Morphometrie der Ganglien und Nervenzellen des Plexus submucosus bei der intestinalen neuronalen Dysplasie des Erwachsenen

Intestinal neuronal dysplasia is a developmental abnormality of the plexus submucosus in the colon. In recent years this initially in children described dysganglionosis has also been found in adults with chronic constipation. The aim of this study was a morphometric characterization of intestinal neuronal dysplasia in the submucous plexus of adults. Biopsies of 10 adults with intestinal neuronal dysplasia were compared with biopsies of 10 healthy controls. Nerve cells and ganglia were stained selectively with a lactate dehydrogenase reaction. Morphometry was made with an optic electronic analysis system. The detection of 6 to 10 giant ganglia with more than 7 nerve cells in 15 biopsy sections proved to be the most characteristic diagnostic indicator of intestinal neuronal dysplasia. The number of giant ganglia in the submucous plexus was 20%. For a quantitative objectivation of intestinal neuronal dysplasia 15 biopsy sections with enough submucosa are necessary.     

Oligoneuronal Hypoganglionosis in Patients with Idiopathic Slow-Transit Constipation

PURPOSE: Several alterations of the enteric nervous system have been described as an underlying neuropathologic correlate in patients with idiopathic slow-transit constipation. To obtain comprehensive data on the structural components of the intramural nerve plexus, the colonic enteric nervous system was investigated in patients with slow-transit constipation and compared with controls by means of a quantitative morphometric analysis. METHODS: Resected specimens were obtained from ten patients with slow-transit constipation and ten controls (nonobstructive neoplasias) and processed for immunohistochemistry with the neuronal marker Protein Gene Product 9.5. The morphometric analysis was performed separately for the myenteric plexus and submucous plexus compartments and included the quantification of ganglia, neurons, glial cells, and nerve fibers. RESULTS: In patients with slow-transit constipation, the total ganglionic area and neuronal number per intestinal length as well as the mean neuron count per ganglion were significantly decreased within the myenteric plexus and external submucous plexus. The ratio of glial cells to neurons was significantly increased in myenteric ganglia but not in submucous ganglia. On statistical analysis, the histopathologic criteria (submucous giant ganglia and hypertrophic nerve fibers) of intestinal neuronal dysplasia previously described in patients with slow-transit constipation were not completely fulfilled. CONCLUSION: The colonic motor dysfunction in slow-transit constipation is associated with quantitative alterations of the enteric nervous system. The underlying defect is characterized morphologically by oligoneuronal hypoganglionosis. Because the neuropathologic alterations primarily affect the myenteric plexus and external submucous plexus, superficial submucous biopsies are not suitable to detect these innervational disorders.     

突触素单克隆抗体免疫组化实验诊断肠神经发育不良症的价值

为探讨肠神经发育不良症（IND）的有效诊断方法，对45例术前诊断为先天性巨结肠（HD）的慢性肠梗阻患儿行巨结肠根治手术，并对其结肠痉挛段标本进行突触素单克隆抗体免疫组化实验。结果显示，16例结肠标本可见粘膜下神经丛过渡增生并有巨神经节（诊断为IND），与正常对照组比较，P＜0．01，其中14例有异位神经节（9例合并HD，下称HAIND），其病理切片中发现有异位的神经节细胞，且部分神经节细胞有变形，空泡化；10例IND（其中8例HAIND）可固有层和粘膜肌染色增强，与正常对照组比较，P＜0．01。认为突触素单克隆抗体免疫组化实验显示粘膜一神经丛过度增生，巨神经节可作为诊断IND的硬性指标；异位神经节，固有层和粘膜肌染色增强可作为诊断IND的辅助指标。在病理切片中应用突触素单克隆抗体免疫组化实验，可清晰显示肠壁神经丛的发育情况，是诊断IND的可靠方法。     

Rectal biopsy for diagnosis of intestinal neuronal dysplasia in children: a prospective multicentre study on interobserver variation and clinical outcome

