Pulmonary vasculature development in congenital diaphragmatic hernia: a novel automated quantitative imaging analysis.

PURPOSE: Impaired fetal lung vasculature determines the degree of pulmonary hypertension in the congenital diaphragmatic hernia (CDH). This study aims to demonstrate the morphometric measurements that differ in pulmonary vessels of fetuses with CDH. METHODS: Nitrofen-induced CDH Sprague-Dawley rat fetuses were scanned with microcomputed tomography. The analysis of the pulmonary vascular tree was performed with artificial intelligence. RESULTS: The number of segments in CDH was significantly lower than that in the control group on the left (U = 2.5, p = 0.004) and right (U = 0, p = 0.001) sides for order 1(O1), whereas there was a significant difference only on the right side for O2 and O3. The pooled element numbers in the control group obeyed Horton's law (R2 = 0.996 left and R2 = 0.811 right lungs), while the CDH group broke it. Connectivity matrices showed that the average number of elements of O1 springing from elements of O1 on the left side and the number of elements of O1 springing from elements of O3 on the right side were significantly lower in CDH samples. CONCLUSION: According to these findings, CDH not only reduced the amount of small order elements, but also destroyed the fractal structure of the pulmonary arterial trees.

Association of amnioinfusion volume at the time of surgery for twin-twin transfusion syndrome and latency to delivery.

OBJECTIVE: To evaluate the impact of amnioinfusion and other peri-operative factors on pregnancy outcomes in the setting of Twin-twin transfusion syndrome (TTTS) treated via fetoscopic laser photocoagulation (FLP). METHODS: Retrospective study of TTTS treated via FLP from 2010 to 2019. Pregnancies were grouped by amnioinfusion volume during FLP (<1 L vs. ≥1 L). The primary outcome was latency from surgery to delivery. An amnioinfusion statistic (AIstat) was created for each surgery based on the volume of fluid infused and removed and the preoperative deepest vertical pocket. Regression analysis was planned to assess the association of AIstat with latency. RESULTS: Patients with amnioinfusion of ≥1 L at the time of FLP had decreased latency from surgery to delivery (61 ± 29.4 vs. 73 ± 28.8 days with amnioinfusion <1 L, p < 0.001) and increased preterm prelabor rupture of membranes (PPROM) <34 weeks (44.7% vs. 33.5%, p = 0.042). Amnioinfusion ≥1 L was associated with an increased risk of delivery <32 weeks (aRR 2.6, 95% CI 1.5-4.5), 30 weeks (aRR 2.4, 95% CI 1.5-3.8), and 28 weeks (aRR 1.9, 95% CI 1.1-2.3). Cox-proportional regression revealed that AIstat was inversely associated with latency (HR 1.1, 95% CI 1.1-1.2). CONCLUSION: Amnioinfusion ≥1 L during FLP was associated with decreased latency after surgery and increased PPROM <34 weeks.

Fetal lung hypoxia and energetic cell failure in the nitrofen-induced congenital diaphragmatic hernia rat model.

PURPOSE: Congenital diaphragmatic hernia (CDH) pathogenesis is poorly understood. We hypothesize that fetal CDH lungs are chronically hypoxic because of lung hypoplasia and tissue compression, affecting the cell bioenergetics as a possible explanation for abnormal lung development. METHODS: To investigate this theory, we conducted a study using the rat nitrofen model of CDH. We evaluated the bioenergetics status using H1 Nuclear magnetic resonance and studied the expression of enzymes involved in energy production, the hypoxia-inducible factor 1α, and the glucose transporter 1. RESULTS: The nitrofen-exposed lungs have increased levels of hypoxia-inducible factor 1α and the main fetal glucose transporter, more evident in the CDH lungs. We also found imbalanced AMP:ATP and ADP:ATP ratios, and a depleted energy cellular charge. Subsequent transcription levels and protein expression of the enzymes involved in bioenergetics confirm the attempt to prevent the energy collapse with the increase in lactate dehydrogenase C, pyruvate dehydrogenase kinase 1 and 2, adenosine monophosphate deaminase, AMP-activated protein kinase, calcium/calmodulin-dependent protein kinase 2, and liver kinase B1, while decreasing ATP synthase. CONCLUSION: Our study suggests that changes in energy production could play a role in CDH pathogenesis. If confirmed in other animal models and humans, this could lead to the development of novel therapies targeting the mitochondria to improve outcomes.

