Massimo Maccherini

Characterization of the Hyperpolarization-Activated Current, If, in Ventricular Myocytes From Human Failing Heart

BACKGROUND Disease-associated electrophysiological alterations may contribute to the increased predisposition to arrhythmias of the hypertrophied or failing myocardium. An I(f)-like current is expressed in rat left ventricular myocytes (LVMs), its amplitude being linearly related to the severity of cardiac hypertrophy. Here, we report the occurrence and electrophysiological properties of I(f) in human LVMs. METHODS AND RESULTS LVMs were isolated from hearts of three male patients undergoing cardiac transplantation for terminal heart failure due to ischemic dilated cardiomyopathy. The patch-clamp technique was used to record I(f), ie, a barium-insensitive, cesium-sensitive, time-dependent increasing inward current elicited on hyperpolarization. Membrane capacitance was 244 +/- 27 pF (n = 25). I(f) occurred in all cells tested; its density measured at -120 mV was 2.1 +/- 0.3 pA/pF. Activation curves of I(f) (n = 24) were fitted by a Boltzmann function; the threshold was -55 mV; midpoint, -70.9 +/- 2.1 mV; slope, -5.4 +/- 0.3 mV; and maximal specific conductance, 19.6 +/- 2.5 pS/pF. I(f) blockade by extracellular cesium was voltage dependent. Reducing extracellular potassium concentration from 25 to 5.4 mmol/L caused a shift of the reversal potential from -12.7 +/- 0.5 to -24.8 +/- 2.1 mV and a 64% decrease of current conductance. CONCLUSIONS I(f) is present in human LVMs. Its electrophysiological characteristics resemble those previously described in hypertrophied rat LVMs and suggest that I(f) could be an arrhythmogenic mechanism in patients with severe heart failure.

Cardiac Growth Factors in Human Hypertrophy Relations With Myocardial Contractility and Wall Stress

The aim of the present study was to investigate whether and which cardiac growth factors are involved in human hypertrophy, whether growth factor synthesis is influenced by overload type and/or by the adequacy of the hypertrophy, and the relationships between cardiac growth factor formation and ventricular function. Cardiac growth factor formation was assessed by measuring aorta-coronary sinus concentration gradient in patients with isolated aortic stenosis (n=26) or regurgitation (n=15) and controls (n=12). Gene expression and cellular localization was investigated in ventricular biopsies using reverse transcriptase-polymerase chain reaction and in situ hybridization. Cardiac hypertrophy with end-systolic wall stress <90 kdyne/cm2 was associated with a selective increased formation of insulin-like growth factor (IGF)-I in aortic regurgitation and of IGF-I and endothelin (ET)-1 in aortic stenosis. mRNA levels for IGF-I and preproET-1 were elevated and mainly expressed in cardiomyocytes. At stepwise analysis, IGF-I formation was correlated to the mean velocity of circumferential fiber shortening (r=0.86, P<0.001) and ET-1 formation to relative wall thickness (r=0.82, P<0. 001). When end-systolic wall stress was >90 kdyne/cm2, IGF-I and ET-1 synthesis by cardiomyocytes was no longer detectable, and only angiotensin (Ang) II was generated, regardless of the type of overload. The mRNA level for angiotensinogen was high, and the mRNA was exclusively expressed in the interstitial cells. Ang II formation was positively correlated to end-systolic stress (r=0.89, P<0.001) and end-diastolic stress (r=0.84, P<0.001). Multivariate stepwise analysis selected end-systolic stress as the most predictive variable and left ventricular end-diastolic pressure as the independent variable for Ang II formation (r=0.93, P<0.001). In conclusion, the present results indicate that the course of human left ventricular hypertrophy is characterized by the participation of different cardiac growth factors that are selectively related both to the type of hemodynamic overload and to ventricular function.

