The amniotic fluid as a source of cells for fetal tissue engineering.

PURPOSE: This study was aimed at determining whether fetal tissue constructs can be engineered from cells normally found in the amniotic fluid. METHODS: A subpopulation of morphologically distinct cells was isolated mechanically from the amniotic fluid of pregnant ewes (n = 5) and expanded selectively. Its lineage was determined by immunofluorescent staining against multiple intermediate filaments and surface antigens. Proliferation rates were determined by both oxidation and total DNA assays and compared with immunocytochemically identical adult and fetal sheep cells. Statistical analysis was by analysis of variance for repeated measures (ANOVA). After expansion, the amniocytes were seeded onto a polyglycolic acid polymer/poly-4-hydroxybutyrate scaffold. The resulting construct was analyzed by both optical and scanning electron microscopy. RESULTS: The immunocytochemical profile of expanded amniocytes was consistent with a mesenchymal, fibroblast/myofibroblast cell lineage. These cells proliferated significantly faster than comparable fetal and adult cells in culture. Amniocyte construct analysis showed dense, confluent layers of cells firmly attached to the scaffold, with no evidence of cell death. CONCLUSIONS: (1) Subpopulations of fetal mesenchymal cells can be isolated consistently from the amniotic fluid. (2) Mesenchymal amniocytes proliferate more rapidly in vitro than comparable fetal and adult cells. (3) Mesenchymal amniocytes attach firmly to polyglycolic acid polymer. The amniotic fluid can be a reliable and practical source of cells for the engineering of select fetal tissue constructs.

Continuous intrapulmonary distension with perfluorocarbon accelerates lung growth in infants with congenital diaphragmatic hernia: initial experience.

BACKGROUND/PURPOSE: The authors have shown previously in an animal model that neonatal lung growth can be accelerated by continuous intrapulmonary distension with a perfluorocarbon (PFC). The authors now describe a preliminary clinical experience with this therapeutic concept in a select group of infants with congenital diaphragmatic hernia (CDH). METHODS: Neonates with very high predicted mortality rate caused by CDH had their lungs completely filled with PFC while on extracorporeal life support (ECLS); (n = 5). A continuous positive pressure of 7 to 10 cm H2O was maintained via the endotracheal tube for 3 to 7 days (mean, 5.6 +/- 0.87 days). The areas of both lungs (L) then were measured daily from digitized chest x-rays and divided by the area of the corresponding L1 vertebrae (V), to create an L/V index, so as to control for variable roentgenographic techniques. Immediately after removal of PFC, blood gas data were collected off ECLS. RESULTS: At the end of continuous pulmonary distension, all patients showed improvements in oxygenation and ventilation. The ipsilateral lungs showed significant increase of the L/V index with time (P =.003) and of L/V's daily change (P <.0001), suggesting accelerated lung growth. Overall survival rate was 40% (2 of 5). Of the 3 patients that had 7 days of distension, 2 survived. CONCLUSIONS: Continuous intrapulmonary distension with PFC for up to 1 week accelerated ipsilateral lung growth, improved gas exchange, and increased survival of CDH infants with profound pulmonary hypoplasia marooned on ECLS. Additional trials of PFC-based pulmonary distension in similar infants are warranted.

Fetal tissue engineering: diaphragmatic replacement.

BACKGROUND/PURPOSE: Prosthetic repair of congenital diaphragmatic hernia has been associated with high complication rates. This study was aimed at applying fetal tissue engineering to diaphragmatic replacement. METHODS: Fetal lambs underwent harvest of skeletal muscle specimens. Once expanded in vitro, fetal myoblasts were suspended in a collagen hydrogel submitted to controlled radial tension. The construct was then placed in a bioreactor. After birth, all animals underwent creation of 2 diaphragmatic defects. One defect was repaired with the autologous-engineered construct placed in between 2 acellular supporting membranes and the other with an identical construct but without any cells. Each animal was its own control (graft, n = 10). Animals were killed at different time-points postimplantation for histologic examination. Statistical analysis was by analysis of variance (ANOVA). RESULTS: Fetal myoblasts expanded up to twice as fast as neonatal cells. Hydrogel-based radial tension enhanced construct architecture by eliciting cell organization within the scaffold. No eventration was present in 4 of 5 engineered constructs but in 0 of 5 acellular grafts (P<.05). At harvest, engineered constructs were thick and histologically resembled normal skeletal muscle, whereas acellular grafts were thin, floppy, and showed low cell density with increased fibrosis. CONCLUSIONS: Unlike acellular grafts, engineered cellular diaphragmatic constructs are anatomically and histologically similar to normal muscle. Fetal tissue engineering may be a viable alternative for diaphragmatic replacement.