A novel surgical toxicological-free model of diaphragmatic hernia in fetal rats.

BACKGROUND: Teratogen-induced congenital diaphragmatic hernia (CDH) rat models are commonly used to study the pathophysiology. We have created a new and reliable surgically induced diaphragmatic hernia (DH) model to obtain a purely mechanical DH rat model, and avoid the confounding teratogen-induced effects on the lung development. METHODS: Fetal DH was surgically created on fetuses at E18.5 and harvested at E21.5 in rats. Four groups were evaluated (n = 16): control (CONT), control exposed to Nitrofen (CONT NIT), DH surgically created (DH SURG), and CDH Nitrofen (CDH NIT). Body weight, total lung weights, and their ratio (BW, TLW, and TLBR) were compared. Air space (AS), parenchyma (PA), total protein, and DNA contents were measured to verify lung hypoplasia. Medial wall thickness (MWT) of pulmonary arterioles was also analyzed. RESULTS: DH SURG showed significant hypoplasia (decreased in total protein and DNA) vs CONT (p < 0.05); DH SURG vs CDH NIT were similar in TLW and TLBR. DH SURG has less AS than CONT (p < 0.05) and similar PA compared to CONT NIT and CDH NIT, MWT were similarly increased in CONT NIT, DH SURG, and CDH NIT. CONCLUSIONS: This novel surgical model generates fetal lung hypoplasia contributing to the study of the mechanical compression effect on fetal lung development in DH. IMPACT: There is a critical need to develop a surgical model in rat to complement the findings of the well-known Nitrofen-induced CDH model. This experimental study is pioneer and can help to understand better the CDH pathophysiological changes caused by herniated abdominal viscera compression against the lung during the final stage of gestation in CDH fetuses, and also to develop more efficient treatments in near future.

Dural substitutes for spina bifida repair: past, present, and future.

PURPOSE: The use of materials to facilitate dural closure during spina bifida (SB) repair has been a highly studied aspect of the surgical procedure. The overall objective of this review is to present key findings pertaining to the success of the materials used in clinical and pre-clinical studies. Additionally, this review aims to aid fetal surgeons as they prepare for open or fetoscopic prenatal SB repairs. METHODS: Relevant publications centered on dural substitutes used during SB repair were identified. Important information from each article was extracted including year of publication, material class and sub-class, animal model used in pre-clinical studies, whether the repair was conducted pre-or postnatally, the bioactive agent delivered, and key findings from the study. RESULTS: Out of 1,121 publications, 71 were selected for full review. We identified the investigation of 33 different patches where 20 and 63 publications studied synthetic and natural materials, respectively. From this library, 43.6% focused on clinical results, 36.6% focused on pre-clinical results, and 19.8% focused on tissue engineering approaches. Overall, the use of patches, irrespective of material, have shown to successfully protect the spinal cord and most have shown promising survival and neurological outcomes. CONCLUSION: While most have shown significant promise as a therapeutic strategy in both clinical and pre-clinical studies, none of the patches developed so far are deemed perfect for SB repair. Therefore, there is an opportunity to develop new materials and strategies that aim to overcome these challenges and further improve the outcomes of SB patients.

Heterogeneous Response in Rabbit Fetal Diaphragmatic Hernia Lungs After Tracheal Occlusion.