Left atrial longitudinal strain by speckle tracking echocardiography correlates well with left ventricular filling pressures in patients with heart failure

BackgroundThe combination of early transmitral inflow velocity and mitral annular tissue Doppler imaging (E/Em ratio) is widely applied to noninvasively estimate left ventricular (LV) filling pressures. However E/Em ratio has a significant gray zone and its accuracy in patients with heart failure is debated. Left atrial (LA) deformation analysis by speckle tracking echocardiography (STE) was recently proposed as an alternative approach to estimate LV filling pressures. This study aimed at exploring the correlation of LA longitudinal function by STE and Doppler measurements with direct measurements of LV filling pressures in patients with heart failure.MethodsA total of 36 patients with advanced systolic heart failure (ejection fraction ≤35%), undergoing right heart catheterization, were studied. Simultaneously to pulmonary capillary wedge pressure (PCWP) determination, peak atrial longitudinal strain (PALS) and mean E/Em ratio were measured in all subjects by two independent operators. PALS values were obtained by averaging all segments (global PALS), and by separately averaging segments measured in the 4-chamber and 2-chamber views.ResultsNot significant correlation was found between mean E/Em ratio and PCWP (R = 0.15). A close negative correlation between global PALS and the PCWP was found (R = -0.81, p < 0.0001). Furthermore, global PALS demonstrated the highest diagnostic accuracy (AUC of 0.93) and excellent sensitivity and specificity of 100% and 93%, respectively, to predict elevated filling pressure using a cutoff value less than 15.1%. Bland-Altman analysis confirmed this close agreement between PCWP estimated by global PALS and invasive PCWP (mean bias 0.1 ± 8.0 mmHg).ConclusionIn a group of patients with advanced systolic heart failure, E/Em ratio correlated poorly with invasively obtained LV filling pressures. However, LA longitudinal deformation analysis by STE correlated well with PCWP, providing a better estimation of LV filling pressures in this particular clinical setting.

Carbon monoxide improves cardiac energetics and safeguards the heart during reperfusion after cardiopulmonary bypass in pigs

Ischemia‐reperfusion injury, a clinical problem during cardiac surgery, involves worsened adenosine trisphosphate (ATP) generation and damage to the heart. We studied carbon monoxide (CO) pretreatment, proven valuable in rodents but not previously tested in large animals, for its effects on pig hearts subjected to cardiopulmonary bypass with cardioplegic arrest. Hearts of CO‐treated pigs showed significantly higher ATP and phosphocreatine levels, less interstitial edema, and apoptosis of cardiomyocytes and required fewer defibrillations after bypass. We conclude that treatment with CO improves the energy status, prevents edema formation and apoptosis, and facilitates recovery in a clinically relevant model of cardiopulmonary bypass surgery.

Everolimus with reduced cyclosporine versus MMF with standard cyclosporine in de novo heart transplant recipients.

Background. Pharmacokinetic modeling supports trough monitoring of everolimus, but prospective data comparing this approach versus mycophenolate mofetil (MMF) in de novo cardiac transplant recipients are currently unavailable. Methods. In a 12-month multicenter open-label study, cardiac transplant patients received everolimus (trough level 3–8 ng/mL) with reduced cyclosporine A (CsA) or MMF (3 g/day) with standard CsA, both with corticosteroids±induction therapy. Results. In total, 176 patients were randomized (everolimus 92, MMF 84). Mean creatinine clearance was 72.5±27.9 and 76.8±32.1 mL/min at baseline, 65.4±24.7 and 72.2±26.2 mL/min at month 6, and 68.7±27.7 and 71.8±29.8 mL/min at month 12 with everolimus and MMF, respectively. The primary endpoint was not met since calculated CrCl at month 6 posttransplant was 6.9 mL/min lower with everolimus, exceeding the predefined margin of 6 mL/min. However, by month 12 the between-group difference had narrowed versus baseline (3.1 mL/min). All efficacy endpoints were noninferior for everolimus versus MMF. The 12-month incidence of biopsy-proven acute rejection International Heart and Lung Transplantation grade more than or equal to 3A was 21 of 92 (22.8%) with everolimus and 25 of 84 (29.8%) with MMF. Adverse events were consistent with class effects including less-frequent cytomegalovirus infection with everolimus (4 [4.4%]) than MMF (14 [16.9%], P=0.01). Conclusion. Concentration-controlled everolimus with reduced CsA results in similar renal function and equivalent efficacy compared with MMF with standard CsA at 12 months after cardiac transplantation.

