子等途径促进细胞凋亡以及通过抑制抗衰老蛋白分子的表达和活性等途径促进细胞衰老。     

雌激素对大鼠心肌细胞钙的调控作用研究

目的　观察雌激素对大鼠心肌细胞钙转运的调控作用。方法　应用膜片钳全细胞记录方法观察L型钙通道电流 (ICa .L) ,共聚焦显微镜系统分析细胞内钙浓度 ([Ca2 + ]i)的变化。结果　应用 10 μmol·L-1和 30 μmol·L-1的 17β Estradiol,分别使ICa .L 抑制到正常的 48 1%± 4 5 %and2 9 3%± 5 2 % (n =5 ,P <0 0 1)。 10 μmol·L-1的 17β Estradiol可升高静息的培养心肌细胞 [Ca2 + ]i,荧光强度 (FI)值由 5 4 2± 13 6增加到 86 5± 15 3(n =6 ,P <0 0 5 ) ;而对具有自发收缩的心肌细胞钙瞬变幅度有明显的抑制作用 ,由给药前的 18 5± 4 3减小到 11 4± 3 9(n =6 ,P <0 0 5 )。17β Estradiol可剂量依赖性地抑制ICa.L。结论　L型钙通道等多种机制参与雌激素对大鼠心肌细胞钙转运的调控作用。     

Spatial and temporal aspects of cAMP signalling in cardiac myocytes

1. beta(1)-Adrenoceptor and M(2) muscarinic receptor regulation of cAMP production plays a pivotal role in autonomic regulation of cardiac myocyte function. However, not all responses are easily explained by a uniform increase or decrease in cAMP activity throughout the entire cell. 2. Adenovirus expression of fluorescence resonance energy transfer (FRET)-based biosensors can be used to monitor cAMP activity in protein kinase A (PKA) signalling domains, as well as the bulk cytoplasmic domain of intact adult cardiac myocytes. 3. Data obtained using FRET-based biosensors expressed in different cellular microdomains have been used to develop a computational model of compartmentalized cAMP signalling. 4. A systems biology approach that uses quantitative computational modelling together with experimental data obtained using FRET-based biosensors has been used to provide evidence for the idea that compartmentation of cAMP signalling is necessary to explain the stimulatory responses to beta(1)-adrenoceptor activation as well as the complex temporal responses to M(2) muscarinic receptor activation.     

Ethanol reduces cardiac myocyte function through activation of the nitric oxide-cyclic GMP pathway

We tested the hypothesis that low-dose ethanol would reduce cardiac myocyte function through increased production in the nitric oxide/cyclic GMP signal transduction pathway, rather than reduced degradation. Ventricular myocytes were isolated from the hearts of 9 rabbits. Myocyte function was studied using a video-edge detector and cyclic GMP levels were measured by radioimmunoassay. Cells were administered 5 and 10 mmol/l ethanol alone or after 10(-6) mol/l N(G)-nitro-L-arginine methyl ester (L-NAME, nitric oxide synthase inhibitor), 10(-6) mol/l 1H-[1,2,4]oxadiazolo[4,3a]quinoxalin-1-one (ODQ, soluble guanylyl cyclase inhibitor) or 10(-5) mol/l zaprinast (cyclic GMP phosphodiesterase inhibitor). Ethanol (10 mmol/l) significantly decreased percent shortening from 10.0 +/- 0.9 to 6.0 +/- 0.2%. Similar decrements occurred in the maximum rate of shortening and relaxation. After L-NAME or ODQ, the decrements in percent shortening, maximum rate of shortening and relaxation caused by ethanol were not significant. After zaprinast, ethanol significantly decreased the maximum rate of shortening and relaxation and percent shortening to 4.3 +/- 0.5. Ethanol (10 mmol/l) significantly increased cyclic GMP from 403 +/- 121 to 529 +/- 128 fmol/10(5) myocytes. Both L-NAME and ODQ lowered cyclic GMP, and ethanol did not affect cyclic GMP after either. Zaprinast raised cyclic GMP, as did its combination with 10 mmol/l ethanol (653 +/- 120). Thus, ethanol both reduced myocyte function and increased cyclic GMP. Blocking nitric oxide production or guanylyl cyclase activity prevent these effects of ethanol, while blocking cyclic GMP degradation did not. This suggests that ethanol acts as a nitric oxide stimulator in ventricular myocytes leading to reduced function and increased cyclic GMP.     

Review article: gastrointestinal sensory and motor disturbances in inflammatory bowel disease - clinical relevance and pathophysiological mechanisms

Background It is well known that inflammation has a profound impact on the neuromuscular apparatus of the gastrointestinal tract during the inflammatory insult and in periods of remission, at the site of inflammation and at distance from this site. The importance of this interaction is illustrated by the higher prevalence of functional gut disorders in patients with inflammatory bowel disease. Aims To document the epidemiological and clinical significance of functional alterations of gut motility and sensitivity in patients with inflammatory bowel disease and to formulate potential pathophysiological mechanisms. Results and conclusions Functional gut disorders occur frequently in patients with inflammatory bowel disease, both during inflammatory episodes and in periods of remission, and have a major impact on their quality of life. The clinical manifestations of these motility and sensitivity disorders vary and are often difficult to treat, mainly because therapeutic guidelines and specific diagnostic tests to distinguish inflammatory bowel disease from functional gut disorders are lacking. Chronic bowel inflammation results in a complicated interaction between neuroendocrine serotonin‐predominant cells of the mucosa, inflammatory cells (particularly mast cells) in the submucosa, the intrinsic and extrinsic innervation and the muscular apparatus including the interstitial cells of Cajal. The outcome of this interaction is a perturbation of gastrointestinal motor function, both locally and at distance from the site of inflammation and during both acute inflammation and remission.