Natriuretic Peptide Processing in Patients with and Without Left Ventricular Dysfunction.

This study aimed to examine the relationship between corin expression and circulating brain natriuretic peptide in patients with left ventricular (LV) dysfunction.Circulating levels of B-type natriuretic peptide (BNP) can be an indicator of LV dysfunction. The 32-amino-acid BNP is cleaved by corin, a cardiac serine protease, from its108-amino-acid pro-brain natriuretic peptide (proBNP) precursor.This study included 25 patients with idiopathic dilated cardiomyopathy (DCMP) and LV dysfunction and 44 heart transplant recipients with normal LV function who underwent diagnostic left and right heart catheterization. Blood samples were used to determine the ratio of plasma proBNP/BNP levels, and LV endomyocardial biopsies were used to determine the expression of NPPB, which encode BNP and corin, respectively, by quantitative reverse transcription-polymerase chain reaction.Patients with DCMP revealed worse hemodynamic profiles and higher plasma proBNP and BNP levels than those of the transplant recipients. Myocardial NPPB expression was higher and CORIN expression was lower in the DCMP patients than in the transplant recipients. CORIN expression significantly correlated with NPPB expression (r = -0.585; P < 0.001), ejection fraction (EF; r = 0.694; P < 0.01), LV end-diastolic pressure (r = -0.373; P < 0.05), and indexed end-diastolic LV volume (r = -0.452; P < 0.001). In addition, the plasma proBNP/BNP levels inversely correlated with the CORIN expression (r = -0.362; P < 0.005).Decreased myocardial CORIN expression and the corresponding higher levels of circulating unprocessed proBNP in DCMP may partly account for the relative BNP resistance observed in patients with LV dysfunction.

α2A-Adrenergic receptor polymorphism potentiates platelet reactivity in patients with stable coronary artery disease carrying the cytochrome P450 2C19*2 genetic variant.

OBJECTIVE: Platelet α2A-adrenergic receptors (ARs) mediate platelet aggregation in response to sympathetic stimulation. The 6.3-kb variant of α2A-AR gene is associated with increased epinephrine-induced platelet aggregation in healthy volunteers. The cytochrome P450 2C19*2 (CYP2C19*2) loss-of-function allele influences P2Y12-mediated platelet inhibition and hence the rate of major adverse cardiovascular events. We assessed the influence of 6.3-kb α2A-AR gene variant on platelet aggregation and its interaction with CYP2C19*2 loss-of-function allele in patients with stable angina on aspirin and clopidogrel (dual antiplatelet therapy). APPROACH AND RESULTS: Aggregation to 5 increasing doses of epinephrine (from 0.156 to 10 µmol/L) was assessed in aggregation units by Multiplate Analyzer and platelet reactivity in P2Y12 reactivity units and % inhibition by VerifyNow P2Y12 assay before percutaneous revascularization. Gene polymorphisms were analyzed with TaqMan Drug Metabolism assay. Of 141 patients, aggregation was higher in 6.3-kb carriers (n=52) when compared with wild types (n=89) at all epinephrine doses (P<0.05) apart from 10 µmol/L (P=0.077). Percentage inhibition was lower (P=0.048) in 6.3-kb α2A-AR carriers. Percentage inhibition was lower (P=0.005) and P2Y12 reactivity units was higher (P=0.012) in CYP2C19*2 allele carriers. Higher P2Y12 reactivity units (P=0.037) and lower percentage inhibition (P=0.009) were observed in carriers of both 6.3-kb α2A-AR variant and CYP2C19*2 allele when compared with wild-type or with either mutation on its own. CONCLUSIONS: The 6.3-kb α2A-AR variant is associated with increased platelet reactivity to epinephrine and has an additive effect along with CYP2C19*2 loss-of-function allele on P2Y12-mediated platelet responses in patients with stable angina on dual antiplatelet therapy.

Characterization of systolic and diastolic function, alongside proteomic profiling, in doxorubicin-induced cardiovascular toxicity in mice.