BACKGROUND: Intestinal neuronal dysplasia (IND) of the colonic submucous plexus is considered to be a congenital malformation of the enteric nervous system causing symptoms resembling those of Hirschsprung's disease. In contrast with the established diagnosis of aganglionosis using enzyme histochemistry, controversy exists over the diagnostic criteria of IND on rectal biopsies previously defined by a consensus report and the causal relation between morphological findings and clinical symptoms. AIMS: The interobserver variability was prospectively investigated with respect to final diagnoses and several histological features in rectal biopsy specimens from children suspected of having colonic motility disturbances. METHODS: 377 biopsy specimens from 108 children aged 4 days to 15 years were independently coded without knowledge of clinical symptoms by three experienced pathologists for 20 histological features, and a final diagnosis was given for every case. Interobserver variation for the different items and the final diagnosis were analysed using Cohen's kappa statistic. Clinical data at biopsy and outcome after 12 months were related to morphological findings. RESULTS: The three pathologists agreed completely with respect to the diagnosis Hirschsprung's disease (kappa = 1), but in only 14% of the children without aganglionosis. In 15 (17%) of the 87 children without aganglionosis, at least one pathologist judged the case as normal, while another diagnosed IND. kappa values were close to the zero value expected by chance for the diagnoses normal and IND. Young age was related to the presence of several morphological features-for example, acetylcholine esterase staining and presence of giant ganglia. Children with chronic constipation diagnosed as having IND, given no other specific diagnosis by any of the pathologists, were significantly younger (median 8.8 months) and had a higher cure rate after one year (60%) than constipated patients considered by all observers to have no histological abnormalities (median 6.1 years, cure rate 23%). CONCLUSIONS: In contrast with Hirschsprung's disease, there is a high interobserver variation with regard to the different morphological features and final diagnosis of IND, based on the criteria and conditions of the previous consensus report. The high frequency of histological "abnormalities" in young infants suggests that some of the features may represent a normal variant of postnatal development rather than a pathological process. Investigations using more refined and morphometric methods in rectal specimens from infants and children without bowel disease are needed to define the normal range of morphological appearance at different ages. These preliminary data indicate that, with current knowledge, rectal biopsy for diagnostic purposes should only be performed in constipated children for diagnosis of Hirschsprung's disease.     

Abnormalities of the enteric nervous system in heterozygous endothelin B receptor deficient (spotting lethal) rats resembling intestinal neuronal dysplasia

BACKGROUND: A homozygous mutation of the endothelin B receptor (EDNRB) gene in spotting lethal (sl/sl) rats leads to Hirschsprung's disease (HSCR) with long segmented aganglionosis. However, the effects on the development of the enteric nervous system (ENS) promoted by a heterozygous mutation of the EDNRB gene are not known. The present study aimed to describe and morphometrically assess the phenotypic abnormalities of the ENS in heterozygous (+/sl) EDNRB deficient rats in comparison with homozygous (sl/sl) EDNRB deficient and wild-type (+/+) rats. METHODS: The distal small intestine, caecum, and colon were obtained from sl/sl, +/sl, and +/+ rats. To demonstrate the three dimensional organisation of the ENS, the intestinal wall was microdissected into wholemounts and incubated against the pan-neuronal marker protein gene product 9.5. Assessment of the ENS included morphometric quantification of ganglionic size and density, the number of nerve cells per ganglia, and the diameter of nerve fibre strands within both the myenteric and submucous plexus. RESULTS: Sl/sl rats were characterised by complete aganglionosis resembling the same histopathological features observed in patients with HSCR. +/sl rats revealed more subtle abnormalities of the ENS: the submucous plexus was characterised by a significantly increased ganglionic size and density, and the presence of hypertrophied nerve fibre strands. Morphometric evaluation of the myenteric plexus did not show statistically significant differences between +/sl and +/+ rats. CONCLUSIONS: In contrast with sl/sl rats, +/sl rats display non-aganglionated malformations of the ENS. Interestingly, these innervational abnormalities resemble the histopathological criteria for intestinal neuronal dysplasia (IND). Although IND has been described in several intestinal motility disorders, the concept of a clearly defined clinical-histopathological entity is still controversially discussed. The present findings support the concept of IND based on clearly defined morphological criteria suggesting a genetic link, and thus may provide a model for human IND. Furthermore, the data underline the critical role of the "gene dose" for the phenotypic effects promoted by the EDNRB/EDN3 system and confirm that the development of the ENS is not an "all or none" phenomenon.     