Prolonged local myometrial blockade prevents preterm labor after fetal surgery in a leporine model.

BACKGROUND/PURPOSE: Postoperative premature labor remains the foremost limiting factor to the development of fetal surgery. Most attempts at controlling this complication have involved the use of drugs delivered systemically to the mother. This study assessed the effects of prolonged local anesthetic blockade of the myometrium on preterm delivery after open fetal surgery. METHODS: Eighteen New Zealand rabbits at 23 days' gestation (term, 31 to 33 days) were divided in three groups. In group I (n = 6), the most proximal fetuses of both uterine horns were submitted to open amputation of a forelimb; in a few animals, one of the uterine horns was empty, hence, only one fetus was manipulated. In groups II (n = 5) and III (n = 7), an identical surgical procedure was performed. In group II, immediately before hysterotomy, the myometrium was injected with 0.5 mL of 0.5% bupivacaine along the incision line. In group III, only saline was injected. In group II, before uterine closure, the incised area of the myometrium was injected with 1.5 mL of a novel suspension of biodegradable polylactic-co-glycolic acid microspheres loaded with 75% w/w bupivacaine and 0.05% w/w dexamethasone. This suspension previously has been shown to provide peripheral nerve blockade for approximately 5 days. In group III, microspheres without any drug were injected. RESULTS: Abortion rates were significantly different among the groups: 83.3% (five of six) for the does in group I, zero in group II, and 71.4% (five of seven) in group III (P < .05). The absence of abortions observed in group II occurred despite the fact that the fetal mortality rate was significantly higher in this group (87.5%, seven of eight fetuses) than in groups I (0) and III (33.3%, 4 of 12 fetuses, P < .05). CONCLUSIONS: Prolonged local blockade of the myometrium with bupivacaine inhibits preterm labor after fetal surgery in rabbits. The high fetal mortality rate observed in this study may be caused by "transplacental" transfer of the local anesthetic to the fetus. Notably, the abortifacient effect of a dead fetus was completely suppressed by the local blockade. Studies using microspheres with local anesthetics that do not cross the placenta, in animal models with longer gestational periods, are warranted.

Fetal response to neutral gas and liquid media for intraamniotic distension.

PURPOSE: This study was aimed at comparing the effects of a neutral liquid and a neutral gas used as intraamniotic media on umbilical blood flow, O2 delivery, blood pressure, acid-base status, and electrolytes in the fetus at escalating intraamniotic pressures. METHODS: Eight fetal lambs underwent invasive monitoring of common umbilical blood flow, blood pressure, blood gases, sodium, and hematocrit, as intraamniotic pressure was raised from 0 to 30 mm Hg. The animals were divided equally in two groups depending on the intraamniotic medium used (group I, warmed saline and group II, air). Maternal systemic blood pressure, O2 saturation, and temperature were kept constant. RESULTS: In each group, a threshold level of intraamniotic pressure was evident, above which there was a significant decrease in the common umbilical artery blood flow, with concomitant fetal hypoxemia and hypercarbia. This intraamniotic pressure threshold was 20 mm Hg in group I (saline), but only 15 mm Hg in group II (air). CONCLUSIONS: Although both a neutral liquid and a neutral gas can safely be used as intraamniotic media, a neutral liquid medium allows for a wider range of safe intrauterine working pressure (0 to 20 mm Hg), as compared with a neutral gas (0-15 mm Hg).

Fetal carotid blood flow during videofetoscopy.

