Hideo A. Baba

Dynamic regulation of MEK/Erks and Akt/GSK-3β in human end-stage heart failure after left ventricular mechanical support: myocardial mechanotransduction-sensitivity as a possible molecular mechanism

OBJECTIVEnLeft ventricular assist devices (LVAD) are used to bridge patients with end-stage heart failure to transplantation. After long-term LVAD support, ventricular function may partially recover, a process called reverse remodeling. As several kinase-mediated signal transduction pathways have been implicated in the development of cardiac hypertrophy and failure, we examined the activities of the Erks, MEKs, Akt, GSK-3 beta, p70S6K, JNKs and p38 under LVAD support as well as during single myocyte strain and whole heart stretch.nnnMETHODSnWestern blotting and immunohistochemistry were performed using phospho-specific antibodies in matched samples from ten patients with end-stage heart failure before and after LVAD. Cyclic strain was performed in rat neonatal cardiac myocytes, and tensile stretch applied to Langendorff-perfused mouse hearts via a left ventricular balloon.nnnRESULTSnThe activity of Erks and Akt in failing hearts dramatically decreased after LVAD support, while that of GSK-3 beta increased. There was an endo/epicardial gradient for Erk activity which persisted after LVAD despite the reduction of total Erk activity. TUNEL-positivity and myocyte size decreased after LVAD, but independently of changes in kinase activity. In cardiomyocytes and Langendorff-perfused mouse hearts both strain/stretch and its relief regulated the activities of Erks, Akt, and GSK-3 beta.nnnCONCLUSIONnErks and Akt/GSK-3 beta are highly responsive to myocyte stretch in vitro and in vivo, and may be sensitive molecular parameters of reverse remodeling under LVAD support.

Cardiac remodeling after long term norepinephrine treatment in rats

Objective: In this study we have tested the hypothesis that degradation of collagen by matrix metalloproteinase 2 (MMP-2) precedes the deposition of extracellular matrix (ECM) after long term norepinephrine (NE) treatment. Methods: Female Sprague–Dawley rats received continuous i.v. infusion of NE (0.1 mg/kg·h) for 1, 2, 3, 4 and 14 days. Heart function and weight as well as expression of cardiac colligin and of collagen I and III were examined. Furthermore, we have assessed the degradation pathway of collagen by measuring the mRNA and activity of myocardial MMP-2 and tissue inhibitor of metalloproteinase 2 (TIMP-2) as well as the protein level of TIMP-2. Results: NE induced hypertrophy predominantly of the left ventricle (LV) in a time-dependent manner. It increased the mRNAs of colligin, collagen I and III, and of MMP-2 and TIMP-2 as well as MMP-2 activity in two phases: In the initial phase, at 3 and 4 days, the mRNA of colligin and of collagen I and III was elevated predominantly in the LV, MMP-2 and TIMP-2 mRNA, as well as TIMP-2 protein and MMP-activity were increased in both ventricles. The second phase, after 14 days, was characterized by a less pronounced increase in colligin, collagen I and III and in MMP-2 activity which occurred exclusively in the LV. Finally, long-term treatment with NE induced a 37% increase in interstitial fibrosis which was shown to occur exclusively in the LV after 14 days. Conclusion: NE treatment induced fibrosis exclusively in the LV which was associated with hypertrophy predominantly of the LV. The elevated MMP-2 activity seems to be necessary for the ECM to adapt to the enlargement of myocytes and to reduce overproduction of collagen.

Reversible activation of nuclear factor-κB in human end-stage heart failure after left ventricular mechanical support

OBJECTIVEnLeft ventricular assist devices (LVAD) have been used to bridge patients with end-stage heart failure to transplantation. Although several reports have suggested that the native ventricular function recovers after long-term LVAD support, a process called reverse remodeling, the underlying biological mechanisms are still unknown. As the transcription factor nuclear factor-kappaB (NF-kappaB) has been shown to be active in the failing human heart, we examined whether its activity is altered under LVAD support, and may thus contribute to the dynamic process of reverse remodeling.nnnMETHODSnThe activity of NF-kappaB was studied in 16 patients with end-stage heart failure (eight with dilated cardiomyopathy, six with ischemic heart disease, one with myocarditis, and one with congenital heart disease) before and after LVAD support by immunohistochemistry using an antibody against active NF-kappaB. Gel-shifts for NF-kappaB DNA-binding activity were performed with paired human myocardial tissue from four patients. The mean cardiomyocyte diameter before and after mechanical unloading was measured with an image analyzer system.nnnRESULTSn15 patients out of 16 showed a significant decrease in the number of NF-kappaB positive cardiomyocyte nuclei after LVAD support in the left ventricular myocardium. The NF-kappaB DNA-binding activity also decreased after LVAD support as measured by gel-shift analysis. While the number of positive cardiomyocytes was significantly higher in the subendocardium than in the subepicardium at the time of LVAD implantation, this difference was no longer present at the time of LVAD explantation. The diameter of cardiomyocytes in the left ventricle decreased significantly as a parameter of structural reverse remodeling.nnnCONCLUSIONnLVAD support decreases the extent of NF-kappaB activation in failing human hearts, suggesting that NF-kappaB may be involved in the process of reverse remodeling.

