Increased utilization of the hybrid procedure is not associated with improved early survival for newborns with hypoplastic left heart syndrome: a single-centre experience.

OBJECTIVES: The primary objectives were to examine utilization of the Hybrid versus the Norwood procedure for patients with hypoplastic left heart syndrome or variants and the impact on hospital mortality. The Hybrid procedure was 1st used at our institution in 2004. METHODS: Review of all subjects undergoing the Norwood or Hybrid procedure between 1 January 1984 and 31 December 2022. The study period was divided into 8 eras: era 1, 1984-1988; era 2, 1989-1993; era 3, 1994-1998; era 4, 1999-2003; era 5, 2004-2008; era 6, 2009-2014; era 7, 2015-2018 and era 8, 2019-2022. The primary outcome was in-hospital mortality. Mortality rates were computed using standard binomial proportions with 95% confidence intervals. Rates across eras were compared using an ordered logistic regression model with and adjusted using the Tukey-Kramer post-hoc procedure for multiple comparisons. In the risk-modelling phase, logistic regression models were specified and tested. RESULTS: The Norwood procedure was performed in 1899 subjects, and the Hybrid procedure in 82 subjects. Use of the Hybrid procedure increased in each subsequent era, reaching 30% of subjects in era 8. After adjustment for multiple risk factors, use of the Hybrid procedure was significantly and positively associated with hospital mortality. CONCLUSIONS: Despite the increasing use of the Hybrid procedure, overall mortality for the entire cohort has plateaued. After adjustment for risk factors, use of the Hybrid procedure was significantly and positively associated with mortality compared to the Norwood procedure.

Contemporary outcomes of open repair of acute complicated type B aortic dissection.

OBJECTIVE: Open repair of acute complicated type B aortic dissection (ACTBAD), required when endovascular repair is not possible, is historically considered high-risk. We analyze our experience with this high-risk cohort compared with the standard cohort. METHODS: We identified consecutive patients undergoing descending thoracic or thoracoabdominal aortic aneurysm (TAAA) repair from 1997 to 2021. Patients with ACTBAD were compared with those having surgery for other reasons. Logistic regression was used to identify associations with major adverse events (MAEs). Five-year survival and competing risk of reintervention were calculated. RESULTS: Of 926 patients, 75 (8.1%) had ACTBAD. Indications included rupture (25/75), malperfusion (11/75), rapid expansion (26/75), recurrent pain (12/75), large aneurysm (5/75), and uncontrolled hypertension (1/75). The incidence of MAEs was similar (13.3% [10/75] vs 13.7% [117/851], P = .99). Operative mortality was 5.3% (4/75) vs 4.8% (41/851) (P = .99). Complications included tracheostomy (8%, 6/75), spinal cord ischemia (4%, 3/75), and new dialysis (2.7%, 2/75). Renal impairment, urgent/emergent operation, forced expiratory volume in 1 second ≤50%, and malperfusion were associated with MAEs, but not ACTBAD (odds ratio: 0.48, 95% confidence interval [CI]: [0.20-1.16], P = .1). At 5 and 10 years, there was no difference in survival (65.8% [95% CI: 54.6-79.2] vs 71.3% [95% CI: 67.9-74.9], P = .42, and 47.3% [95% CI: 34.5-64.7] vs 53.7% [95% CI: 49.3-58.4], P = .29, respectively) or 10-year reintervention (12.5% [95% CI: 4.3-25.3] vs 7.1% [95% CI: 4.7-10.1], P = .17, respectively). CONCLUSIONS: In an experienced center, open repair of ACTBAD can be performed with low rates of operative mortality and morbidity. Outcomes similar to elective repair are achievable even in high-risk patients with ACTBAD. In patients unsuitable for endovascular repair, transfer to a high-volume center experienced in open repair should be considered.

Noonan syndrome associated with hypoplastic left heart syndrome.

Noonan syndrome is an inherited disorder caused by alterations in the RAS-MAPK pathway. There have been several identified genotype-phenotype associations made with respect to congenital cardiac lesions and Noonan syndrome variants, but limited data exist regarding single ventricle disease in this population. Here, we report two patients with PTPN11-related Noonan syndrome and hypoplastic left heart syndrome variants.