BACKGROUND/PURPOSE: Intracranial bleeding has been reported as one of the complications of both open and minimally invasive fetal surgery and putatively attributed to intraoperative fluctuations of carotid blood flow. The aim of this study was to look at fetal carotid blood flow and its relationship with umbilical blood flow, blood pressure, oxygen delivery, and acid-base status in the fetus at various intraamniotic pressures with both liquid and gas media during fetoscopic surgery. METHODS: Six 115- to 130-day-gestation ewes underwent continuous invasive systemic blood pressure monitoring in the descending aorta. A hysterotomy was performed. A 6-mm ultrasonic blood flow probe was placed around the common umbilical artery at its origin from the fetal aorta. This was followed by placement of a double-lumen, 4F catheter in the fetal descending aorta through a femoral artery. A 4-mm ultrasonic blood flow probe was then placed around the fetal left common carotid artery. A pressure-monitoring, multiperforated catheter was placed inside the amniotic cavity. The fetus was repositioned inside the uterus, which was then closed. The abdominal wall was closed loosely. No further manipulation was performed for 1 hour. Intraamniotic pressure was raised from 0 to 30 mm Hg at 5-mm Hg intervals by infusing either warmed saline or medical air. Common umbilical artery and left carotid artery blood flows, blood pressure, blood gases, bicarbonate, sodium, and hematocrit were recorded in all fetuses at each 5-mm Hg interval. Maternal systemic blood pressure, O2 saturation, and temperature were kept constant. RESULTS: Carotid blood flow remained stable within the intra-amniotic pressure range studied (0 to 30 mm Hg), despite the significant drop in common umbilical artery blood flow uniformly observed above 20 mm Hg when saline was infused and above 15 mm Hg when air was infused. There was fetal hypoxemia and hypercarbia concomitant with decreased common umbilical artery blood flow (however, without fetal acidosis, because of compensatory elevation of bicarbonate). Within the intraamniotic pressure range studied, fetal aortic blood pressure, sodium, and hematocrit did not vary significantly, even when there was decreased umbilical blood flow. CONCLUSIONS: Fetal carotid blood flow is protected, possibly autoregulated, remaining stable even after umbilical blood flow decreases as a consequence of elevated intrauterine pressures up to 30 mm Hg during videofetoscopy. These data suggests that perioperative intracranial bleeding related to videofetoscopic surgery is caused by factors other than fluctuations of cerebral blood flow.

Insulin-like growth factor-I gene expression in three models of accelerated lung growth.

BACKGROUND/PURPOSE: We have learned previously that in utero tracheal ligation reverses the structural and physiological effects of surgically created congenital diaphragmatic hernia. In addition, we have discovered that postnatal lung growth similarly can be accelerated using liquid-based airway distension with perfluorocarbon. Another model of accelerated lung growth is that of compensatory growth seen after neonatal pneumonectomy. In all of these models, growth has occurred because of an increase in alveolar number rather than enlargement of preexisting alveoli. However, the molecular mechanisms underlying these processes remain unknown. The purpose of this study was to determine if gene expression could be altered by changes in physical forces in the prenatal and postnatal lung. METHODS: The three models of accelerated lung growth studied were the following: (1) The prenatal group, consisted of fetal lambs (n = 12) that underwent the surgical creation of a left diaphragmatic hernia at 90 days' gestation. Six of these animals also underwent simultaneous tracheal ligation. (2) The PFC group consisted of five neonatal animals that underwent isolation of the superior segment of the right upper lobe, with intrabronchial distension with perfluorocarbon to 7 to 10 mm Hg pressure for a 3-week period. (3) The postpneumonectomy group consisted of four neonatal animals that underwent left pneumonectomy. In the fetal study, lungs were retrieved at term (130 days), and in the postnatal study, lungs were retrieved 3 weeks after initial intervention. In all cases, RNA was extracted from snap-frozen lung samples and Northern blot analysis performed. RESULTS: Insulinlike growth factor-I, insulinlike growth factor-II, and vascular endothelial growth factor gene expression were analyzed by densitometry. Insulinlike growth factor-I gene expression was found to be decreased in association with experimental diaphragmatic hernia (P = .005), but restored to normal with tracheal ligation. Insulinlike growth factor-I gene expression was significantly increased in both postnatal models of accelerated lung growth (P = .022, P = .016). No significant differences were found in insulinlike growth factor-II or vascular endothelial growth factor gene expression. CONCLUSIONS: The authors conclude from these preliminary data that (1) insulin like growth factor-I gene expression is reduced in experimental fetal diaphragmatic hernia and restored to normal by tracheal ligation, and (2) insulinlike growth factor-I gene expression is increased in both the liquid-based airway distension and postpneumonectomy models of accelerated postnatal lung growth. The authors speculate that all of these manipulations exploit a natural pathway essential for normal lung growth.

Continuous intrapulmonary distension with perfluorocarbon accelerates neonatal (but not adult) lung growth.

