Risk-stratified Results among Congenital Diaphragmatic Hernia Patients in Two Large Extracorporeal Membrane Oxygenation Centers in South America.

INTRODUCTION: Mortality related to CDH is high, but with great variability among centers. There are few studies on patients with this condition born in South America which show poor outcomes. The goal of this study is to present the outcome of CDH in several high-volume quaternary centers in South America, ascertain the factors associated with lower mortality in our population, and compare our outcomes to those of the CDH Study Group (CDHSG). METHODS: The data from two South American centers were retrospectively analyzed and compared with contemporary data from other CDHSG participating centers. Patient demographic and clinical characteristics were also evaluated and compared. RESULTS: Between 2013 and 2018, the two South American centers saw 335 patients with CDH with an overall survival rate of 73.1%. Survival for the high, intermediate, and low-risk groups as determined by the Brindle score was 50%, 70%, and 87%, respectively. In our cohort the strongest predictors of mortality were ECMO use and early PaCO2. There were no significant differences in mortality between the two South American centers and the other CDHSG centers when adjusted by risk score, however, the South American centers had higher use of ECMO in the intermediate-risk group. DISCUSSION: Quaternary South American centers had similar outcomes to CDHSG centers worldwide. The availability and coordination of centralized dedicated care allow more efficient use of scarce technical and professional resources in patients with CDH. LEVEL OF EVIDENCE: III.

Congenital diaphragmatic hernia: phosphodiesterase-5 and Arginase inhibitors prevent pulmonary vascular hypoplasia in rat lungs.

BACKGROUND: Severe pulmonary hypoplasia related to congenital diaphragmatic hernia (CDH) continues to be a potentially fatal condition despite advanced postnatal management strategies. OBJECTIVE: To evaluate the effect of the antenatal sildenafil and 2(S)-amino-6-boronohexanoic acid (ABH-Arginase inhibitor) on lung volume, pulmonary vascular development, and nitric oxide (NO) synthesis in a Nitrofen-induced CDH rat model. METHODS: Nitrofen-induced CDH rat model was used. Nitrofen was administrated on embryonic day(E) 9,5. At E14, five intervention groups were treated separately: Nitrofen, Nitrofen+Sildenafil, Nitrofen+ABH, Nitrofen+Sildenafil+ABH and Control. At term, offspring's lungs were weighed, some paraffin-embedded for histology, others snap-frozen to analyze eNOS, Arginase I-II expression, and activity. RESULTS: In CDH-bearing offsprings, ABH or Sildenafil+ABH preserved the total lung/body-weight index (p < 0.001), preventing pulmonary vascular smooth muscle cell hyperproliferation and improving lung morphometry. Sildenafil+ABH increased 1.7-fold the lung nitrite levels (p < 0.01) without changes in eNOS expression. Sildenafil and ABH improved the number of pulmonary vessels. CONCLUSION: These results suggest that in this CDH rat model, the basal activity of Arginase participates in the lung volume and, together with phosphodiesterase-5, regulates NOS activity in the term fetal lung. The combined treatment (Sildenafil+ABH) could revert some of the pulmonary features in CDH by improving the local NO synthesis and preventing smooth muscle cell hyperproliferation. IMPACT: This study presents Arginase inhibition as a new therapeutic target and the importance of the combined antenatal treatment to improve pulmonary vascular development in a congenital diaphragmatic hernia (CDH) rat model. This study shows that the action of an Arginase inhibitor (ABH) enhances the effects already described for sildenafil in this model. These results reinforce the importance of prenatal treatments' synergy in recovering the hypoplastic lung in the Nitrofen-induced CDH rat model.

Early use of combined exogenous surfactant and inhaled nitric oxide reduces treatment failure in persistent pulmonary hypertension of the newborn: a randomized controlled trial.