BACKGROUND: Fetal tracheal occlusion (TO) is an experimental therapeutic approach to stimulate lung growth in the most severe congenital diaphragmatic hernia (CDH) cases. We have previously demonstrated a heterogeneous response of normal fetal rabbit lungs after TO with the appearance of at least two distinct zones. The aim of this study was to examine the fetal lung response after TO in a left CDH fetal rabbit model. METHODS: Fetal rabbits at 25 d gestation underwent surgical creation of CDH followed by TO at 27 d and harvest on day 30. Morphometric analysis, global metabolomics, and fluorescence lifetime imaging microscopy (FLIM) were performed to evaluate structural and metabolic changes in control, CDH, and CDH + TO lungs. RESULTS: Right and left lungs were different at the baseline and had a heterogeneous pulmonary growth response in CDH and after TO. The relative percent growth of the right lungs in CDH + TO was higher than the left lungs. Morphometric analyses revealed heterogeneous tissue-to-airspace ratios, in addition to size and number of airspaces within and between the lungs in the different groups. Global metabolomics demonstrated a slower rate of metabolism in the CDH group with the left lungs being less metabolically active. TO stimulated metabolic activity in both lungs to different degrees. FLIM analysis demonstrated local heterogeneity in glycolysis, oxidative phosphorylation (OXPHOS), and FLIM "lipid-surfactant" signal within and between the right and left lungs in all groups. CONCLUSIONS: We demonstrate that TO leads to a heterogeneous morphologic and metabolic response within and between the right and left lungs in a left CDH rabbit model.

Fetal brain damage in congenital hydrocephalus.

BACKGROUND: Congenital hydrocephalus (HCP) is a developmental brain disorder characterized by the abnormal accumulation of cerebrospinal fluid within the ventricles. It is caused by genetic and acquired factors that start during early embryogenesis with disruption of the neurogerminal areas. As might be expected, early-onset hydrocephalus alters the process of brain development leading to irreparable neurological deficit. A primary alteration of the ependyma/neural stem cells (affecting vesicle trafficking and abnormal cell junctions) leads to its loss or denudation and translocation of neural progenitor cells (NPCs) and neural stem cells (NSCs) into the cerebrospinal fluid (CSF). Under these abnormal conditions, morphological and functional processes, underlying the concept of astroglial reaction, are initiated in an attempt to recover homeostasis in the periventricular zone. This astroglial reaction includes astrocyte hypertrophy, hyperplasia, and development of a new layer with reorganized functional features that resemble the ependyma. Despite decades of research, there is a lack of information concerning the biological basis of the brain abnormalities that are associated with HCP. DISCUSSION: The present review of current literature discusses the neuropathological changes during gestation following the onset of congenital hydrocephalus and the unanswered questions into the pathophysiology of the disease. A better understanding of those missing points might help create novel therapeutic strategies that can reverse or even prevent the ultimate neurological impairment that affects this population and improve long-term clinical outcome.

In Vivo Evaluation of Novel PLA/PCL Polymeric Patch in Rats for Potential Spina Bifida Coverage.

BACKGROUND: Current therapeutic materials for spina bifida repair showed a limited number of options in the market, and none of them have all the requirements as the ideal patch. In fact, sometimes the surgical procedures pose substantial challenges using different patches to fully cover the spina bifida lesion. For this purpose, a tailored patch made of poly (L-lactic acid) and poly (ε-caprolactone) blend was designed and validated in vitro to accomplish all these requirements but was never tested in vivo. MATERIAL AND METHODS: In our present study, the designed patch was analyzed in terms of rejection from the animal when implanted subcutaneously and as a dural substitute in the spinal cord. Inflammatory reaction (Iba1), astrogliosis (GFAP), was analyzed and functional interaction with spinal cord tissue assessing the (%motor-evoked potentials /compound motor action potential) by electrophysiology. RESULTS: No evidence of adverse or inflammatory reactions was observed in both models of subcutaneous implantation, neither in the neural tissue as a dural substitute. No signs of astrogliosis in the neural tissue were observed, and no functional alteration with improvement of the motor-evoked potential's amplitude was detected after 4 wk of implantation as a dural substitute in the rat spinal cord. CONCLUSIONS: Designed patch used as a dural substitute will apparently not produce inflammation, scar formation, or tethering cord and not induce any adverse effect on regular functions of the spinal cord. Further studies are needed to evaluate potential improvements of this novel polymeric patch in the spinal cord regeneration using spina bifida models.

