Satish K. Rajagopal

Epicardial progenitors contribute to the cardiomyocyte lineage in the developing heart

The heart is formed from cardiogenic progenitors expressing the transcription factors Nkx2-5 and Isl1 (refs 1 and 2). These multipotent progenitors give rise to cardiomyocyte, smooth muscle and endothelial cells, the major lineages of the mature heart. Here we identify a novel cardiogenic precursor marked by expression of the transcription factor Wt1 and located within the epicardium—an epithelial sheet overlying the heart. During normal murine heart development, a subset of these Wt1+ precursors differentiated into fully functional cardiomyocytes. Wt1+ proepicardial cells arose from progenitors that express Nkx2-5 and Isl1, suggesting that they share a developmental origin with multipotent Nkx2-5+ and Isl1+ progenitors. These results identify Wt1+ epicardial cells as previously unrecognized cardiomyocyte progenitors, and lay the foundation for future efforts to harness the cardiogenic potential of these progenitors for cardiac regeneration and repair.

Development of heart valves requires Gata4 expression in endothelial-derived cells

Cardiac malformations due to aberrant development of the atrioventricular (AV) valves are among the most common forms of congenital heart disease. At localized swellings of extracellular matrix known as the endocardial cushions, the endothelial lining of the heart undergoes an epithelial to mesenchymal transition (EMT) to form the mesenchymal progenitors of the AV valves. Further growth and differentiation of these mesenchymal precursors results in the formation of portions of the atrial and ventricular septae, and the generation of thin, pliable valves. Gata4, which encodes a zinc finger transcription factor, is expressed in the endothelium and mesenchyme of the AV valves. Using a Tie2-Cre transgene, we selectively inactivated Gata4 within endothelial-derived cells. Mutant endothelium failed to undergo EMT, resulting in hypocellular cushions. Mutant cushions had decreased levels of Erbb3, an EGF-family receptor essential for EMT in the atrioventricular cushions. In Gata4 mutant embryos, Erbb3 downregulation was associated with impaired activation of Erk, which is also required for EMT. Expression of a Gata4 mutant protein defective in interaction with Friend of Gata (FOG) cofactors rescued the EMT defect, but resulted in a decreased proliferation of mesenchyme and hypoplastic cushions that failed to septate the ventricular inlet. We demonstrate two novel functions of Gata4 in development of the AV valves. First, Gata4 functions as an upstream regulator of an Erbb3-Erk pathway necessary for EMT, and second, Gata4 acts to promote cushion mesenchyme growth and remodeling.

Gata4 is required for maintenance of postnatal cardiac function and protection from pressure overload-induced heart failure

An important event in the pathogenesis of heart failure is the development of pathological cardiac hypertrophy. In cultured cardiomyocytes, the transcription factor Gata4 is required for agonist-induced hypertrophy. We hypothesized that, in the intact organism, Gata4 is an important regulator of postnatal heart function and of the hypertrophic response of the heart to pathological stress. To test this hypothesis, we studied mice heterozygous for deletion of the second exon of Gata4 (G4D). At baseline, G4D mice had mild systolic and diastolic dysfunction associated with reduced heart weight and decreased cardiomyocyte number. After transverse aortic constriction (TAC), G4D mice developed overt heart failure and eccentric cardiac hypertrophy, associated with significantly increased fibrosis and cardiomyocyte apoptosis. Inhibition of apoptosis by overexpression of the insulin-like growth factor 1 receptor prevented TAC-induced heart failure in G4D mice. Unlike WT-TAC controls, G4D-TAC cardiomyocytes hypertrophied by increasing in length more than width. Gene expression profiling revealed up-regulation of genes associated with apoptosis and fibrosis, including members of the TGF-β pathway. Our data demonstrate that Gata4 is essential for cardiac function in the postnatal heart. After pressure overload, Gata4 regulates the pattern of cardiomyocyte hypertrophy and protects the heart from load-induced failure.

