Duhamel and transanal endorectal pull-throughs for Hirschsprung disease: a Bayesian network meta-analysis.

BACKGROUND: To comprehensively compare the effects of open Duhamel (OD), laparoscopic-assisted Duhamel (LD), transanal endorectal pull-through (TEPT), and laparoscopic-assisted endorectal pull-through (LEPT) in Hirschsprung disease. METHODS: PubMed, Embase, Cochrane Library, Web of Science, CNKI, WanFang, and VIP were comprehensively searched up to August 4, 2022. The outcomes were operation-related indicators and complication-related indicators. The Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach was used to evaluate the quality of evidence. Network plots, forest plots, league tables and rank probabilities were drawn for all outcomes. For measurement data, weighted mean differences (WMDs) and 95% credibility intervals (CrIs) were reported; for enumeration data, relative risks (RRs) and 95%CrIs were calculated. RESULTS: Sixty-two studies of 4781 patients were included, with 2039 TEPT patients, 1669 LEPT patients, 951 OD patients and 122 LD patients. Intraoperative blood loss in the OD group was more than that in the LEPT group (pooled WMD = 44.00, 95%CrI: 27.33, 60.94). Patients lost more blood during TEPT versus LEPT (pooled WMD = 13.08, 95%CrI: 1.80, 24.30). In terms of intraoperative blood loss, LEPT was most likely to be the optimal procedure (79.76%). Patients undergoing OD had significantly longer gastrointestinal function recovery time, as compared with those undergoing LEPT (pooled WMD = 30.39, 95%CrI: 16.08, 44.94). The TEPT group had significantly longer gastrointestinal function recovery time than the LEPT group (pooled WMD = 11.49, 95%CrI: 0.96, 22.05). LEPT was most likely to be the best operation regarding gastrointestinal function recovery time (98.28%). Longer hospital stay was observed in patients with OD versus LEPT (pooled WMD = 5.24, 95%CrI: 2.98, 7.47). Hospital stay in the TEPT group was significantly longer than that in the LEPT group (pooled WMD = 1.99, 95%CrI: 0.37, 3.58). LEPT had the highest possibility to be the most effective operation with respect to hospital stay. The significantly reduced incidence of complications was found in the LEPT group versus the LD group (pooled RR = 0.24, 95%CrI: 0.12, 0.48). Compared with LEPT, OD was associated with a significantly increased incidence of complications (pooled RR = 5.10, 95%CrI: 3.48, 7.45). Patients undergoing TEPT had a significantly greater incidence of complications than those undergoing LEPT (pooled RR = 1.98, 95%CrI: 1.63, 2.42). For complications, LEPT is most likely to have the best effect (99.99%). Compared with the LEPT group, the OD group had a significantly increased incidence of anastomotic leakage (pooled RR = 5.35, 95%CrI: 1.45, 27.68). LEPT had the highest likelihood to be the best operation regarding anastomotic leakage (63.57%). The incidence of infection in the OD group was significantly higher than that in the LEPT group (pooled RR = 4.52, 95%CrI: 2.45, 8.84). The TEPT group had a significantly increased incidence of infection than the LEPT group (pooled RR = 1.87, 95%CrI: 1.13, 3.18). LEPT is most likely to be the best operation concerning infection (66.32%). Compared with LEPT, OD was associated with a significantly higher incidence of soiling (pooled RR = 1.91, 95%CrI: 1.16, 3.17). Patients with LEPT had the greatest likelihood not to develop soiling (86.16%). In contrast to LD, LEPT was significantly more effective in reducing the incidence of constipation (pooled RR = 0.39, 95%CrI: 0.15, 0.97). LEPT was most likely not to result in constipation (97.81%). LEPT was associated with a significantly lower incidence of Hirschprung-associated enterocolitis (HAEC) than LD (pooled RR = 0.34, 95%CrI: 0.13, 0.85). The OD group had a significantly higher incidence of HAEC than the LEPT group (pooled RR = 2.29, 95%CrI: 1.31, 4.0). The incidence of HAEC was significantly greater in the TEPT group versus the LEPT group (pooled RR = 1.74, 95%CrI: 1.24, 2.45). LEPT was most likely to be the optimal operation in terms of HAEC (98.76%). CONCLUSION: LEPT may be a superior operation to OD, LD and TEPT in improving operation condition and complications, which might serve as a reference for Hirschsprung disease treatment.

