In vivo heat shock preconditioning migrates calcium overload during ischema/reperfusion in the isolated, perfused rat heart

Heat shock (HS) pretreatment of the heart is effective in mitigating the deleterious effects of ischaemia/reperfusion. The main objective of this study was to determine whether the beneficial effect of HS is associated with the preservation of intracellular Ca2+ handling in the ischaemic/reperfused, isolated rat heart. Twenty-four hours after raising body core temperature to 42 °C for 15 min, rat hearts were perfused according to Langendorff and subjected to 30 min ischaemia followed by 20 min reperfusion. Cyclic changes of cytoplasmic calcium ion [Ca2+i] levels were measured by surface fluorometry using Indo-1 AM. Reperfused HS hearts showed improved recovery of contractile function compared with control hearts: end-diastolic pressure: 45±11 vs. 64±22 mm Hg; developed pressure: 72±12 vs. 41±20 mm Hg; maximum rate of pressure increase (+dP/dtmax): 1,513±305 vs. 938±500 mm Hg/s; maximum rate of pressure decrease (–dP/dtmax): –1,354±304 vs. –806±403 mm Hg/s. HS hearts displayed a significantly lower end-diastolic cytosolic [Ca2+] ([Ca2+]i) after reinstallation of flow. The dynamic parameters of the Ca2+i transients, i.e. the maximum rate of increase/decrease (±dCa2+i/dtmax) and amplitude, did not differ between reperfused control and HS hearts. The novel finding of this study is that improved performance of the HS-preconditioned heart after an ischaemic insult is associated with a reduced end-diastolic Ca2+i load, and most likely, preserved Ca2+ sensitivity of the myocardial contractile machinery

Poly(ADP-ribose) polymerase regulates myocardial calcium handling in doxorubicin-induced heart failure

Reactive oxygen and nitrogen species are overproduced in the cardiovascular system in response to the exposure to doxorubicin, a cardiotoxic anticancer compound. Oxidant-induced cell injury involves the activation of the nuclear enzyme poly(ADP-ribose) polymerase (PARP) and pharmacological inhibition of PARP has recently been shown to improve myocardial contractility in doxorubicin-induced heart failure models. The current investigation, by utilizing an isolated perfused heart system capable of beat-to-beat intracellular calcium recording, addressed the following questions: (1) is intracellular calcium handling altered in hearts of rats after 6-week doxorubicin treatment, under baseline conditions, and in response to oxidative stress induced by hydrogen peroxide exposure in vitro; and (2) does pharmacological inhibition of PARP with the phenanthridinone-based PARP inhibitor PJ34 affect the changes in myocardial mechanical performance and calcium handling in doxorubicin-treated hearts under normal conditions and in response to oxidative stress. The results showed a marked elevation in intracellular calcium in the doxorubicin-treated hearts which was normalized by pharmacological inhibition of PARP. PARP inhibition also prevented the myocardial contractile disturbances and calcium overload that developed in response to hydrogen peroxide in the doxorubicin-treated hearts. We conclude that PARP activation contributes to the development of the disturbances in cellular calcium handling that develop in the myocardium in response to prolonged doxorubicin exposur

Poly(ADP-ribose) polymerase regulates myocardial calcium handling in doxorubicin-induced heart failure

Reactive oxygen and nitrogen species are overproduced in the cardiovascular system in response to the exposure to doxorubicin, a cardiotoxic anticancer compound. Oxidant-induced cell injury involves the activation of the nuclear enzyme poly(ADP-ribose) polymerase (PARP) and pharmacological inhibition of PARP has recently been shown to improve myocardial contractility in doxorubicin-induced heart failure models. The current investigation, by utilizing an isolated perfused heart system capable of beat-to-beat intracellular calcium recording, addressed the following questions: (1) is intracellular calcium handling altered in hearts of rats after 6-week doxorubicin treatment, under baseline conditions, and in response to oxidative stress induced by hydrogen peroxide exposure in vitro; and (2) does pharmacological inhibition of PARP with the phenanthridinone-based PARP inhibitor PJ34 affect the changes in myocardial mechanical performance and calcium handling in doxorubicin-treated hearts under normal conditions and in response to oxidative stress. The results showed a marked elevation in intracellular calcium in the doxorubicin-treated hearts which was normalized by pharmacological inhibition of PARP. PARP inhibition also prevented the myocardial contractile disturbances and calcium overload that developed in response to hydrogen peroxide in the doxorubicin-treated hearts. We conclude that PARP activation contributes to the development of the disturbances in cellular calcium handling that develop in the myocardium in response to prolonged doxorubicin exposur