BACKGROUND: The anthracycline doxorubicin (DOX) is a highly effective anticancer agent, especially in breast cancer and lymphoma. However, DOX can cause cancer therapy-related cardiovascular toxicity (CTR-CVT) in patients during treatment and in survivors. Current diagnostic criteria for CTR-CVT focus mainly on left ventricular systolic dysfunction, but a certain level of damage is required before it can be detected. As diastolic dysfunction often precedes systolic dysfunction, the current study aimed to identify functional and molecular markers of DOX-induced CTR-CVT with a focus on diastolic dysfunction. METHODS: Male C57BL/6J mice were treated with saline or DOX (4 mg/kg, weekly i.p. injection) for 2 and 6 weeks (respectively cumulative dose of 8 and 24 mg/kg) (n = 8 per group at each time point). Cardiovascular function was longitudinally investigated using echocardiography and invasive left ventricular pressure measurements. Subsequently, at both timepoints, myocardial tissue was obtained for proteomics (liquid-chromatography with mass-spectrometry). A cohort of patients with CTR-CVT was used to complement the pre-clinical findings. RESULTS: DOX-induced a reduction in left ventricular ejection fraction from 72 ± 2% to 55 ± 1% after 2 weeks (cumulative 8 mg/kg DOX). Diastolic dysfunction was demonstrated as prolonged relaxation (increased tau) and heart failure was evident from pulmonary edema after 6 weeks (cumulative 24 mg/kg DOX). Myocardial proteomic analysis revealed an increased expression of 12 proteins at week 6, with notable upregulation of SERPINA3N in the DOX-treated animals. The human ortholog SERPINA3 has previously been suggested as a marker in CTR-CVT. Upregulation of SERPINA3N was confirmed by western blot, immunohistochemistry, and qPCR in murine hearts. Thereby, SERPINA3N was most abundant in the endothelial cells. In patients, circulating SERPINA3 was increased in plasma of CTR-CVT patients but not in cardiac biopsies. CONCLUSION: We showed that mice develop heart failure with impaired systolic and diastolic function as result of DOX treatment. Additionally, we could identify increased SERPINA3 levels in the mice as well as patients with DOX-induced CVT and demonstrated expression of SERPINA3 in the heart itself, suggesting that SERPINA3 could serve as a novel biomarker.

Coronary Microvascular Dysfunction in Patients With Heart Failure: Characterization of Patterns in HFrEF Versus HFpEF.

BACKGROUND: Coronary microvascular dysfunction (CMD) is involved in heart failure (HF) onset and progression, independently of HF phenotype and obstructive coronary artery disease. Invasive assessment of CMD might provide insights into phenotyping and prognosis of patients with HF. We aimed to assess absolute coronary flow, absolute microvascular resistance, myocardial perfusion, coronary flow reserve, and microvascular resistance reserve in patients with HF with preserved ejection fraction and HF with reduced ejection fraction (HFrEF). METHODS: Single-center, prospective study of 56 consecutive patients with de novo HF with nonobstructive coronary artery disease divided into HF with preserved ejection fraction (n=21) and HFrEF (n=35). CMD was invasively assessed by continuous intracoronary thermodilution and defined as coronary flow reserve <2.5. Left ventricular and left anterior descending artery-related myocardial mass was quantified by echocardiography and coronary computed tomography angiography. Myocardial perfusion (mL/min per g) was calculated as the ratio between absolute coronary flow and left anterior descending artery-related mass. RESULTS: Patients with HFrEF showed a higher left ventricular and left anterior descending artery-related myocardial mass compared with HF with preserved ejection fraction (P<0.010). Overall, 52% of the study population had CMD, with a similar prevalence between the 2 groups. In HFrEF, CMD was characterized by lower absolute microvascular resistance and higher absolute coronary flow at rest (functional CMD; P=0.002). CMD was an independent predictor of a lower rate of left ventricular reverse remodeling at follow-up. In patients with HF with preserved ejection fraction, CMD was mainly due to higher absolute microvascular resistance and lower absolute coronary flow during hyperemia (structural CMD; P≤0.030). CONCLUSIONS: Continuous intracoronary thermodilution allows the definition and characterization of patterns with distinct CMD in patients with HF and could identify patients with HFrEF with a higher rate of left ventricular reverse remodeling at follow-up.

Myocardial sodium-glucose cotransporter 2 expression and cardiac remodelling in patients with severe aortic stenosis: The BIO-AS study.