Increased Fibronectin Impairs the Function of Excitatory/Inhibitory Synapses in Hirschsprung Disease.

Although approximately 50% of cases have a known genetic defect, the precise pathogenesis of Hirschsprung disease (HSCR) is still unclear. We recently reported that expression of fibronectin (FN), which is involved in the migration, colonization, and differentiation of enteric neural crest cells (ENCCs), is increased in aganglionic colonic segments obtained from patients. We hypothesized that abnormally high levels of FN might play a role in the etiology of HSCR. Here, to test this hypothesis, we investigated aganglionic, transitional, and ganglionic colon segments from 63 children with HSCR and distal colon from thirty healthy Wistar rats at embryonic day 20, in addition to in vitro studies with PC12 Adh neural crest cells. We measured the protein and mRNA expression levels of FN, together with a panel of excitatory (VGLUT1, GluA1, GluN1, PSD-95, and NL-1) and inhibitory (GAD67, GABA AR-α1, NL-2, and SLC32) synaptic markers. Expression of all these synaptic markers was significantly decreased in aganglionic colon, compared to ganglionic colon, whereas expression of FN was significantly increased. In a neural crest cell line, PC12 Adh, knockdown of FN with small-interfering RNA increased the expression of synaptic markers. Co-culture of colons from embryonic day 20 rats with RGD recombinant protein, which contains the RGD motif of FN, reduced the expression of excitatory and inhibitory synaptic markers. These results are consistent with the idea that the etiology of HSCR involves aberrant overexpression of FN, which may impair synaptic function and enteric nervous system development, leading to motor dysfunction of intestinal muscles.

Aberrant Distributions of Collagen I, III, and IV in Hirschsprung Disease.

BACKGROUND: Hirschsprung disease (HSCR) is the most common congenital gut motility disorder, involving a severe anomaly of the enteric nervous system, and is characterized by functional intestinal obstruction due to lack of intrinsic innervation (aganglionosis) in the distal bowel. OBJECTIVE: The aim of this study was to examine the distribution patterns of collagens I (Col I), III (Col III), and IV (Col IV) in the enteric nervous system of HSCR patients, to determine whether or not collagen levels are altered in the aganglionic bowel. METHODS: We measured the expression levels of Col I, Col III, and Col IV in colonic muscle from 129 children with HSCR. The localizations of the 3 collagens and myenteric ganglia were assessed morphologically by immunofluorescence staining. western blots and real-time fluorescence quantitative polymerase chain reaction were performed to examine the relative levels of these collagens in aganglionic, transitional, and ganglionic colon segments. RESULTS: Immunoreactivities of Col I and Col III were high around and within myenteric ganglia in the ganglionic segment, moderate in the transitional segment, and weak in the aganglionic segment. Col IV immunoreactivity showed the opposite pattern, being lowest in the ganglionic segment and highest in the aganglionic segment. CONCLUSION: Col I and Col III are decreased and Col IV is increased in the distal colon of patients with HSCR.

The down-regulation of neuroligin-2 and the correlative clinical significance of serum GABA over-expression in Hirschsprung's disease.