Comparative study of intracisternal kaolin injection techniques to induce congenital hydrocephalus in fetal lamb.

PURPOSE: Kaolin (aluminum silicate) has been used to generate hydrocephalus by direct cisterna magna injection in animal models. The aim of the present study is to compare which method of Kaolin injection into fetal cisterna magna is feasible, safer, and more effective to induce hydrocephalus in fetal lambs. METHODS: Twenty-five well-dated pregnant ewes at gestational 85-90 days (E85-90) were used to compare three different kaolin injection puncture techniques into the fetal cisterna magna. Group 1, ultrasound guidance in a maternal percutaneous transabdominal (TA); group 2, without opening the uterus in a transuterine (TU) technique; group 3, by occipital direct access after exteriorizing fetal head (EFH); and group 4, control group, was normal fetal lambs without injection. The fetal lambs were assessed using lateral ventricle diameter ultrasonographic measurements prior the kaolin injection and on the subsequent days. We analyzed the effectivity, mortality, and fetal losses to determine the best technique to create hydrocephalus in fetal lamb. RESULTS: After fetal intracisternal kaolin (2%, 1mL) injection, lateral ventricle diameters increased progressively in the three different interventional groups compared with the normal values of the control group (p ≤ 0.05). We observed that the transabdominal method had a 60% of fetal losses, considering failure of injection and mortality, compared with the 12.5% in the open group (EFH), and 0% for the transuterine group. CONCLUSIONS: Based on our study, we believe that both, open uterine (EFH) and transuterine approaches are more effective and safer than the transabdominal ultrasound-guided method to induce hydrocephalus.

Proteomic profiling of tracheal fluid in an ovine model of congenital diaphragmatic hernia and fetal tracheal occlusion.

Congenital diaphragmatic hernia (CDH) occurs in ~1:2,000 pregnancies and is associated with substantial morbidity and mortality. Fetal tracheal occlusion (TO) is an emerging therapy that improves lung growth and reduces mortality, although substantial respiratory compromise persists in survivors. In this study, we used tracheal fluid in a fetal sheep model of CDH with TO for proteomic analysis with subsequent validation of findings in sheep lung tissue. We found that the proteomic profiles of CDH tracheal fluid was most similar to control lung and CDH/TO lung most similar to TO lung. Among 118 proteins altered in CDH, only 11 were reciprocally regulated in CDH/TO. The most significantly altered pathways and processes were cell proliferation, phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin signaling, inflammation, and microtubule dynamics. CDH suppressed and TO promoted cell proliferation and AKT-related signaling cascades. By Western blot analysis and immunohistochemistry, epithelial PCNA and phosphorylated AKT were decreased in CDH and increased in TO and CDH/TO lungs. The Wnt target Axin2 was decreased threefold in CDH lung compared with control without a significant increase in CDH/TO lung. Cilia-related pathways were among the most dysregulated with CDH lung having a nearly twofold increase in acetylated α-tubulin and a relative increase in the number of ciliated cells. While TO improves lung growth and patient survival in CDH, the procedure substantially alters many processes important in lung development and cell differentiation. Further elucidation of these changes will be critical to improving lung health in infants with CDH treated with TO.

Fetal tracheal occlusion in mice: a novel transuterine method.