AIM: Cardiac remodelling plays a major role in the prognosis of patients with aortic stenosis (AS) and could impact the benefits of aortic valve replacement. Our study aimed to evaluate the expression of sodium-glucose cotransporter 2 (SGLT2) gene and protein in patients with severe AS stratified in high gradient (HG) and low flow-low gradient (LF-LG) AS and its association with cardiac functional impairments. METHODS AND RESULTS: Gene expression and protein levels of main biomarkers of cardiac fibrosis (galectin-3, sST2, serpin-4, procollagen type I amino-terminal peptide, procollagen type I carboxy-terminal propeptide, collagen, transforming growth factor [TGF]-ß), inflammation (growth differentiation factor-15, interleukin-6, nuclear factor-κB [NF-κB]), oxidative stress (superoxide dismutase 1 [SOD1] and 2 [SOD2]), and cardiac metabolism (sodium-hydrogen exchanger, peroxisome proliferator-activated receptor [PPAR]-α, PPAR-γ, glucose transporter 1 [GLUT1] and 4 [GLUT4]) were evaluated in blood samples and heart biopsies of 45 patients with AS. Our study showed SGLT2 gene and protein hyper-expression in patients with LF-LG AS, compared to controls and HG AS (p < 0.05). These differences remained significant even after adjusting for age, gender, body mass index, history of diabetes mellitus, arterial hypertension, and coronary artery disease. SGLT2 gene expression was positively correlated with: (i) TGF-ß (r = 0.72, p < 0.001) and collagen (r = 0.73, p < 0.001) as markers of fibrosis; (ii) NF-κB (r = 0.36, p < 0.01) and myocardial interleukin-6 (r = 0.68, p < 0.001) as markers of inflammation: (iii) SOD2 (r = -0.38, p < 0.006) as a marker of oxidative stress; (iv) GLUT4 (r = 0.33, p < 0.02) and PPAR-α (r = 0.36, p < 0.01) as markers of cardiac metabolism. CONCLUSION: In patients with LF-LG AS, SGLT2 gene and protein were hyper-expressed in cardiomyocytes and associated with myocardial fibrosis, inflammation, and oxidative stress.

Differential expression of epigenetic modifiers in early and late cardiotoxic heart failure reveals DNA methylation as a key regulator of cardiotoxicity.

Background: Anthracycline-induced cardiotoxicity is a well-known serious clinical entity. However, detailed mechanistic insights on how short-term administration leads to late and long-lasting cardiotoxicity, are still largely undiscovered. We hypothesize that chemotherapy provokes a memory effect at the level of epigenomic DNA modifications which subsequently lead to cardiotoxicity even years after cessation of chemotherapy. Methods: We explored the temporal evolution of epigenetic modifiers in early and late cardiotoxicity due to anthracyclines by means of RNA-sequencing of human endomyocardial left ventricular biopsies and mass spectrometry of genomic DNA. Based on these findings, validation of differentially regulated genes was obtained by performing RT-qPCR. Finally, a proof-of-concept in vitro mechanistic study was performed to dissect some of the mechanistic aspects of epigenetic memory in anthracycline-induced cardiotoxicity. Results: Correlation of gene expression between late and early onset cardiotoxicity revealed an R 2 value of 0.98, demonstrating a total of 369 differentially expressed genes (DEGs, FDR < 0.05). of which 72% (n = 266) were upregulated, and 28% of genes, (n = 103) downregulated in later as compared to earlier onset cardiotoxicity. Gene ontology analysis showed significant enrichment of genes involved in methyl-CpG DNA binding, chromatin remodeling and regulation of transcription and positive regulation of apoptosis. Differential mRNA expression of genes involved in DNA methylation metabolism were confirmed by RT-qPCR in endomyocardial biopsies. In a larger biopsy cohort, it was shown that Tet2 was more abundantly expressed in cardiotoxicity biopsies vs. control biopsies and vs. non-ischemic cardiomyopathy patients. Moreover, an in vitro study was performed: following short-term doxorubicin treatment, H9c2 cells were cultured and passaged once they reached a confluency of 70%-80%. When compared to vehicle-only treated cells, in doxorubicin-treated cells, three weeks after short term treatment, Nppa, Nppb, Tet1/2 and other genes involved in active DNA demethylation were markedly upregulated. These alterations coincided with a loss of DNA methylation and a gain in hydroxymethylation, reflecting the epigenetic changes seen in the endomyocardial biopsies. Conclusions: Short-term administration of anthracyclines provokes long-lasting epigenetic modifications in cardiomyocytes both in vivo and in vitro, which explain in part the time lapse between the use of chemotherapy and the development of cardiotoxicity and, eventually, heart failure.

Doxorubicin-induced cardiovascular toxicity: a longitudinal evaluation of functional and molecular markers.