Changes of peripheral A2A adenosine receptors in chronic heart failure and cardiac transplantation

Peripheral blood mononuclear cells of chronic heart failure (CHF) patients produce great amounts of pro‐inflammatory cytokines, indicating that circulating cells are activated and could mirror changes occurring in inflammatory cells infiltrating the failing heart. Adenosine is a regulatory metabolite acting through four membrane receptors that are linked to adenylyl cyclase: activation of the A2A receptor subtype has been reported to inhibit cytokine release. Changes of the adenosinergic system may play a role in CHF development. Here we report an increase of A2A receptor expression, density, and coupling to adenylyl cyclase in blood circulating cells of CHF patients. A2A receptor up‐regulation was also found in the explanted hearts of these patients, suggesting that changes of peripheral adenosine receptors mirror changes occurring in the disease target organ. In a cohort of patients followed longitudinally after heart transplantation, alterations of peripheral A2A adenosine receptor progressively normalized to control values within 6 months, suggesting that improvement of cardiac performance is accompanied by progressive restoration of a normal adenosinergic system. These results validate the importance of the A2A receptor in human diseases characterized by a marked inflammatory/immune component and suggest that the evaluation of this receptor in peripheral blood cells may be useful for monitoring hemodynamic changes and the efficacy of pharmacological and non‐pharmacological treatments in CHF patients.

Speckle tracking echocardiography as a new technique to evaluate right ventricular function in patients with left ventricular assist device therapy.

BACKGROUND Right ventricular (RV) systolic function has a critical role in determining the clinical outcome and the success of using left ventricular assist devices (LVADs) in patients with refractory heart failure. RV deformation analysis by speckle tracking echocardiography (STE) has recently allowed a deeper analysis of RV longitudinal function. The aim of the study was to observe RV function by STE in patients with advanced heart failure before and after LVAD implantation. METHODS Transthoracic echo Doppler was performed in 10 patients referred for LVAD therapy at baseline and with serial echocardiograms after LVAD implantation. In a sub-group of 4 patients, an echocardiographic evaluation was also made after intra-aortic balloon pump (IABP) support was initiated and before LVAD implantation. All echocardiographic images were analyzed off-line to calculate the free wall RV longitudinal strain (RVLS). RESULTS Three patients who presented the lowest free wall RVLS values at baseline, showed a progressive decline of RVLS after LVAD implant, presenting finally RV failure; however, patients with higher values of RVLS at baseline presented a further and overt increase of strain values in the course of follow-up. The overall performance for the prediction of RV failure after LVAD implant was greatest for free wall RVLS (area under the curve, 0.93). For the sub-group receiving the IABP as an intermediate step, only 2 patients with an increase of RVLS after IABP implantation also showed an increase of RVLS levels, after subsequent LVAD implantation. The 2 patients without an increase of RVLS after IABP implantation also presented with RV failure after LVAD therapy. CONCLUSIONS This study of 10 patients indicates that the new parameter of RVLS, representing RV myocardial deformation, may have important clinical implications for the selection and management of LVAD patients. A large multicenter study is required to confirm these observations and to quantify the clinical significance of changes in RVLS value.