Radiographic and histologic characterisation of white matter injury in a sheep model of CHD.

Nearly one in five children with CHD is born with white matter injury that can be recognised on postnatal MRI by the presence of T1 hyperintense lesions. This pattern of white matter injury is known to portend poor neurodevelopmental outcomes, but the exact aetiology and histologic characterisation of these lesions have never been described. A fetal sheep was cannulated at gestational age 110 days onto a pumpless extracorporeal oxygenator via the umbilical vessels and supported in a fluid environment for 14.5 days. The fetus was supported under hypoxic conditions (mean oxygen delivery 16 ml/kg/day) to simulate the in utero conditions of CHD. At necropsy, the brain was fixed, imaged with MRI, and then stained to histologically identify areas of injury. Under hypoxemic in utero conditions, the fetus developed a T1 hyperintense lesion in its right frontal lobe. Histologically, this lesion was characterised by microvascular proliferation and astrocytosis without gliosis. These findings may provide valuable insight into the aetiology of white matter injury in neonates with CHD.

Chronic hypoxemia induces mitochondrial respiratory complex gene expression in the fetal sheep brain.

Objective: The molecular pathways underlying hypoxemia-induced alterations in neurodevelopment of infants with congenital heart disease have not been delineated. We used transcriptome analysis to investigate differential gene expression induced by hypoxemia in an ovine artificial-womb model. Methods: Mid-gestation fetal sheep (median [interquartile range] 109 [107-112] days' gestation) were cannulated via the umbilical vessels, attached to a pumpless, low-resistance oxygenator circuit, and incubated in a sterile, fluid environment for 22 [21-23] days. Fetuses were maintained with an oxygen delivery of 20-25 mL/kg/min (normoxemia, n = 3) or 14-16 mL/kg/min (hypoxemia, n = 4). Transcriptional profiling by RNA sequencing was carried out on left frontal brains and hypoxemia-regulated genes were identified by differential gene expression analysis. Results: A total of 228 genes whose expression was up or down regulated by ≥1.5-fold (false discovery rate ≤0.05) were identified. The majority of these genes were induced in hypoxemic animals compared to normoxemic controls, and functional enrichment analysis identified respiratory electron transport as a pathway strongly upregulated in the brain during chronic hypoxemia. Further examination of hypoxemia-induced genes showed robust induction of all 7 subunits of the mitochondrial NADH:ubiquinone oxidoreductase (complex I). Other hypoxemia-induced genes included cytochrome B, a component of complex III, and ATP6, ATP8, both of which are components of complex V. Conclusions: Chronic fetal hypoxemia leads to upregulation of multiple mitochondrial respiratory complex genes critical for energy production and reactive oxygen species generation, including complex I. These data provide valuable insight into potential pathways involved in chronic hypoxemia-induced neuropathology and offers potential therapeutic targets for fetal neuroprotection in fetuses with congenital heart defects.

Chronic foetal hypoxaemia does not cause elevation of serum markers of brain injury.

OBJECTIVES: The objective of this study was to investigate changes in serum biomarkers of acute brain injury, including white matter and astrocyte injury during chronic foetal hypoxaemia. We have previously shown histopathological changes in myelination and neuronal density in fetuses with chronic foetal hypoxaemia at a level consistent with CHD. METHODS: Mid-gestation foetal sheep (110 ± 3 days gestation) were cannulated and attached to a pumpless, low-resistance oxygenator circuit, and incubated in a sterile fluid environment mimicking the intrauterine environment. Fetuses were maintained with an oxygen delivery of 20-25 ml/kg/min (normoxemia) or 14-16 ml/kg/min (hypoxaemia). Myelin Basic Protein and Glial Fibrillary Acidic Protein serum levels in the two groups were assessed by ELISA at baseline and at 7, 14, and 21 days of support. RESULTS: Based on overlapping 95% confidence intervals, there were no statistically significant differences in either Myelin Basic Protein or Glial Fibrillary Acidic Protein serum levels between the normoxemic and hypoxemic groups, at any time point. No statistically significant correlations were observed between oxygen delivery and levels of Myelin Basic Protein and Glial Fibrillary Acidic Protein. CONCLUSION: Chronic foetal hypoxaemia during mid-gestation is not associated with elevated serum levels of acute white matter (Myelin Basic Protein) or astrocyte injury (Glial Fibrillary Acidic Protein), in this model. In conjunction with our previously reported findings, our data support the hypothesis that the brain dysmaturity with impaired myelination found in fetuses with chronic hypoxaemia is caused by disruption of normal developmental pathways rather than by direct cellular injury.

