Emily J. Tsai

Detection of Cardiac Allograft Rejection and Response to Immunosuppressive Therapy With Peripheral Blood Gene Expression

Background—Assessment of gene expression in peripheral blood may provide a noninvasive screening test for allograft rejection. We hypothesized that changes in peripheral blood expression profiles would correlate with biopsy-proven rejection and would resolve after treatment of rejection episodes. Methods and Results—We performed a case-control study nested within a cohort of 189 cardiac transplant patients who had blood samples obtained during endomyocardial biopsy (EMB). Using Affymetrix HU133A microarrays, we analyzed whole-blood expression profiles from 3 groups: (1) control samples with negative EMB (n=7); (2) samples obtained during rejection (at least International Society for Heart and Lung Transplantation grade 3A; n=7); and (3) samples obtained after rejection, after treatment and normalization of the EMB (n=7). We identified 91 transcripts differentially expressed in rejection compared with control (false discovery rate <0.10). In postrejection samples, 98% of transcripts returned toward control levels, displaying an intermediate expression profile for patients with treated rejection (P<0.0001). Cluster analysis of the 40 transcripts with >25% change in expression levels during rejection demonstrated good discrimination between control and rejection samples and verified the intermediate expression profile of postrejection samples. Quantitative real-time polymerase chain reaction confirmed significant differential expression for the predictive markers CFLAR and SOD2 (UniGene ID No. 355724 and No. 384944). Conclusions—These data demonstrate that peripheral blood expression profiles correlate with biopsy-proven allograft rejection. Intermediate expression profiles of treated rejection suggest persistent immune activation despite normalization of the EMB. If validated in larger studies, expression profiling may prove to be a more sensitive screening test for allograft rejection than EMB.

2013 ACCF/AHA Guideline for the Management of Heart Failure

Jeffrey L. Anderson, MD, FACC, FAHA, Chair; Alice K. Jacobs, MD, FACC, FAHA, Immediate Past Chair[‡‡][1]; Jonathan L. Halperin, MD, FACC, FAHA, Chair-Elect; Nancy M. Albert, PhD, CCNS, CCRN, FAHA; Biykem Bozkurt, MD, PhD, FACC, FAHA; Ralph G. Brindis, MD, MPH, MACC; Mark A. Creager, MD, FACC,

Cardiac Fibroblast Glycogen Synthase Kinase-3β Regulates Ventricular Remodeling and Dysfunction in Ischemic Heart

Background— Myocardial infarction–induced remodeling includes chamber dilatation, contractile dysfunction, and fibrosis. Of these, fibrosis is the least understood. After myocardial infarction, activated cardiac fibroblasts deposit extracellular matrix. Current therapies to prevent fibrosis are inadequate, and new molecular targets are needed. Methods and Results— Herein we report that glycogen synthase kinase-3&bgr; (GSK-3&bgr;) is phosphorylated (inhibited) in fibrotic tissues from ischemic human and mouse heart. Using 2 fibroblast-specific GSK-3&bgr; knockout mouse models, we show that deletion of GSK-3&bgr; in cardiac fibroblasts leads to fibrogenesis, left ventricular dysfunction, and excessive scarring in the ischemic heart. Deletion of GSK-3&bgr; induces a profibrotic myofibroblast phenotype in isolated cardiac fibroblasts, in post–myocardial infarction hearts, and in mouse embryonic fibroblasts deleted for GSK-3&bgr;. Mechanistically, GSK-3&bgr; inhibits profibrotic transforming growth factor-&bgr;1/SMAD-3 signaling via interactions with SMAD-3. Moreover, deletion of GSK-3&bgr; resulted in the significant increase of SMAD-3 transcriptional activity. This pathway is central to the pathology because a small-molecule inhibitor of SMAD-3 largely prevented fibrosis and limited left ventricular remodeling. Conclusions— These studies support targeting GSK-3&bgr; in myocardial fibrotic disorders and establish critical roles of cardiac fibroblasts in remodeling and ventricular dysfunction.