Right Atrial Speckle Tracking Analysis as a Novel Noninvasive Method for Pulmonary Hemodynamics Assessment in Patients with Chronic Systolic Heart Failure

Background: The right atrium (RA) plays multiple roles in the cardiac cycle. The reservoir phase of the RA is a dynamic rather than a static phase of cardiac cycle and RA deformation is dependent on pulmonary pressures exerted on the right ventricle and, therefore, backwards on the RA. The purpose of this study was to assess the accuracy and the clinical applicability of the speckle tracking echocardiography (STE) evaluation of the RA in predicting the invasive systolic pulmonary artery pressure (SPAP) in patients with systolic heart failure (HF) undergoing right heart catheterization (RHC). Methods: Thirty‐one hemodynamically stable, in‐clinic HF patients who were undergoing RHC were included. Doppler echocardiography and RHC catheterization were simultaneously performed. Echocardiographic measures and STE where obtained as peak atrial longitudinal strain (PALS), RA strain rate, and time to peak longitudinal strain (TPLS). RA PALS was inversely correlated with invasively assessed SPAP (r =–0.81; P < 0.001) while RA strain directly correlated with SPAP (r = 0.82; P < 0.001). RA PALS and strain rate retained this correlation even after nitroprusside challenge test (r =–0.81; P < 0.001 and r = 0.91; P < 0.001, respectively). Area under the curve optimal cutoffs for predicting the SPAP > 50 mmHg were for RA PALS 10.3% (AUC:0.93, sensitivity: 100%, specificity: 78%). Conclusion: RA STE showed a significant correlation with pulmonary pressure. RA assessment with STE can predict pulmonary artery hypertension in HF patients. This result is consistent with nitroprusside challenge test. Although RA STE is not routinely used, its evaluation may implement right heart evaluation in HF patients. (Echocardiography 2011;28:658‐664)

Capillary electrophoresis in the evaluation of ischemic injury: simultaneous determination of purine compounds and glutathione.

An understanding of tissue energy metabolism and antioxidant status is of major interest in the field of organ preservation for transplantation. Nucleotide and glutathione are indicators of cell damage occurring during ischemia and reperfusion. A high performance capillary electrophoresis (HPCE) method with UV detection (185 nm) for the simultaneous analysis of intracellular free ribonucleotides, nucleosides, bases and glutathione (oxidized and reduced form) in myocardial tissues is described. The method does not involve thiol derivatization. The separations were carried out in an uncoated fused‐silica capillary, 60 cm long, 52.5 cm to detector, 75 μm ID, with 20 mM Na‐borate buffer, pH 10.00, at 20 kV voltage and reading at 185 nm. Injection was hydrostatic for 12 s and total analysis time was 20 min. The technique enables optimum separation of all the compounds examined and has a resolution similar to that of HPLC analysis, with the advantage of fast simultaneous measurement of cell nucleotide metabolism and redox state, not possible with HPLC.

Predictive role of pretransplant serum CXCL10 for cardiac acute rejection.

Background. The detection of acute rejection in heart transplantation remains an important feature of transplant management, especially in the early phase. Frequent surveillance with endomyocardial biopsy is necessary, even though it is an invasive procedure and carries a certain risk. Hence, noninvasive biomarkers able to predict acute rejection could be a further helpful tool in patient management. The interferon-&ggr;-inducible chemokine CXCL10 is required for initiation and development of graft failure caused by acute or chronic rejection. It has been reported that CXCL10 serum level is predictive of graft loss in kidney graft recipients. In the present study, we investigated whether pretransplant CXCL10 serum level may be a predictive noninvasive biomarker in heart transplant (HTx) recipients, as well. Methods. Sera from 143 patients undergoing orthotopic heart transplantation were collected before surgery and tested for CXCL10 and CCL22 and compared with serum samples from healthy subjects. Results. We found that basal CXCL10 serum levels in HTx recipients were significantly higher than in healthy subjects, whereas no difference was seen in CCL22 levels. Among HTx recipients, CXCL10 serum levels of rejectors were significantly higher than in nonrejectors. Our results showed that CXCL10 was a significant independent risk factor of several variables and had the highest predictive value for early acute heart rejection, with 160 pg/mL cutoff value. Conclusions. In HTx recipients, measurement of pretransplant CXCL10 serum levels could be a clinically useful tool for predicting cardiac acute rejection, especially in the early posttransplant period.