Intestinal neuronal dysplasia]

Intestinal neuronal dysplasia (IND) type B is a disease of the submucosal plexus of intestine manifesting chronic intestinal obstruction or severe chronic constipation. IND is one of intestinal dysganglionoses and clinically closely associated with Hirschsprung's disease. Until recently, it is not fully clear whether IND is a congenital malformation or an acquired secondary condition related to some gastrointestinal problems. However, recently published data and consensus reports have enhanced our understanding of the pathogenesis and management of IND. The aim of this paper was to review the current state of knowledge regarding the controversial issues of IND including the etiology, classification, diagnostic criteria, and available therapeutic intervention.     

Differential changes in intrinsic innervation and interstitial cells of Cajal in small bowel atresia in newborns

AIM: To investigate morphological changes of the enteric nervous system (ENS) and the interstitial cells of Cajal (ICCs) in small bowel atresia.METHODS: Resected small bowel specimens from affected patients (n = 7) were divided into three parts (proximal, atretic, distal). Standard histology and enzyme immunohistochemistry anti-S100, anti-protein gene product (PGP) 9.5, anti-neurofilament (NF), antic-kit-receptor (CD117) was carried out on conventional paraffin sections of the proximal and distal part. RESULTS: The neuronal and glial markers (PGP 9.5, NF, S-100) were expressed in hypertrophied ganglia and nerve fibres within the myenteric and submucosal plexuses. Furthermore, the submucous plexus contained typical giant ganglia. The innervation pattern of the proximal bowel resembled intestinal neuronal dysplasia. The density of myenteric ICCs was clearly reduced in the proximal bowel, whereas a moderate number of muscular ICCs were found. The anti-CD117 immunore- action revealed additional numerous mast cells. The distal bowel demonstrated normal morphology and density of the ENS, the ICCs and the mast cells.CONCLUSION: The proximal and distal bowel in small bowel atresia revealed clear changes in morphology and density of the ENS and ICCs.     

Histological studies on Hirschsprung's disease and its allied disorders in childhood

BACKGROUND/AIMS: To obtain accurate diagnosis for Hirschsprung's disease (HD) and its allied disorders such as hypoganglionosis (Hypo) and intestinal neuronal dysplasia (IND) in childhood patients with chronic constipation, we studied the histology of childhood patients with refractory constipation accompanied by abdominal distension and pain. METHODOLOGY: Based on clinical signs and symptoms noted on admission, all of 109 patients (60 males and 49 females, aged 2-15 years with a mean age of 9.8 years) were suspected to have chronic refractory constipation. To obtain accurate histological diagnosis in childhood patients with chronic refractory constipation, we performed rectal biopsies on these patients. Tissue samples were frozen and 12-microm sections were stained with acetylcholinesterase (AChE) by the method of Karnovsky and Roots, and with nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase by the modified Scherer-Singler's method. RESULTS: On the basis of histological studies using rectal biopsies, 20 cases were diagnosed with Hypo, 5 with HD, 2 with intestinal neuronal dysplasia (IND), and 82 with normal findings. The incidence of normal cases was significantly greater than that of allied disorders of HD including both Hypo and IND (P<0.0001). The incidence of Hypo was also significantly greater than that of Hypo and IND (P<0.01, P<0.0001, respectively). Both HD and IND could be diagnosed by rectal mucosal biopsies with AChE staining. However, accurate diagnosis of Hypo could be made only through examination of the submucosal and myenteric plexuses by NADPH-diaphorase staining in full-thickness rectal specimens. CONCLUSIONS: We were able to obtain accurate diagnosis of childhood patients with HD and IND by rectal mucosal biopsy with AChE staining. On the other hand, accurate histological diagnosis in patients with Hypo could also be obtained by NADPH-diaphorase staining in full-thickness rectal specimens. That is to say, it is easier for the investigator to detect the cholinergic fiber and ganglion cell in the gut wall using NADPH-diaphorase staining than by using AChE staining.     