OBJECTIVE: To evaluate whether combined surfactant with inhaled nitric oxide (iNO) use will prevent newborns with hypoxemic respiratory failure (HRF) from developing an Oxygenation Index (OI) > 40. METHODS: 100 term newborns with acute HRF (OI ≥ 20) were randomized to: Surfactant+iNO: received iNO plus up to two doses of surfactant or iNO-Controls: received iNO+placebo. Main outcome was the development of severe HRF (OI > 40) despite iNO use. RESULTS: Baseline mean ± SD OI was 37.4 ± 14 for the Surfactant+iNO group and 38.2 ± 16 for the controls. Infants receiving surfactant+iNO improved their oxygenation faster, resulting in lower OI at 24 h: 12.9 ± 9 vs 18.7 ± 11 of controls, p < 0.05; and a lower proportion developing OI > 40: 24%(12/50) vs 50%(25/50) of controls, p < 0.02. Fewer infants receiving surfactant+iNO presented the combined outcome of death or ECMO: 16%(8/50) compared to 36%(18/50) of controls, p < 0.05. CONCLUSIONS: Early use of combined surfactant+iNO improves oxygenation preventing the progression to severe HRF. This may reduce mortality and ECMO need. TRIAL REGISTRATION NUMBER: ISRCTN13727958.

Infection profile in neonatal patients during extracorporeal membrane oxygenation.

OBJECTIVES: To describe risk factors for acquired infection during neonatal extracorporeal membrane oxygenation and to examine the predictive value of inflammatory markers in the diagnosis of infection. METHODS: A retrospective study was conducted with data for patients under 30 days supported with extracorporeal membrane oxygenation from 2003 to April 2016, in a neonatal intensive care unit. RESULTS: Our study included 160 neonatal patients, the average age of connection was 8.5 days and the duration of extracorporeal membrane oxygenation support was 9.7 days. The incidence of confirmed infection was 23%. Patients with confirmed infection present more frequently: vaginal delivery, lower birth weight, female sex, diagnosis of congenital diaphragmatic hernia, and longer duration of extracorporeal membrane oxygenation. When comparing the group of patients with confirmed infection and suspicion of infection, there were no significant differences in the inflammatory markers. When calculating the slope for each one, the difference in white blood cell count slope 72 h before the infection is significant; in patients with confirmed infection, the count of white blood cell increases (slope: 0.25), versus the group of patients with suspected infection in whom the count decreases (slope: -0.39). No differences were found in other variables. CONCLUSION: Our study describes that the factors that increase the risk of infection are lower birth weight, vaginal birth, duration of extracorporeal membrane oxygenation, and a positive trend of white blood cell 72 h prior to infection/suspicion. Further studies are necessary to include or definitively rule out the use of these factors and the biomarkers as predictors of infection in neonatal patients supported with extracorporeal membrane oxygenation.

Neonatal and pediatric extracorporeal membrane oxygenation in developing Latin American countries / Oxigenação por membrana extracorpórea neonatal e pediátrica em países emergentes da América Latina

Abstract Objective: To review the principles of neonatal-pediatric extracorporeal membrane oxygenation therapy, prognosis, and its establishment in limited resource-limited countries in Latino America. Sources: The PubMed database was explored from 1985 up to the present, selecting from highly-indexed and leading Latin American journals, and Extracorporeal Life Support Organization reports. Summary of the findings: Extracorporeal membrane oxygenation provides “time” for pulmonary and cardiac rest and for recovery. It is used in the neonatal-pediatric field as a rescue therapy for more than 1300 patients with respiratory failure and around 1000 patients with cardiac diseases per year. The best results in short- and long-term survival are among patients with isolated respiratory diseases, currently established as a standard therapy in referral centers for high-risk patients. The first neonatal/pediatric extracorporeal membrane oxygenation Program in Latin America was established in Chile in 2003, which was also the first program in Latin America to affiliate with the Extracorporeal Life Support Organization. New extracorporeal membrane oxygenation programs have been developed in recent years in referral centers in Argentina, Colombia, Brazil, Mexico, Perú, Costa Rica, and Chile, which are currently funding the Latin American Extracorporeal Life Support Organization chapter. Conclusions: The best results in short- and long-term survival are in patients with isolated respiratory diseases. Today extracorporeal membrane oxygenation therapy is a standard therapy in some Latin American referral centers. It is hoped that these new extracorporeal membrane oxygenation centers will have a positive impact on the survival of newborns and children with respiratory or cardiac failure, and that they will be available for an increasing number of patients from this region in the near future.