Attrition between the superior cavopulmonary connection and the Fontan procedure in hypoplastic left heart syndrome.

OBJECTIVE: We investigated the incidence and predictors of failure to undergo the Fontan in children with hypoplastic left heart syndrome who survived superior cavopulmonary connection. METHODS: The cohort consists of all patients with hypoplastic left heart syndrome who survived to hospital discharge after superior cavopulmonary connection between 1988 and 2017. The primary outcome was attrition, which was defined as death, nonsuitability for the Fontan, or cardiac transplantation before the Fontan. Subjects were excluded if they were awaiting the Fontan, were lost to follow-up, or underwent biventricular repair. The study period was divided into 4 eras based on changes in operative or medical management. Attrition was estimated with 95% confidence intervals, and predictors were identified using adjusted, logistic regression models. RESULTS: Of the 856 hospital survivors after superior cavopulmonary connection, 52 died, 7 were deemed unsuitable for Fontan, and 12 underwent or were awaiting heart transplant. Overall attrition was 8.3% (71/856). Attrition rate did not change significantly across eras. A best-fitting multiple logistic regression model was used, adjusting for superior cavopulmonary connection year and other influential covariates: right ventricle to pulmonary artery shunt at Norwood (P < .01), total support time at superior cavopulmonary connection (P < .01), atrioventricular valve reconstruction at superior cavopulmonary connection (P = .02), performance of other procedures at superior cavopulmonary connection (P = .01), and length of stay after superior cavopulmonary connection (P < .01). CONCLUSIONS: In this study spanning more than 3 decades, 8.3% of children with hypoplastic left heart syndrome failed to undergo the Fontan after superior cavopulmonary connection. This attrition rate has not decreased over 30 years. Use of a right ventricle to pulmonary artery shunt at the Norwood procedure was associated with increased attrition.

Contrast-Enhanced Brain Ultrasound Perfusion Metrics in the EXTra-Uterine Environment for Neonatal Development (EXTEND): Correlation With Hemodynamic Parameters.

OBJECTIVES: Contrast-enhanced ultrasound (CEUS) can provide quantitative perfusion metrics and may be useful to detect cerebral pathology in neonates and premature infants, particularly in extrauterine environments. The effect of hemodynamics on cerebral perfusion metrics is unknown, which limits the clinical application of this technology. We aimed to determine associations between systemic hemodynamics and concurrently measured brain perfusion parameters in an animal model of extrauterine support. METHODS: Nine fetal lambs were transferred to an extrauterine support device. Lumason® ultrasound contrast (0.1-0.3 ml) was administered via the umbilical vein and 90-second cine clips were obtained. Time-intensity-curves (TICs) were generated and time-dependent and area-under-curve (AUC) parameters were derived. Associations between brain perfusion metrics and hemodynamics including heart rate (HR) and mean arterial pressure (MAP) were evaluated by multilevel linear mixed-effects models. RESULTS: Eighty-six ultrasound examinations were performed and 72 examinations were quantifiable. Time-dependent measurements were independent of all hemodynamic parameters (all p ≥.05). Oxygen delivery and mean blood flow were correlated with AUC measurements (all p ≤.01). Physiologic HR and MAP were not correlated with any measurements (all p ≥.05). CONCLUSION: Detected aberrations in time-dependent CEUS measurements are not correlated with hemodynamic parameters and are thought to reflect the changes in cerebral blood flow, thus providing a promising tool for evaluation of brain perfusion. CEUS brain perfusion parameters are not correlated with physiologic HR and MAP, but AUC-dependent measurements are correlated with oxygen delivery and blood flow, suggesting that CEUS offers additional value over standard monitoring. Overall, these findings enhance the applicability of this technology.

