NOX5 is expressed aberrantly but not a critical pathogenetic gene in Hirschsprung disease

Background Hirschsprung disease (HSCR) is a congenital disorder characterized by the absence of intramural ganglion cells in the distal gastrointestinal tract (GI), which results in tonic contraction of the aganglionic gut segment and functional intestinal obstruction. Recent studies have suggested NADPH oxidase 5 (NOX5) as a candidate risk gene for HSCR. In this study, we examined the function of NOX5 to verify its role in the development of the enteric nervous system (ENS). Methods HSCR tissue specimens (n = 10) were collected at the time of pull-through surgery and control specimens (n = 10) were obtained at the time of colostomy closure in patients. The NOX5 expression in aganglionic and ganglionic segments of HSCR colon and normal colon were analyzed by immunohistochemistry (IHC), western blot and real-time quantitative PCR (qPCR). The gene expression levels and spatiotemporal expression spectrum of NOX5 in different development stages of zebrafish embryo were determined using qPCR and in-situ hybridization (ISH). The enteric nervous system in NOX5 Morpholino (MO) knockdown and wild type (WT) zebrafish embryo was analyzed by whole-mount immunofluorescence (IF). Intestinal transit assay was performed to analyze the gastrointestinal motility in NOX5 knockdown and control larvae. Results NOX5 is strongly expressed in the ganglion cells in the proximal segment of HSCR colons and all segments of normal colons. Moreover, the expression of NOX5 is markedly decreased in the aganglionic segment of HSCR colon compared to the ganglionic segment. In zebrafish, NOX5 mRNA level is the highest in the one cell stage embryos and it is decreased overtime with the development of the embryos. Interestingly, the expression of NOX5 appears to be enriched in the nervous system. However, the number of neurons in the GI tract and the GI motility were not affected upon NOX5 knockdown. Conclusions Our study shows that NOX5 markedly decreased in the aganglionic segment of HSCR but didn’t involve in the ENS development of zebrafish. It implies that absence of intestinal ganglion cells may lead to down-regulation of NOX5.

THE VALUE OF INTRAOPERATIVE EXPRESS BIOPSY IN DETERMINING THE LEVEL OF COLON RESECTION IN THE SURGICAL TREATMENT OF HIRSCHSPRUNG'S DISEASE

Introduction. Hirschsprung's disease is a congenital malformation characterized by the absence of intramural ganglia in the intestinal wall. The goals of the surgical treatment of HD are to remove the aganglionic segment of the intestine and restore the patency of the gastrointestinal tract, bringing down the normally innervated intestine to the anus while maintaining the function of the sphincters. An incorrect determination of the length of the affected segment of the colon can lead to non-radical surgical treatment with a persistent aganglionic zone. The use of an intraoperative express biopsy to determine the resection area during radical surgery will help to solve this problem. Purpose: determination of the level of agangliosis of the colon in the surgical treatment of Hirschsprung's disease. Methods: From 2010 to 2019, 49 patients aged 3 weeks to 16 years underwent an intraoperative express biopsy to determine the area of bowel resection. The use of intraoperative express biopsy made it possible to objectively and reliably determine the area of bowel resection, to reduce the number of complications associated with errors in resection of the aganglionic segment. Conclusion. When performing radical surgery for Hirschsprung's disease, an intraoperative express biopsy is required to determine the level of coloanal anastomosis in order to avoid errors in determining the aganglionic region and the transitional zone of the intestine.

A novel third space endoscopic procedure, per-rectal endoscopic myotomy, for Hirschsprung’s disease: Medium and long-term outcomes

Introduction Hirschsprung’s disease (HSCR) is congenital aganglionosis affecting hindgut and presents with constipation. Surgical pull-through is current standard but has morbidity. Per-rectal endoscopic myotomy (PREM) is a novel third space endoscopy technique for treating SS-HSCR. Methods Retrospective series of SS-HSCR patients diagnosed on history, contrast enema, rectal biopsies, anorectal manometry and treated by PREM. Aganglionic segment mapped pre-PREM. PREM performed using third space endoscopy principles. Objective – to compare stool frequency and unit laxative (UL) usage pre and post-PREM. Results N = 9; duration 4-years. Mean age – 7.5 (± 5.2) years; 7 males. Mean aganglionic segment length – 6.3 (± 4.4) cm. Mean procedure time – 96.1 (±37.9) minutes. Mean LOS – 2.5 (±0.7) days. Median follow-up –17 months (9 – 58). Stool frequency – pre – 1/4.4 (±1.5) vs. post – 1/1.2 (±0.4) days (p = 0.0004). Mean UL usage – pre – 5.4 (±4.9) vs. post – 0.4 (±0.7) units (p = 0.0002). No laxatives in 6 (66%). Single AE (anal stenosis) – dilatation. Conclusions PREM is a safe and effective minimally invasive procedure for SS-HSCR and provides long-term response.