AIMS: Apart from cardiotoxicity, the chemotherapeutic doxorubicin (DOX) induces vascular toxicity, represented by arterial stiffness and endothelial dysfunction. Both parameters are of interest for cardiovascular risk stratification as they are independent predictors of future cardiovascular events in the general population. However, the time course of DOX-induced cardiovascular toxicity remains unclear. Moreover, current biomarkers for cardiovascular toxicity prove insufficient. Here, we longitudinally evaluated functional and molecular markers of DOX-induced cardiovascular toxicity in a murine model. Molecular markers were further validated in patient plasma. METHODS AND RESULTS: DOX (4 mg/kg) or saline (vehicle) was administered intra-peritoneally to young, male mice weekly for 6 weeks. In vivo cardiovascular function and ex vivo arterial stiffness and vascular reactivity were evaluated at baseline, during DOX therapy (Weeks 2 and 4) and after therapy cessation (Weeks 6, 9, and 15). Left ventricular ejection fraction (LVEF) declined from Week 4 in the DOX group. DOX increased arterial stiffness in vivo and ex vivo at Week 2, which reverted thereafter. Importantly, DOX-induced arterial stiffness preceded reduced LVEF. Further, DOX impaired endothelium-dependent vasodilation at Weeks 2 and 6, which recovered at Weeks 9 and 15. Conversely, contraction with phenylephrine was consistently higher in the DOX-treated group. Furthermore, proteomic analysis on aortic tissue identified increased thrombospondin-1 (THBS1) and alpha-1-antichymotrypsin (SERPINA3) at Weeks 2 and 6. Up-regulated THBS1 and SERPINA3 persisted during follow-up. Finally, THBS1 and SERPINA3 were quantified in plasma of patients. Cancer survivors with anthracycline-induced cardiotoxicity (AICT; LVEF < 50%) showed elevated THBS1 and SERPINA3 levels compared with age-matched control patients (LVEF ≥ 60%). CONCLUSIONS: DOX increased arterial stiffness and impaired endothelial function, which both preceded reduced LVEF. Vascular dysfunction restored after DOX therapy cessation, whereas cardiac dysfunction persisted. Further, we identified SERPINA3 and THBS1 as promising biomarkers of DOX-induced cardiovascular toxicity, which were confirmed in AICT patients.

T Cell and Antibody Response Following Double Dose of BNT162b2 mRNA Vaccine in SARS-CoV-2 Naïve Heart Transplant Recipients.

Introduction: Preliminary studies have suggested a low post-vaccination antibody response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in heart transplant(HTx)recipients. Although many studies have focused on the role of antibodies in vaccine-induced protection against SARS-CoV-2, the role of T cell immunity is less well characterized. To date, data regarding seroconversion and T cell response after mRNA SARS-CoV-2 vaccination in patients undergoing HTx are scarce. Therefore, the present study aimed to assess the specific memory humoral and cellular responses after two doses of the BNT162b2 vaccine in HTx recipients. Methods: Blood was drawn from heart transplant (HTx) recipients at two pre-specified time points after the first and second vaccine doses to measure both the anti-SARS-CoV-2 antibody response against the spike protein and the SARS-CoV-2-reactive T cell response. Results: Our study included 34 SARS-CoV-2 naïve HTx recipients (mean age, 61 ± 11 years). The mean time from transplantation to the first vaccine dose is 10 ± 10 years. Subgroup analysis (n = 21) demonstrated that after the first vaccine dose, only 14% had antibodies and 19% had a SARS-CoV-2-reactive T-cell response, which increased to 41% and 53%, respectively, after the second dose. Interestingly, 20% of patients with no antibodies after the second dose still had a positive SARS-CoV-2-reactive T cell response. The percentage of patients with positive S-IgG antibody titers was significantly higher 5 years after transplantation (18% 0-5 years post-TX vs. 65% 5 years post-TX, p = 0.013). Similarly, 5 years after heart transplantation, the percentage of patients with a T cell response was significantly higher (35% 0-5 years post-TX vs. 71% 5 years post-TX, p = 0.030). Conclusions: In SARS-CoV-2 naïve HTx recipients, post-vaccination antibody titers but also SARS-CoV-2 specific T cell response are low. Therefore, the protection from SARS-CoV-2 that is generally attributed to vaccination should be regarded with caution in HTx recipients.

Hemodynamic Response to Acute Volume Load and Endomyocardial NO-synthase Gene Expression in Heart Transplant Recipients.

A pulmonary capillary wedge pressure (PCWP) >18 mm Hg following volume load has been proposed as a partition value for the detection of heart failure with preserved ejection fraction. As hemodynamic changes in filling pressures (FP) have been attributed to a nitric oxide (NO)-mediated rightward shift of the pressure-volume relationship, we investigated the hemodynamic response to volume load in heart transplant recipients (HTx) and examined the role of inducible NO synthase (iNOS) gene expression on diastolic function changes. Methods: In 36 HTx, FPs were measured before and after volume load, following which Starling curves were constructed using PCWP and cardiac index (CI). Patients were categorized into those with normal (group A, n = 21) and abnormal hemodynamics (group B, n = 15, PCWP >15 mm Hg at rest or >18 mm Hg following volume load). For the establishment of the potential role of NO, endomyocardial iNOS gene expression level was measured. Results: Except for PCWP (P < 0.001) and mean pulmonary artery pressure (P < 0.001) no differences in age, baseline characteristics, and ejection fraction were observed between both groups, and volume load significantly increased PCWP in both groups (group A: P < 0.001 and group B: P < 0.001) without any change in heart rate. Interestingly, volume load significantly increased CI in group A (P < 0.001) but not in group B (P = 0.654), and the Starling curves revealed a higher CI at any given PCWP in group A together with significantly higher iNOS gene expression (P = 0.009). Conclusions: In HTx, volume load increases FP and unmasks the presence of left ventricular diastolic dysfunction. Interestingly, following saline load group B shows a blunted Starling response, with higher PCWP and lack of CI increase at any given PCWP. The higher iNOS gene expression level in group A suggests a potential role of NO as mediator of diastolic function.