Extracorporeal membrane oxygenation for the support of infants, children, and young adults with acute myocarditis: A review of the Extracorporeal Life Support Organization registry*

Objective: To describe survival outcomes for pediatric patients supported with extracorporeal membrane oxygenation for severe myocarditis and identify risk factors for in-hospital mortality. Design: Retrospective review of Extracorporeal Life Support Organization registry database. Setting: Data reported to Extracorporeal Life Support Organization from 116 extracorporeal membrane oxygenation centers. Patients: Patients ≤18 yrs of age supported with extracorporeal membrane oxygenation for myocarditis during 1995 to 2006. Interventions: None. Measurements and Main Results: Of 19,348 reported pediatric extracorporeal membrane oxygenation uses from 1995 to 2006, 260 runs were for 255 patients with a diagnosis of myocarditis (1.3%). Survival to hospital discharge was 61%. Seven patients (3%) underwent heart transplantation and six patients survived to discharge. Of 100 patients not surviving to hospital discharge, extracorporeal membrane oxygenation support was withdrawn in 70 (70%) with multiple organ failure as the indication in 58 (83%) patients. In a multivariable model, female gender (adjusted odds ratio, 2.3, 95% confidence interval, 1.3–4.2), arrhythmia on extracorporeal membrane oxygenation (adjusted odds ratio, 2.7, 95% confidence interval, 1.5–5.1), and renal failure requiring dialysis (adjusted odds ratio, 5.1, 95% confidence interval, 2.3–11.4) were associated with increased odds of in-hospital mortality. Conclusion: Extracorporeal membrane oxygenation is a valuable tool to rescue children with severe cardiorespiratory compromise related to myocarditis. Female gender, arrhythmia on extracorporeal membrane oxygenation, and need for dialysis during extracorporeal membrane oxygenation were associated with increased mortality.

Gata4 Is Essential for the Maintenance of Jejunal-Ileal Identities in the Adult Mouse Small Intestine

ABSTRACT Gata4, a member of the zinc finger family of GATA transcription factors, is highly expressed in duodenum and jejunum but is nearly undetectable in distal ileum of adult mice. We show here that the caudal reduction of Gata4 is conserved in humans. To test the hypothesis that the regional expression of Gata4 is critical for the maintenance of jejunal-ileal homeostasis in the adult small intestine in vivo, we established an inducible, intestine-specific model that results in the synthesis of a transcriptionally inactive Gata4 mutant. Synthesis of mutant Gata4 in jejuna of 6- to 8-week-old mice resulted in an attenuation of absorptive enterocyte genes normally expressed in jejunum but not in ileum, including those for the anticipated targets liver fatty acid binding protein (Fabp1) and lactase-phlorizin hydrolase (LPH), and a surprising induction of genes normally silent in jejunum but highly expressed in ileum, specifically those involved in bile acid transport. Inactivation of Gata4 resulted in an increase in the goblet cell population and a redistribution of the enteroendocrine subpopulations, all toward an ileal phenotype. The gene encoding Math1, a known activator of the secretory cell fate, was induced ∼75% (P < 0.05). Gata4 is thus an important positional signal required for the maintenance of jejunal-ileal identities in the adult mouse small intestine.

A Risk-Prediction Model for In-Hospital Mortality After Heart Transplantation in US Children

We sought to develop and validate a quantitative risk‐prediction model for predicting the risk of posttransplant in‐hospital mortality in pediatric heart transplantation (HT). Children <18 years of age who underwent primary HT in the United States during 1999–2008 (n = 2707) were identified using Organ Procurement and Transplant Network data. A risk‐prediction model was developed using two‐thirds of the cohort (random sample), internally validated in the remaining one‐third, and independently validated in a cohort of 338 children transplanted during 2009–2010. The best predictive model had four categorical variables: hemodynamic support (ECMO, ventilator support, VAD support vs. medical therapy), cardiac diagnosis (repaired congenital heart disease [CHD], unrepaired CHD vs. cardiomyopathy), renal dysfunction (severe, mild‐moderate vs. normal) and total bilirubin (≥ 2.0, 0.6 to <2.0 vs. <0.6 mg/dL). The C‐statistic (0.78) and the Hosmer–Lemeshow goodness‐of‐fit (p = 0.89) in the model‐development cohort were replicated in the internal validation and independent validation cohorts (C‐statistic 0.75, 0.81 and the Hosmer–Lemeshow goodness‐of‐fit p = 0.49, 0.53, respectively) suggesting acceptable prediction for posttransplant in‐hospital mortality. We conclude that this risk‐prediction model using four factors at the time of transplant has good prediction characteristics for posttransplant in‐hospital mortality in children and may be useful to guide decision‐making around patient listing for transplant and timing of mechanical support.