Ultrashort-segment Hirschsprung disease in a 4-year-old female

Hirschsprung disease (HSCR) is characterized by the absence of neuronal ganglion cells in a distal portion of the intestinal tract [1]. In 1691, Frederick Ruysch described the disease as congenital megacolon. HSCR-associated congenital anomalies have been reported in 5–32% of affected patients [2]. The clinical symptoms of HSCR are usually evident in the neonatal period. However, in some cases where the extent of the aganglionic segment is short, symptoms may become clinically relevant later in childhood [3]. HSCR is one of the most difficult diseases to identify in pediatric surgery due to its multiple clinical, histological and radiological variations [2, 3]. The goal of surgical management is to remove the aganglionic segment and reconstruct the intestinal tract through techniques such as Swenson, Duhamel and Soave [4]. The following case consists of a 4-year-old patient with a chronic presentation of constipation secondary to ultrashort-segment Hirschsprung disease.

NOX5 is Expressed Aberrantly but Not a Critical Pathogenetic Gene in Hirschsprung Disease

Background:Hirschsprung disease (HSCR) is a congenital disorder characterized by the absence of intramural ganglion cells in the distal gastrointestinal tract (GI), which results in tonic contraction of the aganglionic gut segment and functional intestinal obstruction. Recent studies have suggested NADPH oxidase 5 (NOX5) as a candidate risk gene for HSCR. In this study, we examined the function of NOX5 to verify its role in the development of enteric nervous system (ENS).Methods: HSCR tissue specimens (n = 10) were collected at the time of pull-through surgery and control specimens (n = 10) were obtained at the time of colostomy closure in patients. The NOX5 expression in aganglionic and ganglionic segments of HSCR colon and normal colon were analyzed by immunohistochemistry (IHC), western blot and RT-qPCR. The gene expression levels and spatiotemporal expression spectrum of NOX5 in different development stages of zebrafish embryo were investigated using real-time quantitative PCR (RT-qPCR) and in-situ hybridization (ISH). The enteric nervous system in NOX5 Morpholino (MO) knockdown and wild type (WT) zebrafish embryo was analyzed by whole-mount immunofluorescence (IF). Intestinal transit assay was performed to analyze the gastrointestinal motility in NOX5 knockdown and control larvae.ResultsNOX5 is strongly expressed in the ganglion cells in the proximal segment of HSCR colons and all segments of normal colons. Moreover, the expression of NOX5 is markedly decreased in the aganglionic segment of HSCR colon compared to the ganglionic segment. In zebrafish, NOX5 mRNA level is the highest in one cell stage embryos and it is decreased by the development of the embryos. Interestingly, the expression of NOX5 appears to be enriched in the nervous system. However, the number of neurons in the GI tract and the GI motility were not affected upon NOX5 knockdown.ConclusionsNOX5 may play a role in the early development of zebrafish embryo, but not required for the ENS development and loss of NOX5 didn’t affect the GI motility in zebrafish. Nevertheless, we demonstrated that NOX5 specifically expressed in the ganglionic cell in colon tissue and markedly decreased in the aganglionic segment.

A Hirschsprung Pull-through, “with a Twist”

Hirschsprung disease is the most common neurocristopathy in children, resulting in the congenital loss of enteric ganglia. Surgery, which involves resecting the aganglionic segment and restoring bowel continuity, usually results in a good outcome; however, some patients suffer from multiple episodes of enterocolitis and other obstructive symptoms. A contrast enema, examination under anesthesia, and rectal biopsy can identify the cause of obstruction in many cases, including a rare explanation, a twist of the pull-through, a case of which we present here.

Surgery, Surgical Pathology, and Postoperative Management of Patients With Hirschsprung Disease

Endorectal pullthrough surgery is integral in the treatment of patients with Hirschsprung disease. Several different surgical procedures exist, which share as common goals to excise the aganglionic segment and upstream transition zone and attach ganglionic bowel just proximal to the anal canal. The operation requires collaboration between surgeon and pathologist to localize ganglionic bowel and prevent retention of transition zone. Intraoperative frozen sections are extremely important, first to establish that ganglion cells are present and subsequently to exclude features of transition zone (partial circumferential aganglionosis, myenteric hypoganglionosis, and submucosal nerve hypertrophy) at the proximal surgical (anastomotic) margin. Postoperative histopathological analysis of resection specimens should be tailored to document distal aganglionosis, document the length of the aganglionic segment and its proximity to the anastomotic margin, and confirm that transition zone has been resected completely. Adherence to the recommendations described in this review will reduce the likelihood of transition zone pullthrough and should decrease the incidence of persistent postoperative obstructive symptoms.

CONGENITAL AGANGLIONOSIS;

… Objective : To report early and late outcome after a one stage operation andcomparison with others similar work and to see feasibility of this procedures in our limitedresources. Patients and method: We have performed transanal pull through in department ofPaediatric Surgery, Nishtar Hospital Multan during May 2010 to May 2013. Fifty patients out of 117were selected, older than one year, non marasmic and diagnosed as short segmentaganglionosis by barium enema were selected. Transanal Swenson’s pull through wasperformed, aganglionic segment was decided on gross appearance (narrow) due to lack offrozen section facility, anastomosis made with 4/0 vicryl. Results: Procedure completed on anaverage within one hour, without any transfusion. Feeding was started from day one, manageablecomplications were observed in 25% patients, no mortality seen. Hospital stay was on anaverage 6 days. Conclusions: Transanal one stage pull through is very practical, can be donewithout facility of frozen section.