In vivo heat shock preconditioning migrates calcium overload during ischema/reperfusion in the isolated, perfused rat heart

Heat shock (HS) pretreatment of the heart is effective in mitigating the deleterious effects of ischaemia/reperfusion. The main objective of this study was to determine whether the beneficial effect of HS is associated with the preservation of intracellular Ca2+ handling in the ischaemic/reperfused, isolated rat heart. Twenty-four hours after raising body core temperature to 42 °C for 15 min, rat hearts were perfused according to Langendorff and subjected to 30 min ischaemia followed by 20 min reperfusion. Cyclic changes of cytoplasmic calcium ion [Ca2+i] levels were measured by surface fluorometry using Indo-1 AM. Reperfused HS hearts showed improved recovery of contractile function compared with control hearts: end-diastolic pressure: 45±11 vs. 64±22 mm Hg; developed pressure: 72±12 vs. 41±20 mm Hg; maximum rate of pressure increase (+dP/dtmax): 1,513±305 vs. 938±500 mm Hg/s; maximum rate of pressure decrease (–dP/dtmax): –1,354±304 vs. –806±403 mm Hg/s. HS hearts displayed a significantly lower end-diastolic cytosolic [Ca2+] ([Ca2+]i) after reinstallation of flow. The dynamic parameters of the Ca2+i transients, i.e. the maximum rate of increase/decrease (±dCa2+i/dtmax) and amplitude, did not differ between reperfused control and HS hearts. The novel finding of this study is that improved performance of the HS-preconditioned heart after an ischaemic insult is associated with a reduced end-diastolic Ca2+i load, and most likely, preserved Ca2+ sensitivity of the myocardial contractile machinery

β-Adrenergic activation reveals impaired cardiac calcium handling at early stage of diabetes

Cardiac function is known to be impaired in diabetes. Alterations in intracellular calcium handling have been suggested to play a pivotal role. This study aimed to test the hypothesis that ß-adrenergic activation can reveal the functional derangements of intracellular calcium handling of the 4-week diabetic heart. Langendorff perfused hearts of 4-week streptozotocin-induced diabetic rats were subjected to the ß-adrenoceptor agonist isoproterenol. Cyclic changes in [Ca2+]i levels were measured throughout the cardiac cycle using Indo-1 fluorescent dye. Based on the computational analysis of the [Ca2+]i transient the kinetic parameters of the sarcoplasmic reticulum Ca2+-ATPase and the ryanodine receptor were determined by minimizing the squared error between the simulated and the experimentally obtained [Ca2+]i transient. Under unchallenged conditions, hemodynamic parameters were comparable between control and diabetic hearts. Isoproterenol administration stimulated hemodynamic function to a greater extent in control than in diabetic hearts, which was exemplified by more pronounced increases in rate of pressure development and decline. Under unchallenged conditions, [Ca2+]i amplitude and rate of rise and decline of [Ca2+]i as measured throughout the cardiac cycle were comparable between diabetic and control hearts. Differences became apparent under ß-adrenoceptor stimulation. Upon ß-activation the rate-pressure product showed a blunted response, which was accompanied by a diminished rise in [Ca2+]i amplitude in diabetic hearts. Computational analysis revealed a reduced function of the sarcoplasmic reticulum Ca2+-ATPase and Ca2+-release channel in response to ß-adrenoceptor challenge. Alterations in Ca2+i handling may play a causative role in depressed hemodynamic performance of the challenged heart at an early stage of diabetes