BACKGROUND: Fetal tracheal occlusion (TO) is an emerging surgical therapy in congenital diaphragmatic hernia that improves the fetal lung growth. Different animal models of congenital diaphragmatic hernia and TO present advantages and disadvantages regarding ethical issues, cost, surgical difficulty, size, survival rates, and available genetic tools. We developed a minimally invasive murine transuterine TO model, which will be useful in defining how TO impacts lung molecular biology, cellular processes, and overall lung physiology. MATERIALS AND METHODS: Time-mated C57BL/6 mice underwent laparotomy at embryonic day 16.5 (E16.5) with transuterine TO performed on two fetuses in each uterine horn. At E18.5, dams were sacrificed and fetuses harvested. The lungs of the TO fetuses were compared with the nonmanipulated counterparts by morphometric and histologic analysis. RESULTS: Successful TO was confirmed in 16 of 20 TO fetuses. Twelve of them survived to E18.5 (75%). Fetal weights were comparable, but lung weights were significantly greater in TO (28.41 ± 5.87 versus 23.38 ± 3.09, P = 0.043). Lung to body weight ratio was also greater (0.26 ± 0.003 versus 0.22 ± 0.002, P = 0.006). E18.5 TO lungs demonstrated dilated central and distal airspaces with increased cellularity. DNA/protein and DNA/lung weight ratios were elevated while protein/lung weight ratio was lower in TO compared to control. CONCLUSIONS: Mice fetal transuterine TO is feasible with comparable outcomes to other current animal models. The increase in the lung weight, lung to body weight ratio and the DNA/protein ratio indicate organized lung growth rather than edema or cell hypertrophy.

Successful endoscopic third ventriculostomy in children depends on age and etiology of hydrocephalus: outcome analysis in 51 pediatric patients.

PURPOSE: Endoscopic third ventriculostomy (ETV) has become the method of choice in the treatment of hydrocephalus. Age and etiology could determine success rates (SR) of ETV. The purpose of this study is to assess these factors in pediatric population. METHODS: Retrospective study on 51 children with obstructive hydrocephalus that underwent ETV was performed. The patients were divided into three groups per their age at the time of the treatment: < 6, 6-24, and > 24 months of age. All ETV procedures were performed by the same neurosurgeon. RESULTS: Overall SR of ETV was 80% (40/51) for all etiologies and ages. In patients < 6 months of age SR was 56.2% (9/16), while 6-24 months of age was 88.9% (16/18) and > 24 months was 94.1% (16/17) (p = 0.012). The highest SR was obtained on aqueductal stenosis. SR of posthemorrhagic, postinfectious, and spina bifida related hydrocephalus was 60% (3/5), 50% (1/2), and 14.3% (1/7), respectively. While SR rate at the first ETV attempt was 85.3%, it was 76.9% in patients with V-P shunt performed previously (p = 0.000). CONCLUSIONS: Factors indicating a potential failure of ETV were young age and etiology such as spina bifida, other than isolated aqueductal stenosis. ETV is the method of choice even in patients with former shunting. Fast healing, distensible skulls, and lower pressure gradient in younger children, all can play a role in ETV failure. Based on our experience, ETV could be the first method of choice for hydrocephalus even in children younger than 6 months of age.

Excessive Reversal of Epidermal Growth Factor Receptor and Ephrin Signaling Following Tracheal Occlusion in Rabbit Model of Congenital Diaphragmatic Hernia.

Congenital diaphragmatic hernia (CDH) causes severe pulmonary hypoplasia from herniation of abdominal contents into the thorax. Tracheal occlusion (TO) for human CDH improves survival, but morbidity and mortality remain high, and we do not fully understand the cellular pathways and processes most severely impacted by CDH and TO. We created a left diaphragmatic hernia (DH) in rabbit fetuses with subsequent TO and collected left lung sections for NextGen mRNA sequencing. DH, TO, and DHTO fetuses had comparable body and organ growth to control except for lower lung weights in DH (p<0.05). Of 13,687 expressed genes, DHTO had 687 differentially expressed genes compared to DH, but no other group-group comparison had more than 10. Considering genes in combination, many of the genes reduced in DH were more highly expressed in DHTO than in control. Benchmarking fetal rabbit lung gene expression to published lung development data, both DH and DHTO lungs were more highly correlated with the gene expression of immature lung. DNA synthesis was upregulated in DHTO compared to DH and ribosome and protein synthesis pathways were downregulated. DH reduced total and epithelial cell proliferation by half and two-thirds respectively, and DHTO increased proliferation by 2.5 and 3.4-fold respectively. Signaling pathways downregulated by DH and upregulated in DHTO were epidermal growth factor receptor signaling, ephrin signaling, and cell migration; however, levels of ephrin and EGFR signaling in DHTO exceeded that of control. Identification and inhibition of the ligands responsible for this dysregulated signaling could improve lung development in CDH.