Off-Pump Explant of a Left Ventricular Assist Device Using a Recovery Plug in a Pediatric Patient.

Though the numbers remain small, the use of continuous-flow left ventricular assist devices as a bridge to recovery in pediatric patients has been increasing. Select patients may have sufficient myocardial recovery to allow for device removal. Here, we describe a 13-year old requiring left ventricular assist device implantation for myocarditis who was referred for explant of the device after recovery. This was performed via thoracotomy, without cardiopulmonary bypass, using a newly developed titanium recovery plug that is custom designed to fit the HeartMate 3.

Quantitative contrast-enhanced ultrasound of the brain on twin fetal lambs maintained by the extrauterine environment for neonatal development (EXTEND): initial experience.

BACKGROUND: With the development of an artificial environment to support the extremely premature infant, advanced imaging techniques tested in this extrauterine system might be beneficial to evaluate the fetal brain. OBJECTIVE: We evaluated the feasibility of (a) performing contrast-enhanced ultrasound (CEUS) and (b) quantifying normal and decreased brain perfusion in fetal lambs maintained on the extrauterine environment for neonatal development (EXTEND) system. MATERIALS AND METHODS: Twin premature fetal lambs (102 days of gestational age) were transferred to the EXTEND system. Twin B was subjected to sub-physiological flows (152 mL/kg/min) and oxygen delivery (15.9 mL/kg/min), while Twin A was maintained at physiological levels. We administered Lumason contrast agent into the oxygenator circuit and performed serial CEUS examinations. We quantified perfusion parameters and generated parametric maps. We also recorded hemodynamic parameters, serum blood analysis, and measurements across the oxygenator. Postmortem MRIs were performed. RESULTS: No significant changes in hemodynamic variables were attributable to CEUS examinations. On gray-scale images, Twin B demonstrated ventriculomegaly and progressive parenchymal volume loss culminating in hydranencephaly. By CEUS, Twin B demonstrated decreased peak enhancement and decreased overall parenchymal perfusion when compared to Twin A by perfusion parameters and parametric maps. Changes in perfusion parameters were detected immediately following blood transfusion. Postmortem MRI confirmed ultrasonographic findings in Twin B. CONCLUSION: In this preliminary experience, we show that CEUS of the brain is feasible in fetal lambs maintained on the EXTEND system and that changes in perfusion can be quantified, which is promising for the application of CEUS in this extrauterine system supporting the premature infant.

Fetoscopic insufflation modeled in the extrauterine environment for neonatal development (EXTEND): Fetoscopic insufflation is safe for the fetus.

BACKGROUND: Minimally invasive fetal surgery, or fetoscopy, is an alternative to open fetal surgery to repair common birth defects like myelomeningocele. Although this hysterotomy-sparing approach reduces maternal morbidity, the effects of in utero insufflation on the fetus are poorly understood. Our purpose was to determine the optimal fetal insufflation conditions. METHODS: Fetal sheep at gestational age 104 to 107 days were studied under insufflation conditions in utero and ex utero. The ex utero fetuses were cannulated via their umbilical vessels into a support device, the EXTra-uterine Environment for Neonatal Development (EXTEND). EXTEND fetuses were exposed to four different insufflation conditions for four hours: untreated carbon dioxide (CO2) (n=5), warm humidified (whCO2) (n=4), whCO2 with the umbilical cord exposed (n=3), and whCO2 without amniotic fluid (skin and cord exposed) (n=3). RESULTS: In utero insufflation led to significant increases in fetal CO2 and reductions in fetal pH. Ex utero insufflation with whCO2 did not lead to changes in fetal blood gas measurements or cerebral perfusion parameters. Insufflation with whCO2 with an exposed umbilical cord led to reduced umbilical blood flow. CONCLUSIONS: Insufflation with warm humidified CO2 with an amniotic fluid covered umbilical cord is well tolerated by the fetus without significant changes in hemodynamics or cerebral perfusion parameters. TYPE OF STUDY: Basic science LEVEL OF EVIDENCE: N/A.

