Dataset Information

Noncanonical WNT Activation in Human Right Ventricular Heart Failure.

ABSTRACT: Background: No medical therapies exist to treat right ventricular (RV) remodeling and RV failure (RVF), in large part because molecular pathways that are specifically activated in pathologic human RV remodeling remain poorly defined. Murine models have suggested involvement of Wnt signaling, but this has not been well-defined in human RVF. Methods: Using a candidate gene approach, we sought to identify genes specifically expressed in human pathologic RV remodeling by assessing the expression of 28 WNT-related genes in the RVs of three groups: explanted nonfailing donors (NF, n = 29), explanted dilated and ischemic cardiomyopathy, obtained at the time of cardiac transplantation, either with preserved RV function (pRV, n = 78) or with RVF (n = 35). Results: We identified the noncanonical WNT receptor ROR2 as transcriptionally strongly upregulated in RVF compared to pRV and NF (Benjamini-Hochberg adjusted P < 0.05). ROR2 protein expression correlated linearly to mRNA expression (R 2 = 0.41, P = 8.1 × 10-18) among all RVs, and to higher right atrial to pulmonary capillary wedge ratio in RVF (R 2 = 0.40, P = 3.0 × 10-5). Utilizing Masson's trichrome and ROR2 immunohistochemistry, we identified preferential ROR2 protein expression in fibrotic regions by both cardiomyocytes and noncardiomyocytes. We compared RVF with high and low ROR2 expression, and found that high ROR2 expression was associated with increased expression of the WNT5A/ROR2/Ca2+ responsive protease calpain-?, cleavage of its target FLNA, and FLNA phosphorylation, another marker of activation downstream of ROR2. ROR2 protein expression as a continuous variable, correlated strongly to expression of calpain-? (R 2 = 0.25), total FLNA (R 2 = 0.67), calpain cleaved FLNA (R 2 = 0.32) and FLNA phosphorylation (R 2 = 0.62, P < 0.05 for all). Conclusion: We demonstrate robust reactivation of a fetal WNT gene program, specifically its noncanonical arm, in human RVF characterized by activation of ROR2/calpain mediated cytoskeleton protein cleavage.

SUBMITTER: Edwards JJ

PROVIDER: S-EPMC7575695 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Noncanonical WNT Activation in Human Right Ventricular Heart Failure.

Edwards Jonathan J JJ Brandimarto Jeffrey J Hu Dong-Qing DQ Jeong Sunhye S Yucel Nora N Li Li L Bedi Kenneth C KC Wada Shogo S Murashige Danielle D Hwang Hyun Tae V HTV Zhao Mingming M Margulies Kenneth B KB Bernstein Daniel D Reddy Sushma S Arany Zoltan Z

Frontiers in cardiovascular medicine 20201007

<b>Background:</b> No medical therapies exist to treat right ventricular (RV) remodeling and RV failure (RVF), in large part because molecular pathways that are specifically activated in pathologic human RV remodeling remain poorly defined. Murine models have suggested involvement of Wnt signaling, but this has not been well-defined in human RVF. <b>Methods:</b> Using a candidate gene approach, we sought to identify genes specifically expressed in human pathologic RV remodeling by assessing the ...[more]

PMID: 33134326

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Project description:AimThe severity of cardiac impairment in acute heart failure (AHF) predicts outcome, but challenges remain to identify prognostically important non-invasive parameters of cardiac function. Left ventricular ejection fraction (LVEF) is relevant, but only in those with reduced LV systolic function. We aimed to assess the standard and advanced parameters of left and right ventricular (RV) function from echocardiography in predicting long-term outcomes in AHF.MethodsA total of 418 consecutive AHF patients presenting over 12 months were prospectively recruited and underwent bedside echocardiography within 24 h of recruitment. We retrospectively assessed 8 RV and 5 LV echo parameters of the cardiac systolic function to predict 2-year mortality, using both guideline-directed and study-specific cutoffs, based on the maximum Youden indices via ROC analysis. For the RV, these were the tricuspid annular plane systolic excursion, RV fractional area change, tissue Doppler imaging (TDI) peak tricuspid annular systolic wave velocity, both peak- and end-systolic RV free wall global longitudinal strain (RV GLS) and strain rate (mean RV GLSR), RV ejection fraction (RVEF) derived from a 2D ellipsoid model and the ratio of the TAPSE to systolic pulmonary artery pressure (SPAP). For the LV, these were the LVEF, mitral regurgitant ΔP/Δt (MR dP/dt), the lateral mitral annular TDI peak systolic wave velocity, LV GLS, and the LV GLSR.ResultsA total of 7/8 parameters of RV systolic function were predictive of 2-year outcome, with study cutoffs like international guidelines. A cutoff of < -1.8 s-1 mean RV GLSR was associated with worse outcome compared to > -1.8 s-1 [HR 2.13 95% CI 1.33-3.40 (p = 0.002)]. TAPSE:SPAP of > 0.027 cm/mmHg (vs. < 0.027 cm/mmHg) predicted worse outcome [HR 2.12 95% CI 1.53-2.92 (p < 0.001)]. A 3-way comparison of 2-year mortality by LVEF from the European Society of Cardiology (ESC) guideline criteria of LVEF > 50, 41-49, and < 40% was not prognostic [38.6% vs. 30.9 vs. 43.9% (p = 0.10)]. Of the 5 parameters of LV systolic function, only an MR dP/dt cutoff of < 570 mmHg was predictive of adverse outcome [HR 1.63 95% CI 1.01-2.62 (p = 0.047)].ConclusionWith cutoffs broadly like the ESC guidelines, we identified RV dysfunction to be associated with adverse prognosis, whereas LVEF could not identify patients at risk.