Gene expression profiling of brain cortex microvessels may support brain vasodilation in acute liver failure rat models.

Development of brain edema in acute liver failure can increase intracranial pressure, which is a severe complication of the disease. However, brain edema is neither entirely cytotoxic nor vasogenic and the specific action of the brain microvasculature is still unknown. We aimed to analyze gene expression of brain cortex microvessels in two rat models of acute liver failure. In order to identify global gene expression changes we performed a broad transcriptomic approach in isolated brain cortex microvessels from portacaval shunted rats after hepatic artery ligation (HAL), hepatectomy (HEP), or sham by array hybridization and confirmed changes in selected genes by RT-PCR. We found 157 and 270 up-regulated genes and 143 and 149 down-regulated genes in HAL and HEP rats respectively. Western blot and immunohistochemical assays were performed in cortex and ELISA assays to quantify prostaglandin E metabolites were performed in blood of the sagittal superior sinus. We Identified clusters of differentially expressed genes involving inflammatory response, transporters-channels, and homeostasis. Up-regulated genes at the transcriptional level were associated with vasodilation (prostaglandin-E synthetase, prostaglandin-E receptor, adrenomedullin, bradykinin receptor, adenosine transporter), oxidative stress (hemoxygenase, superoxide dismutase), energy metabolism (lactate transporter) and inflammation (haptoglobin). The only down-regulated tight junction protein was occludin but slightly. Prostaglandins levels were increased in cerebral blood with progression of liver failure. In conclusion, in acute liver failure, up-regulation of several genes at the level of microvessels might suggest an involvement of energy metabolism accompanied by cerebral vasodilation in the cerebral edema at early stages.

Immunomodulatory and antioxidant function of albumin stabilises the endothelium and improves survival in a rodent model of chronic liver failure.

BACKGROUND & AIMS: Liver failure is characterized by endothelial dysfunction, which results in hemodynamic disturbances leading to renal failure. Albumin infusion improves hemodynamics and prevents renal dysfunction in advance liver failure. These effects are only partly explained by the oncotic properties of albumin. This study was designed to test the hypothesis that albumin exerts its beneficial effects by stabilising endothelial function. METHODS: In vivo: systemic hemodynamics, renal function, markers of endothelial dysfunction (ADMA) and inflammation were studied in analbuminaemic and Sprague-Dawley rats, 6-weeks after sham/bile duct ligation surgery. In vitro: human umbilical vein endothelial cells were stimulated with LPS with or without albumin. We studied protein expression and gene expression of adhesion molecules, intracellular reactive oxygen species, and cell stress markers. RESULTS: Compared to controls, analbuminaemic rats had significantly greater hemodynamic deterioration after bile duct ligation, resulting in worse renal function and shorter survival. This was associated with significantly greater plasma renin activity, worse endothelial function, and disturbed inflammatory response. In vitro studies showed that albumin was actively taken up by endothelial cells. Incubation of albumin pre-treated endothelial cells with LPS was associated with significantly less activation compared with untreated cells, decreased intracellular reactive oxygen species, and markers of cell stress. CONCLUSIONS: These results show, for the first time, that absence of albumin is characterised by worse systemic hemodynamics, renal function and higher mortality in a rodent model of chronic liver failure and illustrates the important non-oncotic properties of albumin in protecting against endothelial dysfunction.

Profile of a Multivariate Observation under Destructive Sampling-A Monte Carlo Approach to a Case of Spina Bifida.