A Rabbit Model for Optimization of Amniotic Fluid Components in the EXTrauterine Environment for Newborn Development (EXTEND) System.

In this model article, we present a protocol for continuous amniotic fluid exchange in rabbits using a novel system to test the effects of growth factor-deficient, artificial amniotic fluid on bowel development. BACKGROUND: Ideally, the EXTrauterine Environment for Neonatal Development (EXTEND) will provide physiologic support to the extreme premature infant. An important component of that environment is the amniotic fluid. Thus, we developed an animal model to study the growth factors found within amniotic fluid and inform design of a synthetic fluid to optimize fetal development. METHODS: We designed a model of amniotic fluid exchange within the pregnant rabbit, continuously removing the natural fluid from around 2 fetuses per doe and replacing it with a physiologic electrolyte solution during the final 100 h of gestation. Two fetuses from the contralateral uterine horn were used as sham-operated controls. Thirty-eight fetuses were analyzed, 19 in each group. We analyzed the fetal growth and bowel development. RESULTS: Ultrasound after 100 h of exchange showed equivalent fluid volumes, p = 0.63. Cultures were negative for bacterial colonization. Final fluid protein concentrations were 11.6% that of control fluid (mean 1,451 ± 224.2 vs. 12,491 ± 849.2 µg/mL). There was no significant difference in fetal growth, with experimental weights 91.4% of control weights, p = 0.07. Fetal bowel weights (90.1%, p = 0.16) and lengths (94.2%, p = 0.49) were also not significantly less compared to controls. There was no significant difference in villous height or crypt depth measurements between the groups, and absorptive capacity of the bowel was not different between groups, p = 0.44. CONCLUSION: This animal model allows for manipulation of the components of amniotic fluid. Marked reduction of natural amniotic fluid proteins during gestation does not appear to significantly impair fetal growth or bowel development. Further work with this model will assess the importance of amniotic fluid components for normal development to inform design of a synthetic fluid for use during EXTEND.

The EXTrauterine Environment for Neonatal Development Supports Normal Intestinal Maturation and Development.

BACKGROUND AND AIMS: The Extra-Uterine Environment for Neonatal Development (EXTEND) aims to avoid the complications of prematurity, such as NEC. Our goal was to determine if bowel development occurs normally in EXTEND-supported lambs, with specific emphasis on markers of immaturity associated with NEC. METHODS: We compared terminal ileum from 17 pre-term lambs supported on EXTEND for 2- 4 weeks to bowel from age-matched fetal lambs that developed in utero. We evaluated morphology, markers of epithelial integrity and maturation, enteric nervous system structure, and bowel motility. RESULTS: EXTEND-supported lamb ileum had normal villus height, crypt depth, density of mucin-containing goblet cells, and enteric neuron density. Expression patterns for I-FABP, activated caspase-3 and EGFR were normal in bowel epithelium. Transmural resistance assessed in Ussing chambers was normal. Bowel motility was also normal as assessed by ex vivo organ bath and video imaging. However, Peyer's patch organization did not occur normally in EXTEND ileum, resulting in fewer circulating B cells in experimental animals. CONCLUSION: EXTEND supports normal ileal epithelial and enteric nervous system maturation in pre-term lambs. The classic morphologic changes and cellular expression profiles associated with NEC are not seen. However, immune development within the EXTEND supported lamb bowel does not progress normally.

Prenatal hypoxemia alters microglial morphology in fetal sheep.

