Neonatal outcomes of congenital diaphragmatic hernia in full term versus early term deliveries: A systematic review and meta-analysis.

This systematic review and meta-analysis aimed to review the optimal timing of delivery at term for neonates with prenatally diagnosed congenital diaphragmatic hernia (CDH). We reviewed the literature up to December 19, 2022 using MEDLINE and the Cochrane Library databases. The inclusion criteria were original articles, comparative studies of CDH neonates delivered at an early term (37-38 weeks of gestation) and at full term (39 weeks of gestation or later), and comparative studies investigating outcomes of CDH neonates. Six studies met the inclusion criteria, including 985 neonates delivered at an early term and 629 delivered at full term. The cumulative rate of survival to discharge showed no significant difference between CDH neonates delivered at an early term (395/515; 76.7%) or at full term (345/467; 73.9%) (risk ratio [RR] 1.01; 95% confidence interval [CI], 0.89-1.16; p = 0.85). Furthermore, the number of neonates requiring oxygen therapy at discharge was not significantly different between CDH neonates delivered at an early term (32/370; 8.6%) and at full term (14/154; 9.1%) (RR, 0.99; 95% CI, 0.36-2.70; p = 0.99). Therefore, the optimal timing of delivery at term for neonates with CDH remains unclear.

The comorbidities of recurrent inguinal hernia in children: A systematic review.

BACKGROUND: Higher risk of recurrence has been reported in pediatric inguinal hernia patients with specific comorbidities. The purpose of this systematic review was to investigate which comorbidities predispose to recurrent pediatric inguinal hernias (RPIHs). METHODS: A comprehensive search of six databases was performed, reviewing the literature to date on RPIHs and the co-occurrence of comorbidities. English-language publications were considered for inclusion. The primary surgical technique (e.g., Potts procedure or laparoscopic repair) was not considered. RESULTS: Fourteen articles published between 1967 and 2021 fulfilled the inclusion criteria and did not meet the exclusion criteria. They reported a total of 86 patients diagnosed with RPIHs with 99 comorbidities. Thirty-six percent of patients had conditions with increased intra-abdominal pressure, such as ventriculoperitoneal shunt for hydrocephalus, posterior urethral valves, bladder exstrophy, seizure disorder, asthma, using continuous positive airway pressure for respiratory distress syndrome, and gastroesophageal reflux disease. Twenty-eight percent of patients had diseases with weakness of the anterior abdominal wall, specifically mucopolysaccharidosis, giant omphalocele, Ehlers-Danlos syndrome, connective-tissue disorders, and segmental spinal dysgenesis. CONCLUSIONS: The main comorbidities of RPIHs were conditions with increased intra-abdominal pressure and weakness of the anterior abdominal wall. Although these comorbidities are rare, the risk of recurrence must be noted.

Safety evaluation of a stepwise tracheostomy decannulation program in pediatric patients.

PURPOSE: In the event of failed tracheostomy decannulation, patients might have a tragic course of events. We retrospectively evaluated our stepwise tracheostomy decannulation program and examined its safety. METHODS: A 12-year retrospective study of pediatric patients was conducted. The decannulation program was performed on patients who had airway patency by laryngobronchoscopy and whose cannula could be capped during the day. A stepwise decannulation program was performed: continuous 48-h capping trial during hospitalization (Phase 1), removal of the tracheostomy tube for 48 h during hospitalization (Phase 2), and outpatient observation (Phase 3). If a persistent tracheocutaneous fistula existed, the fistula was closed by surgery (Phase 4). RESULTS: The 77 patients in the study underwent 86 trials. The age at the first time of the decannulation program was 6.5 ± 3.6 years. Sixteen trials failed (18.6%): 8 trials in Phase 1, 2 trials in Phase 2, 4 trials in Phase 3, and 2 trials in Phase 4. Most decannulation failures were due to desaturation in Phase 1/2 and dyspnea in Phase 3/4. The time to reintubation after decannulation was 15-383 days in Phase 3/4. CONCLUSIONS: Patients could fail at every phase of the program, suggesting that a stepwise decannulation program contributes to safety.

Angiographic diagnosis for accurate assessment of congenital porto-systemic shunt and extrahepatic portal vein obstruction in children.