Perioperative factors associated with prolonged mechanical ventilation after complex congenital heart surgery.

Objective: To evaluate perioperative factors associated with prolonged mechanical ventilation in children undergoing complex cardiac surgery for congenital heart disease. Design: Retrospective chart review. Setting: A tertiary care pediatric cardiac intensive care. Interventions: None. Measurements and Main Results: This retrospective cohort study included all patients undergoing complex cardiac surgical procedures (Risk Adjustment in Congenital Heart Surgery-1 category ≥3) at our institution during 2003. We defined prolonged mechanical ventilation as need for mechanical ventilation for ≥7 days (90th percentile of duration of mechanical ventilation for the whole cohort). Multivariate logistic regression analyses were used to determine independent relationships between perioperative factors and prolonged mechanical ventilation. A total of 362 patients were admitted to the cardiac intensive care unit after a cardiac surgical procedure of Risk Adjustment in Congenital Heart Surgery-1 ≥3 level of complexity and survived to hospital discharge. Median age was 242 days (range, 4 days−14.4 yrs), the median duration of mechanical ventilation was 1.5 days (range, 0–7 days), and 41 patients (11%) were ventilated for ≥7 days. Age of <30 days at surgery, higher Pediatric Risk of Mortality III score at the time of cardiac intensive care unit admission, the presence of major noncardiac structural anomalies, healthcare-associated infections, noninfectious pulmonary complications (pleural effusions and pneumothorax), and the need for reintervention were all independently associated with prolonged mechanical ventilation. Conclusions: Younger age, greater severity of illness at postoperative admission, healthcare-associated infections, noninfectious pulmonary complications, and the need for reintervention are associated with prolonged mechanical ventilation after complex cardiac surgery. Future studies and quality improvement initiatives should focus on those risk factors that are modifiable to promote early extubation in children recovering from complex congenital heart surgery.

Extracorporeal membrane oxygenation for the support of adults with acute myocarditis.

Objectives: To characterize survival outcomes for adult patients with acute myocarditis supported with extracorporeal membrane oxygenation and identify risk factors for in-hospital mortality. Design: Retrospective review of Extracorporeal Life Support Organization registry database. Setting: Data reported to Extracorporeal Life Support Organization by 230 extracorporeal membrane oxygenation centers. Patients: Patients 16 years old or older supported with extracorporeal membrane oxygenation for myocarditis during 1995 to 2011. Interventions: None. Measurements and Main Results: There were 150 separate runs of extracorporeal membrane oxygenation for 147 patients with a diagnosis of acute myocarditis in the Extracorporeal Life Support Organization database from 1995 through 2011. Survival to hospital discharge was 61%. Nine patients underwent heart transplantation, and transplant-free survival to discharge was 56%. Extracorporeal membrane oxygenation was deployed during extracorporeal cardiopulmonary resuscitation in 31 patients (21% of the cohort). In a multivariate model evaluating pre–extracorporeal membrane oxygenation and extracorporeal membrane oxygenation support factors, pre–extracorporeal membrane oxygenation arrest (adjusted odds ratio, 2.4; 95% CI, 1.1–5.0) and need for higher extracorporeal membrane oxygenation flows at 4 hours post–extracorporeal membrane oxygenation cannulation (odds ratio, 2.8; 95% CI, 1.1–7.3) were associated with increased odds of in-hospital mortality. In a second multivariate model evaluating adverse events while on extracorporeal membrane oxygenation, central nervous system injury (odds ratio, 26.5; 95% CI, 7.3–96.6), renal failure (odds ratio, 3.6; 95% CI, 1.4–9.3), arrhythmia (odds ratio, 5.8; 95% CI, 2.2–15.1), and hyperbilirubinemia (odds ratio, 9.1; 95% CI, 2.6–31.8) were associated with increased odds of in-hospital mortality. Conclusions: Extracorporeal membrane oxygenation can be used effectively in adults with myocarditis to support the circulation while awaiting myocardial recovery. Early extracorporeal membrane oxygenation deployment prior to cardiac arrest may be associated with better outcomes.