Volume overload induces differential spatiotemporal regulation of myocardial soluble guanylyl cyclase in eccentric hypertrophy and heart failure

Nitric oxide activation of soluble guanylyl cyclase (sGC) blunts the cardiac stress response, including cardiomyocyte hypertrophy. In the concentric hypertrophied heart, oxidation and re-localization of myocardial sGC diminish cyclase activity, thus aggravating depressed nitric oxide-cyclic guanosine monophosphate (NO-cGMP) signaling in the pressure-overloaded failing heart. Here, we hypothesized that volume-overload differentially disrupts myocardial sGC activity during early compensated and late decompensated stages of eccentric hypertrophy. To this end, we studied the expression, redox state, subcellular localization, and activity of sGC in the left ventricle of dogs subjected to chordal rupture-induced mitral regurgitation (MR). Unoperated dogs were used as Controls. Animals were studied at 4weeks and 12months post chordal rupture, corresponding with early (4wkMR) and late stages (12moMR) of eccentric hypertrophy. We found that the sGC heterodimer subunits relocalized away from caveolae-enriched lipid raft microdomains at different stages; sGCβ1 at 4wkMR, followed by sGCα1 at 12moMR. Moreover, expression of both sGC subunits fell at 12moMR. Using the heme-dependent NO donor DEA/NO and NO-/heme-independent sGC activator BAY 60-2770, we determined the redox state and inducible activity of sGC in the myocardium, within caveolae and non-lipid raft microdomains. sGC was oxidized in non-lipid raft microdomains at 4wkMR and 12moMR. While overall DEA/NO-responsiveness remained intact in MR hearts, DEA/NO responsiveness of sGC in non-lipid raft microdomains was depressed at 12moMR. Caveolae-localization protected sGC against oxidation. Further studies revealed that these modifications of sGC were also reflected in caveolae-localized cGMP-dependent protein kinase (PKG) and MAPK signaling. In MR hearts, PKG-mediated phosphorylation of vasodilator-stimulated phosphoprotein (VASP) disappeared from caveolae whereas caveolae-localization of phosphorylated ERK5 increased. These findings show that differential oxidation, re-localization, and expression of sGC subunits distinguish eccentric from concentric hypertrophy as well as compensated from decompensated heart failure.

Vasopressin Antagonists for Patients With Acute Heart Failure: Interpreting New Clinical and Translational Data

Elevated levels of arginine vasopressin (AVP) are associated with a worse prognosis and the development of hyponatremia in patients with heart failure (HF). This observation led to the development of AVP receptor antagonists for the treatment of HF patients. Although AVP receptor antagonists increase serum sodium, their overall benefits in patients with HF have been at best modest, and recent data raise concerns about their safety in patients with acute HF and low serum sodium.

Cardiac Fibroblast GSK-3β Regulates Ventricular Remodeling and Dysfunction in Ischemic Heart

Background— Myocardial infarction–induced remodeling includes chamber dilatation, contractile dysfunction, and fibrosis. Of these, fibrosis is the least understood. After myocardial infarction, activated cardiac fibroblasts deposit extracellular matrix. Current therapies to prevent fibrosis are inadequate, and new molecular targets are needed. Methods and Results— Herein we report that glycogen synthase kinase-3&bgr; (GSK-3&bgr;) is phosphorylated (inhibited) in fibrotic tissues from ischemic human and mouse heart. Using 2 fibroblast-specific GSK-3&bgr; knockout mouse models, we show that deletion of GSK-3&bgr; in cardiac fibroblasts leads to fibrogenesis, left ventricular dysfunction, and excessive scarring in the ischemic heart. Deletion of GSK-3&bgr; induces a profibrotic myofibroblast phenotype in isolated cardiac fibroblasts, in post–myocardial infarction hearts, and in mouse embryonic fibroblasts deleted for GSK-3&bgr;. Mechanistically, GSK-3&bgr; inhibits profibrotic transforming growth factor-&bgr;1/SMAD-3 signaling via interactions with SMAD-3. Moreover, deletion of GSK-3&bgr; resulted in the significant increase of SMAD-3 transcriptional activity. This pathway is central to the pathology because a small-molecule inhibitor of SMAD-3 largely prevented fibrosis and limited left ventricular remodeling. Conclusions— These studies support targeting GSK-3&bgr; in myocardial fibrotic disorders and establish critical roles of cardiac fibroblasts in remodeling and ventricular dysfunction.