A biodegradable hybrid polymer patch was invented at the University of Cincinnati to cover gaps on the skin over the spinal column of a growing fetus, characterized by the medical condition spina bifida. The inserted patch faces amniotic fluid (AF) on one side and cerebrospinal fluid on the other side. The goal is to provide a profile of the roughness of a patch over time at 0, 4, 8, 12, and 16 weeks with a 95% confidence band. The patch is soaked in a test tube filled with either amniotic fluid (AF) or phosphate-buffered saline (PBS) in the lab. If roughness is measured at any time point for a patch, the patch is destroyed. Thus, it is impossible to measure roughness at all weeks of interest for any patch. It is important to assess the roughness of a patch because the rougher the patch is, the faster the skin grows under the patch. We use a model-based approach with Monte Carlo simulations to estimate the profile over time with a 95% confidence band. The roughness profiles are similar with both liquids. The profile can be used as a template for future experiments on the composition of patches.

Brain magnetic resonance in experimental acute-on-chronic liver failure.

BACKGROUND & AIM: Acute-on-chronic liver failure is the term that refers to sustained liver injury with acute decompensation, usually induced by a precipitating factor. A common link between ensuing failures of various organs is impairment of the vascular supply, which may also induce vasogenic oedema in the brain. The aim of this study was to perform magnetic resonance (MR) study of the brain in a rat model combining bile duct ligation (BDL) and lipopolysaccharide (LPS) administration to investigate brain oedema in liver failure. METHODS: Bile duct-ligated rats underwent in vivo brain MR imaging at 4, 5 and 6 weeks, and after superimposed administration of LPS. The MR techniques applied enabled assessment of brain metabolites, and intra- or extracellular water distribution. Brain water content was assessed by gravimetry. RESULTS: MR spectroscopy showed an increase in brain glutamine and a decrease in myo-inositol and choline in relation to progression of liver disease. BDL rats showed a slight, progressive increase in the amount of cortical brain water that was significant after LPS injection. These changes did not modify the apparent diffusion coefficient, supporting a mixed origin of brain oedema (vasogenic and cytotoxic). CONCLUSIONS: The mechanisms leading to the development of brain oedema in an experimental liver disease model were related to the time course of liver failure and to pro-inflammatory stimuli. MR findings support the presence of cytotoxic and vasogenic mechanisms in induced brain oedema in BDL rats exposed to LPS.

Creation of a novel synthetic amniotic fluid for use in fetal therapy with in vitro testing on human amniotic membranes.

BACKGROUND: Normal saline or lactated Ringer's solutions are usually infused at the time of fetal interventions; however, the effect of these fluids on the amniotic membranes has never been assessed. Given both the significant differences between the composition of normal saline solution, lactated Ringer's solution, and amniotic fluid and the significant risk of prematurity after fetal interventions, an investigation is warranted. OBJECTIVE: This study aimed to evaluate the effect of current amnioinfusion fluids on the human amnion compared with a novel synthetic amniotic fluid. STUDY DESIGN: Amniotic epithelial cells from term placentas were isolated and cultured per protocol. A synthetic amniotic fluid was created with similar electrolyte, pH, albumin, and glucose concentrations to human amniotic fluid, termed "Amnio-well." The cultured human amniotic epithelium was exposed to normal saline solution, lactated Ringer's solution, and Amnio-well. As a control, 1 group of cells remained in culture media. Cells were evaluated for apoptosis and necrosis. A second analysis to examine if cells could be "rescued" was performed, wherein the cells were allowed to remain in the culture media for an additional 48 hours after amnioinfusion. Subsequently, tissue testing with human amniotic membrane explants was evaluated similarly. Immunofluorescent intensity studies were undertaken to evaluate reactive oxygen species-mediated cell damage. Real-time quantitative polymerase chain reaction was used to evaluate gene expression in apoptotic pathways. RESULTS: With simulated amnioinfusion, 44%, 52%, and 89% of amniotic epithelial cells were alive after exposure to normal saline solution, lactated Ringer's solution, and Amnio-well, respectively, compared with 85% in control (P<.001). After amnioinfusion and attempted cell rescue, 21%, 44%, 94%, and 88% of cells were alive after exposure to normal saline solution, lactated Ringer's solution, Amnio-well, and control, respectively (P<.001). In simulated amnioinfusion with full-thickness tissue explants, 68%, 80%, 93%, and 96% of cells were viable in normal saline solution, lactated Ringer's solution, Amnio-well, and control, respectively (P<.001). In culture, reactive oxygen species production was higher in normal saline solution, lactated Ringer's solution, and Amnio-well than in control (4.9-, 6.6-, and 1.8-fold higher, respectively, P<.001); however, this could be mitigated in Amnio-well by adding ulin-A-statin and ascorbic acid. Gene expression data revealed abnormal signaling in the p21 and BCL2/BAX pathways with normal saline solution compared with control (P=.006 and P=.041); changes were not seen with Amnio-well. CONCLUSION: In vitro, normal saline and lactated Ringer's solutions caused increased amniotic membrane reactive oxygen species and cell death. The use of a novel fluid similar to human amniotic fluid led to the normalization of cellular signaling and less cell death.