BACKGROUND: Congenital porto-systemic shunt (CPSS) is a rare disease and can cause fatal complications. Accurate angiographic assessment is mandatory for proper treatment. Although technically difficult, we developed assessment techniques and assessed their accuracy. One technique came from evaluating patients with extrahepatic portal vein obstruction (EHPVO). METHODS: We conducted a single center retrospective study to evaluate the efficacy of angiographic diagnostic procedure for the assessment of CPSS and EHPVO, and its impact on patients' subsequent interventions and clinical course. Eight patients with CPSS and two patients with EHPVO who underwent diagnostic angiography were included. Assessment of the intrahepatic portal vein was performed in all patients. The route of the shunt, and portal vein pressure under shunt occlusion, were also evaluated for patients with CPSS. Evaluation was first attempted with a balloon angiographic catheter (standard method). Three additional techniques were performed as needed: (i) direct wedge-catheter injection without balloon inflation, (ii) use of occlusion balloon in two patients, and (iii) hybrid angiography with sheath placement directly into the superior mesenteric vein. RESULTS: The standard method was sufficient in four patients. On the other hand, all three techniques were required in two patients each. One lost contact during follow up, but all other patients underwent optimal intervention. There were no complications related to the angiographic procedure. CONCLUSIONS: Use of direct wedge-catheter injection without balloon inflation, occlusion balloon, and hybrid catheterization improved the diagnostic yield in patients with CPSS or EHPVO.

Pancreaticobiliary maljunction without biliary dilatation in pediatric patients.

PURPOSE: Pancreaticobiliary maljunction (PBM) without biliary dilatation is a condition in which dilatation of the bile duct is not seen in patients with PBM. Recently, the Japanese Study Group on Pancreaticobiliary Maljunction (JSGPM) published new diagnostic criteria for PBM. In these criteria, biliary dilatation is defined according to the standard diameter at each age. We reviewed cases of pediatric patients with PBM without biliary dilatation. METHODS: From 1992 to 2019, 134 patients with PBM were treated in our institution. Among these, 7 patients were retrospectively diagnosed with PBM without biliary dilatation. The clinical information was retrospectively assessed in these patients. RESULTS: Of the seven patients, six were female. All patients had symptoms similar to those of patients with congenital biliary dilatation. In all seven patients, the diagnosis of PBM was made before definitive surgery. Six patients had type B PBM, and one had type D PBM. All patients underwent extrahepatic bile duct resection and hepaticojejunostomy, and their symptoms resolved. One patient experienced postoperative complications of anastomotic leakage followed by anastomotic stricture. CONCLUSION: The present report revealed important clinical features of this entity. However, there are still some issues that need to be discussed, and further research is needed.

Remote ischemic conditioning causes CD4 T cells shift towards reduced cell-mediated inflammation.

PURPOSE: Necrotizing enterocolitis (NEC) is a gastrointestinal disease in neonates that is associated with immune-mediated intestinal inflammation. Remote ischemic conditioning (RIC) applied to a limb has been shown to be protective against experimental NEC. In this study, we explore the immune cell-mediated response involved in NEC and the immunomodulatory effects of RIC in an experimental mouse model of the disease. METHODS: NEC was induced in C57BL/6 mice (ethical approval #58119) pups on postnatal day5 (p5) using gavage hyperosmolar formula, lipopolysaccharide, and hypoxia. RIC consisted of 4 cycles of 5 min ischemia followed by 5 min reperfusion of the right hindlimb during NEC induction on p6 and p8. Breastfed mice were used as control. The mice were sacrificed on p9, with ileal tissue evaluated for inflammatory cytokines and by characterization of T-cell populations. RESULTS: NEC mice had increased number of CD4+ cells indicating an accumulation of T-cells in the mesenchyme of the NEC ileum. Compared to control, NEC pups had upregulated expression pro-inflammatory cytokines (GATA3, IFNγ, IL1ß, IL6, IL17, IL22, and TNFα) and reduced anti-inflammatory cytokine (TGFß). In NEC, there was also a shift in the balance of Treg/Th17 cells towards Th17. Compared to NEC alone, RIC during the course of NEC resulted in reduction of pro-inflammatory cytokines (GATA3, IFNγ, IL1ß, IL6, IL17, IL22, and TNFα), increase in anti-inflammatory cytokine TGFß and concomitant shift back of Th17 cells towards Treg cells. CONCLUSION: In experimental NEC, remote ischemic conditioning reduces the production of pro-inflammatory markers and increases the production of anti-inflammatory markers. In addition, during NEC, RIC reverses the imbalance of Treg/Th17 providing support for its effect on cell-mediated inflammation. RIC is a non-invasive physical maneuver that can have a significant beneficial effect in reducing the inflammation seen in NEC.