Initial Right Ventricular Dysfunction Severity Identifies Severe Peripartum Cardiomyopathy Phenotype With Worse Early and Overall Outcomes: A 24‐Year Cohort Study

Background Outcomes in peripartum cardiomyopathy (PPCM) vary. We sought to determine whether severity of left or right ventricular dysfunction (RVD) at PPCM diagnosis differentially associates with adverse outcomes. Methods and Results We conducted a single‐center retrospective cohort study of 53 patients with PPCM. The primary outcome was a composite of left ventricular assist device implantation, cardiac transplantation, or death. We used Kaplan‐Meier curves to examine event‐free survival and Cox proportional hazards models to examine associations of left ventricular (LV) ejection fraction <30%, LV end‐diastolic diameter ≥60 mm, and moderate‐to‐severe RVD at PPCM diagnosis with the primary outcome. Median (interquartile range) follow‐up time was 3.6 (1.4–7.3) years. Seventeen patients (32%) experienced the primary outcome, of whom 11 had moderate‐to‐severe RVD at time of PPCM diagnosis. Overall event‐free survival differed by initial RVD severity and LV ejection fraction <30%, but not by LV end‐diastolic diameter ≥60 mm. In univariable analyses, LV ejection fraction <30% and moderate‐to‐severe RVD were associated with the outcome (hazard ratios [95% confidence intervals] of 4.85 [1.11–21.3] and 4.26 [1.47–11.6], respectively). In a multivariable model with LV ejection fraction <30%, LV end‐diastolic diameter ≥60 mm, and moderate‐to‐severe RVD, only moderate‐to‐severe RVD was independently associated with the outcome (hazard ratio [95% confidence interval], 3.21 [1.13–9.10]). Although most outcomes occurred within the first year, nearly a third occurred years after PPCM diagnosis. Conclusions Initial moderate‐to‐severe RVD is associated with a more advanced cardiomyopathy phenotype and increased risk of adverse outcomes in PPCM, within and beyond the first year of diagnosis. By identifying a worse PPCM phenotype, initial moderate‐to‐severe RVD may prompt earlier consideration of advanced heart replacement therapies.

Response to “Clarification of Enrolled Subjects in Tolvaptan HF Trials”

We appreciate the efforts of Blais et al.1 in their letter, “Clarification of Enrolled Subjects in Tolvaptan HF Trials,” to correct factual errors in our recent article that discussed the role of V2-selective arginine vasopressin receptor (AVPV2-R) antagonists in the treatment of heart failure (HF).2 We agree that we erred by not counting the patients with hyponatremia in both SALT-1 and SALT-2 who received the AVP-V2-R antagonist tolvaptan. We also agree that, in contrast to the time line provided by Finley et al.,3 two of the dose-ranging studies with tolvaptan were performed prior to rather than “subsequent to” the completion of the SALT trials. Finally, as Blais et al. point out, the information presented regarding serum sodium levels in patients with HF enrolled in the EVEREST trial (tolvaptan) was derived from Lanfear et al.4 The initial report from the EVEREST trial5 provided only the number of patients with a serum sodium level ≤137. Lanfear et al. provided far more useful data—both the median serum sodium level and the interquartile range (140 mEq/dl: 137, 142)—all of which were within the normal range. Thus, we stand by our original contention that only a small number of patients enrolled in studies with tolvaptan had an acute exacerbation of HF accompanied by clinically significant hyponatremia (<135 mEq/l). Unfortunately, Blais et al. missed the primary point of our discussion: can we use available data to reconcile the neutral effects of tolvaptan on clinical outcomes with the increased mortality seen early after the administration of lixivaptan in the BALANCE trial? We posit three explanations for this disparity: (i) lixivaptan-specific maladaptive off-target effects; (ii) the statistical vagaries associated with an underpowered clinical trial; and/or (iii) substantive differences in the demographics of patients in the various trials. We stand by our hypothesis that the increased mortality in the BALANCE trial was due to enrichment for patients with an acute exacerbation of HF and significant hyponatremia. This hypothesis is supported by the data we presented originally but is now supported by new preclinical data demonstrating that activation of the cardiac AVP-V1A-R significantly attenuates the ability of β-adrenergic receptor agonists to increase cardiac contractility in vitro, ex vivo, and in vivo.6 These results, if applicable to humans, suggest that an increase in circulating AVP, now recognized to occur with the administration of all AVP-V2-R antagonists, could limit the ability of endogenous or exogenous adrenergic drive to increase contractility in patients with an acute exacerbation of HF and compromised cardiac reserve. Until this hypothesis is tested in humans, we remain concerned about the use of these agents in this fragile patient population.

