The autosomal dominant mutation in the human alphaB-crystallin gene inducing a R120G amino acid exchange causes a multisystem, protein aggregation disease including cardiomyopathy. The pathogenesis of cardiomyopathy in this mutant (hR120GCryAB) is poorly understood. Here, we show that transgenic mice overexpressing cardiac-specific hR120GCryAB recapitulate the cardiomyopathy in humans and find that the mice are under reductive stress. The myopathic hearts show an increased recycling of oxidized glutathione (GSSG) to reduced glutathione (GSH), which is due to the augmented expression and enzymatic activities of glucose-6-phosphate dehydrogenase (G6PD), glutathione reductase, and glutathione peroxidase. The intercross of hR120GCryAB cardiomyopathic animals with mice with reduced G6PD levels rescues the progeny from cardiac hypertrophy and protein aggregation. These findings demonstrate that dysregulation of G6PD activity is necessary and sufficient for maladaptive reductive stress and suggest a novel therapeutic target for abrogating R120GCryAB cardiomyopathy and heart failure in humans.

Isolated single coronary artery is a rare congenital anomaly occuring in approximately 0.024% of the population. This entity can be diagnosed during life only by coronary angiography. Ten patients with isolated single coronary artery are reported. Based on angiographic analysis, a new classification is proposed, according to the site of origin and anatomical distribution of the branches. Typical angina did not occur with single coronary artery in the absence of coexisting coronary artery disease or aortic stenosis. No correlation was apparent between the type of anomalous patterns and the symptoms of angina.

Adult human enteroviral heart disease is often associated with the detection of enteroviral RNA in cardiac muscle tissue in the absence of infectious virus. Passage of coxsackievirus B3 (CVB3) in adult murine cardiomyocytes produced CVB3 that was noncytolytic in HeLa cells. Detectable but noncytopathic CVB3 was also isolated from hearts of mice inoculated with CVB3. Sequence analysis revealed five classes of CVB3 genomes with 5 termini containing 7, 12, 17, 30, and 49 nucleotide deletions. Structural changes (assayed by chemical modification) in cloned, terminally deleted 5-nontranslated regions were confined to the cloverleaf domain and localized within the region of the deletion, leaving key functional elements of the RNA intact. Transfection of CVB3 cDNA clones with the 5-terminal deletions into HeLa cells generated noncytolytic virus (CVB3/TD) which was neutralized by anti-CVB3 serum. Encapsidated negative-strand viral RNA was detected using CsCl-purified CVB3/TD virions, although no negative-strand virion RNA was detected in similarly treated parental CVB3 virions. The viral protein VPg was detected on CVB3/TD virion RNA molecules which terminate in 5 CG or 5 AG. Detection of viral RNA in mouse hearts from 1 week to over 5 months postinoculation with CVB3/TD demonstrated that CVB3/TD virus strains replicate and persist in vivo. These studies describe a naturally occurring genomic alteration to an enteroviral genome associated with long-term viral persistence.The six serotypes of the group B coxsackieviruses (CVB1-6) are enteroviruses (Picornaviridae, species HEV-B) (53). The CVB genome is a single-stranded RNA molecule, 7,400 nucleotides (nt) in length, that is encapsidated within an icosahedral shell (74). The 11 viral proteins (83) are encoded by a single open reading frame which is flanked on the 5Ј and 3Ј termini by nontranslated regions (NTRs) (20). The CVB induce numerous human illnesses, including inflammatory heart disease, pancreatitis, and aseptic meningitis and may also trigger the onset of type 1 diabetes (5,31,72,81,82). The CVB were recognized as causes of human heart disease shortly after their description early in the 1950s (26, 27) and remain the enteroviruses most commonly associated with human cardiomyopathies on the grounds of isolation and serology (5,6,34,62,88). Meta-analysis shows an association between enteroviruses and myocarditis in 23% of cases (7), although this association is more variable in cases of dilated cardiomyopathy, a serious disease that often leads to a failing heart (61). Mouse models of CVB-induced pancreatitis (94, 110), myocarditis (45,52,110), myositis (104, 105), and rapid-onset type 1 diabetes (31) which facilitate the study of these diseases have been developed.Enterovirus infections are generally considered to be acute events, with symptoms and virus titers peaking within a few days postinoculation (p.i.) and with virus being cleared by the adaptive immune response (17). However, enterovirus infections can persist under conditions of immunodeficiency (48,51,...

Intracellular [Na+] ([Na+]i) is regulated in cardiac myocytes by a balance of Na+ influx and efflux mechanisms. In the normal cell there is a large steady state electrochemical gradient favoring Na+ influx. This potential energy is used by numerous transport mechanisms, including Na+ channels and transporters which couple Na+ influx to either co- or counter-transport of other ions and solutes. Six sarcolemmal Na+ influx pathways are discussed in relatively quantitative terms: Na+ channels, Na+/Ca2+ exchange, Na+/H+ exchange, Na+/Mg2+ exchange, Na+/HCO3- cotransport and Na+/K+/2Cl- cotransport. Under normal conditions Na+/Ca2+ exchange and Na+ channels are the dominant Na+ influx pathways, but other transporters may become increasingly important during altered conditions (e.g. acidosis or cell volume stress). Mitochondria also exhibit Na+/Ca2+ antiporter and Na+/H+ exchange activity that are important in mitochondrial function. These coupled fluxes of Na+ with Ca2+, H+ and HCO3- make the detailed understanding of [Na+]i regulation pivotal to the understanding of both cardiac excitation-contraction coupling and pH regulation. The Na+/K+-ATPase is the main route for Na+ extrusion from cells and [Na+]i is a primary regulator under physiological conditions. [Na+]i is higher in rat than rabbit ventricular myocytes and the reason appears to be higher Na+ influx in rat with a consequent rise in Na+/K+-ATPase activity (rather than lower Na+/K+-ATPase function in rat). This has direct functional consequences. There may also be subcellular [Na+]i gradients locally in ventricular myocytes and this may also have important functional implications. Thus, the balance of Na+ fluxes in heart cells may be complex, but myocyte Na+ regulation is functionally important and merits focused attention as in this issue.