BACKGROUND/PURPOSE: We have previously demonstrated that experimental fetal tracheal ligation reverses the structural and physiological effects of pulmonary hypoplasia associated with congenital diaphragmatic hernia. The purpose of this study was to determine if lung growth could be similarly accelerated postnatally by continuous liquid-based intrapulmonary distension. METHODS: Ten neonatal lambs were divided into two experimental groups. Five neonatal animals underwent a right thoracotomy with isolation of the anterior superior segment of the right upper lobe. A pressure monitoring catheter was introduced and perfluorocarbon (PFC) was instilled into the segment. Animals were subjected to a 21-day distention period with continuous maintenance of 7 to 10 mm Hg intrabronchial pressure. Five other neonatal animals used as age- and weight-matched controls were killed immediately after distension with PFC to 7 to 10 mm Hg. To evaluate the effect of age on postnatal growth, identical procedures were performed on seven mature sheep. Four adult animals underwent a 21-day distension with PFC, and three animals were killed immediately after PFC distension. RESULTS: Neonatal animals who underwent distension showed a significant acceleration of lung growth based on right upper lobe volume to body weight ratio (P = .0019), total alveolar number (P = .003), and total alveolar surface area (P = .006), when compared with controls. Alveolar growth was attributed to an increased alveolar number rather than increased alveolar size based on a normal histological appearance, normal airspace fraction (P = NS), and normal alveolar numerical density (P = NS). In contrast, no significant differences in lung growth or maturation indices were present in adult animals. CONCLUSIONS: From this preliminary data we conclude: (1) Liquid-based airway distension does accelerate postnatal lung growth, (2) lung architecture remains normal during this period of accelerated growth, (3) adult sheep do not respond to liquid-based airway distension with lung growth, and (4) prolonged exposure to intrapulmonary PFC appears to be safe. We speculate that stretch is the stimulus for lung growth because there are no known growth factors present in PFC.

Videofetoscopically assisted fetal tissue engineering: skin replacement.

BACKGROUND/PURPOSE: Treatment of several congenital anomalies is frequently hindered by lack of enough tissue for surgical reconstruction in the neonatal period. The purposes of this study were (1) introduction of a novel concept in perinatal surgery, involving minimally invasive harvest of fetal tissue, which is then processed through tissue engineering techniques in vitro while pregnancy is allowed to continue, so that, at delivery, the newborn can benefit from having autologous, expanded tissue promptly available for surgical implantation at birth; (2) analysis of the progress of an engineered fetal skin graft with time, after implantation in the neonate; and (3) study of the effects of current tissue engineering techniques on fetal keratinocytes and fetal dermal fibroblasts. METHODS: Ten 90- to 95-day-gestation fetal lambs underwent surgical creation of two large paramedian excisional skin defects on the posterior body wall. Subsequently, fetal skin specimens no larger than 1.5 x 1.5 cm were videofetoscopically harvested. Fetal keratinocytes and dermal fibroblasts were then separately cultivated and expanded in vitro for 45 to 50 days, resulting in a total of approximately 250 to 300 million cells. Seven to 10 days before fetal delivery, all cells were seeded in two layers on a 16 to 20-cm2, 3-mm thick biodegradable polyglycolic acid polymer matrix. One to 4 days after delivery, the autologous engineered skin was implanted over one of two previously created skin defects. The second skin defect region received an absorbable polymer scaffold without cells as a control. If necessary, the original skin wounds were further amplified before implantation. Each animal provided at least one time-point for histological analysis of both types of repair through excisional biopsies performed at weekly intervals, up to 8 weeks postimplantation. Normal skin specimens were also used as controls. RESULTS: Fetal and neonatal survival rates were 100%. Based on previous postnatal skin engineering studies, fetal dermal fibroblasts multiplied significantly faster in vitro (approximately fivefold) than expected. Fetal keratinocytes multiplied at expected postnatal rates. The engineered grafts induced faster epithelization of the wound (partial at 1 week and complete between 2 and 3 weeks postoperatively) than did the acellular ones (partial at 3 weeks and complete between 3 and 4 weeks postoperatively). Analysis of skin architecture showed a higher level of epidermal organization and less dermal scarring in the wounds that received the engineered, cell-implanted polymer scaffold. CONCLUSIONS: (1) Videofetoscopically assisted fetal tissue engineering is a viable method for obtaining expanded autologous tissue for prompt surgical reconstruction at birth. (2) Fetal skin can be expanded and engineered in vitro at faster rates than expected postnatally, with current tissue engineering techniques. (3) Engineered autologous fetal skin induces a faster and more organized healing of neonatal skin defects than that observed with second intention. This concept may prove useful for the treatment of certain human neonatal conditions such as giant neoplasias, ectopia cordis, and other body wall defects.

Videofetoscopically assisted fetal tissue engineering: bladder augmentation.