Neurochemical features of endomorphin-2-containing neurons in the submucosal plexus of the rat colon

AIM:To investigate the distribution and neurochemical phenotype of endomorphin-2(EM-2)-containing neurons in the submucosal plexus of the rat colon.METHODS:The mid-colons between the right and left flexures were removed from rats,and transferred into Kreb's solution. For whole-mount preparations,the mucosal,outer longitudinal muscle and inner circularmuscle layers of the tissues were separated from the submucosal layer attached to the submucosal plexus. The whole-mount preparations from each rat mid-colon were mounted onto seven gelatin-coated glass slides,and processed for immunofluorescence histochemical double-staining of EM-2 with calcitonin gene-related peptide(CGRP),choline acetyltransferase(Ch AT),nitric oxide synthetase(NOS),neuron-specific enolase(NSE),substance P(SP) and vasoactive intestinal peptide(VIP). After staining,all the fluorescence-labeled sections were observed with a confocal laser scanning microscope. To estimate the extent of the co-localization of EM-2 with CGRP,Ch AT,NOS,NSE,SP and VIP,ganglia,which have a clear boundary and neuronal cell outline,were randomly selected from each specimen for this analysis. RESULTS:In the submucosal plexus of the mid-colon,many EM-2-immunoreactive(IR) and NSE-IR neuronal cell bodies were found in the submucosal plexus of the rat mid-colon. Approximately 6 ± 4.2 EM-2-IR neurons aggregated within each ganglion and a few EM-2-IR neurons were also found outside the ganglia. The EM-2-IR neurons were also immunopositive for Ch AT,SP,VIP or NOS. EM-2-IR nerve fibers coursed near Ch AT-IR neurons,and some of these fibers were even distributed around Ch AT-IR neuronal cell bodies. Some EM-2-IR neuronal cell bodies were surrounded by SP-IR nerve fibers,but many long processes connecting adjacent ganglia were negative for EM-2 immunostaining. Long VIP-IR processes with many branches coursed through the ganglia and surrounded the EM-2-IR neurons. The percentages of the EM-2-IR neurons that were also positive for Ch AT,SP,VIP or NOS were approximately 91% ± 2.6%,36% ± 2.4%,44% ± 2.5% and 44% ± 4.7%,respectively,but EM-2 did not co-localize with CGRP. CONCLUSION:EM-2-IR neurons are present in the submucosal plexus of the rat colon and express distinct neurochemical markers.     

Intestinal neuronal dysplasia

Intestinal neuronal dysplasia type B (IND B) is currently defined as a disease of the submucous plexus of the intestine. The aetiology of IND B remains largely obscure. The congenital origin of IND B is supposed; nevertheless, the findings of IND B associated with chronic intestinal obstruction support the notion that this disease could be caused by a reaction of the enteral nervous system to intestinal obstruction or inflammatory disease either in the fetal or the postnatal period. The treatment of IND type B has no unified concept of treatment. The ultimate clinical diagnosis of IND B should be based on a definitive histological diagnosis relating to clinical symptoms, the course of treatment and long-term follow-up of patients with this dysfunction of intestinal motility, despite the fact that no correlations of the clinical picture, radiological investigation and anorectal manometric studies with IND B have been found so far.     

Nerve cell density in submucous plexus throughout the gut of cat and opossum

Quantitative differences in submucous plexus density were sought in cat and opossum gut by examining full-thickness whole mounts of the submucosa stained with silver, and counting ganglia per square centimeter and nerve cell bodies per ganglion in order to compute density of innervation (nerve cell bodies per square centimeter). In the cat, the nerve cell bodies per square centimeter in the 12 named regions were as follows: proximal esophagus, 0; mid-esophagus, 0; distal esophagus, 0; fundus, 84; gastric antrum, 18; duodenum, 5831; jejunum, 4632; ileum, 3191; proximal colon, 1275; mid-colon, 689; distal colon, 359; rectum, 144. In the opossum, values were as follows: proximal esophagus, 37; mid-esophagus, 52; distal esophagus, 84; duodenum, 1812; jejunum, 2234; ileum, 1488; proximal colon, 206; mid-colon, 197; distal colon, 121; rectum, 61. Adequate specimens could not be obtained from opossum stomach. Differences were due more to variations in distribution density of ganglia than in ganglionic size. The relatively dense submucous plexus of the intestine probably is related to the capacity of the intestinal mucosa for peptide secretion as well as to its absorptive function.     