Calcium homeostasis in cardiac myocytes results from the integrated function of transsarcolemmal Ca2+ influx and efflux pathways modulated by membrane potential and from intracellular Ca2+ uptake and release caused predominantly by SR function. These processes can be importantly altered in different disease states as well as by pharmacological agents, and the resulting changes in systolic and diastolic [Ca2+]i can cause clinically significant alterations in contraction and relaxation of the heart. It may be anticipated that a rapid increase in our understanding of the pathophysiology of Ca2+ homeostasis in cardiac myocytes will be forthcoming as the powerful new tools of molecular and structural biology are used to investigate the regulation of Ca2+ transport systems.

Abstract-Mouse myocyte contractility and the changes induced by pressure overload are not fully understood. We studied contractile reserve in isolated left ventricular myocytes from mice with ascending aortic stenosis (AS) during compensatory hypertrophy (4-week AS) and the later stage of early failure (7-week AS) and from control mice. ] o , 25°C), the amplitude of myocyte shortening and peak-systolic [Ca 2ϩ ] i in 7-week AS were not different from those of controls, whereas contraction, relaxation, and the decline of [Ca 2ϩ ] i transients were slower. In response to the challenge of high [Ca 2ϩ ] o , fractional cell shortening was severely depressed with reduced peak-systolic [Ca 2ϩ ] i in 7-week AS compared with controls. In response to rapid pacing stimulation, cell shortening and peak-systolic [Ca 2ϩ ] i increased in controls, but this response was depressed in 7-week AS. In contrast, the responses to both challenge with high [Ca 2ϩ ] o and rapid pacing in 4-week AS were similar to those of controls. Although protein levels of Na ϩ -Ca 2ϩ exchanger were increased in both 4-week and 7-week AS, the ratio of SR Ca 2ϩ -ATPase to phospholamban protein levels was depressed in 7-week AS compared with controls but not in 4-week AS. This was associated with an impaired capacity to increase sarcoplasmic reticulum Ca 2ϩ load during high work states in 7-week AS myocytes. In hypertrophied failing mouse myocytes, depressed contractile reserve is related to an impaired augmentation of systolic [Ca 2ϩ ] i and SR Ca 2ϩ load and simulates findings in human failing myocytes. (Circ Res. 2000;87:588-595.) Key Words: myocytes Ⅲ contractility Ⅲ sarcoplasmic reticulum Ⅲ Ca 2ϩ -ATPase Ⅲ heart failure Ⅲ hypertrophy T he regulation of contractility is under intense investigation with the use of transgenic mice. 1,2 The mouse cardiovascular system differs from that of humans and larger mammals, including the rapid heart rate, faster myofibrillar ATPase activity and sarcoplasmic reticulum (SR) Ca 2ϩ uptake, 2,3 and higher mechanical performance per unit ventricular mass. 4 To interpret findings in genetically manipulated mice, it is critical to understand the properties of normal mouse myocyte contractility and the changes induced by clinically relevant stimuli such as chronic load. We reported that mice with ascending aortic stenosis (AS) develop compensated hypertrophy (4-week AS) and the later stage of early heart failure (7-week AS). 4,5 The aim of the present study was to examine contractile reserve in myocytes from AS mice in transition from hypertrophy to early heart failure. We measured myocyte contraction and [Ca 2ϩ ] i transients in left ventricular (LV) myocytes from normal, 4-week AS, and 7-week AS mice in response to the challenge of stepped increases in [Ca 2ϩ ] o . In separate experiments, we examined the response to rapid pacing stimulation. Although only subtle abnormalities in the time course of contraction and the [Ca 2ϩ ] i transients are present at baseline, contractile reserve is depressed in ...

The P2X4 receptor is a newly identified receptor expressed in the heart cell. Its function was elucidated with cardiac transgenic (TG) expression of the receptor by using the myocardium-specific a-myosin heavy chain promoter. The presence of the transgene was determined by polymerase chain reaction by using primers specific to the receptor and the vector linker region, by Southern blotting of the genomic DNA, and by immunoblotting and immunohistochemistry of both isolated cardiac myocytes and intact hearts. In intact heart study, the P2X4 receptor TG mouse exhibited significantly elevated basal cardiac contractility with greater rates of contraction and relaxation, left ventricular developed pressure, and cardiac output compared with nontransgenic (NTG) animals but showed no evidence of hypertrophy or heart failure. The TG heart also showed a greater increase of cardiac contractility in response to the P2X receptor agonist 2-methylthioATP, consistent with overexpression of a functional P2X4 receptor with consequent increase in the receptor-mediated response. In isolated cardiac cell study, the TG heart cell showed a similar level of basal contraction amplitude as the NTG heart cell while exhibiting a threefold greater increase in contractility during stimulation by 2-methylthioATP. Thus, an increased responsiveness of the overexpressed P2X4 receptor to endogenous ATP is responsible for the enhanced basal cardiac performance in the intact TG heart. The sustained enhanced contractile function with no associated heart pathology in the P2X4 receptor TG mouse suggests a novel physiologic role of the P2X4 receptor, that of stimulating the cardiac contractility.

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