Outcomes of Neonates Undergoing Extracorporeal Membrane Oxygenation Support Using Centrifugal Versus Roller Blood Pumps

BACKGROUND Advances in centrifugal blood pump technology have led to increased use of centrifugal pumps in extracorporeal membrane oxygenation (ECMO) circuits. Their efficacy and safety in critically ill neonates remains unknown. Blood cell trauma leading to hemolysis may result in end-organ injury in critically ill neonates receiving centrifugal pump ECMO. We hypothesized that neonates undergoing ECMO support using centrifugal pumps were at increased odds of hemolysis and subsequent end-organ injury. METHODS Children 30 days of age or younger who received support with venoarterial ECMO and were reported to the Extracorporeal Life Support Registry during 2007 to 2009 underwent propensity score matching (Greedy matching 1:1) using pre-ECMO support characteristics. RESULTS A total of 1,592 neonates receiving ECMO (centrifugal pump = 163 and roller pump = 1,492) were identified. Significant differences in demographic, presupport, and cannulation variables were present before matching. One hundred seventy-six neonates who were supported using either centrifugal (n = 88) or roller pumps (n = 88) were matched using propensity scoring. No significant differences in demographic, presupport, or cannulation variables were present after matching. Neonates undergoing support using centrifugal pumps had increased odds of hemolysis (odds ratio [OR], 7.7 [2.8-21.2]), hyperbilirubinemia (OR, 20.8 [2.7-160.4]), hypertension (OR, 3.2 [1.3-8.0]), and acute renal failure (OR, 2.4 [1.1-5.6]). Survival to discharge was not different between pump types. CONCLUSIONS Use of ECMO using centrifugal pumps is associated with increased odds of hemolysis that likely contributes to other end-organ injury. Research into the optimal use of centrifugal pumps and strategies to prevent support-related complications need to be investigated.

Acute kidney injury after Fontan completion: Risk factors and outcomes

OBJECTIVE Acute kidney injury (AKI) is a predictor of outcomes in heterogeneous populations of children undergoing cardiac surgery. We investigated its causes and consequences in a cohort undergoing Fontan completion, hypothesizing that central venous pressure is independently associated with development of AKI. METHODS In this retrospective cohort study of patients undergoing Fontan (n = 211), univariable and multivariable analyses identified factors associated with AKI within 3 days of surgery. Secondary analyses identified factors associated with hospital length of stay, and examined effects of perioperative kidney injury on follow-up renal function. RESULTS Acute kidney injury occurred in 42% of cases (n = 89), with the following independent risk factors: mean renal perfusion (mean arterial minus central venous) pressure on postoperative day zero (per mm Hg; adjusted odds ratio [AOR] 0.83; P < .001); preoperative atrioventricular valve regurgitation > mild (AOR 6.78; P = .02); bypass time (per 10 minutes, AOR 1.08; P = .04); peak inotrope score on postoperative day zero (per point, AOR 1.17; P < .001); and preoperative pulmonary vascular resistance (per Wood unit, AOR 1.69; P = .04). Central venous pressure was not independently associated with AKI. Moderate and severe (but not mild) AKI were independently associated with prolonged hospital length of stay (adjusted hazard ratios, 0.56; P = .004, and .41; P = .006, respectively). Perioperative injury was not associated with longer-term renal dysfunction. CONCLUSIONS Acute kidney injury is common after Fontan completion and has several potentially modifiable risk factors. Moderate-to-severe injury is associated with longer hospital length of stay but not with renal dysfunction at follow-up.