BACKGROUND/PURPOSE: Treatment of several congenital anomalies is frequently hindered by lack of enough tissue for surgical reconstruction in the neonatal period. Minimally invasive harvest of fetal tissue, which is then processed through tissue engineering techniques in vitro while pregnancy is allowed to continue so that at delivery a newborn with a prenatally diagnosed congenital anomaly can benefit from having autologous, expanded tissue promptly available for surgical reconstruction at birth. This concept was applied to a bladder defect. METHODS: Bladder exstrophy was surgically created in ten 90- to 95-day gestation fetal lambs, which were divided in two groups. In group I, a small fetal bladder specimen was harvested through a minimally invasive technique (videofetoscopy). Urothelial and smooth muscle cells were then separately cultivated and expanded in vitro for 55 to 60 days, resulting in a total of approximately 200 million cells. Seven to 10 days before delivery, the cells were seeded in two layers in a 16- to 20-cm2, 3-mm thick biodegradable polyglycolic acid polymer matrix. One to 4 days after delivery, autologous engineered tissue was used for surgical augmentation of the exstrophic bladder. In group II, no harvest was performed, and the bladder exstrophy was primarily closed after delivery. In both groups, a catheter was left inside the bladder for 3 weeks, at which time a cystogram was performed and the catheter then removed. In all animals, at 60 days, another cystogram was performed and urodynamic studies of the bladder were performed. The bladder was then removed for histological analysis. RESULTS: Fetal survival rate was 100%. One newborn died immediately after the implantation of the engineered bladder from an anesthetic accident. The other nine (four in group I and five in group II) survived. One of the animals from group I lost its bladder catheter prematurely and had a urinary leak detected only at the time of death. There were no other complications. The engineered bladders were more compliant (P < .05) and had greater capacity pressures greater than 20 mm Hg (P < .05) than those closed primarily. Histological analysis of the engineered tissue showed a multilayered urothelial lining on the luminal side and overlying layers of smooth muscle cells surrounded by connective tissue. CONCLUSIONS: Videofetoscopically assisted fetal bladder engineering may be a viable alternative for prompt bladder reconstruction at birth. The architecture of autologous engineered fetal bladder tissue resembles that of native bladder. This concept may prove useful for the treatment of certain human neonatal conditions such as bladder and cloacal exstrophies.

Management of biliary atresia in the liver transplantation era: a 15-year, single-center experience.

BACKGROUND/PURPOSE: The aim of this study was to address the perioperative aspects of hepatoportoenterostomy (HPE) for biliary atresia (BA), through the study of a 15-year, single-center experience of the management of this disease. METHODS: One hundred twenty-seven patients were divided into three groups, depending on the variant of HPE performed: group A (n = 53) underwent HPE with external diversion of the Roux-en-Y anastomosis; group B (n = 54) underwent HPE with a long (35 to 40 cm) Roux-en-Y anastomosis, without diversion; and group C (n = 20) underwent the same kind of HPE as group B, but with a modified, "super extensive" dissection of the porta hepatitis. Eleven children in group B had an intussusception type antireflux valve in the Roux-en-Y loop. The porta hepatitis of 105 children was histologically classified in types I to III and correlated with rate of postoperative bile flow and age at surgery. Liver transplantation was performed after HPE in 20 patients. RESULTS: Overall, biliary drainage was achieved in 72.5% of the children after HPE and 26.8% of all patients became jaundice free. Porta hepatitis type III was associated with a significantly higher rate of biliary drainage post-HPE then types I and II. There was no difference in the rate of bile drainage, relative number of jaundice-free patients, and mean number of episodes of cholangitis per year among surgical groups A, B, C. In group A, 43.7% of the patients had complications related to the stoma. The actuarial survival of children who underwent HPE followed by liver transplantation was 85%. CONCLUSIONS: (1) There is no correlation between type of porta hepatis and age at surgery for BA; (2) type III porta hepatis is associated with higher rates of bile drainage post-HPE; (3) children older than 16 weeks with BA should still be considered for HPE; (4) in these older patients, factors other than the type of porta hepatis, possibly the degree of liver damage, play a role in the lower rate of bile drainage observed; (5) external diversion of the Roux-en-Y in HPE is not beneficial and is detrimental because of stoma-related complications; (6) an antireflux valve in the Roux-en-Y loop does not reduce the incidence of cholangitis post-HPE; (7) Surgical reexploration does not restore biliary drainage; and (8) sequential therapy with HPE followed by liver transplantation remains the treatment of choice for BA.

ECMO in evolution: the impact of changing patient demographics and alternative therapies on ECMO.