The goal of this study was to investigate the expression level of neuroligin-2 in different colon tissue segments of children with Hirschsprung's disease (HSCR) and the correlative clinical significance of serum Gamma-Aminobutyric Acid (serum GABA) in HSCR. Neuroligin-2 was assessed by Immunohistochemistry staining method on routine paraffin section from different colon tissue segments of HSCR (ganglionic colonic segment, transitional colonic segment and aganglionic colonic segment). Western-blot analysis and real-time fluorescence quantitative PCR(qRT-PCR) were applied to compare and evaluate the expression levels of neuroligin-2 from three segments of HSCR, and we used Enzyme-linked Immunosorbent Assay (ELISA) method to detect and compare the serum GABA between HSCR and non-HSCR. Immunohistochemistry staining demonstrated that intensive neuroligin-2 staining was detected in the ganglion cells in the ganglionic colonic and transitional colonic segments from the HSCR children; however, neuroligin-2 staining was down-regulated significantly in the aganglionic colonic segments. The expression levels of neuroligin-2 mRNA and protein in the aganglionic colonic segment were decreased compared to the ganglionic colonic segment and transitional colonic segment (P < 0.05). And the level of serum GABA was significantly higher in HSCR than that in non-HSCR. The expression of neuroligin-2 varies from different segments of HSCR. The down-regulation of neuroligin-2 in aganglionic colonic segments may be correlated with the excessive intestine contraction and further result in HSCR. The over-expression of serum GABA may be considered as a new diagnostic method of HSCR.

Expression of neurexin and neuroligin in the enteric nervous system and their down-regulated expression levels in Hirschsprung disease.

To investigate the expression levels of neurexins and neuroligins in the enteric nervous system (ENS) in Hirschsprung Disease (HSCR). Longitudinal muscles with adherent mesenteric plexus were obtained by dissection of the fresh gut wall of mice, guinea pigs, and humans. Double labeling of neurexin I and Hu (a neuron marker), neuroligin 1 and Hu, neurexin I and synaptophysin (a presynaptic marker), and neuroligin 1 and PSD95 (a postsynaptic marker) was performed by immunofluorescence staining. Images were merged to determine the relative localizations of the proteins. Expression levels of neurexin and neuroligin in different segments of the ENS in HSCR were investigated by immunohistochemistry. Neurexin and neuroligin were detected in the mesenteric plexus of mice, guinea pigs, and humans with HSCR. Neurexin was located in the presynapse, whereas neuroligin was located in the postsynapse. Expression levels of neurexin and neuroligin were significant in the ganglionic colonic segment of HSCR, moderate in the transitional segment, and negative in the aganglionic colonic segment. The expressions of neurexin and neuroligin in the transitional segments were significantly down-regulated compared with the levels in the normal segments (P < 0.05). Expression levels of neurexin and neuroligin in ENS are significantly down-regulated in HSCR, which may be involved in the pathogenesis of HSCR.

Effect of green tea on reward learning in healthy individuals: a randomized, double-blind, placebo-controlled pilot study.

BACKGROUND: Both clinical and preclinical studies revealed that regular intake of green tea reduced the prevalence of depressive symptoms, as well as produced antidepressant-like effects in rodents. Evidence proposed that disturbed reward learning has been associated with the development of anhedonia, a core symptom of depression. However, the relationship between green tea and reward learning is poorly investigated. Our goal was to test whether chronic treatment with green tea in healthy subjects affects the process of reward learning and subsequently regulates the depressive symptoms. METHODS: Seventy-four healthy subjects participated in a double-blind, randomized placebo-controlled study with oral administration of green tea or placebo for 5weeks. We used the monetary incentive delay task to evaluate the reward learning by measurement of the response to reward trial or no-reward trial. We compared the reaction time of reward responsiveness between green tea and placebo treatment. Furthermore, we selected Montgomery-Asberg depression rating scale (MADRS) and 17-item Hamilton Rating Scale for Depression (HRSD-17) to estimate the depressive symptoms in these two groups. RESULTS: The results showed chronic treatment of green tea increased reward learning compared with placebo by decreasing the reaction time in monetary incentive delay task. Moreover, participants treated with green tea showed reduced scores measured in MADRS and HRSD-17 compared with participants treated with placebo. CONCLUSIONS: Our findings reveal that chronic green tea increased the reward learning and prevented the depressive symptoms. These results also raised the possibility that supplementary administration of green tea might reverse the development of depression through normalization of the reward function.

A single-center study of thoracoscopic surgery in the treatment of pediatric mediastinal neurogenic tumors.

