Background-The failing myocardium is characterized by decreased force production, slowed relaxation, and depressed responses to ␤-adrenergic stimulation. In some heart failure patients, heart function is so poor that a left ventricular assist device (LVAD) is inserted as a bridge to transplantation. In the present research, we investigated whether circulatory support with an LVAD influenced the functional properties of myocytes from the failing heart. Methods and Results-Myocytes were isolated from human explanted failing hearts (HF-myocytes) and failing hearts with antecedent LVAD support (HF-LVAD-myocytes). Studies of myocyte function indicated that the magnitude of contraction was greater (9.6Ϯ0.7% versus 6.9Ϯ0.5% shortening), the time to peak contraction was significantly abbreviated (0.37Ϯ0.01 versus 0.75Ϯ0.04 seconds), and the time to 50% relaxation was reduced (0.55Ϯ0.02 versus 1.45Ϯ0.11 seconds) in the HF-LVAD-myocytes compared with the HF-myocytes (PϽ0.05). The HF-LVAD-myocytes had larger contractions than the HF-myocytes at all frequencies of stimulation tested. The negative force-frequency relationship of the HF-myocytes was improved in HF-LVAD-myocytes but was not reversed. Responses to ␤-adrenergic stimulation (by isoproterenol) were greater in HF-LVAD-myocytes versus HF-myocytes. Conclusions-The results of the study strongly support the idea that circulatory support with an LVAD improves myocyte contractile properties and increases ␤-adrenergic responsiveness. (Circulation. 1998;97:2316-2322.)

Our objective was to determine the respective roles of the sarcoplasmic reticulum (SR) and the Na+/Ca2+ exchanger in the small, slowly decaying Ca2+ transients of failing human ventricular myocytes. Left ventricular myocytes were isolated from explanted hearts of patients with severe heart failure (n=18). Cytosolic Ca2+, contraction, and action potentials were measured by using indo-1, edge detection, and patch pipettes, respectively. Selective inhibitors of SR Ca2+ transport (thapsigargin) and reverse-mode Na+/Ca2+ exchange activity (No. 7943, Kanebo Ltd) were used to define the respective contribution of these processes to the Ca2+ transient. Ca2+ transients and contractions induced by action potentials (AP transients) at 0.5 Hz exhibited phasic and tonic components. The duration of the tonic component was determined by the action potential duration. Ca2+ transients induced by caffeine (Caf transients) exhibited only a phasic component with a rapid rate of decay that was dependent on extracellular Na+. The SR Ca2+-ATPase inhibitor thapsigargin abolished the phasic component of the AP Ca2+ transient and of the Caf transient but had no significant effect on the tonic component of the AP transient. The Na+/Ca2+ exchange inhibitor No. 7943 eliminated the tonic component of the AP transient and reduced the magnitude of the phasic component. In failing human myocytes, Ca2+ transients and contractions exhibit an SR-related, phasic component and a slow, reverse-mode Na+/Ca2+ exchange-related tonic component. These findings suggest that Ca2+ influx via reverse-mode Na+/Ca2+ exchange during the action potential may contribute to the slow decay of the Ca2+ transient in failing human myocytes.

A previously uncharacterized type of sickled cell was found in venous blood ofpatients with sickle cell disease when blood was collected without exposure to air and fixed immediately with 1% glutaraldehyde solution equilibrated with 5% oxygen. These cells were either elongated, resembling irreversibly sickled cells (ISCs), or nonelongated, with a raisinlike shape. Both types assumed a normal discoidal shape upon full oxygenation. Since these cells exist only under partially oxygenated conditions, they are described as partially oxygenated sickled cells (POSCs). POSCs are morphologically distinct from partially deoxygenated sickled cells formed during deoxygenation by having rounded edges, while the latter have sharp edges. Transmission electron microscopy of POSCs revealed various amounts of misaligned Hb S polymers. Investigations in vitro demonstrated the formation of POSC-like cells by partial oxygenation of deoxygenated cells. Since POSCs contain intracellular fibers and sickle readily upon deoxygenation, they may have clinical and pathological significance.Sickle cell disease was so named because of the abnormal morphology of red blood cells (RBCs) observed by microscopic examination offixed and stained blood smears obtained from affected patients (1). Subsequently, relationships were found between oxygen and RBC sickling (2) and between intracellular polymerization of abnormal hemoglobin and cell deformation (3-5). Demonstration of sickling of RBCs from patients with sickle cell disease (SS cells) in vitro is highly dependent upon the method of deoxygenation. For example, classic sickle-or crescent-shaped cells are formed after slow deoxygenation (6-9), whereas SS cells with a granular or mosaic appearance result from rapid deoxygenation with nitrogen (6) or sodium dithionite (7). These differences may be explained by the number and size of domains of polymerized Hb S formed in the cells (10). During morphologic studies of SS cells, we noticed that the number and shape of sickled cells in venous blood were altered by the oxygenation of blood during and after blood collection. To avoid postcollection artifacts, we developed a method to collect blood under venous oxygen pressure without exposure to air. This enabled us to discover two additional types of reversibly sickled cells, elongated sickled cells with a shape like irreversibly sickled cells (ISCs) and shrunken cells with a raisin-like appearance. Since these cells exist only under partially oxygenated conditions, they are described as partially oxygenated sickled cells (POSCs) (11). We report our investigation of the morphologic properties and analysis by electron microscopy ofintracellular polymers of POSCs found in venous blood of patients with sickle cell disease. MATERIALS AND METHODSPreparation of Blood Samples. To minimize sickling during blood drawing (12), blood was drawn from the antecubital vein after release of the tourniquet. A conventional 5-ml plastic disposable syringe was used by the technique similar to that used for collectio...

Defects in myocyte contraction and relaxation are key features of human heart failure. Sodium/calcium exchanger-mediated contribution to contraction and relaxation were separated from other mechanisms [L-type calcium current, sarco(endo)plasmic reticulum (SR) Ca2+-ATPase] based on voltage, temperature, and selective blockers. Rod-shaped left ventricular myocytes were isolated from failed human explants ( n = 29) via perfusion with collagenase-containing Krebs solution. Action potentials using perforated patch and contractions using an edge detector were recorded at 0.5–1.5 Hz in Tyrode solution at 25°C and 37°C. Contraction duration was dependent on action potential (AP) duration at 37°C but not at 25°C, suggesting the role of the exchanger in relaxation and linking myocyte relaxation to the repolarization phase of the AP. Voltage-clamp experiments from −50 to +10 mV for 1,500 ms in Tyrode or Na+- and K+-free solutions after conditioning pulses triggered biphasic contractions that included a rapid SR-mediated component and a slower voltage-dependent exchanger-mediated component. We used thapsigargin to block the SR, which eliminated the rapid component, and we used an exchanger blocker, Kanebo 7943, which eliminated the slow component. The exchanger was shown to contribute to contraction through reverse-mode exchange, as well as to play a key role in relaxation of human ventricular myocytes.

The increases in phasic contraction magnitude observed in high-Na cells compared to Na-free cells were most likely due to increased SR Ca loading resulting from increased reverse-mode Na-Ca exchange. Our results also suggest that tonic contractions in high-Na cells were mediated by Ca entry via reverse-mode Na-Ca exchange and were not the result of either SR Ca release or L-type Ca channel activity.