The incidence of neonatal extracorporeal membrane oxygenation (ECMO) is decreasing nationally. This decrease is presumed to be a result of the emergence of alternative technologies such as high-frequency oscillatory ventilation (HFOV), nitric oxide (NO), and surfactant therapy as well as others. The purposes of the present report were to determine just how rapidly the demographics of ECMO are changing and to determine the impact of competing technologies on ECMO use. The authors reviewed their entire ECMO experience of 455 cases (370 neonatal, 38 pediatric, and 47 cardiac). The neonatal cases also were separated into diagnostic groups: MAS (meconium aspiration syndrome), PPHN (persistent pulmonary hypertension of the newborn), RDS (respiratory distress syndrome), and sepsis. To allow statistical comparison, the patients were divided into four chronological groups, of equal 3-year duration, spanning the 12 years that ECMO has been available. The results of the analysis demonstrated four principle findings. (1) The total number of patients receiving ECMO per year was declining (P = .0001). This decline was attributable to a reduction in the total number of neonatal patients, with the exception of cases of congenital diaphragmatic hernia. (2) The complexity of each ECMO run was increasing, as evidenced by substantial increases in mean ECMO duration per patient and an increase in the incidence of patient complications on ECMO (P = .0001). (3) There has been a significant decrease in the overall survival rate for patients treated with ECMO (P = .0001). (4) The ECMO population mix has shifted away from straightforward neonatal cases and toward the more complex pediatric and cardiac cases. This demographic shift has occurred as a result of improvements in pre-ECMO management of neonatal patients, and is primarily responsible for the findings noted above. However, there also has been a worsening of condition severity within each diagnostic group, which also is partly responsible for the changes noted. If these trends continue, pediatric, cardiac, and CDH patients will likely account for the majority of ECMO patients. Consequently, existing ECMO centers must be prepared to adapt to the changing demographics by evolving programs that support pediatric, cardiac, and adult patients, in addition to neonates. Furthermore, the complexity associated with transporting these unstable older patients and the likelihood that the number of active ECMO centers will decline may require remaining ECMO centers to develop long-distance ECMO transport capabilities.

Resection of a massive sacrococcygeal teratoma using hypothermic hypoperfusion: a novel use of extracorporeal membrane oxygenation.

A 33-week-gestation infant with a massive sacrococcygeal teratoma weighted 4,000 g, but the actual weight of the infant was approximately 1,500 g. With the potential for massive blood loss and impaired lung compliance during resection, some type of cardiopulmonary support was necessary. Resection was undertaken with the assistance of venoarterial extracorporeal membrane oxygenation (ECMO) and hypothermic hypoperfusion. Immediately after removal of the tumor, which weighted 2,420 g, the infant was decannulated from ECMO, and the carotid artery was primarily reconstructed end-to-end. The amount of intraoperative blood loss was 550 mL Postoperatively, the child weighted 1,580 g. Follow-up head ultrasound results were normal, and the patient has done well. This is the first reported case in which ECMO with hypothermic hypoperfusion was used for resection of a massive tumor. This experience shows that ECMO is both useful and safe as a means of temporary cardiopulmonary support for resection of massive tumors in infants.

Fate of the reconstructed carotid artery after extracorporeal membrane oxygenation.

Reconstruction of the right common carotid artery has been shown to be feasible in neonates after extracorporeal membrane oxygenation (ECMO). However, the long-term outcome after carotid artery reconstruction (CAR) remains unknown. The purpose of this study was to evaluate the natural progression of the anastomotic site after CAR. Between February 1990 and June 1993, 201 patients received ECMO. All veno-arterial (VA) ECMO patients (n = 172) were considered candidates for reconstruction unless a significant neurological event (ie, intracranial hemorrhage, stroke) had occurred; the duration of ECMO exceeded 10 days, making carotid mobilization difficult; or the patient's prognosis was deemed poor. Reconstruction was performed by excising the arteriotomy site, followed by primary end-to-end anastomosis. Reconstruction was abandoned and the artery ligated if an intimal flap, arterial thrombosis, or excessive tension was encountered. After reconstruction all patients had early carotid ultrasonography and either head computed tomography (CT) or magnetic resonance imaging (MRI). Subsequent ultrasound examinations were performed at approximately 6-month intervals. Diameter index (DI) (a measure of anastomotic narrowing) was calculated using ultrasound by dividing the anastomotic diameter by the diameter of the carotid artery 5 mm proximal to the anastomosis. Forty-three of 172 VA ECMO patients (25%) had successful reconstruction. Long-term follow-up data were available on 27 patients. These 27 patients had 39 ultrasound examinations, with an average follow-up time of 7.3 months (range, 4 days to 29 months). All carotid arteries were patent. Linear regression analysis showed significant improvement in the DI with time (P = .0001, r2 = .382).(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental fetal tracheal ligation and congenital diaphragmatic hernia: a pulmonary vascular morphometric analysis.