Circulating SERPINA3 improves prognostic stratification in patients with a de novo or worsened heart failure.

AIMS: We investigated the prognostic relevance of serpin peptidase inhibitor, clade A member 3 (SERPINA3) in patients admitted with a de novo or worsened heart failure (HF). METHODS AND RESULTS: In the first stage, 83 HF-related left ventricular (LV) transcripts were examined in patients with congestive cardiomyopathy (CCMP, n = 44) who died within 5 years and compared with age-matched and haemodynamically matched CCMP survivors (n = 39) and controls with normal LV function (n = 17). Among 14 differentially expressed transcripts, myocardial gene and circulating SERPINA3 levels were up-regulated in non-survivors vs. survivors (2.40 ± 3.66 vs. 0.36 ± 0.22 units, P < 0.01 and 334.7 ± 138.7 vs. 228.2 ± 83.1 µg/mL, P < 0.01, respectively). While no significant transmyocardial gradient was detected, cytokine stimulation of human endothelial cells induced SERPINA3 secretion. In an independent validation cohort with a de novo or worsened HF (n = 387), circulating SERPINA3 levels > 316 µg/mL were associated with increased all-cause mortality {hazard ratio [HR] [95% confidence interval (CI)]: 2.4 [1.5-3.9], P = 0.0002} and its composite with unplanned cardiovascular readmission [HR (95% CI): 2.0 (1.2-3.3), P = 0.004]. Patients with elevated SERPINA3 levels and elevated either N-terminal pro brain natriuretic peptide or ST2 showed worse freedom from both endpoints. In a multivariate analysis, including established clinical risk factors, SERPINA3 remained independent predictor of all-cause mortality together with age, gender, ST2, glomerular filtration, and pulmonary capillary wedge pressure. CONCLUSION: In patients with a de novo or worsened HF, increased SERPINA3 levels > 316 µg/mL are associated with increased mortality or unplanned cardiac readmission. Elevated SERPINA3 levels on top of established clinical predictors appear to identify a subgroup of HF patients at higher mortality risk. Prospective studies should further validate its value in prognostic stratification of HF.

Low MicroRNA-126 Levels in Right Ventricular Endomyocardial Biopsies Coincide With Cardiac Allograft Vasculopathy in Heart Transplant Patients.

Endothelium-enriched microRNAs (miRs) are involved in the development of cardiac allograft vasculopathy (CAV). Recently, serum-derived miR-126-3p and -5p, known endothelial microRNAs with a crucial function in angiogenesis and re-endothelialization, provided additional predictive power for cardiac allograft vasculopathy in addition to clinical predictors. However, their myocardial expression in and relationship with CAV are still unknown. Our study aim was to investigate the expression of endomyocardial microRNA-126-3p and microRNA-126-5p levels in heart transplant recipients and their relationship with allograft vasculopathy. METHODS: We studied 39 heart transplant recipients, 21 with proven allograft vasculopathy (CAV+) and 18 without allograft vasculopathy (CAV-) with serial coronary angiograms. Additionally, 8 patients with end-stage native coronary artery disease (CAD) were added to the study to investigate disease specificity of the microRNA signature. The mRNA levels of miR-126-3p and miR-126-5p were determined by qRT-PCR in the right ventricular endomyocardial biopsies obtained at baseline and during routine follow-up. RESULTS: MiR-126-3p levels were significantly lower in the CAV+ group compared to the CAV- group at follow-up, while miR-126-5p levels were unaltered. This was in stark contrast to native CAD patients in whom miR-126-3p and -5p levels were significantly higher. qPCR levels of miR-126 targets are differentially regulated in CAV versus ischemic cardiomyopathy and are influenced by the administration of immunosuppressive agents in endothelial cells. CONCLUSIONS: Our data provide evidence for a distinct microRNA signature in heart transplantation patients with allograft vasculopathy. In contrast to CAD patients, lower miR-126-3p levels coincide with the development of cardiac allograft vasculopathy. Further studies in a larger patient population are warranted to determine if the serial measurement of myocardial microRNA-126 products could help in risk assessment and early detection of CAV.