OBJECTIVE: To study the feasibility, safety, and efficacy of thoracoscopic surgery in the treatment of pediatric mediastinal neurogenic tumors, and summarize the treatment experiences and surgical skills. METHODS: A single-center retrospective analysis of 37 patients with pediatric mediastinal neurogenic tumors was conducted. Clinical charactersistics and postoperative complications were all analyzed. RESULTS: All the operations were successfully completed. There was no statistically significant difference in tumor diameter between the two groups (p > 0.05). The open surgery group had an average operation time of 96.5 ± 32.38 min, while the thoracoscopic surgery group had an average operation time of 78.3 ± 24.51 min (p < 0.05). The thoracoscopic surgery group had significantly lower intraoperative blood loss than the open surgery group (p < 0.05). In addition, the duration of the postoperative thoracic drainage tube was 5.43 ± 0.76 days in the open surgery group, which was longer than the 2.38 ± 0.87 days in the thoracoscopic surgery group (p < 0.05). Furthermore, the postoperative length of hospital stay was an average of 10.23 ± 1.43 days for the open surgery group, longer than for the thoracoscopic surgery group (4.36 ± 0.87 days) (p < 0.05). CONCLUSIONS: Thoracoscopic surgery has several advantages in the treatment of pediatric mediastinal neurogenic tumors and is worthy of clinical popularization and application. For giant mediastinal malignant neurogenic tumors, puncture biopsy and adjuvant chemotherapy can be performed before surgery to lessen the tumor volume and enlarge the operation space, which would reduce bleeding and complications.

Risk factors for Hirschsprung disease-associated enterocolitis: a systematic review and meta-analysis.

BACKGROUND: The incidence of Hirschsprung disease (HSCR) is nearly 1/5000 and patients with HSCR are usually treated through surgical intervention. Hirschsprung disease-associated enterocolitis (HAEC) is a complication of HSCR with the highest morbidity and mortality in patients. The evidence on the risk factors for HAEC remains inconclusive to date. METHODS: Four English databases and four Chinese databases were searched for relevant studies published until May 2022. The search retrieved 53 relevant studies. The retrieved studies were scored on the Newcastle-Ottawa Scale by three researchers. Revman 5.4 software was employed for data synthesis and analysis. Stata 16 software was employed for sensitivity analysis and bias analysis. RESULTS: A total of 53 articles were retrieved from the database search, which included 10 012 cases of HSCR and 2310 cases of HAEC. The systematic analysis revealed anastomotic stenosis or fistula [ I2 =66%, risk ratio (RR)=1.90, 95% CI 1.34-2.68, P <0.001], preoperative enterocolitis ( I2 =55%, RR=2.07, 95% CI 1.71-2.51, P <0.001), preoperative malnutrition ( I2 =0%, RR=1.96, 95% CI 1.52-2.53, P <0.001), preoperative respiratory infection or pneumonia ( I2 =0%, RR=2.37, 95% CI 1.91-2.93, P <0.001), postoperative ileus ( I2 =17%, RR=2.41, 95% CI 2.02-2.87, P <0.001), length of ganglionless segment greater than 30 cm ( I2 =0%, RR=3.64, 95% CI 2.43-5.48, P <0.001), preoperative hypoproteinemia ( I2 =0%, RR=1.91, 95% CI 1.44-2.54, P <0.001), and Down syndrome ( I2 =29%, RR=1.65, 95% CI 1.32-2.07, P <0.001) as the risk factors for postoperative HAEC. Short-segment HSCR ( I2 =46%, RR=0.62, 95% CI 0.54-0.71, P <0.001) and transanal operation ( I2 =78%, RR=0.56, 95% CI 0.33-0.96, P =0.03) were revealed as the protective factors against postoperative HAEC. Preoperative malnutrition ( I2 =35 % , RR=5.33, 95% CI 2.68-10.60, P <0.001), preoperative hypoproteinemia ( I2 =20%, RR=4.17, 95% CI 1.91-9.12, P <0.001), preoperative enterocolitis ( I2 =45%, RR=3.51, 95% CI 2.54-4.84, P <0.001), and preoperative respiratory infection or pneumonia ( I2 =0%, RR=7.20, 95% CI 4.00-12.94, P <0.001) were revealed as the risk factors for recurrent HAEC, while short-segment HSCR ( I2 =0%, RR=0.40, 95% CI 0.21-0.76, P =0.005) was revealed as a protective factor against recurrent HAEC. CONCLUSION: The present review delineated the multiple risk factors for HAEC, which could assist in preventing the development of HAEC.