Multiple endocrine neoplasia 2B: Differential increase in enteric nerve subgroups in muscle and mucosa

Multiple endocrine neoplasia 2B(MEN2B) is a rare syndrome caused by an activating mutation of the RET gene, leading to enteric gangliomatosis. This child presented with constipation at 1-mo old, was diagnosed with MEN2 B by rectal biopsy at 4 mo, had thyroidectomy at 9 mo and a colectomy at 4 years. We studied the extent of neuronal and nerve fibre proliferation and which classes of enteric nerves are affected by examining the colon with multiple neuronal antibodies. Resected transverse colon was fixed, frozen, sectioned and processed for fluorescence immunohistochemistry labelling with antibodies against TUJ1, Hu, ChAT, NOS, VIP, SP and CGRP and cKit. Control transverse colon was from the normal margin of Hirschsprung(HSCR) colon(4-year-old) and a child with familial adenomatous polyposis(FAP, 12 year). Myenteric ganglia were increased in size to as wide as the circular muscle. There was a large increase in nerve cells and nerve fibres. ChAT-, NOS-, VIP-and SP-immunoreactive nerve fibres all increased in the myenteric ganglia. NOS-IR nerves preferentially increased in the muscle, while VIP and SP increased in submucosal ganglia and mucosal nerve fibres. The density of ICC was normal. RET overactivation in MEN2B lead to a large increase in intrinsic nerve fibres in the myenteric and submucosal ganglia, with a relative increase in NOS-IR nerve fibres in the circular muscle and VIP and SP in the submucosal ganglia and mucosa. The changes were associated with severe constipation resulting in colectomy at 4 years.     

Intestinal neuronal dysplasia type B: A still little known diagnosis for organic causes of intestinal chronic constipation

Intestinal neuronal dysplasia type B(IND-B) is a controversial entity among the gastrointestinal neuromuscular disorders. It may occur alone or associated with other neuropathies, such as Hirschsprung's disease(HD). Chronic constipation is the most common clinical manifestation of patients. IND-B primarily affects young children and mimics HD, but has its own histopathologic features characterized mainly by hyperplasia of the submucosal nerve plexus. Thus, IND-B should be included in the differential diagnoses of organic causes of constipation. In recent years, an increasing number of cases of IND-B in adults have also been described, some presenting severe constipation since childhood and others with the onset of symptoms at adulthood. Despite the intense scientific research in the last decades, there are still knowledge gaps regarding definition, pathogenesis, diagnostic criteria and therapeutic possibilities for IND-B. However, in medical practice, we continue to encounter patients with severe constipation or intestinal obstruction who undergo to diagnostic investigation for HD and their rectal biopsies present hyperganglionosis in the submucosal nerve plexus and other features, consistent with the diagnosis of IND-B. This review critically discusses aspects related to the disease definitions, pathophysiology and genetics, epidemiology distribution, clinical presentation, diagnostic criteria and therapeutic possibilities of this still little-known organic cause of intestinal chronic constipation.     

Das neurogene Megakolon: Liegt immer ein Morbus Hirschsprung zugrunde?

The development of megacolon in adults is attributed to malformations of the enteric nervous system apart from mechanic, metabolic, endocrinologic, pharmacologic, neurologic, infectious or systemic causes. Hirschsprung’s disease is considered to represent the most acknowledged form of intestinal innervation disorders underlying the formation of megacolon. In order to evaluate this association, morphologic alterations of the enteric nervous system were examined in patients (age: 19 to 67 years) with megacolon. From the resected colonic segments conventional serial sections and whole-mount preparations were obtained and submitted to immunohistochemical procedures for Protein Gene Product 9.5 as a neuronal marker allowing the 2-dimensional assessment of the architecture of the intramural nervous plexus layers. Whereas complete aganglionosis was diagnosed in only 25% of the cases examined, thus resembling classic Hirschsprung’s disease, the remaining colonic segments showed other forms of intestinal neuronal malformations: 1. Hypoganglionosis of varying severity, 2. intestinal neuronal dysplasia characterised by submucosal giant ganglia and concomitant hypertrophy of nerve fibers, 3. heterotopic ganglia located ectopically within the longitudinal muscle layer as well as within the lamina propria mucosae. In contrast to conventional histologic sections, whole-mount preparations allowed a more subtle assessment of the morphologic alterations of intramural nervous plexus from the normal to the pathologic area and, therefore, a more precise diagnostic classification of intestinal innervation disorders. It could be shown that neurogenic megacolon in adults is not only caused by aganglionosis but also by non-aganglionic innervation disorders. The findings implicate that in adolescent and adult patients suffering from intestinal motility disorders and a concomitant development of megacolon hypoganglionic conditions and intestinal neuronal dysplasia have to be taken into consideration in regards to the diagnostic and therapeutic approach.     