Procedural microvascular activation in long lesions treated with bioresorbable vascular scaffolds or everolimus-eluting stents: the PROACTIVE trial.

AIMS: Significant platelet activation after long stented coronary segments has been associated with periprocedural microvascular impairment and myonecrosis. In long lesions treated either with an everolimus-eluting bioresorbable vascular scaffold (BVS) or an everolimus-eluting stent (EES), we aimed to investigate (a) procedure-related microvascular impairment, and (b) the relationship of platelet activation with microvascular function and related myonecrosis. METHODS AND RESULTS: Patients (n=66) undergoing elective percutaneous coronary intervention (PCI) in long lesions were randomised 1:1 to either BVS or EES. The primary endpoint was the difference between groups in changes of pressure-derived corrected index of microvascular resistance (cIMR) after PCI. Periprocedural myonecrosis was assessed by high-sensitivity cardiac troponin T (hs-cTnT), platelet reactivity by high-sensitivity adenosine diphosphate (hs-ADP)-induced platelet reactivity with the Multiplate Analyzer. Post-dilatation was more frequent in the BVS group, with consequent longer procedure time. A significant difference was observed between the two groups in the primary endpoint of ΔcIMR (p=0.04). hs-ADP was not different between the groups at different time points. hs-cTnT significantly increased after PCI, without difference between the groups. CONCLUSIONS: In long lesions, BVS implantation is associated with significant acute reduction in IMR as compared with EES, with no significant interaction with platelet reactivity or periprocedural myonecrosis.

Endomyocardial upregulation of beta1 adrenoreceptor gene expression and myocardial contractile reserve following cardiac resynchronization therapy.

BACKGROUND: Congestive heart failure (CHF) is associated with a blunted force-frequency relation (FFR) and myocardial contractile reserve (MCR) partially from a downregulation of beta1-adrenoreceptors (beta1-AR). We investigated whether acute and chronic cardiac resynchronization therapy (CRT) was capable of reversing the blunted FFR and MCR and if this was associated with upregulation of beta1-AR. METHODS AND RESULTS: Left ventricle dP/dtmax was invasively measured in 10 CHF patients (New York Heart Association class > or =3; ejection fraction <25%) during incremental dual chamber (DDD)-CRT pacing at 70, 90, 110, and 130 beats/min, with and without continuous infusion of intravenous dobutamine, immediately after CRT implantation (BL) and 4 months later (FU). In a subgroup of 5 patients, serial left ventricle beta1 and beta2-AR gene expression was measured using reverse transcriptase-polymerase chain reaction. Four months after the initiation of resynchronization therapy, DDD-CRT pacing results in a significant upward shift of the heart rate versus LV dP/dtmax relationship (P < .01) with force frequency amplification as evidenced by the steeper slope of the force frequency response (P = .04). Infusion of dobutamine recruits myocardial contractile reserve and increases the heart rate versus LV dP/dtmax relationship at BL and at FU (both P < .05). However, only at follow-up was an additional force frequency amplification noticed (P < .05) during dobutamine infusion. This observation was paralleled by a significant upregulation of beta1-AR gene expression (P = .02). CONCLUSIONS: Chronic CRT is associated with a partial restoration of the FFR and with a recruitment in myocardial contractile reserve, which is paralleled by upregulation of beta1-AR.

Early and late effects of cardiac resynchronization therapy on force-frequency relation and contractility regulating gene expression in heart failure patients.