A predictive scoring system for small diaphragmatic defects in infants with congenital diaphragmatic hernia.

PURPOSE: To develop a predictive score for small diaphragmatic defects in infants with congenital diaphragmatic hernia (CDH) for determining thoracoscopic surgery indication. METHODS: The Japanese CDH Study Group cohort was randomly divided into derivation (n = 397) and validation (n = 396) datasets. Using logistic regression, a prediction model and weighted scoring system for small diaphragmatic defects were created from derivation dataset and validated with validation dataset. RESULTS: Six weighted variables were selected: no hydramnios, 1 point; 1 min Apgar score of 5-10, 1 point; apex type of the lung (left lung is detected radiographically in apex area), 1 point; oxygenation index < 8, 1 point; abdominal nasogastric tube (tip of the nasogastric tube is detected radiographically in the abdominal area), 2 points; no right-to-left flow of ductus arteriosus, 1 point. In validation dataset, rates of small diaphragmatic defects for Possible (0-3 points), Probable (4-5 points), and Definite (6-7 points) groups were 36%, 81%, and 94%, respectively (p < 0.001). Additionally, sensitivity, specificity, positive predictive value, and C statistics were 0.78, 0.79, 0.88, 0.76, and 0.45, 0.94, 0.94, 0.70 for Probable and Definite groups, respectively. CONCLUSION: Our scoring system effectively predicted small diaphragmatic defects in infants with CDH.

Prenatal predictors of mortality in fetuses with congenital diaphragmatic hernia: a systematic review and meta-analysis.

PURPOSE: This study aimed to evaluate prenatal predictors of mortality in fetuses with congenital diaphragmatic hernia (CDH). METHODS: A systematic literature search was performed to identify relevant observational studies that evaluated the ability of lung-to-head ratio (LHR), observed-to-expected LHR (o/e-LHR), observed-to-expected total fetal lung volume (o/e-TFLV), lung-to-thorax transverse area ratio (L/T ratio), intrathoracic herniation of the liver and the stomach, and side of diaphragmatic hernia, using a threshold for the prediction of mortality in fetuses with CDH. Study quality was assessed using the QUADAS-2 tool. Hierarchical summary receiver operating characteristic curves were constructed. RESULTS: A total of 50 articles were included in this meta-analysis. The QUADAS-2 tool identified a high risk of bias in more than one domain scored in all parameters. Among those parameters, the diagnostic odds ratio of mortality with o/e-LHR < 25%, o/e-TFLV < 25%, and L/T ratio < 0.08 were 11.98 [95% confidence interval (CI) 4.65-30.89], 11.14 (95% CI 5.19-23.89), and 10.28 (95% CI 3.38-31.31), respectively. The predictive values for mortality were similar between the presence of liver herniation and retrocardiac fetal stomach position. CONCLUSIONS: This systematic review suggests that o/e-LHR, o/e-TFLV, and L/T ratio are equally good predictors of neonatal mortality in fetuses with isolated CDH.

The efficacy of the postnatal nasogastric tube position as a prognostic marker of left-sided isolated congenital diaphragmatic hernia.

PURPOSE: The prenatal diagnosis of the stomach position in congenital diaphragmatic hernia (CDH) has been a reliable prognostic factor, but few studies have focused on the postnatal position. We therefore evaluated the significance of the nasogastric (NG) tube position just after birth. METHODS: The Japanese CDH Study Group database enrolled 1037 CDH neonates over 15 years. In our multicenter retrospective study, 464 cases of left-sided isolated CDH with prenatal diagnoses were divided into two groups: NG tube below the diaphragm (BD; n = 190) or above the diaphragm (AD; n = 274). The primary outcome was the 90-day survival rate, and the secondary outcomes were mechanical ventilation duration, hospitalization duration, and recurrence rate. RESULTS: The BD group had a significantly higher 90-day survival rate (98.4 vs. 89.4%, p < 0.001), shorter mechanical ventilation (11 vs. 19 days, p < 0.001), shorter hospitalization (38 vs. 59 days, p < 0.001), and lower recurrence rate (p = 0.002) than the AD group. A multivariate analysis showed that BD (adjusted odds ratio, 3.68; 95% confidence interval 1.02-13.30) was a favorable prognostic factor for the 90-day survival. CONCLUSION: The assessment of the NG tube position revealed it to be a reliable prognostic factor of left-sided isolated CDH. Therefore, it should be included as a routine assessment.