Resumo Objetivo: Analisar os fundamentos, o prognóstico e o estabelecimento da terapia de oxigenação por membrana extracorpórea ECMO neonatal-pediátrica em países da América Latina com recursos limitados. Fontes: A base de dados PubMed foi explorada de 1985 até hoje, selecionamos os principais periódicos da América Latina e relatos da Organização de Suporte de Vida Extracorpóreo. Resumo dos achados: A oxigenação por membrana extracorpórea proporciona “tempo” para descanso pulmonar e cardíaco e para recuperação. Ela é usada no campo neonatal-pediátrico como terapia de resgate, com mais de 1.300 pacientes com insuficiência respiratória e cerca de 1.000 pacientes com cardiopatias por ano. Os melhores resultados de sobrevida de curto e longo prazo são de pacientes com doenças respiratórias isoladas, o que estabelece uma terapia padrão em centros de encaminhamento para pacientes de alto risco. O primeiro programa de oxigenação por membrana extracorpórea neonatal/pediátrico na América Latina foi estabelecido no Chile em 2003, que também foi o primeiro programa na América Latina a se afiliar à Organização de Suporte de Vida Extracorpóreo. Novos programas de oxigenação por membrana extracorpórea foram desenvolvidos nos últimos anos em centros de encaminhamento em Argentina, Colômbia, Brasil, México, Peru, Costa Rica e Chile, que atualmente fundam a seção da América Latina da Organização de Suporte de Vida Extracorpóreo. Conclusões: Os melhores resultados de sobrevida de curto e longo prazo são de pacientes com doenças respiratórias isoladas. Atualmente, a terapia de oxigenação por membrana extracorpórea é uma terapia padrão em alguns centros de encaminhamento da América Latina. Esperamos que esses novos centros de oxigenação por membrana extracorpórea tenham um impacto positivo sobre a sobrevida de neonatos e crianças com insuficiência respiratória ou cardíaca e que estejam disponíveis para um número cada vez maior de pacientes de nossa região no futuro próximo.

Neonatal and pediatric extracorporeal membrane oxygenation in developing Latin American countries.

OBJECTIVE: To review the principles of neonatal-pediatric extracorporeal membrane oxygenation therapy, prognosis, and its establishment in limited resource-limited countries in Latino America. SOURCES: The PubMed database was explored from 1985 up to the present, selecting from highly-indexed and leading Latin American journals, and Extracorporeal Life Support Organization reports. SUMMARY OF THE FINDINGS: Extracorporeal membrane oxygenation provides "time" for pulmonary and cardiac rest and for recovery. It is used in the neonatal-pediatric field as a rescue therapy for more than 1300 patients with respiratory failure and around 1000 patients with cardiac diseases per year. The best results in short- and long-term survival are among patients with isolated respiratory diseases, currently established as a standard therapy in referral centers for high-risk patients. The first neonatal/pediatric extracorporeal membrane oxygenation Program in Latin America was established in Chile in 2003, which was also the first program in Latin America to affiliate with the Extracorporeal Life Support Organization. New extracorporeal membrane oxygenation programs have been developed in recent years in referral centers in Argentina, Colombia, Brazil, Mexico, Perú, Costa Rica, and Chile, which are currently funding the Latin American Extracorporeal Life Support Organization chapter. CONCLUSIONS: The best results in short- and long-term survival are in patients with isolated respiratory diseases. Today extracorporeal membrane oxygenation therapy is a standard therapy in some Latin American referral centers. It is hoped that these new extracorporeal membrane oxygenation centers will have a positive impact on the survival of newborns and children with respiratory or cardiac failure, and that they will be available for an increasing number of patients from this region in the near future.