Altered calcium handling is an early sign of streptozotocin-induced diabetic cardiomyopathy

The main objective of the present study was to determine alterations of calcium handling in the diabetic rat heart during the transition from adaptive to maladaptive phase of cardiomyopathy. By inhibiting the nuclear enzyme poly(ADP-ribose) polymerase (PARP), we also investigated the possible role of this enzyme in the sequence of pathological events. Six weeks after induction of type I diabetes by injection of streptozotocin in rats, the hearts were perfused according to Langendorff. Intracellular-free calcium (Ca2+i) levels were measured by surface fluorometry using Indo-1 AM. Cyclic changes in Ca2+i concentrations and hemodynamic parameters were measured simultaneously. The hearts were challenged by infusion of isoproterenol. Six weeks of diabetes resulted in reduced inotropy and lusitropy. The diabetic hearts (DM) expressed a significantly elevated end-diastolic Ca2+i level (control, 111±20 vs DM, 221±35 nM). The maximal transport capacity of SERCA2a and conductance of RyR2 were reduced. These changes were not accompanied by major alterations in the tissue content of SERCA2a, RyR2, phospholamban and Na+/Ca2+ exchanger. In response to ß-adrenergic activation, SERCA2a transport capacity and RyR2 conductance were stunted in the DM hearts. Inhibition of PARP induced minor changes in the mechanical function and calcium handling of the DM hearts. In conclusion, the observed changes in contractility and in Ca2+i handling are most likely attributable to functional disturbances of SERCA2a and RyR2 in this transitional phase of diabetes. At this stage of diabetes, PARP does not appear to play a significant pathogenetic role in the alterations in contractile function and calcium handlin

Altered calcium handling is an early sign of streptozotocin-induced diabetic cardiomyopathy

The main objective of the present study was to determine alterations of calcium handling in the diabetic rat heart during the transition from adaptive to maladaptive phase of cardiomyopathy. By inhibiting the nuclear enzyme poly(ADP-ribose) polymerase (PARP), we also investigated the possible role of this enzyme in the sequence of pathological events. Six weeks after induction of type I diabetes by injection of streptozotocin in rats, the hearts were perfused according to Langendorff. Intracellular-free calcium (Ca2+i) levels were measured by surface fluorometry using Indo-1 AM. Cyclic changes in Ca2+i concentrations and hemodynamic parameters were measured simultaneously. The hearts were challenged by infusion of isoproterenol. Six weeks of diabetes resulted in reduced inotropy and lusitropy. The diabetic hearts (DM) expressed a significantly elevated end-diastolic Ca2+i level (control, 111±20 vs DM, 221±35 nM). The maximal transport capacity of SERCA2a and conductance of RyR2 were reduced. These changes were not accompanied by major alterations in the tissue content of SERCA2a, RyR2, phospholamban and Na+/Ca2+ exchanger. In response to ß-adrenergic activation, SERCA2a transport capacity and RyR2 conductance were stunted in the DM hearts. Inhibition of PARP induced minor changes in the mechanical function and calcium handling of the DM hearts. In conclusion, the observed changes in contractility and in Ca2+i handling are most likely attributable to functional disturbances of SERCA2a and RyR2 in this transitional phase of diabetes. At this stage of diabetes, PARP does not appear to play a significant pathogenetic role in the alterations in contractile function and calcium handlin

Quantitative measurement of the mechanical properties of human antibodies with sub-10-nm resolution in a liquid environment

The nanomechanical properties of single human immunoglobulin G and M antibodies were measured in a liquid environment using a fast force-volume technique with sub-10-nm spatial resolution. The ultrastructural details of these molecules were resolved in the images. Simultaneously, important physical properties, including elasticity, adhesion, and deformation were measured. The dimensions and adsorption of the immunoglobulin M antibodies onto the substrate indicated that they are highly flexible. The antibodies were characterized by a low elastic stiffness (34 +/- 10 MPa) and high deformability (1.5 +/- 0.5 nm)