Human myenteric plexus: confirmation of unfamiliar structures in adults and neonates

To define the myenteric plexus along the human gastrointestinal tract, we studied three neonatal and six adult specimens, postmortem, by silver impregnation. There were no clear differences between the neonatal and the adult gastrointestinal tracts. In the body of the esophagus, the plexus was sparse, with few ganglia; 30%-40% of fascicular intersections were devoid of ganglia. In the lower 5 cm, the esophagus had thick bundles of nerve fibers ("shunt fascicles"), which crossed the gastroesophageal junction and radiated to the periphery of the stomach through several branches. The plexus in the stomach was uniform, with intermediate and intrafascicular ganglia. A thick nerve bundle encircled the pylorus and gave branches on either side to the antrum and the duodenum. Shunt fascicles in the stomach did not cross the pylorus but extended to the distal antrum. In the duodenum and proximal jejunum, the plexus was regular, but in the mid-small intestine, the longitudinal interganglionic fascicles were more prominent than the circumferential fascicles. Distally, this pattern was reversed; circumferential fascicles were more prominent and ganglia were dense in the terminal ileum. Thin, short shunt fascicles were scattered along the entire small intestine, becoming more abundant in the terminal ileum. Short, thick shunt fascicles traveled proximally from the ileocecal junction for about 25-30 cm. As in the stomach, shunt fascicles did not cross the ileocecal junction, but a thick nerve bundle encircled it. In the cecum and proximal colon, the plexus was sparse with large intermediate and intrafascicular ganglia. In the rectum and distal colon, the plexus was dense, with parafascicular and intrafascicular ganglia. Long ascending nerves extended from the distal rectum into the midcolon. In addition, there were short, thick nerve bundles in the rectum that traveled proximally.     

Analysis of the RET, GDNF, EDN3, and EDNRB genes in patients with intestinal neuronal dysplasia and Hirschsprung disease

BACKGROUND: Hirschsprung disease (HSCR) is a frequent congenital disorder with an incidence of 1 in 5000 live births, characterised by the absence of parasympathetic intramural ganglion cells in the hindgut resulting in intestinal obstruction in neonates and severe constipation in infants and adults. Intestinal neuronal dysplasia (IND) shares clinical features with HSCR but the submucosal parasympathetic plexus is affected. IND has been proposed as one of the most frequent causes of chronic constipation and is often associated with HSCR. METHODS: We examined 29 patients diagnosed with sporadic HSCR, 20 patients with IND, and 12 patients with mixed HSCR/IND for mutations in the coding regions of the RET, GDNF, EDNRB, and EDN3 genes. The entire coding regions were analysed by single strand conformational polymorphism and DNA sequencing. RESULTS: Only three RET mutations were detected in patients with HSCR. In patients with IND or a mixed HSCR/IND phenotype, no mutations in these genes were observed. While HSCR and HSCR/IND showed over representation of a specific RET polymorphism in exon 2, IND exhibited a significantly lower frequency comparable with that of controls. CONCLUSIONS: The mutation frequency found in our sporadic HSCR patients (10%) and the allelic distribution of RET polymorphisms are comparable with earlier published data. A significantly different allelic distribution in an established HSCR associated polymorphism argues against common genetic pathways for HSCR and IND.     

Localisation of endothelin like immunoreactivity in adult and developing human gut.

The distribution of immunoreactivity for the potent vasoconstrictor endothelin-1 was studied in adult and developing human gut using antisera to endothelin-1 (1-21) and the C terminus of big endothelin-1. The coexistence of these peptides with other neuropeptides was investigated using comparative immunocytochemistry. Endothelin-1 like immunoreactivity was detected in extracts of adult (range 20-60 fmol/g wet weight) and fetal (33 fmol/g) gastrointestinal tract and was shown by chromatography to be the predominant isoform of endothelin present in both. It was localised by immunocytochemistry to ganglion cells in the submucous and myenteric plexuses and to scattered nerves, whereas big endothelin-1 like immunoreactivity was found in the submucous plexus only. Colocalisation studies showed immunoreactivity for both endothelin-1 and vasoactive intestinal peptide in the same ganglion cells of the submucous plexus. Although endothelin-1 immunoreactivity was not detected by immunocytochemistry in the fetal human gut until the 32nd week of gestation, big endothelin-1 was found as early as 11 weeks in the developing neural structures and epithelial cells. The latter were shown to be endocrine cells by their immunoreactivity for chromogranin. Our results indicate that endothelin is a neuropeptide found in adult human gut which shows transient expression in endocrine cells during development.     