Sildenafil stimulates and dexamethasone inhibits pulmonary vascular development in congenital diaphragmatic hernia rat lungs.

BACKGROUND: A predictor of neonatal mortality in infants with congenital diaphragmatic hernia (CDH) is disrupted pulmonary vascular development, clinically expressed as pulmonary hypertension. OBJECTIVE: To determine if prenatal corticosteroids and phosphodiesterase-5 (PDE-5) inhibitors have a beneficial effect on pulmonary vascular development in CDH lungs. METHODS: We induced CDH in fetal rats by giving nitrofen. We then exposed them to dexamethasone or to sildenafil. We separated them into three groups: (1) DEX, 4 pregnant rats received dexamethasone at days E16, E18 and E20; (2) SILD, 4 pregnant rats received sildenafil and L-arginine between E14 and E22, and (3) placebo. We then analyzed the lung of each fetus with CDH at E22. We examined the number of arterioles and arteries, and their percent of medial wall thickness (%MWT). RESULTS: We obtained 30 CDH-positive fetuses. We analyzed 3,560 arterioles and 211 arteries. SILD showed a significant increase in the number of arterioles, but no significant increase in the number of arteries. No change was noted in the arteriolar %MWT. In contrast, DEX showed significant decreases in the number of arterioles and arteries and a significant increase in %MWT. CONCLUSIONS: PDE-5 inhibitors may improve pulmonary arteriolar development in fetuses with CDH. In contrast, prenatal corticosteroids could have deleterious effects on arteriolar and arterial development in CDH lungs.

Blood component usage during extracorporeal membrane oxygenation: experience in 98 patients at a Latin-American tertiary hospital.

BACKGROUND/AIMS: Hemorrhagic complications during ECMO may affect a large proportion of the patients depending on the clinical setting. To guarantee optimal delivery of blood products to these patients, blood banks require updated information on the transfusion requirements. Few studies to date provide this information. This work assesses transfusion requirements in neonates and children during ECMO during the past 9 years. METHODS: We reviewed blood bank and hospital records of patients who underwent ECMO at our institution between May 2003 and May 2012. Data obtained included age, weight, diagnosis, type, length of ECMO, and daily transfusion requirements during ECMO. Descriptive and non-parametric inferential statistic analyses were performed. Our series included 98 patients. RESULTS: Mean time of patients on ECMO was 9.2 days, with the longest treatment spanning 22 days. Mean daily transfusion requirements were 39.5 ml/kg of RBC, 12.9 ml/kg of plasma, 34.3 ml/kg of platelets and 1.4 ml/kg of cryoprecipitate. Patients who underwent ECMO due to cardiac disease or congenital diaphragmatic hernia (CDH) required significantly higher transfusion volumes of plasma (p<0.05), platelets (p< 0.05) and cryoprecipitate (p<0.05) when compared to patients underwent ECMO due to respiratory disease. Concomitant with the aging of ECMO circuits, patients showed increased requirements of RBC, plasma, and CRYO around the seventh day of the ECMO run. This effect was not observed for platelets, which remained nearly consistent around 2.2 transfusions/day. CONCLUSIONS: ECMO patients required significant transfusion support, which was particularly higher among patients who underwent ECMO due to cardiac disease or congenital diaphragmatic hernia.

