ヒルシュスプルング病（全結腸型又は小腸型）

Mutation in paired-like homeobox 2B (PHOX2B) is used as the diagnostic marker of Congenital Central Hypoventilation Syndrome (CCHS). The mutant gene/protein affects neural crest cells embryonic development which leads to congenital central hypoventilation syndrome (CCHS). When individuals also have Hirschsprung's disease (HSCR) with CCHS it is known as Haddad syndrome (HS). Previous studies on CCHS/HS have mainly focused on the conformational dynamics of the mutant protein and have remained controversial. Here we performed RNA-sequencing on the patient derived neuroepithelial stem cells (NESCs), pertinent to the neurodevelopmental phenotype in CCHS/HS, and found that the PHOX2B-PARM has a profound impact on the transcriptional profile of the cells. The single copy of PHOX2B-PARM in heterozygote cells led to >8 fold differentially expressed genes. This include genes e.g., STMN2, L1CAM, ONECUT2 and NFASC, that are reported to have role in neurodevelopment. In the patient cells there was a significant enrichment of genes related to neuronal development and synapse organization mainly driven by L1CAM interactions and synaptogenesis signaling pathway. Our results not only highlight the use of a suitable model of CCHS/HS but also provide a clear path for future experimental validation.

Extracellular vesicles (EVs) are lipid bilayer-delimited nanoparticles released by cells to act as mediators of intercellular communication during organ development, injury, and repair. EVs carry cargo (bioactive proteins, lipids, and nucleic acids) that reflects the status of the parent cell and is transferred to recipient cells to regulate biological processes, such as inflammation, immune responses, and tissue regeneration. These properties have made EVs promising tools for investigating disease pathogenesis, improving diagnostic and prognostic accuracy, and developing cell-free regenerative therapies for conditions characterized by dysregulation of multiple biological pathways. EVs are particularly relevant in diseases that affect the pediatric population where pathogenesis often remains poorly understood, access to affected tissues is limited, and treatment options are frequently inadequate. This review summarizes current evidence on EV applications in fetal and neonatal disorders, including necrotizing enterocolitis, congenital diaphragmatic hernia, and bronchopulmonary dysplasia, and highlights emerging data in biliary atresia, spina bifida, short bowel syndrome, and Hirschsprung's disease. In this age group, human milk and amniotic fluid represent particularly attractive biologically accessible sources of EVs, combining therapeutic potential with feasibility of clinical application. Building on robust preclinical evidence, the field is now advancing toward clinical translation, but several aspects still need to be addressed such as cargo heterogeneity, scalability of production, dosing, biodistribution, safety, and regulatory standardization. Herein, we discuss the translational challenges and future directions that will shape the clinical application of EVs in perinatal conditions.

PURPOSE: A nationwide simulated Hirschsprung's Soave pull-through surgery (HPTS) workshop using a lamb model was set up to train pediatric surgeons. Operative confidence, skills acquisition and clinical application were evaluated. METHODS: The model (< 5 USD) comprised fresh lamb anorectum with terminal perianal muscle and skin preserved and a 20-cm rectum, inserted and fixed onto a mini basket, which allowed dissection using monopolar diathermy. Each trainee performed the five-step Soave procedure twice under supervision. Operative confidence for each step was measured pre- and post-workshop using a 10-point Likert scale, with model realism and utility similarly rated. A 3-6-month follow-up survey assessed clinical translation. RESULTS: Seventeen participants and 14 trainers from nine centres facilitated completion of 34 simulated procedures. Median overall confidence improved from 2.5 to 7.0 (P < 0.0005), with the greatest gains obtained in submucosal dissection (2.0 to 8.0), cuff management (2.0 to 8.0) and anastomosis (3.0 to 8.0) (all P < 0.001). Model realism and training utility were rated 9/10 and 9/10, respectively, by 25 respondents (93% response rate). Follow-up survey (12/17; 71%) showed 100% agreement that the workshop improved ability to assist or perform HPTS. CONCLUSION: This structured simulation-based HPTS workshop, using a validated low-cost model, significantly improved procedural confidence which translated into behavioural change in clinical practice. The model offers high fidelity, scalability and ethical training.

OBJECTIVES: To evaluate the efficacy and safety of minimally invasive sacral neuromodulation (SNM) and noninvasive enteral neuromodulation (ENM) in children with refractory gastrointestinal motility disorders (GMD). MATERIALS AND METHODS: This prospective exploratory trial enrolled pediatric patients with GMD between 2019 and 2024 at a single tertiary referral center. Children with inflammatory bowel disease or mechanical causes of GMD were excluded. Participants received either SNM through an implanted device or ENM through surface electrodes. Stimulation was delivered at 14 Hz, 210 μs pulse width, with individualized intensity (median 1.0 mA for SNM; 6.0 mA for ENM). Primary outcomes were abdominal pain, fecal incontinence, defecation frequency, and stool consistency. Treatment success was defined as clinically significant improvement in at least two of these four domains. Quality of life was assessed at baseline and 12 weeks. Safety outcomes were monitored over a 12-month follow-up. RESULTS: Of 70 eligible patients, 48 completed the study (18 SNM; 30 ENM). Diagnoses included Hirschsprung disease, functional constipation, and congenital neuronal malformations. Severe comorbidities were more frequent in the SNM group (45%) than the ENM group (3%; p = 0.0018). Treatment success was observed in 80% (24/30) of the ENM cohort and 83% (15/18) of SNM cohort. No significant differences were found between groups for individual outcomes. No major complications occurred. Minor adverse events were comparable (ENM 27%; SNM 17%). CONCLUSIONS: Both SNM and ENM are effective and safe options for treating pediatric GMD and may be considered within a multimodal therapeutic approach. CLINICAL TRIAL REGISTRATION: This trial is registered at clinicaltrials.gov (ID NCT04713085, title "Sacral Neuromodulation in Children and Adolescents"). Web link: https://clinicaltrials.gov/study/NCT04713085.

Hirschsprung disease (HSCR) is a complex developmental disorder of the enteric nervous system, primarily driven by regulatory variants within enhancer elements of the RET gene. To investigate how these variants lead to aganglionosis, we developed a humanized mouse model by inserting an intact 77kb human RET genomic locus into the Rosa26 safe-harbor locus. Utilizing "big DNA" synthetic biology and Bxb1-mediated recombination, we integrated the complete human locus including all exons, introns, and upstream regulatory elements which we validated via nanopore and short-read sequencing. Functional analysis confirmed in vivo human RET expression; however, our initial HSCR-associated "sensitive" haplotype expressed at only 21% of wild-type levels. This significant reduction proved insufficient to rescue the viability when endogenous mouse Ret was deleted. We identified that this deficiency is partially driven by five risk SNPs within established enhancers. Specifically, using CRISPR/Cas9 to restore a conserved So×10 binding site (converting a sensitive SNP to a protective one) increased RET expression by 1.9-fold and restored transcription factor binding. This study provides a robust framework for modeling human-specific regulatory disorders and demonstrates the critical impact of non-coding variation on disease pathogenesis.