BACKGROUND: Heart failure is associated with a reduction in left ventricular (LV) contractility as evidenced by a blunted force-frequency response (FFR) and downregulation of contractility regulating genes. OBJECTIVE: This study sought to investigate whether cardiac resynchronization therapy (CRT) is capable of reversing the blunted FFR and the downregulation of contractility regulating genes. METHODS: Twenty heart failure patients underwent echocardiographic examination during incremental AAI and DDD-CRT pacing at 70, 90, and 110 beats/min, immediately after and 4 months after CRT implantation. The FFR was determined from the ratio of the LV systolic pressure/end systolic volume index at given heart rate. In a subgroup of 6 patients with idiopathic dilated cardiomyopathy, serial LV dP/dtmax was invasively measured during both pacing modes and serial LV endomyocardial biopsies were taken to measure sarcoplasmatic reticulum calcium ATPase 2alpha (SERCA2alpha), phospholamban (PLN), sarcolemmal sodium calcium exchanger (NCX), beta1-adrenoreceptor (beta1-AR), and apelin (APL) gene expression using reverse-transcriptase polymerase chain reaction. RESULTS: Acutely, DDD-CRT pacing was associated with a decrease in dyssynchrony (P <.01) and increase in diastolic filling time (P <.01) at all heart rates paralleled by an upward shift of the FFR (P <.01) without force-frequency amplification. A greater upward shift of the FFR was noticed during DDD-CRT as compared with AAI (P <.01) after 4 months. In addition, CRT was associated with a significant force-frequency amplification at follow-up as evidenced from the steeper slope of the FFR relationship (P = .039). This was associated with a significant upregulation of SERCA2alpha P = .01), PLN (P = .01), their ratio (P = .01), ratio of SERCA/NCX (P = .02), beta1-AR (P = .03), and APL (P = .01) mRNA levels. CONCLUSION: CRT is associated with an acute upward shift in the FFR without force-frequency amplification related to restored synchronicity and increased filling time of the LV. Only chronic CRT is associated with force-frequency amplification in parallel to upregulation of contractility regulating genes.

Differences in von Willebrand factor function in type 2A von Willebrand disease and left ventricular assist device-induced acquired von Willebrand syndrome.

BACKGROUND: Patients with von Willebrand disease (VWD) type 2A or acquired von Willebrand syndrome (aVWS) as a consequence of implantation of left ventricular assist devices (LVAD) are both characterized by a loss of von Willebrand factor (VWF) function. Loss of VWF function is however more severe in VWD type 2A than in LVAD patients. OBJECTIVES: To compare VWF function in patients with VWD type 2A and LVAD-induced aVWS to highlight the differences in VWF activity and to stress the importance of VWF multimer analysis for correct diagnosis of aVWS in LVAD patients. PATIENTS/METHODS: Plasma samples from nine VWD type 2A, nine LVAD patients, and 20 healthy donors (HD) were analyzed for VWF function (VWF:CB/VWF:Ag and VWF:RCo/VWF:Ag) and loss of high molecular weight (HMW) VWF multimers. RESULTS: A severely impaired VWF function was indeed confirmed in all VWD 2A patients. HMW VWF multimers were severely reduced compared to HD (0% [0, 12.29] vs 34.19% [31.68, 38.88] for HD, P < 0.001) and this loss was reflected by VWF:CB/VWF:Ag and VWF:RCo/VWF:Ag ratios <0.7. In contrast, VWF function was less affected in LVAD patients. Although HMW VWF multimers were reduced in all patients (20.31% [15.84, 21.71], vs 34.19% [31.68, 38.88] for HD, P < 0.001), six out of nine LVAD patients had normal VWF:CB/VWF:Ag or VWF:RCo/VWF:Ag ratios (>0.7). CONCLUSIONS: VWF:CB/VWF:Ag or VWF:RCo/VWF:Ag analysis allows detection of impaired VWF function in VWD type 2A but not always in LVAD-induced aVWS patients. In contrast, VWF multimeric analysis allows detection of the loss of HMW VWF multimers in both groups of patients. Hence, performing VWF multimer analysis is crucial to detect aVWS in LVAD patients.

T2238C Atrial Natriuretic Peptide Gene Variant and the Response to Antiplatelet Therapy in Stable Ischemic Heart Disease Patients.

The T2238C variant of the ANP gene is associated with higher risk of major cardiovascular events. The purpose of this study is to investigate if this polymorphism influences the response to antiplatelet agents and it is responsible of increased platelet reactivity, thus contributing to the adverse outcome. In patients undergoing elective percutaneous coronary intervention (PCI), loaded with antiplatelets, blood samples were withdrawn for genotyping, platelet reactivity assessment and for troponin T measurement to investigate the association between the polymorphism with residual platelet reactivity and with the incidence of PPMI. No significant differences in platelet reactivity nor in PPMI incidence were observed between groups. Nevertheless, higher ARU, PRU, and % PI were detected in diabetic patients, with PRU significantly higher in carriers versus non-carriers. We observed increased residual platelet reactivity exclusively in diabetic carriers of the T2238C variant undergoing elective PCI, suggesting the need of a more effective platelet inhibition in this category of patients.

Time-dependent effects on coronary remodeling and epicardial conductance after intracoronary injection of enriched hematopoietic bone marrow stem cells in patients with previous myocardial infarction.