The authors have previously shown that fetal tracheal ligation (TL) reverses the pulmonary hypoplasia in experimental diaphragmatic hernia (DH) by accelerating fetal alveolar growth. The purpose of this study was to determine if growth of the accompanying macroscopic and microscopic pulmonary vasculature is also accelerated. Eighteen fetal lambs were divided into three experimental groups: diaphragmatic hernia (DH), DH and simultaneous tracheal ligation (DH/TL), and sham-operated controls (C). Animals were delivered near term, the lungs retrieved, and pulmonary capillary growth (5 to 50 microns in diameter) evaluated by standard morphometric techniques. Capillary ultrastructure was evaluated by electron microscopy. Nine additional fetal lambs of the same gestational age were equally divided into the same three groups and their lungs analyzed by pulmonary arteriography for evaluation of large vessel growth (< 100-microns diameter). Computer digital analysis of angiogram lung slices showed that the total area of large vessels was increased in DH/TL lungs when compared with DH lungs and decreased in DH lungs when compared with C lungs (P = .003); however, the ratio of large vessel area per unit of lung area was similar in all groups. Microscopic morphometry of the capillary bed showed that the total number of capillaries was increased in DH/TL lungs over both DH and C lungs (P = .0001); however, the number of capillaries per alveolus (cap/alv) was similar in all groups. In DH/TL lungs, electron microscopy showed normal capillary wall structure and normal thickness of the capillary-alveolar interface, whereas in DH lungs, capillary structure was abnormal and the capillary-alveolar interface was thickened.(ABSTRACT TRUNCATED AT 250 WORDS)

Congenital diaphragmatic hernia and associated anomalies: their incidence, identification, and impact on prognosis.

The general concept of the association of congenital diaphragmatic hernia (CDH) with other anomalies has been well described. This study is aimed at assessing the distribution of the associated anomalies (AA) by organ system, their influence on prognosis, and the practical signs that should prompt a diagnostic search. One hundred and sixty-six high-risk patients with CDH (symptomatic within the first 6 hours of life) were treated in this institution in the past decade. Sixty-five patients (39.2%) were found to have one or more AA, and 101 had isolated CDH. Of patients with anomalies, cardiac (excluding patent foramen ovale and patent ductus arteriosus) was the most frequent type of AA (63%). Hypoplastic heart syndrome was the most common defect. Many patients had multiple AA. For purposes of analysis, the patients were divided into three groups: isolated CDH, cardiac anomalies, and all other anomalies. The groups were compared with respect to several common clinical and laboratory variables, as well as survival. The frequency and timing of antenatal diagnosis were also noted. The analysis led to the following conclusions. (1) AA are present in more than one third of high-risk patients with CDH; in this group, cardiac lesions predominate. (2) High-risk CDH infants with AA have significantly lower APGAR scores and a lower BPDPO2 (best postductal PO2 before ECMO or surgery) than those with isolated CDH. This is even more evident in the group with cardiac AA. In such patients, a careful search for an undetected AA, especially cardiac, is warranted.(ABSTRACT TRUNCATED AT 250 WORDS)

Surgically produced congenital diaphragmatic hernia in fetal rabbits.

Animal models of congenital diaphragmatic hernia (CDH) still are indispensable for the evolution of knowledge related to this disease and to fetal surgery in general. The lamb has provided the most reliable experimental design thus far. Considering the possible advantages of using rabbits (rather than lambs) namely lower costs, no need of special veterinary facilities, smaller body size, year-round availability, higher number of fetuses per pregnancy, and shorter gestational period, a successful model of CDH was developed in fetal rabbits. Sixteen pregnant New Zealand rabbits underwent hysterotomy and fetal operation. Group 1 (6 does) underwent surgery on gestational day 20 and group 2 (10 does) on gestational day 24 or 25. The normal full gestation time is 31 to 33 days. In group 1, the CDH was created by transabdominal puncture and dilatation of the fetal diaphragm. In group 2, the CDH was created through open thoracotomy, either left or right. The fetuses were delivered by cesarean section on gestational day 30. The overall fetal survival rate was 0 for group 1 and 70% for group 2. All operated fetuses in group 2 that were born alive had CDH. The histological morphometric examinations (radial alveolar count after sustained lung expansion) of the normal and operated fetuses in group 2 showed pulmonary hypoplasia in all the lungs on the same side as the CDH (statistical analysis was performed using the Neuman-Keuls test and analysis of variance; the significance level was set at .05).(ABSTRACT TRUNCATED AT 250 WORDS)

Antenatal diagnosis of isolated congenital diaphragmatic hernia is not an indicator of outcome.