Non-Targeted Metabolic Profiling of Cerebellum in Spina Bifida Fetal Rats.

Spina bifida, known more commonly as myelomeningocele, is a neural tube defect that results in herniation of the cerebellum through the foramen magnum into the central canal as part of the Chiari II malformation. Effects stemming from the herniated cerebellum and its metabolic profile have not been extensively studied. The objective of this study is to examine the metabolic effects of this disease on the cerebellum in utero through the utilization of a retinoid acid-induced Spina bifida rat model. Analysis of this model at mid-late (day 15) and term (day 20) of gestation in comparison to both non-exposed and retinoic acid-exposed non-myelomeningocele controls, the observed metabolic changes suggest that mechanisms of oxidative stress and energy depletion are at play in this neuro tissue. These notable mechanisms are likely to result in further damage to neural tissue as the fetus grows and the compressed cerebellum develops and herniates more due to myelomeningocele.

Fetal Endoscopic Third Ventriculostomy Is Technically Feasible in Prenatally Induced Hydrocephalus Ovine Model.

BACKGROUND: Congenital obstructive hydrocephalus generates progressive irreversible fetal brain damage by ventricular enlargement and incremental brain tissue compression that leads to maldevelopment and poor clinical outcomes. Intrauterine treatments such as ventriculo-amniotic shunting have been unsuccessfully tried in the eighties. OBJECTIVE: To assess if prenatal endoscopic third ventriculostomy (ETV) is feasible in a large animal model and optimize this technique for ventricular decompression and potential arrest of fetal brain damage in fetal lambs. METHODS: We generated hydrocephalus in 50 fetal lambs by injecting a polymeric agent into the cisterna magna at midgestation (E85). Subsequently, 3 weeks later (E105), fetal ETV was performed using a small rigid fetoscope. The endoscopy entry point was located anterior to the coronal suture, 7 mm from the midline. RESULTS: We obtained clear visualization of the enlarged lateral ventricles by endoscopy in the hydrocephalic fetal lambs. The floor of the third ventricle was bluntly perforated and passed with the scope for a successful ETV. Total success was achieved in 32/50 cases (64%). Causes of failure were blurred vision or third ventricle obliteration by BioGlue in 10/50 (20%) cases, anatomic misdirection of the endoscope in 5 (10%) cases, 2 cases of very narrow foramen of Monro, and 1 case of choroid plexus bleeding. If we exclude the cases artificially blocked by the polymer, we had a successful performance of prenatal-ETV in 80% (32/40) of hydrocephalic fetuses. CONCLUSION: Despite the inherent difficulties arising from ovine brain anatomy, this study shows that innovative fetal ETV is technically feasible in hydrocephalic fetal lambs.