6'-O-Galloylpaeoniflorin Exerts Inhibitory Bioactivities in Human Neuroblastoma Cells via Modulating AMPK/miR-489/XIAP Pathway.

Although therapies against neuroblastoma (NBM) have advanced, the patients still suffer from poor prognoses due to distal metastasis or the occurrence of multidrug resistance. Accumulating evidence has proved that chemicals derived from natural products possess potent anti-NBM properties or can be used as adjuvants for chemotherapy. In the present study, we demonstrated that 6'-O-galloylpaeoniflorin (GPF), a galloylated derivative of paeoniflorin isolated from the roots of Paeonia lactiflora Pall, exerted significant inhibitory effects on proliferation and invasion of SH-SY5Y cells (an NBM cell line) and enhanced the sensitivity of SH-SY5Y cells to cisplatin in vitro. Further studies showed that GPF treatment upregulated miR-489 in NBM cells via activating AMP-activated protein kinase (AMPK). We also demonstrated that similar to GPF treatment, miR-489 exhibited a significant anti-NBM capacity. Further studies showed that miR-489 directly targeted the X-linked inhibitor of apoptosis protein (XIAP). Overall, our results indicated that GPF possessed an evident anti-NBM capacity dependent on AMPK/miR-489/XIAP pathway, providing an emerging strategy for clinical treatment of NBM.

Administration of a novel penicillamine-bound membrane: a preventive and therapeutic treatment for abdominal adhesions.

BACKGROUND: Adhesions formation is a significant postsurgical complication. At present, there is no effective method for preventing adhesions formation 1, although barrier products such as Dextran (Dex) 2 and sodium hyaluronate (SH) 3 have proved the most clinically successful 456, This study is designed to investigate the preventive and therapeutic potential of a novel penicillamine-bound membrane for abdominal adhesions formation. METHODS: 150 rats were involved in the present study. All animals were randomly divided into 6 groups (1 vehicle group and 5 test groups respectively treated with dextran, sodium hyaluronate, penicillamine, penicillamine-bound membrane or non-penicillamine-bound membrane). The occurrence, grade and score of abdominal adhesions were compared between the different groups. The breaking strength of incision was compared between the vehicle group and the penicillamine, membrane with/without penicillamine - treated groups. Expression of collagen type I was compared between the vehicle and penicillamine-treated group. The occurrence of adhesions was compared between the Dextran (Dex), sodium hyaluronate (SH), penicillamine-treated group and membrane with or without penicillamine- treated groups. RESULTS: Penicillamine and penicillamine-bound membrane had significant preventive effects on abdominal adhesions formation, better than dextran, sodium hyaluronate and non-penicillamine-bound membrane. However, neither of them influenced incision healing, although they insignificantly decreased the breaking strength of the incision. Penicillamine-bound membrane, which can be loaded locally and more efficaciously, shows greater advantages than penicillamine. CONCLUSIONS: Penicillamine-bound membrane can be applied as an effective therapeutic intervention for abdominal adhesions with inconsequential side effects.

Role of GDNF, GFRα1 and GFAP in a Bifidobacterium-Intervention Induced Mouse Model of Intestinal Neuronal Dysplasia.