Heart-directed Expression of a Human Cardiac Isoform of cAMP-Response Element Modulator in Transgenic Mice

The transcriptional activation mediated by cAMP-response element (CRE) and transcription factors of the CRE-binding protein (CREB)/CRE modulator (CREM) family represents an important mechanism of cAMP-dependent gene regulation possibly implicated in detrimental effects of chronic β-adrenergic stimulation in end-stage heart failure. We studied the cardiac role of CREM in transgenic mice with heart-directed expression of CREM-IbΔC-X, a human cardiac CREM isoform. Transgenic mice displayed atrial enlargement with atrial and ventricular hypertrophy, developed atrial fibrillation, and died prematurely. In vivo hemodynamic assessment revealed increased contractility of transgenic left ventricles probably due to a selective up-regulation of SERCA2, the cardiac Ca2+-ATPase of the sarcoplasmic reticulum. In transgenic ventricles, reduced phosphorylation of phospholamban and of the CREB was associated with increased activity of serine-threonine protein phosphatase 1. The density of β1-adrenoreceptor was increased, and messenger RNAs encoding transcription factor dHAND and small G-protein RhoB were decreased in transgenic hearts as compared with wild-type controls. Our results indicate that heart-directed expression of CREM-IbΔC-X leads to complex cardiac alterations, suggesting CREM as a central regulator of cardiac morphology, function, and gene expression.

Reversal of metallothionein expression is different throughout the human myocardium after prolonged left-ventricular mechanical support

OBJECTIVESnWe examined the distribution of metallothionein (MT), a stress-inducible protein, and the cardiomyocyte diameter in human hearts after left-ventricular assist device (LVAD) support.nnnBACKGROUNDnRemodeling in end-stage heart failure is characterized by myocyte hypertrophy and alterations of several inducible proteins. LVADs used as a bridge to cardiac transplantation unload the left ventricle and may lead to a reversal of the remodeling, but little is known about the pathophysiology of this process.nnnMETHODSnThe immunoreactivity for MT and the cardiomyocyte diameter was analyzed in left-ventricular tissue specimens of 17 patients with end-stage heart failure before and after LVAD support.nnnRESULTSnMT positive cells were mainly located sub-endocardially in vacuolized cardiomyocytes and in small vessels throughout the myocardium. During LVAD support, MT-positive myocytes decreased in the sub-endocardial (p < 0.008) and sub-epicardial region (p < 0.003), MT-positive vessels decreased similarly (p < 0.003). Cardiomyocyte diameter decreased significantly only in the sub-endocardium (p < 0.03). Hearts of patients supported longer than 88 days (= median) showed substantially lower MT reactivity at the time of LVAD explantation as compared to patients supported less than 88 days.nnnCONCLUSIONnOur results suggest that unloading of the left ventricle during prolonged LVAD support leads to regression of cellular hypertrophy and a decrease of MT expression. The preferential reduction of MT-positive vacuolized cardiomyocytes in the sub-endocardium is comparable with the concept of greatest reduction of wall stress in this area of the myocardium and may be due to the improvement of myocardial blood flow and the energy balance.

Expression of Heat Shock Proteins 72/73 in Human Peritoneal Mesothelial Cells in vivo and in vitro

Leukocyte accumulation during peritonitis leads to an injurious microenvironment which is involved in the host defense reaction but is also thought to cause peritoneal damage. We tested the hypothesis that mesothelial cells (MC) respond to the injurious microenvironment during peritonitis by an increased expression of heat shock proteins (HSP 72/73), a basic way by which cells are protected against injury. Comparison of resting MC and activated MC during peritonitis in vivo by means of immunohistochemistry revealed an increased expression of HSP 72/73. As assessed by Western immunoblotting, incubation of MC in vitro with tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) caused a time-dependent induction of HSP 72/73 expression, which was maximal 6 h after stimulation. We suggest that the increased HSP 72/ 73 expression of MC during peritonitis is in part induced by TNF-α and IL-1β and may exert a cell-protective function, lessening MC damage during peritonitis.

Surgical Management of Extensive Lipomatous Hypertrophy of the Right Atrium

Lipomatous hypertrophy of the right atrium is a rare lesion that is usually limited to the interatrial septum. The authors report on a patient that suffered from an extensive lipomatous hypertrophy, which protruded into the right and left atrium as well as the superior vena cava and caused inflow obstruction. Therapeutic management is discussed.