Clinical guidelines for the treatment of congenital diaphragmatic hernia.

Congenital diaphragmatic hernia (CDH) is a birth defect of the diaphragm in which abdominal organs herniate through the defect into the thoracic cavity. The main pathophysiology is respiratory distress and persistent pulmonary hypertension because of pulmonary hypoplasia caused by compression of the elevated organs. Recent progress in prenatal diagnosis and postnatal care has led to an increase in the survival rate of patients with CDH. However, some survivors experience mid- and long-term disabilities and complications requiring treatment and follow-up. In recent years, the establishment of clinical practice guidelines has been promoted in various medical fields to offer optimal medical care, with the goal of improvement of the disease' outcomes, thereby reducing medical costs, etc. Thus, to provide adequate medical care through standardization of treatment and elimination of disparities in clinical management, and to improve the survival rate and mid- and long-term prognosis of patients with CDH, we present here the clinical practice guidelines for postnatal management of CDH. These are based on the principles of evidence-based medicine using the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach. The recommendations are based on evidence and were determined after considering the balance among benefits and harm, patient and society preferences, and medical resources available for postnatal CDH treatment.

Thoracoscopic repair of congenital diaphragmatic hernia in neonates: findings of a multicenter study in Japan.

PURPOSE: We compared the efficacy of thoracoscopic repair (TR) with that of open repair (OR) for neonatal congenital diaphragmatic hernia (CDH). METHODS: The subjects of this multicenter retrospective cohort study were 524 infants with left-sided isolated CDH, diagnosed prenatally, and treated at one of 15 participating hospitals in Japan between 2006 and 2018. The outcomes of infants who underwent TR and those who underwent OR were compared, applying propensity score matching. RESULTS: During the study period, 57 infants underwent TR and 467 underwent OR. Ten of the infants who underwent TR required conversion to OR for technical difficulties and these patients were excluded from the analysis. The survival rate at 180 days was similar in both groups (TR 98%; OR 93%). Recurrence developed after TR in 3 patients and after OR in 15 patients (TR 7%, OR 3%, p = 0.40). The propensity score was calculated using the following factors related to relevance of the surgical procedure: prematurity (p = 0.1), liver up (p < 0.01), stomach position (p < 0.01), and RL shunt (p = 0.045). After propensity score matching, the multivariate analysis adjusted for severity classification and age at surgical treatment revealed a significantly shorter hospital stay (odds ratio 0.50) and a lower incidence of chronic lung disease (odds ratio 0.39) in the TR group than in the OR group. CONCLUSIONS: TR can be performed safely for selected CDH neonates with potentially better outcomes than OR.

Optimal timing of surgery in infants with prenatally diagnosed isolated left-sided congenital diaphragmatic hernia: a multicenter, cohort study in Japan.

PURPOSE: To date, there is no compelling evidence of the optimal timing of surgery for congenital diaphragmatic hernia (CDH). This study aimed to establish the optimal timing of surgery in neonates with isolated left-sided CDH. METHODS: This multicenter cohort study enrolled 276 patients with isolated left-sided CDH at 15 institutions. Based on the timing of surgery, the patients were classified into four groups [< 24 h (G1), 24-47 h (G2), 48-71 h (G3), and ≥ 72 h (G4)]. The clinical outcomes were compared among the groups using a logistic regression model, after adjusting for potential confounders, such as disease severity. RESULTS: Multivariate analyses showed that G2 also had a lower mortality rate than the other groups. In mild and severe cases, there were no significant differences in mortality across the four groups. In moderate cases, G2 resulted in significantly increased survival rates, compared with G1. CONCLUSION: The study results suggest that surgery within 24 h of birth is not recommended for patients with moderate severity CDH, that there is no benefit in the delay of surgery for more than 72 h in mild severity CDH patients, and that there is no definite optimal time to perform surgery in severe cases of CDH.