Clinical utility of early amplitude integrated EEG in monitoring term newborns at risk of neurological injury / Utilidade clínica de um monitoramento por EEG de amplitude integrada precoce em recém-nascidos a termo com risco de lesão neurológica

OBJECTIVE: to test the clinical utility of an early amplitude-integrated electroencephalography (aEEG) to predict short-term neurological outcome in term newborns at risk of neurology injury. METHODS: this was a prospective, descriptive study. The inclusion criteria were neonatal encephalopathy, neurologic disturbances, and severe respiratory distress syndrome. Sensitivity, specificity, positive and negative predictive values, and likelihood ratio (LR) were calculated. Clinical and demographic data were analyzed. Neurological outcome was defined as the sum of clinical, electroimaging, and neuroimaging findings. RESULTS: ten of the 21 monitored infants (48%) presented altered short-term neurologic outcome. The aEEG had 90% sensitivity, 82% specificity, 82% positive predictive value, and 90% negative predictive value. The positive LR was 4.95, and the negative LR was 0.12. In three of 12 (25%) encephalopathic infants, the aEEG allowed for a better definition of the severity of their condition. Seizures were detected in eight infants (38%), all subclinical at baseline, and none had a normal aEEG background pattern. The status of three infants (43%) evolved and required two or more drugs for treatment. CONCLUSIONS: in infants with encephalopathy or other severe illness, aEEG disturbances occur frequently. aEEG provided a better classification of the severity of encephalopathy, detected early subclinical seizures, and allowed for monitoring of the response to treatment. aEEG was a useful tool at the neonatal intensive care unit for predicting poor short-term neurological outcomes for all sick newborn. .

OBJETIVO: testar a utilidade clínica do aEEG precoce em recém-nascidos a termo com risco delesão neurológica, para prever resultados neurológicos de curto prazo. MÉTODOS: estudo prospectivo e descritivo. Os critérios de inclusão foram encefalopatia neonatal, distúrbios neurológicos e bebês com SARA grave. Sensibilidade, especificidade, valor preditivo positivo e negativo e razão de verossimilhança foram calculados. Dados clínicos edemográficos foram analisados. O resultado neurológico foi definido como a soma de conclusões clínicas, de eletro e de neuroimagem. RESULTADOS: dentre os 21 neonatos monitorados, dez (48%) apresentaram resultado neurológico de curto prazo alterado. O aEEG apresentou sensibilidade de 90%, especificidade de 82%, valor preditivo positivo de 82% e valor preditivo negativo de 90%. A VR positiva foi de 4,95, e a RV negativa de 0,12. Em três dos 12 (25%) neonatos com encefalopatia foi possível definir melhora gravidade de sua condição pelo aEEG. Foram detectadas convulsões em oito neonatos (38%), todas subclínicas no início do estudo, e nenhum apresentou um padrão histórico normal no aEEG. O estado de três neonatos (43%) evoluiu e exigiu dois ou mais medicamentos para tratamento. CONCLUSÕES: em neonatos com encefalopatia ou outra doença grave, os distúrbios no aEEGocorrem com mais frequência. O aEEG forneceu uma classificação melhor da gravidade da encefalopatia, detectou convulsões subclínicas precoces e permitiu que fosse feito o monitoramento da resposta ao tratamento. O aEEG é uma ferramenta útil para prever resultados neurológicos de curto prazo em todos os bebês doentes na UTIN. .

Clinical utility of early amplitude integrated EEG in monitoring term newborns at risk of neurological injury.

OBJECTIVE: to test the clinical utility of an early amplitude-integrated electroencephalography (aEEG) to predict short-term neurological outcome in term newborns at risk of neurology injury. METHODS: this was a prospective, descriptive study. The inclusion criteria were neonatal encephalopathy, neurologic disturbances, and severe respiratory distress syndrome. Sensitivity, specificity, positive and negative predictive values, and likelihood ratio (LR) were calculated. Clinical and demographic data were analyzed. Neurological outcome was defined as the sum of clinical, electroimaging, and neuroimaging findings. RESULTS: ten of the 21 monitored infants (48%) presented altered short-term neurologic outcome. The aEEG had 90% sensitivity, 82% specificity, 82% positive predictive value, and 90% negative predictive value. The positive LR was 4.95, and the negative LR was 0.12. In three of 12 (25%) encephalopathic infants, the aEEG allowed for a better definition of the severity of their condition. Seizures were detected in eight infants (38%), all subclinical at baseline, and none had a normal aEEG background pattern. The status of three infants (43%) evolved and required two or more drugs for treatment. CONCLUSIONS: in infants with encephalopathy or other severe illness, aEEG disturbances occur frequently. aEEG provided a better classification of the severity of encephalopathy, detected early subclinical seizures, and allowed for monitoring of the response to treatment. aEEG was a useful tool at the neonatal intensive care unit for predicting poor short-term neurological outcomes for all sick newborn.