OBJECTIVE: Neuroimmune cells, particularly microglia and astrocytes, play a critical role in neurodevelopment. Neurocognitive delays are common in children with congenital heart disease, but their etiology is poorly understood. Our objective was to determine whether prenatal hypoxemia, at levels common in congenital heart disease, induced neuroimmune activation to better understand the origins of neurobehavioral disorders in congenital heart disease. METHODS: Eight fetal sheep at gestational age 109 ± 3 days (term ∼145 days) were cannulated onto a pumpless extracorporeal oxygenator via the umbilical vessels and supported in a fluid environment for 22 ± 2 days under normoxic (n = 4) or hypoxic (n = 4) conditions. Control fetuses (n = 7) were harvested at gestational age 133 ± 4 days. At necropsy, brains were stained with ionized calcium-binding adaptor molecule 1 and glial fibrillary acidic protein antibodies to quantify microglia and astrocytes, respectively, in gray and white matter in frontotemporal and cerebellar sections. Microglia were classified into 4 morphologic types based on cell shape. Data were analyzed with 1-way analysis of variance or Fisher exact test, as appropriate. RESULTS: Oxygen delivery was significantly reduced in hypoxic fetuses (15.6 ± 1.8 mL/kg/min vs 24.3 ± 2.3 mL/kg/min; P < .01). Rates of apoptosis were similar in hypoxic, normoxic, and intrauterine control animals in all examined areas. There were also no differences between groups in area occupied by glial fibrillary acidic protein-labeled astrocytes or ionized calcium-binding adaptor molecule 1-labeled microglia in all examined areas. However, round microglia were significantly increased in hypoxic animals compared with normoxic animals (33% vs 6%; P < .01) and control animals (33% vs 11%; P < .01). CONCLUSIONS: Prenatal hypoxemia altered microglial morphology without significant gliosis. Additional studies characterizing these mechanisms may provide insight into the origins of neurobehavioral disabilities in children with congenital heart disease.

The Effects of Nitric Oxide in Oxygenator Sweep Gas During Extracorporeal Circulation in a Neonatal Ovine Model.

Extracorporeal membrane oxygenation is a life-saving intervention, but bleeding complications are frequent. Given that the combination of platelet loss and dysfunction is a major contributor to this acquired bleeding diathesis, efforts to combat these phenomena are of great clinical importance. In this study, we investigated the effects of nitric oxide (NO) added to the sweep gas of an extracorporeal circuit in a neonatal ovine model. Eight lambs (age 9.6 ± 1.9 days) were cannulated via the neck vessels and maintained on a pumpless arteriovenous extracorporeal membrane oxygenation circuit with blood flow restricted to 100 ml/min for 72 hours. All animals were heparinized, and a subset (n = 4) also received NO in the sweep gas at a concentration of 200 ppm. We observed no adverse effects from NO administration, and methemoglobin levels remained unchanged. Platelet counts significantly declined in all animals over the course of the study; however, mean counts were higher in the NO-treated group, and this difference was statistically significant at 24 hours (62 ± 3% vs. 32 ± 7% of baseline, P < 0.01). Likewise, mean plasma levels of beta-thromboglobulin, a marker of platelet activation, were lower in the NO-treated group, and this difference was also significant at the 24 hour time point (9.5 ± 2.2 vs. 19.7 ± 6.5 pg/mL/10 platelets, P < 0.05). We conclude that 200 ppm NO can be safely blended into the oxygenator sweep gas of a low-flow extracorporeal circuit and that it may transiently attenuate platelet consumption and activation.

Inhaled Nitric Oxide Is Associated with Improved Oxygenation in a Subpopulation of Infants with Congenital Diaphragmatic Hernia and Pulmonary Hypertension.