Meso-Rex bypass versus portosystemic shunt for the management of extrahepatic portal vein obstruction in children: systematic review and meta-analysis.

PURPOSE: Extrahepatic portal vein obstruction (EHPVO) is a major cause of non-cirrhotic portal hypertension in children. Surgical procedures for EHPVO include portosystemic shunts (PSS) and meso-Rex bypass (MRB). We conducted a systematic review and meta-analysis to compare the effectiveness of MRB versus PSS in EHPVO patients. METHODS: A systematic literature search was performed using four databases. Articles reporting EHPVO and comparing patients who received MRB and PSS were included in the analysis. RESULTS: We retrieved 851 papers, of which five observational studies met the inclusion criteria. There was no difference in shunt complications, mortality, or gastrointestinal bleeding after surgery between MRB and PSS in the meta-analysis. MRB had increased shunt complications compared with PSS in the non-comparative studies. MRB had a potential advantage over PSS in long-term prognosis in one comparative study. Overall, the quality of the evidence was low. CONCLUSIONS: Based on available data, our meta-analysis indicates that MRB does not increase shunt complications, mortality, or gastrointestinal bleeding after surgery. The present study did not reveal superiority for either MRB or PSS. The paucity of well conducted trials in this area justifies future multicenter studies and studies that examine long-term outcomes.

Nasogastric decompression after intestinal surgery in children: a systematic review and meta-analysis.

PURPOSE: Postoperative nasogastric decompression has been routinely used after intestinal surgery. However, the role of nasogastric decompression in preventing postoperative complications and promoting the recovery of bowel function in children remains controversial. This systematic review aimed to assess whether routine nasogastric decompression is necessary after intestinal surgery in children. METHODS: A systematic review was conducted following the PRISMA guideline. Literature search was performed in electronic databases including PubMed, Embase, CENTRAL, and Web of science. Studies comparing outcomes between children who underwent intestinal surgery with postoperative nasogastric tube (NGT) placement (NGT group) and without postoperative NGT placement (no NGT group) were included. RESULTS: Six studies were eligible for inclusion criteria including two randomized controlled trials (RCT) and four comparative observational studies. The overall rate of postoperative anastomotic leak was 0.6% (1/179) in NGT group and 0.9% (2/223) in no NGT group. The overall rate of wound dehiscence was 2.4% (4/169) in NGT group and 1.6% (4/245) in no NGT group. Meta-analysis of two RCTs in children undergoing elective intestinal surgery showed significant increase of mild vomiting in no NGT group compared with NGT group (OR 3.54 95% CI 1.04, 11.99) but no significant difference in persistent vomiting requiring NGT reinsertion (OR 3.11 95% CI 0.47, 20.54), abdominal distension (OR 2.36 95% CI 0.34, 16.59), NGT reinsertion (OR 3.11 95% CI 0.47, 20.54), wound infection (OR 1.63 95% CI 0.49, 5.48) and time to return of bowel movement (MD - 0.14 95% CI - 0.45, 0.17). There was no incidence of anastomotic leak in these 2 RCTs. However, there was an incidence of NGT-related discomfort in NGT group, which ranged from 30 to 100% of children studied. CONCLUSION: Routine postoperative nasogastric decompression can be omitted in children undergoing intestinal surgery due to no benefit in preventing postoperative complications while increasing patient discomfort.

Risk factors of dumping syndrome after fundoplication for gastroesophageal reflux in children.

PURPOSE: In postoperative cases of fundoplication, the gastric emptying ability is promoted and sometimes exhibits dumping syndrome. Dumping syndrome often goes unrecognized in children. Furthermore, the risk factors for postoperative dumping syndrome are unknown. This study aimed to investigate the risk factors of developing dumping syndrome after fundoplication. METHODS: A retrospective chart review of all consecutive patients between January 2003 and March 2018 (190 patients) who had fundoplication at our clinic was conducted. Regarding the risk factors of dumping syndrome, gender, age and body weight at the time of surgery, neurological impairment, severe scoliosis, microgastria, chromosomal abnormalities, complex cardiac anomalies, gastrostomy, and laparoscopic surgery were retrospectively studied. RESULTS: 17 patients (9%) developed dumping syndrome post-operatively. Multivariate analysis showed that significant risk factors for dumping syndrome included: undergoing surgery within 12 months of age (adjusted OR 10.3, 95% CI 2.6-45.2), severe scoliosis (adjusted OR 19.3, 95% CI 4.4-91.1), and microgastria (adjusted OR 26.5, 95% CI 1.4-896.4). CONCLUSIONS: We identified that: age at fundoplication being within 12 months of age, severe scoliosis, and microgastria were risk factors for dumping syndrome after fundoplication, and that this information should be explaining to the family before conducting the fundoplication.