Survival of newborn infants with severe respiratory failure before and after establishing an extracorporeal membrane oxygenation program.

BACKGROUND: Severe hypoxic respiratory failure is a leading cause of neonatal mortality in Chile. Extracorporeal membrane oxygenation improves survival in neonates with hypoxic respiratory failure. OBJECTIVE: To determine the impact of the establishment of a Neonatal Extracorporeal Membrane Oxygenation Program on the outcome of newborns with severe hypoxic respiratory failure in a developing country. DESIGN/PATIENTS: Data of newborns (birthweight > 2,000 g and gestational age ≥ 35 wk) with hypoxic respiratory failure and oxygenation index greater than 25 were compared before and after extracorporeal membrane oxygenation was available. Extracorporeal membrane oxygenation was initiated in infants with refractory hypoxic respiratory failure who failed to respond to inhaled nitric oxide/high-frequency oscillatory ventilation. MAIN RESULTS: Data from 259 infants were analyzed; 100 born in the pre-extracorporeal membrane oxygenation period and 159 born after the extracorporeal membrane oxygenation program was established. Patients were similar in terms of risk factors for death for both periods except for a higher oxygenation index and a greater proportion of outborn infants during the extracorporeal membrane oxygenation period. Survival significantly increased from 72% before extracorporeal membrane oxygenation to 89% during the extracorporeal membrane oxygenation period (p < 0.01). During the extracorporeal membrane oxygenation period, 98 of 159 patients (62%) with hypoxic respiratory failure were rescued using inhaled nitric oxide/high-frequency oscillatory ventilation, whereas 61 (38%) did not improve; 52 of these 61 neonates were placed on extracorporeal membrane oxygenation. Extracorporeal membrane oxygenation survival rate to discharge was 85%. After adjusting for potential confounders, the severity of the pretreatment oxygenation index, a late arrival to the referral center, the presence of a pneumothorax, and the diagnosis of a diaphragmatic hernia were significantly associated with the need for extracorporeal membrane oxygenation or death. CONCLUSIONS: The establishment of an extracorporeal membrane oxygenation program was associated with a significant increase in the survival of newborns more than or equal to 35 weeks old with severe hypoxic respiratory failure.

Chylothorax in congenital diaphragmatic hernia.

PURPOSE: Following surgical repair of congenital diaphragmatic hernia (CDH), chylothorax can be present in 7-28% of the cases. It has been associated with prenatal diagnosis, the use of ECMO and prosthetic patches during reparatory surgery. The objective is to present a neonatal unit experience in handling this complication and the search for predictive factors for its appearance in our patients. METHODS: A retrospective study was carried out between 2003 and 2009. RESULTS: We found 65 patients with CDH, of which 7 (10.8%) developed a chylothorax, 5 responded to drainage and diet restriction. Octreotide (OCT) was used in two cases that did not respond to the usual treatment. OCT was successful with one patient, while surgical resolution of the chylothorax was necessary in the other. Two patients died, none of them during treatment of chylothorax. We did not find a significant association between chylothorax and the variables studied in these patients. CONCLUSIONS: Chylothorax is a common complication following CDH repair. We have a low rate of this complication in our institution. Conservative management is an appropriate approach for all patients; OCT could be an alternative to avoid surgery. We did not find any predictive factors for chylothorax in our series.

Improvement of survival in infants with congenital diaphragmatic hernia in recent years: effect of ECMO availability and associated factors.