Bone marrow (BM) cells may interact with coronary endothelium and modulate coronary atherosclerosis. We investigated the time course of coronary luminal loss and changes in conductance after intracoronary injection of enriched hematopoietic BM stem cells in patients with previous myocardial infarction (MI). Among 24 patients with acute MI, 13 were randomized to early (<7 days) and 11 to late (4 months) intracoronary injection of CD133+ cells after the infarction. Segmental quantitative coronary angiography and fractional flow reserve (FFR) measurements of the infarct-related (IR) artery (A) and contralateral artery (control) were performed. In the early group, at 4 months, cumulative luminal loss (LL) of the minimal luminal diameter (MLD) of the IRA distal to the stented segment was -0.39 (-0.51-0.10) mm (p < 0.05 vs. control). There was no further change in LL between 4 and 8 months [-0.09 (-0.26-0.15) mm]. In parallel, FFR decreased at 4 months [-0.16 (-0.26-0.001), p < 0.05 vs. control] but slightly increased from 4 to 8 months follow-up [+0.05 (-0.10-0.09)]. In the late group, LL of the MLD of the IRA distal to the stented segments was -0.12 (-0.47-0.07) mm (NS vs. control) at 4 months and further -0.07 (-0.25-0.05) mm (NS) between 4 and 8 months. At 8 months, the total LL of the MLD in the early and late group was only slightly higher compared to control [-0.34 (-0.48--0.16), -0.36 (-0.69--0.09), and -0.12 (-0.39-0.05) mm, respectively, NS]. Early intracoronary administration of hematopoietic BM stem cells in patients with previous MI is associated with accelerated luminal loss and reduced conductance of the infarct-related artery.

Relationship of asymmetric dimethylarginine (ADMA) with extent and functional severity of coronary atherosclerosis.

BACKGROUND: Elevated serum levels of asymmetric dimethylarginine (ADMA) are associated with endothelial dysfunction and atherogenesis. In patients with suspected coronary artery disease (CAD), we assessed the correlation of serum ADMA levels with extent and functional significance of coronary atherosclerosis. METHODS: We enrolled 281 patients with suspected CAD undergoing coronary angiogram. Angiographic CAD severity was evaluated by Bogaty score. In patients with angiographic evidence of at least one intermediate coronary stenosis (≥50% diameter stenosis), functional significance was assessed by fractional flow reserve (FFR). Blood samples were collected in all patients prior to coronary angiography for measurement of serum ADMA levels. RESULTS: We observed across tertiles of ADMA levels increasingly higher values of both Stenosis Score (2.25±1.70 vs. 2.89±1.99 vs. 2.95±1.82, p=0.016) and Extent Index (0.52±0.32 vs. 0.61±0.39 vs. 0.72±0.47, p=0.003). The association between ADMA levels and Extent Index remained significant after multivariate adjustment (p=0.005). Patients with FFR ≤0.80 in at least one vessel (n=113) had significantly higher ADMA levels compared with patients without functionally significant CAD (0.51 [0.43-0.64] vs. 0.46 [0.39-0.58]µmol/L, p=0.005). Serum ADMA levels were independent predictors of abnormal FFR after adjustment for extent score (odds ratio 7.35, 95% confidence interval 1.05-56.76, p=0.046). CONCLUSIONS: Serum ADMA levels are independent predictors of coronary atherosclerosis extent and functional significance of CAD.

Monocyte-platelets aggregates as cellular biomarker of endothelium-dependent coronary vasomotor dysfunction in patients with coronary artery disease.

Monocyte-platelet aggregates (MPA) are increased in patients with acute coronary syndrome. We investigated whether MPA are associated with the presence of functionally significant coronary stenoses or with coronary arterial endothelial dysfunction. One hundred forty five patients undergoing elective coronary angiography were prospectively enrolled. Functional significance of coronary stenosis was assessed by fractional flow reserve (FFR). Thirty randomly selected patients underwent pacing protocol to evaluate Coronary endothelium-dependent vasomotor function (CVF). Whole blood was drawn to evaluate MPA. In patients with FFR ≤ 0.8 (FFRpos, n = 75), MPA did not significantly differ from FFR >0.8 patients (FFRneg, n = 70) (38.1% [25.7-56.6] vs. 34.0% [20.5-49.9], p = 0.08). CVF was similar in FFRpos and FFRneg patients (percent vessel diameter change, %VDC = 7.19 % [6.01-10.9] vs. 8.0 % [0.81-9.80], p = 0.78). Yet, patients with abnormal CVF showed higher MPA as compared to patients with preserved CVF (28.3% [28.8-53.4] vs. 20.5 % [17.0-32.9], p = 0.01). Moreover, MPA was inversely correlated with %VDC (R2 = 0.26, p < 0.01). MPA levels are significantly higher in patients with abnormal coronary vasomotor function regardless of the presence of functionally significant coronary stenosis.