Best pre-ductal PaO2 prior to extracorporeal membrane oxygenation as predictor of mortality in patients with congenital diaphragmatic hernia: a retrospective analysis of a Japanese database.

PURPOSE: Predicting lethal pulmonary hypoplasia in infants with congenital diaphragmatic hernia (CDH) before extracorporeal membrane oxygenation (ECMO) initiation is difficult. This study aimed to predict lethal pulmonary hypoplasia in patients with CDH prior to ECMO. METHODS: This was a multicenter cohort study involving neonates prenatally diagnosed with isolated unilateral CDH (born 2006-2020). Patients who required ECMO due to respiratory insufficiency were included in this study. Patients who underwent ECMO due to transient disorders were excluded from analysis. Blood gas analysis data within 24 h of birth were compared between survivors and non-survivors. Predictive abilities were assessed for factors with significant differences. RESULTS: Overall, 34 patients were included (18 survivors and 16 non-survivors). The best pre-ductal PaO2 was significantly lower in non-survivors than in survivors (50.4 [IQR 30.3-64.5] vs. 67.5 [IQR 52.4-103.2] mmHg, respectively; p = 0.047). A cutoff PaO2 of 42.9 mmHg had a sensitivity, specificity, and positive predictive value of 50.0%, 94.4%, and 88.9%, respectively, to predict mortality. CONCLUSION: The best PaO2 within 24 h after birth predicted mortality following ECMO initiation. This should be shared to families and caregivers to optimize the best interests of the infants with CDH.

Long-term follow-up for anorectal function after anorectoplasty in patients with high/intermediate imperforate anus: a single center experience.

PURPOSE: The aim of this study was to evaluate the clinical outcomes and postoperative anorectal function in the patients with high/intermediate imperforate anus (HIA/IIA) treated in our institution. In addition, we report our novel procedure, laparoscopically assisted anorectoplasty with anterior perineal incision (LAARP-API), which allows direct approach to the anterior edge of the puborectal sling and muscle complex, and is useful for the creation of a pulling-through route for the rectal pouch. METHODS: From 1976 to 2016, 22 patients with HIA and 43 patients with IIA underwent sacroperineal/sacroabdominoperineal pull-through anorectoplasty (SP/SAP), Potts procedure (Potts), SP with API (SP-API) or LAARP-API. Clinical data and anorectal function of those patients were retrospectively evaluated using the Japanese clinical score. RESULTS: Of the 22 cases of HIA, 15 were treated by SAP, 2 were SP and 5 were LAARP-API. Of the 43 cases of IIA, one was treated by SAP, 31 were SP, two were Potts and nine were SP-API. The mean score of anorectal function of HIA/IIA both increased with age. In IIA, the score after SP-API was significantly higher than the score after SP. CONCLUSION: Long-term outcomes of our anorectoplasty for HIA/IIA are good with excellent anorectal function score.

Lysosomal overloading and necrotizing enterocolitis.

PURPOSE: Intestinal absorption in premature infants occurs via direct epithelial cellular endocytosis and degradation by intracellular lysosomes. Autophagy is a mechanism by which cytoplasmic organelles contribute to lysosomal degradation. However, excessive autophagy can lead to cell death. The purpose of this study was to investigate whether autophagy and endocytosis are present in the small intestinal mucosa during experimental necrotizing enterocolitis (NEC). METHODS: NEC was induced by gavage feeding of hyperosmolar formula, lipopolysaccharide and hypoxia between P5 and P9 (ethical approval 44032). Breastfed mice were used as control. Distal ileum was harvested on P5, P7 and P9 and analyzed for intestinal epithelial cellular morphology as well as autophagy/lysosomal activity, and cell death. Groups were compared using Student's t test. RESULTS: During NEC, giant lysosomes were present in the intestinal villi, with some exceeding their degradation ability leading to their rupture. The NEC group had significantly increased inflammation and autophagy activity, decreased lysosome activity, and increased apoptosis compared to control. CONCLUSIONS: NEC induction causes excessive autophagy and endocytosis leading to lysosomal overloading, lysosomal membrane permeabilization and rupture which results in cell death. These novel findings may help to clarify the pathogenesis of NEC. Reduction of lysosome overload and assisting in their degradation capability may reduce the burden of NEC.