OBJECTIVES: To determine which patients with congenital diaphragmatic hernia (CDH) and pulmonary hypertension (PH) benefit from inhaled nitric oxide (iNO) treatment by comparing characteristics and outcomes of iNO responders to nonresponders. STUDY DESIGN: We performed a retrospective chart review of infants with CDH treated at our center between 2011 and 2016. In a subset of patients, iNO was initiated for hypoxemia or echocardiographic evidence of extrapulmonary right to left shunting. Initial post-treatment blood gases were reviewed, and patients were classified as responders (increased PaO2 >20 mm Hg) or nonresponders. Baseline characteristics, echocardiograms and outcomes were compared between groups with Fisher exact tests and Mann-Whitney t tests, as appropriate. RESULTS: During the study period, 95 of 131 patients with CDH (73%) were treated with iNO. All patients with pretreatment echocardiograms (n = 90) had echocardiographic evidence of PH. Thirty-eight (40%) patients met treatment response criteria. Responders had significant improvements in PaO2 (51 ± 3 vs 123 ± 7 mm Hg, P < .01), alveolar-arterial gradient (422 ± 30 vs 327 ± 27 mm Hg, P < .01), and PaO2 to FiO2 ratio (82 ± 10 vs 199 ± 15 mm Hg, P < .01). Nonresponders were more likely to have left ventricular systolic dysfunction (27% vs 8%, P = .03) on echocardiogram. Responders were less likely to require extracorporeal membrane support (50 vs 24%, P = .02). CONCLUSIONS: iNO treatment is associated with improved oxygenation and reduced need for ECMO in a subpopulation of patients with CDH with PH and normal left ventricular systolic function.

Premature Lambs Exhibit Normal Mitochondrial Respiration after Long-Term Extrauterine Support.

BACKGROUND: In an effort to mitigate the major morbidities and mortality associated with extreme prematurity, we have developed an EXTrauterine Environment for Neonatal Development (EXTEND) designed to provide physiologic support of extremely premature infants. OBJECTIVES: We have previously shown that long-term, physiologic support of premature fetal lambs is possible with EXTEND, but in this study, we sought to demonstrate bioenergetic equipoise at the tissue level. METHODS: Four premature fetal lambs were delivered by hysterotomy at gestational ages (GA) of 105-107 days (term ∼145 days), cannulated via the umbilical vessels, and transitioned to support on EXTEND for 3-4 weeks. Five control fetuses were age-matched to the GA of experimental fetuses at the time of study end (128-134 days GA) and immediately sacrificed after hysterotomy. Mitochondria were isolated from the heart, liver, kidney, and skeletal muscle of fetuses at the time of sacrifice, and oxygen consumption rates (OCRs) were measured. RESULTS: There were no differences in basal mitochondrial OCR between EXTEND and control fetuses for heart, kidney, or skeletal muscle. For liver, the basal OCR was higher in EXTEND fetuses compared to controls. There were no differences in physiologic maximal OCR or reserve capacity for any tissue analyzed. CONCLUSIONS: Fetal lambs supported by EXTEND demonstrate physiologic mitochondrial function as evidenced by adequate basal and physiologic maximal cellular respiration as well as preserved reserve capacity.

Erythropoietin Prevents Anemia and Transfusions in Extremely Premature Lambs Supported by an EXTrauterine Environment for Neonatal Development (EXTEND).

BACKGROUND: We recently developed an EXTrauterine Environment for Neonatal Development (EXTEND) that provides physiologic support for premature lambs. Here, we assess the efficacy of exogenous erythropoietin (EPO) to prevent anemia and transfusions on EXTEND. MATERIALS AND METHODS: Lambs were cannulated at 0.7 gestation and supported on EXTEND for up to 4 weeks. The lambs were divided into three groups: (1) No EPO, (2) Low EPO (300 U kg-1 per day), and (3) High EPO (800 U kg-1 per day). Daily hematocrit and weekly complete blood count were assessed. RESULTS: The mean percentage change in hematocrit from baseline was significantly different between the groups (No EPO -23.6 ± 7.8% vs. Low EPO -16.6 ± 6.4% vs. High EPO +2.6 ± 6.6%; p = 0.02). This occurred despite a greater median number of blood transfusions in the No EPO group (5 vs. 1 vs. 0; p = 0.02). EPO administration was associated with a higher mean corpuscular volume (MCV; p < 0.01) and reticulocyte count (p = 0.02). The High EPO group was comparable to in utero control fetuses with respect to hematocrit (p = 0.49), MCV (p = 0.24), and reticulocyte count (p = 0.68). CONCLUSIONS: EPO (800 U kg-1 per day) prevents anemia, eliminates transfusions, and restores normal red blood cell indices in premature lambs supported by EXTEND.