OBJECTIVE: To investigate the effects of glial cell-derived neurotrophic factor (GDNF), GDNF family receptor alpha 1 (GFRα1), and glial fibrillary acidic protein (GFAP) on colonic motility in a mouse model of intestinal neuronal dysplasia by intervention with Bifidobacterium and to explore the influence of Bifidobacterium on enteric glial cells (EGCs). METHODS: Western blotting and qRT-PCR were employed to detect the expression of GFRα1 and GFAP in colonic tissues of mice with or without Tlx2 mutations, and ELISA was used to detect the expression of GDNF in serum. IHC was used to detect the appearance of the ganglion cells. Subsequently, Tlx2 homozygous mutant (Tlx2-/-) mice were treated with Bifidobacterium. Colonic motility was measured before and after intervention by measuring the glass bead expelling time. The variations in abdominal circumference and GDNF, GFRα1, and GFAP expression were measured. In addition, 16SrRNA gene sequencing was performed to detect the abundance of the intestinal microbiota. RESULTS: The mRNA and protein expression of GFRα1 and GFAP was decreased in the colonic tissues of Tlx2-/- mice and GDNF expression was decreased in serum compared with Tlx2+/- and WT mice. After confirming the colonization of Bifidobacterium by 16S rRNA gene sequencing, the expelling time and abdominal distension were ameliorated, and the expression of GFAP, GDNF, and GFRα1 was increased. CONCLUSIONS: The expression of GDNF, GFRα1, and GFAP is associated with colonic motility. The altered expression of EGC-related factors suggested that Bifidobacterium may be involved in the EGC activation process. The amelioration of IND symptoms after intervention with Bifidobacterium prompted the elicitation of adjuvant therapy.

Effect of Neuroligin1 and Neurexin1 on the Colonic Motility in a Mouse Model of Neuronal Intestinal Dysplasia.

AIM: To investigate the expressions of neuroligin1 (NL1) and neurexin1 (NX1) in a mouse model of neuronal intestinal dysplasia (Tlx2-/- mice) and to explore their effects on colonic motility. METHODS: Immunohistochemistry staining was employed to explore the histological appearances of NL1, NX1, the presynaptic marker of glutamatergic synapses VGLUT1, and the subunit of NMDA receptors of NR1 in the colon of mice with or without Tlx2 mutation. Western blotting and qRT-PCR were performed to detect their relative expressions in the colon. Colonic motility was measured by a glass bead technique. Then, the Tlx2-/- mice were intervened by Huperzine A. Variations on expressions of NL1, NX1, VGLUT1, and NR1 and variations on colonic motility were measured. Additionally, serum concentrations of Glu were measured by ELISA. RESULTS: Immunohistochemistry staining reveals that NL1, NX1, VGLUT1, and NR1 were mainly concentrated in the myenteric plexus of ENS. Compared to those in WT and Tlx2+/- mice, expressions of NL1 and NX1 in colon of Tlx2-/- mice were upregulated with increased VGLUT1 and NR1 abundances and impaired colonic motility (P < 0.05). After intervention, the upregulated expressions of NL1 and NX1 were decreased with a correlated reduce of VGLUT1 and NR1 and a recovery of the impaired colonic motility (P < 0.05). After intervention, the upregulated expressions of NL1 and NX1 were decreased with a correlated reduce of VGLUT1 and NR1 and a recovery of the impaired colonic motility (P < 0.05). After intervention, the upregulated expressions of NL1 and NX1 were decreased with a correlated reduce of VGLUT1 and NR1 and a recovery of the impaired colonic motility (. CONCLUSION: NL1 and NX1 are closely related to the colonic motility through their effects of targeting the formation of glutamatergic synapses and may be involved in the pathogenesis of NID. The variations of serum Glu seem to be a potential and less painful auxiliary measure for colonic motility and NID.

Comparative Clinical Study Between Modified Ureteral Orthotopic Reimplantation and Cohen Method Under Pneumovesicum in Pediatric Patients With Hydroureteronephrosis.