A novel model of injured liver ductal organoids to investigate cholangiocyte apoptosis with relevance to biliary atresia.

PURPOSE: The fibrogenic process in cholangiopathic diseases such as biliary atresia (BA) involves bile duct injury and apoptosis of cholangiocytes, which leads to the progression of liver fibrosis into liver cirrhosis and can result in end-staged liver disease. Recent advances in the development of organoids or mini-organ structures have allowed us to create an ex vivo injury model of the bile duct that mimics bile duct injury in BA. The aim of this experimental study was to develop a novel model of injured intrahepatic cholangiocytes as this can be relevant to BA. Our new model is important for studying the pathophysiological response of bile ducts to injury and the role of cholangiocytes in initiating the fibrogenic cascade. In addition, it has the potential to be used as a tool for developing new treatment strategies for BA. METHODS: Liver ductal organoids were generated from the liver of healthy neonatal mouse pups. Intrahepatic bile duct fragments were isolated and cultured in Matrigel dome. Injury was induced in the organoids by administration of acetaminophen in culture medium. The organoids were then evaluated for fibrogenic cytokines expression, cell apoptosis marker and cell proliferation marker. RESULTS: Organoids generated from intrahepatic bile duct fragments organized themselves into single-layer epithelial spheroids with lumen on the inside mimicking in vivo bile ducts. After 24-h exposure to acetaminophen, cholangiocytes in the organoids responded to the injury by increasing expression of fibrogenic cytokines, transforming growth factor beta-1 (TGF-ß1) and platelet-derived growth factor-BB (PDGF-BB). This fibrogenic response of injured organoids was associated with increased cholangiocyte apoptosis and decreased cholangiocyte proliferation. CONCLUSION: To our knowledge this is the first description of cholangiocyte injury in the organoids derived from intrahepatic bile ducts. Our injury model demonstrated that cholangiocyte apoptosis and its fibrogenic response may play a role in initiation of the fibrogenic process in cholangiopathic diseases such as BA. These findings are important for the development of novel therapy to reduce cholangiocyte apoptosis and to halt the early fibrogenic cascade in liver fibrogenesis. This novel injury model can prove very valuable for future research in biliary atresia.

The role of autophagy in intestinal epithelial injury.

PURPOSE: Autophagy is a natural mechanism aimed to degrade and recycle cellular components within cells. Previous studies reported that autophagy in the intestinal epithelium can be activated and that excessive autophagy can have negative consequences. However, the mechanism by which autophagy is regulated during intestinal epithelial injury remains unclear. This study aimed to investigate the mechanism of autophagy regulation during intestinal epithelial cells (IEC) injury. METHODS: Rat IEC18 were exposed to hypoxia and Lipopolysaccharide (LPS) (200 µg/ml) to induce injury. IEC18 were treated with autophagy initiation inhibitor, Wortmannin or with autophagy degradation inhibitor, Bafilomycin A1 were added for 24 h. We assessed the number and diameter of autophagic vacuoles, Cell viability, inflammation and apoptosis. RESULTS: Hypoxia and LPS administration increased the number and diameter of autophagic vacuoles in IEC18. Wortmannin administration reduced the number and diameter of autophagic vacuoles. On the contrary, Bafilomycin A1 administration increased the number of autophagic vacuoles. Cell viability increased following administration of Wortmannin and decreased following administration of Bafilomycin A1. CONCLUSIONS: We found that accumulation of autophagic vacuoles which characterize excessive or incomplete autophagy was detrimental to cell survival. This was shown by an increase in the number and size of the autophagic vacuoles with Bafilomycin A1treatment after hypoxia and LPS stressors relative to hypoxia and LPS alone. Conversely, there was a decrease in the number of autophagic vacuoles with Wortmannin treatment after hypoxia and LPS stressors relative to hypoxia and LPS alone. Therefore, reducing autophagosomes accumulation may represent a novel therapeutic strategy for intestinal injury.