Early proliferative changes in hearts of hypertensive Goldblatt rats: an immunohistochemical and flow-cytometrical study

Hyperplasia of myocytes in cardiac adaptation is a rare event in the mammalian cardiac muscle. Recent findings support the concept that proliferation of myocytes in the adult mammalian heart may be induced after a prolonged increase in pressure load on the myocardium. To determine whether short-term hypertension leads to hyperplasia of myocyte nuclei in the rat heart renal hypertension was produced in 12 Wistar rats. As soon as hypertension occurred, bromodeoxyuridine (BrdU) (50 mg/kg/day) was injected intraperitoneally on three subsequent days. Twelve sham-operated rats served as controls. After 3 days, the left cardiac ventricle was excised and double-staining with anti-BrdU antibody and propidium iodide was performed to determine the phase of cell-cycle of the BrdU-positive cells by flow-cytometry. Immunohistochemical double-staining with desmin, smooth muscle actin, vimentin, and BrdU was done to classify the BrdU-positive cells.Most of the BrdU-positive cells were in the G0/G1-phase of the cellcycle, suggesting cell proliferation or DNA-repair have taken place; polyploidy was not observed. In the hypertensive group (4.62%±2.36) significantly more cells incorporated BrdU than in the control group (1.46%±0.96). Immunohistochemically, the majority of the BrdU-positive cells consisted of fibrocytes, smooth muscle cells, and endothelial cells. Only 0.35%±0.26 of cardiac myocytes in the normotensive group showed positive BrdU-staining compared to 0.48%±0.32 in the hypertensive group. This difference was statistically not significant.This study showed that early after onset of hypertension proliferation of non-myocytes, but not of myocytes occurred. DNA synthesis is limited almost completely to the interstitial cells and does not occur in any significant extent in cardiac myocytes. In conclusion, hyperplasia of cardiac myocytes is not observed at carly stages of hypertension, but it may develop at a late stage of cardiac adaptation.

Metallothionein: Localization in Human Transplant Endomyocardium, Relation to Cytokines and Allograft Function

BACKGROUNDnThe aim of this study was to investigate the role of metallothionein in cardiac transplants in relation to cytokines and allograft function. Recent studies have revealed an association of allograft dysfunction with elevated proinflammatory cytokines independent of cellular rejection. In animal experiments, cytokines induced overexpression of metallothionein, a low-molecular-weight protein implicated in cellular stress response.nnnMETHODSnIn 105 consecutive biopsies from 15 patients during the first 3 months after heart transplantation, metallothionein expression was investigated immunohistochemically. Its relation to serum interleukin-6, tumor necrosis factor-alpha, interleukin-2 (IL-2), soluble interleukin-2 receptor rejection, and echocardiographic parameters was determined. Forty-three biopsies of 12 patients with idiopathic ventricular tachycardia served as controls.nnnRESULTSnMetallothionein expression was demonstrated in small vessels, cardiomyocytes, fibrocytes, and interstitial round cells. A positive relation between interleukin-6 levels and the number of metallothionein-positive small vessels (p < 0.028) was observed. Patients with lower serum IL-2 levels showed significantly higher numbers of metallothionein-positive small vessels (p < 0.043). Grafts with prolonged ischemic time (>150 minutes) showed a significantly higher myocardial metallothionein score (p < 0.021). Metallothionein expression was associated with lower fractional shortening, larger left ventricular end-systolic diameter, and lower mean arterial pressure but not with acute cellular rejection.nnnCONCLUSIONSnMetallothionein expression is associated with elevated interleukin-6 and decreased interleukin-2 serum levels and left ventricular allograft dysfunction in the absence of rejection.

Intracardiac Tuberculoma Causing “Idiopathic” Ventricular Tachycardia in a Patient Without Detectable Heart Disease

A 37-year-old man presented with frequent monomorphic ventricular tachycardia (VT; cycle length 340 msec, right bundle branch block morphology, superior axis, R-S interval . 80 msec) (panel A), with a second VT morphology inducible during electrophysiologic study (panel B). There was no clinical sign of infection or chronic disease. Results of echocardiography, right and left heart angiography, routine laboratory tests, and chest X-ray were normal. Antiarrhythmic drugs (beta blocker, sotalol 640 mg/day, and x8f ecainide alone or in combination) did not suppress VT inducibility or recurrence. Although extensive right and left ventricular endocardial mapping was performed, presystolic activity was not detected during VT. Earliest endocardial activation (5 msec before onset of QRS) was mapped to the left mid-lateral wall. Radiofrequency catheter ablation at this site was not successful, suggesting a nonendocardial VT origin. During open chest mapping, we identix8e ed a left mid-lateral epicardial area with presystolic activation during VT (30 msec before onset of QRS) and fractionated electrograms during sinus rhythm. Applying pressure in this region terminated the VT. Palpation revealed a single nodular mass (1.3-cm diameter) in this area. Excising this mass suppressed VT inducibility. Histopathologic examination showed a nodule with granulomatous inx8f ammation containing giant cells (panel C, arrows indicate granulomatous inx8f ammation); polymerase chain reaction conx8e rmed presence of Mycobacterium tuberculosis. The patient died 4 days postoperatively, most likely due to tuberculous sepsis. “Idiopathic” VT may be the x8e rst clinical manifestation of tuberculosis. J Cardiovasc Electrophysiol, Vol. 12, p. 118, January 2001.