Project description:Right ventricular heart failure (RVF) associated with pulmonary hypertension (PH) is characterized by a distinct gene expression pattern when compared with functional compensatory hypertrophy. Carvedilol treatment after RVF has been established reduces right ventricle (RV) hypertrophy and improves the RV function. In addition, carvedilol treatment has been shown to alter the gene expression of select genes. We sought to identify, on a genome-wide basis, the effect of carvedilol on gene expression. RVF was induced in male Sprague-Dawley rats by the combination of VEGF-receptor blockade and chronic hypoxia; thereafter, one group was treated with carvedilol. RNA was isolated from the RV and subjected to microarray analysis. A prediction analysis of the carvedilol-treated RVs showed that carvedilol treated RVs most resembled in their expression pattern the RVF pattern. However, an analysis beyond the boundaries of the prediction set revealed a small set of genes associated with carvedilol reversal of RVF. Pathway analysis of this set of genes revealed expression changes of genes involved in cardiac hypertrophy, mitochondrial dysfunction, protein ubiquitination, and sphingolipid metabolism. Genes encoding proteins in the cardiac hypertrophy and protein ubiquitination pathways were downregulated in the RV by carvedilol, while genes encoding proteins in the mitochondrial dysfunction and sphingolipid metabolism pathways were upregulated by carvedilol. Male Sprague-Dawley rats (Harlan Laboratories Inc., Indianapolis, IN) weighing 200g were injected subcutaneously with SU5416 suspended in 0.5% (w/v) carboxymethylcellulose sodium, 0.9% (w/v) sodium chloride, 0.4% (w/v) polysorbate 80, and 0.9% (v/v) benzyl alcohol in deionized water. Rats were given a single injection of SU5416 (20 mg/kg) at the beginning of the 6-week experiment. The animals were then exposed to chronic hypoxia (simulated altitude of 5,000 m in a nitrogen dilution chamber) for four weeks; thereafter the animals were kept at the altitude of Richmond, VA (sea level) for another two weeks. Hypoxia-only rats were exposed to chronic hypoxia for four weeks. Carvedilol (15 mg/kg; Sigma-Aldrich, St. Louis, MO) was dissolved in 20% dimethyl sulfoxide and water and administered once daily per oral gavage for 4 weeks, beginning after return to room air breathing. Six rats per treatment group were randomly assigned. Rats were euthanized with Euthanasia-III Solution (390 mg/ml pentobarbital sodium, 50 mg/ml phenytoin sodium; Med-Pharmex, Inc., Pomona, CA) and hearts were removed. The RV was dissected from the LV and both ventricles were snap-frozen in liquid nitrogen. Total RNA was isolated from approximately 30 mg of snap-frozen rat heart tissue using the Qiagen RNeasy Mini Kit (Qiagen, Valencia, CA). Hearts were homogenized with Buffer RLT and beta-mercaptoethanol in an MP FastPrepM-BM-.-24 Lysing Matrix D tube (MP Biomedicals LLC, Solon, OH), and then RNA was isolated and purified following the manufacturerM-bM-^@M-^Ys protocol. RNA concentration was determined using a NanoDrop ND-1000 (Thermo Fisher Scientific, Wilmington, DE). All samples had an A260/A280 ratio between 1.9 and 2.1. The amplification and hybridization process is as follows. 500 ng of total RNA was amplified and labeled with Cyanine-5 and 500 ng of universal rat reference RNA (Stratagene, Santa Clara, CA) was amplified and labeled with Cyanine-3 using the Agilent QuickAmp Labeling kit (Agilent Technologies Inc., Santa Clara, CA) to produce labeled cRNA following the manufacturerM-bM-^@M-^Ys protocol. After amplification and labeling, the dye incorporation was determined using a NanoDrop ND-1000 spectrophotometer (Thermo Fisher Scientific, Wilmington, DE). All ratios were greater than 8.0 pmole dye per mg cRNA per the manufacturerM-bM-^@M-^Ys recommendation. 825 ng of sample and 825 ng of reference RNA were combined and incubated with an Agilent Whole Rat Genome 4x44k microarray slide (Agilent Technologies Inc., Wilmington, DE) for 17 hours at 65M-BM-0C. Following hybridization, slides were washed following the manufacturerM-bM-^@M-^Ys protocol and scanned using an Axon GenePix 4200A scanner (Axon Instruments, Union City, CA) at a resolution of 5 mM. The raw data were generated using GenePix Pro 5.0 software (Axon Instruments, Union City, CA). Raw expression data files were uploaded into R and normalized using the Bioconductor marray package by the Lowess normalization algorithm, then exported to BRB Array Tools for analysis.

Dataset Information

Noncanonical WNT Activation in Human Right Ventricular Heart Failure.

Publications

Noncanonical WNT Activation in Human Right Ventricular Heart Failure.

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