Cardiac Fibroblast Glycogen Synthase Kinase-3β Regulates Ventricular Remodeling and Dysfunction in Ischemic HeartCLINICAL PERSPECTIVE

Background— Myocardial infarction–induced remodeling includes chamber dilatation, contractile dysfunction, and fibrosis. Of these, fibrosis is the least understood. After myocardial infarction, activated cardiac fibroblasts deposit extracellular matrix. Current therapies to prevent fibrosis are inadequate, and new molecular targets are needed. Methods and Results— Herein we report that glycogen synthase kinase-3&bgr; (GSK-3&bgr;) is phosphorylated (inhibited) in fibrotic tissues from ischemic human and mouse heart. Using 2 fibroblast-specific GSK-3&bgr; knockout mouse models, we show that deletion of GSK-3&bgr; in cardiac fibroblasts leads to fibrogenesis, left ventricular dysfunction, and excessive scarring in the ischemic heart. Deletion of GSK-3&bgr; induces a profibrotic myofibroblast phenotype in isolated cardiac fibroblasts, in post–myocardial infarction hearts, and in mouse embryonic fibroblasts deleted for GSK-3&bgr;. Mechanistically, GSK-3&bgr; inhibits profibrotic transforming growth factor-&bgr;1/SMAD-3 signaling via interactions with SMAD-3. Moreover, deletion of GSK-3&bgr; resulted in the significant increase of SMAD-3 transcriptional activity. This pathway is central to the pathology because a small-molecule inhibitor of SMAD-3 largely prevented fibrosis and limited left ventricular remodeling. Conclusions— These studies support targeting GSK-3&bgr; in myocardial fibrotic disorders and establish critical roles of cardiac fibroblasts in remodeling and ventricular dysfunction.

Chronic beta-adrenergic blockade prevents volume overload-induced re-localization and oxidation of soluble guanylyl cyclase

Background While b-adrenergic blockade is a cornerstone of heart failure therapy, its therapeutic role in chronic mitral regurgitation remains questionable. Animal studies and a small clinical trial have demonstrated cardiac functional improvement with b1-adrenoceptor blocker metoprolol in chronic mitral regurgitation [1,2]. How b1AR-blockade halts functional decline of the volume-overloaded, eccentric hypertrophied heart is not well understood; anti-oxidant effects of b-blockade (bB) may play a role. We recently demonstrated that volume-overload cardiac stress induces re-localization and microdomain-specific oxidation of the nitric oxide receptor soluble guanylyl cyclase (sGC) in the failing heart [3,4]. Given that nitric oxide-cyclic guanosine monophosphate (NO-cGMP) modulates cardiac contractility and protects against cardiac hypertrophy, we hypothesized that b1AR-blockade prevents oxidation of sGC and promotes myocardial NO-cGMP signaling in a microdomain-specific fashion.