BACKGROUND/PURPOSE: Survival of patients with congenital diaphragmatic hernia (CDH) depends both on non-modifiable congenital conditions and on modifiable pre and postnatal management. ECMO improves survival up to 80% in neonates with CDH in the best ECMO centers worldwide. The first Neonatal ECMO Program in Chile was started in our University in 2003. Our objective is to determine the impact of a Neonatal ECMO Program in a level III NICU on newborns with CDH. METHODS: Data of all newborns with CDH admitted to our NICU was separated into two groups: pre ECMO (1996-2003) and ECMO (2003-2007). Crude and adjusted odds ratios for 24 months survival were estimated by logistic regression. RESULTS: Data of 46 newborns with CDH was analysed, 20 in the pre ECMO and 26 in the ECMO period. Patient characteristics were similar in both groups; however, 24-month survival increased significantly from 25% (5/20) in the pre ECMO period to 77% (20/26) in the ECMO period (P = 0.001). Adjusted odds ratios for 24-month survival were 26.98 for OI or= 7 and 17.5 for ECMO availability. CONCLUSIONS: The establishment of an ECMO program was associated with a significant increase in long-term survival for infants with CDH.

[Extracorporeal membrane oxygenation (ECMO): consolidation of a neonatal-pediatric program in Chile and report of three cases]. / Oxigenación con membrana extracorpórea en pacientes pediátricos: comunicación de los 3 primeros casos tratados.

Extracorporeal membrane oxygenation (ECMO) improves survival in neonatal and pediatric patients with reversible severe respiratory or cardiac failure, in whom intensive treatment fails. Since 1999, a multidisciplinary team is trained to form the first neonatal-pediatric ECMO center in Chile, according to the norms of the Extracorporeal Life Support Organization (ELSO). During 2003 the first three patients were admitted to the program: a male newborn with pulmonary hypertension, a 38 days old female operated for a total anomalous pulmonary venous connection and a 3 months old male with a severe pneumonia caused by respiratory syncytial virus. They remained in ECMO for five, seven and nine days respectively and all survived to the procedure. No neurological complications were observed after one and a half year of follow up. This consolidates the first national neonatal-pediatric ECMO program, associated to ELSO. Up to date, twelve patients have been admitted to the program.

Formation and remodeling of the coronary vascular bed in the embryonic avian heart.

To study the formation of the coronary vessels in the developing avian heart, we stained developmentally staged quail hearts with the endothelial specific antibody QH-1. QH-1 reacted with individual cells in the proepicardial organ in Hamburger and Hamilton stage (HH) 17 embryos only after it had contacted the heart. In HH18-26 hearts, individual QH-1+ cells accumulated over the surface of the atria and ventricles. The first endothelial vessels appeared in the dorsal atrioventricular groove in HH23 hearts. CD45+ hematopoietic precursors accumulated on the heart surface, demonstrating the close temporal relationship of hematopoiesis with vasculogenesis during heart development. However, CD45 expression preceded association of these cells with the vasculature, suggesting hematopoietic commitment precedes formation of blood islands in the coronary vasculature. Endothelial tubules first appeared on the dorsal and then the ventral aspects of the heart, coalescing into large sinusoids. These sinusoids remodeled into compact muscularized vessels by HH35. Smooth muscle cell markers were first expressed at HH27 and only in association with developing vasculature. We did not observe markers of smooth muscle differentiation in the proepicardium, but it remains uncertain whether cells in the proepicardium are committed to this cell fate. Our data support a strictly vasculogenic mechanism for the formation of the coronary vessels and blood islands.

Mechanisms of embryonic coronary artery development.

Coronary artery development is a complex vasculogenic process that begins shortly after heart looping. Coronary vasculogenesis is regulated by the myocardium, but is spatially and temporally dependent on the epicardium and its precursor, the proepicardial organ, for the provision of coronary vascular progenitor cells. Better understanding of the mechanisms of coronary artery development may clarify mechanisms of disease and suggest new potential therapies for disorders of the coronary vasculature.