Challenges in the diagnosis of intestinal neuronal dysplasia type B: A look beyond the number of ganglion cells

Intestinal neuronal dysplasia type B (IND-B) is a controversial condition among gastrointestinal neuromuscular disorders. Constipation is its most common clinical manifestation in patients. Despite intense scientific research, there are still knowledge gaps regarding the diagnostic criteria for IND-B in the histopathological analysis of rectal biopsies. The guidelines published in the past three decades have directed diagnostic criteria for quantifying the number of ganglion cells in the nervous plexus of the enteric nervous system. However, it is very complex to distinguish numerically what is pathological from what is normal, mainly because of the difficulty in determining a reliable control group composed of healthy children without intestinal symptoms. Thus, a series of immunohistochemical markers have been proposed to assist in the histopathological analysis of the enteric nervous system. Several of these markers facilitate the identification of other structures of the enteric nervous system, in addition to ganglion cells. These structures may be related to the etiopathogenesis of IND-B and represent new possibilities for the histopathological diagnosis of this disease, providing a view beyond the number of ganglion cells. This review critically discusses the aspects related to the disease definitions and diagnostic criteria of this organic cause of constipation.     

Origins of peptide and norepinephrine nerves in the mucosa of the guinea pig small intestine

Norepinephrine, acetylcholine, and certain peptides are contained in mucosal nerves and have potent effects on transepithelial water and electrolyte fluxes. It is difficult to ascribe roles for these nerves as their sources are unknown. The present studies were undertaken to determine the origins of nerve fibers that are found in the mucosa of the guinea pig small intestine and which contain one of the following substances: vasoactive intestinal peptide, substance P, somatostatin, neuropeptide Y, cholecystokinin, or norepinephrine. Nerve fiber origins were ascertained by making lesions to sever pathways through which the nerves could reach the mucosa. The lesioning operations were homotopic autotransplants of short (2 cm) segments of intestine; myectomies, in which a 5-10-mm length of intestine was stripped of longitudinal muscle and myenteric plexus; and extrinsic denervation, in which nerves reaching the intestine through the mesentery were severed. The results of these studies, considered along with previously published work, led to the upcoming conclusions. Nerve fibers in the mucosa showing immunoreactivity for vasoactive intestinal peptide, somatostatin, cholecystokinin, and neuropeptide Y arise from cell bodies in the overlying submucous plexus. Substance P fibers arise in part from the overlying submucous plexus and in part from the overlying myenteric plexus. Mucosal norepinephrine fibers arise from extrinsic sympathetic ganglia. Enkephalin, gastrin-releasing peptide, and 5-hydroxytryptamine, which are in some enteric nerves, are not found in submucous nerve cells and few, if any, fibers containing these substances supply the mucosa. Thus, the mucosa receives a dense nerve supply, much of which arises locally from submucous ganglia.     

Morphological study on the motility of the human colon

We tried to make a clear three-dimensional picture of the autonomic nerves in the wall of the human colon, using a Golgi method rarely applied to human materials. At autopsy, sigmoid colon without mucosal lesions were collected from 16 males after sudden death from apoplexy, head injury, or myocardial infarction. These materials were fixed in 10% formalin, impregnated with a modified Golgi method and embedded in celloidin. Then three-dimensional serial sections were made and observed with a light microscope. Many fine nerve fibers formed a plexus in the subserosa, muscular layer, submucosa, and mucosa. The myenteric plexus was made up of rectangular meshes of nerve fiber bundles. However, unlike myenteric plexus, no regular mesh was found in the submucosal plexus. Further, nerve fibers connecting myenteric and submucosal plexus were observed. It may be concluded from these findings that there exist nerve pathways regulating intestinal motility between myenteric and submucosal plexus.