It has been reported previously that infants diagnosed with congenital diaphragmatic hernia (CDH) antenatally have a much poorer prognosis than those diagnosed postnatally. The authors identified 173 high-risk (symptomatic within the first 6 hours of life) infants with CDH treated in the past decade. Seventy-seven cases were diagnosed antenatally and 96 were diagnosed postnatally. The survival rate was slightly worse for the antenatal group (34% v 48% for the postnatal group; P = .04). However, 59 of the 173 patients (34%) had other life-threatening congenital anomalies. Among the 114 patients with isolated CDH, the survival rate increased to 59% in the antenatal group and 63% in the postnatal group--a difference that was not significant. The timing of antenatal diagnosis (> or < 25 weeks) had no impact on the statistics (P = .08). The only parameter consistently different between all groups and subgroups was the Apgar score at 5 minutes, which was always better in the antenatal group (P < .02), possibly reflecting more efficient resuscitation. Of the 59 patients with other life-threatening anomalies (42 cardiac), there was one survivor among the 34 in the antenatal group and only two among the 25 in the postnatal group, despite the fact that 39 patients had undergone resuscitation, which included ECMO in 25.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental fetal tracheal ligation reverses the structural and physiological effects of pulmonary hypoplasia in congenital diaphragmatic hernia.

Infants with congenital diaphragmatic hernia (DH) and profound pulmonary hypoplasia are currently unsalvageable. The authors previously demonstrated that tracheal ligation (TL) accelerates fetal lung growth and reverses the pulmonary hypoplasia of fetal nephrectomy. The purpose of this study was to determine if the pulmonary hypoplasia of experimental DH could be similarly reversed and, if so, whether the resulting lungs would show better function than those of their DH counterparts. Eighteen fetal lambs were divided into three experimental groups of six animals each. In group 1, DH was created at 90 days' gestation. In group 2, DH was created at 90 days' gestation and TL performed during the same operation. Group 3 consisted of sham-operated controls. These animals were delivered near full-term, and their lungs analyzed by standard morphometric techniques. Ten additional fetal lambs were divided into two experimental groups of five animals each. In group 4, DH was created at 90 days' gestation. In group 5, DH was created at 90 days' gestation and TL performed 20 days later, at 110 days' gestation. These animals were pressure-ventilated via tracheostomy over a 2-hour period in which PaO2, PaCO2, and compliance were measured. Intratracheal pressure (ITP) was measured at the time of delivery in all groups. Upon retrieval, DH animals had abdominal viscera in the chest and small lungs; in contrast, DH/TL animals had the herniated viscera reduced from the chest by enlarged lungs. DH/TL lungs showed markedly increased growth, with significant increases in lung volume:body weight ratio (LV:BW; P = .0001), alveolar surface area (ALV.SA; P = .0001), and alveolar number (ALV#) (P = .0001) when compared with those of the DH or control group. This growth was associated with a normal maturation pattern based on histological appearance, normal airspace fraction, and normal alveolar numerical density. ITP in the DH/TL group was increased when compared with that of DH and control animals (P = .0001). Total lung DNA and protein were both elevated in the DH/TL animals (P = .0001). However, the DNA:protein ratio remained normal, suggesting lung growth had occurred through cell proliferation, not by hypertrophy. When ventilated over a range of settings, DH/TL lungs were more compliant (P = .0001) and achieved higher PaO2s (P < .003) and lower PaCO2s (P = .0001) than their DH counterparts. From these data, the authors conclude: (1) Experimental fetal DH produces hypoplastic lungs that are not capable of adequate gas exchange with conventional ventilation.(ABSTRACT TRUNCATED AT 400 WORDS)

[Liver transplantation in children: initial experience of the Instituto da Criança of the Hospital das Clínicas of Universidade de São Paulo]. / Transplante de fígado em criança: experiência inicial do Instituto da Criança do Hospital das Clínicas da Universidade de São Paulo.

The initial experience at the Instituto da Criança do Hospital das Clínicas--School of Medicine of São Paulo University with liver transplantation in children is presented. A staff experienced in the management of children, including pediatric surgeons, hepatologists, critical care specialists, anesthesiologists, and other has been joined to draw therapeutic protocols. Afterward, more than 100 experimental liver transplant were performed in animals of medium size (dogs and pigs). From September, 1989 to July 1991, 12 liver transplants were performed on 9 children (3 retransplants) ranging in age from 2.5 to 17 years, being 5 boys and 4 girls. The donors had been selected according to the ABO blood group and body weight. Just once a blood A+ recipient received a liver from a blood O+ donor. The regular postoperative immunosuppression consisted of triple therapy with cyclosporin, prednisone and azathioprine. The postoperative stay in the Intensive Care Unit ranged from 3 to 24 days, according to the necessity of ventilatory support. These was no intraoperative mortality, arterial or venous thromboses, or early biliary complications. The overall survival is 78% (7/9). Primary non-function of the graft was the cause of death in two of our children. Although the number of cases is still small our results are comparable to those of the best liver transplant centers in the world.