Purpose: To report our initial experience with a modified ureteral orthotopic reimplantation technique under pneumovesicum and compare the outcomes vs. those obtained with the Cohen technique under pneumovesicum for the correction of primary obstructive megaureter (POM) or vesicoureteral reflux(VUR) in pediatric patients. Methods: A total of 46 patients (38 POM and 8 VUR; mean age: 16.24 months) treated with modified ureteral orthotopic reimplantation (OR) and 43 patients (34 POM and 9 VUR; mean age: 22.98 months) treated with Cohen reimplantation (CR) under pneumovesicum were included. We compared the results perioperatively and during follow-up. Results: The mean operative time was significantly shorter in the OR group (OR: 86.86 and 108.18 vs. CR: 95.14 and 124.29 min for unilateral and bilateral cases, respectively). The mean postoperative hospital stay (OR: 5.02 vs. CR: 5.07 days), blood loss (OR: 3.67 vs. CR: 3.84 ml), and follow-up time (OR: 23.17 vs. CR: 23.37 months) did not exhibit significant differences between the two groups. One patient converted to open surgery in the CR group, whereas there was no conversion in the OR group. Postoperative febrile urinary tract infection occurred in two cases in each group. Both infections were controlled using antibiotics. All patients in both groups showed improved hydroureteronephrosis, and all patients with VUR showed reflux resolution post-surgery. Conclusions: Our modified ureteral orthotopic reimplantation technique under pneumovesicum can be safely and effectively performed, achieving a high success rate that is equivalent to that obtained through the Cohen technique under pneumovesicum. Moreover, it involves a simpler procedure and shorter operation time.

Abundance and Significance of Neuroligin-1 and Neurexin II in the Enteric Nervous System of Embryonic Rats.

Aim. To investigate the abundance of neuroligin-1 and neurexin II in the enteric nervous system (ENS) of rats on different embryonic days and to explore their potential significance. Methods. The full-thickness colon specimens proximal to the ileocecal junction of rats on embryonic days 16, 18, and 20 and of newborns within 24 hours (E16, E18, E20, and Ep0) were studied, respectively. qRT-PCR was applied for detecting the expressions of neuroligin-1 and neurexin II on mRNA, and western blotting was employed for detecting their further expressions on the whole tissue. Finally, the histological appearance of neuroligin-1 and neurexin IIα was elucidated using immunohistochemical staining. Results. qRT-PCR showed that the neuroligin-1 and neurexin II mRNA expressions of groups E16, E18, E20, and Ep0 increased gradually with the growth of embryonic rats (P < 0.05). Western blotting confirmed the increasing tendency. In immunohistochemical staining, proteins neuroligin-1 and neurexin IIα positive cells concentrated mostly in the myenteric nerve plexus of the colon and their expressions depend on the embryonic time. Conclusion. Neuroligin-1 and neurexin II were both expressed in the ENS and have temporal correlation with the development of ENS, during which neuronal intestinal malformations (NIM) may occur due to their disruptions and consequent abnormal ENS development.

Expression and significance of neuroligins in myenteric cells of Cajal in Hirschsprung's disease.

PURPOSE: The aim of this study was to investigate the expression and significance of neuroligins in myenteric cells of Cajal (ICC-MY) in Hirschsprung's disease (HSCR). METHODS: Longitudinal muscle with adherent myenteric plexus (LMMP) from surgical excision waste colon of HSCR children were prepared by peeling off the mucous layer, sub-mucosal layer and circular muscle. Neuroligins, c-Kit (c-Kit-immunoreactivity representing ICC) and their relationship were assessed by double labeling immunofluorescence staining. ICC-MY were dissociated and cultured from LMMP by enzymolysis method, and were purified and analyzed using a combination of magnetic-activated cell sorting (MACS) and flow cytometry (FCM). Western-blot analysis was applied to compare and evaluate the expression levels of neuroligins in ICC-MY which were dissociated from different segments of HSCR (ganglionic colonic segment, transitional colonic segment and aganglionic colonic segment). RESULTS: Neuroligins and c-Kit were expressed on the same cells (ICC-MY); ICC-MY were dissociated, cultured and purified. For HSCR, neuroligins were expressed significantly in ICC-MY from ganglionic colonic segments, moderately in those from transitional colonic segments and down-regulated significantly in those from aganglionic colonic segments. CONCLUSIONS: Neuroligins were expressed in ICC-MY of human beings, and the expression varies from different segments of HSCR. This abnormal expression might play an important role in the pathogenesis of this disease through affecting the synaptic function of ICC-MY.