慢传输型便秘结肠神经病理改变的意义

目的研究慢传输型便秘结肠肌间神经丛的神经丝蛋白和S-100蛋白的病理改变,探索结肠动力减弱的原因,为临床治疗提供理论依据.方法采用免疫组织化学方法研究33例结肠慢传输型便秘患者(STC组)和25例非便秘性结肠(对照组)的升结肠、横结肠、降结肠、乙状结肠的肌间神经丛内神经丝蛋白和S-100蛋白的表达,利用计算机图象分析系统作定量分析,并与病程及年龄作直线相关性分析.所得数据用t检验进行统计学处理.结果对照组结肠肌间神经丛内神经丝蛋白和S-100蛋白的含量在各段之间无显著性差别(0.09±0.03vs0.10±0.02,P＞0.05),STC组结肠各段与对照组比较,神经丝蛋白的平均光密度值明显高于对照组(0.12±0.03vs0.09±0.02,P＜0.01);S-100蛋白的含量及平均光密度值明显高于对照组(0.10±0.04vs0.08±0.03,P＜0.01).神经丝蛋白和S-100蛋白的改变随着病程的延长而增加,二者呈直线相关(r=0.75).结论慢传输型便秘结肠肌间神经丛存在着全结肠性退行性病理改变,表现为神经丝蛋白的堆积聚集和神经间质的增生,且随着病程的延长而加重,这是造成结肠动力减弱的主要原因.提示,手术切除结肠的范围应是全结肠或次全结肠.     

慢传输型便秘乙状结肠VIP,SP免疫组化研究

目的探讨慢传输型便秘（ＳＴＣ）的神经病理学基础。方法应用半定量免疫细胞组织化学的方法，对１４例ＳＴＣ和１１例非梗阻性直肠腺癌患者的乙状结肠标本进行研究，主要观察肠壁内血管活性肠肽（ＶＩＰ）和Ｐ物质（ＳＰ）的变化。结果常规ＨＥ染色下，两组结肠肌间神经丛无明显改变；免疫组化见ＳＴＣ患者乙状结肠壁内ＶＩＰ含量减少（Ｐ＜０．０５）；ＳＰ含量明显降低（Ｐ＜０．００１）；而粘膜层内无明显变化。结论ＳＴＣ患者结肠壁存在明显的神经病理学变化，其结肠传输减慢可能与肠壁内ＶＩＰ和ＳＰ能神经元数量减少和／或功能障碍有关。     

慢传输型便秘肠神经系统一氧化氮合酶和P物质的分布意义

目的研究结肠一氧化氮（ＮＯ）和Ｐ物质（ＳＰ）在慢传输型便秘（ＳＴＣ）发病中的作用．方法应用一氧化氮合酶（ＮＯＳ）和ＳＰ的兔多克隆抗体，对手术切除的１５例ＳＴＣ患者（男２例，女１３例，年龄２６岁～６８岁）和１１例对照组患者乙状结肠标本进行免疫组化染色和半定量分析．结果ＳＴＣ患者乙状结肠肌间神经丛ＮＯＳ免疫反应性明显升高（χ２＝１５６３，Ｐ＜００１），而ＳＰ免疫反应性明显降低（χ２＝１３４４，Ｐ＜００１）；粘膜下神经丛ＮＯＳ及ＳＰ免疫反应性与对照组相差不显著．结论ＳＴＣ结肠神经系统ＮＯＳ和ＳＰ免疫反应性的异常变化，可能是其结肠传输减慢的神经病理基础     

慢传输型便秘结肠神经病理改变及其临床意义

目的研究慢传输型便秘结肠肌间神经丛的神经丝蛋白和S100蛋白的病理改变,探索结肠动力减弱的原因,为临床治疗提供理论依据.方法采用免疫组织化学方法研究33例结肠慢传输型便秘患者(STC组)和25例非便秘性结肠(对照组)的升结肠、横结肠、降结肠、乙状结肠的肌间神经丛内神经丝蛋白和S-100蛋白的表达,利用计算机图象分析系统作定量分析,并与病程及年龄作直线相关性分析.所得数据用t检验进行统计学处理.结果对照组结肠肌间神经丛内神经丝蛋白和S-100蛋白的含量在各段之间无显著性差别(P>0.05),STC组结肠各段与对照组比较,神经丝蛋白的含量无明显减少(P>0.05),但出现堆积聚集现象,平均光密度值明显高于对照组(P<0.01);S100蛋白的含量及平均光密度值明显高于对照组(P<0.01).神经丝蛋白和S-100蛋白的改变随着病程的延长而增加,二者呈直线相关(P<0.02)结论慢传输型便秘结肠肌间神经丛存在着全结肠性退行性病理改变,表现为神经丝蛋白的堆积聚集和神经间质的增生,且随着病程的延长而加重,这是造成结肠动力减弱的主要原因.提示,手术切除结肠的范围应是全结肠或次全结肠.     

胃电节律失常大鼠胃窦肌间神经丛胆碱能神经的改变

目的：探讨大鼠胃窦肌间神经丛胆碱能神经，氮能神经含量变化与胃电节律失常的关系。方法：63例大鼠随机分为正常对照组，胃电节律失常模型组和白芍组。饲养4周后记录并分析胃电信号，测定胃窦肌间神经丛胆碱能神经含量。结果：模型组胃电节律失常明显增加，胃窦肌间神经丛胆碱能神经含量减少；经白芍治疗后，胃电节律失常明显减少，胃窦肌间神经丛胆碱能神经含量恢复正常。结论：胃窦肌间神经丛的胆碱能神经与胃电节律关系密切，当胆碱能神经减少时，胃电节律失常明显增加。     

胆囊收缩素在胃电节律失常中的作用及其神经病理学机制

目的：探讨胆囊收缩素（CCK）在胃电节律失常中的作用及其神经学机制。方法：在建立胃窦肌间神经丛铺片方法的基础上,用酶组织化学与免疫细胞化学方法,观察胃电节律失常大鼠胃窦肌间神经丛内胆碱能（Ach）神经、一氧化氮合酶（NOS）神经及CCK神经的变化。结果：模型组和CCK组大鼠均出现胃电节律失常,异常节律指数及慢波频率变异系数均显著高于正常组（P＜0．01）;模型组和CCK组NOS神经显著增加,Ach神经含量显著减少（P＜0．01）。结论：外源性及内源性CCK增加,能诱发胃电节律失常。CCK通过激活NOS,产生胃电节律失常。胃窦肌间神经丛神经中CCK及NOS神经含量异常增加,Ach神经减少是发生胃电节律失常的神经病理学机制之一。     

慢传输型便秘大鼠结肠壁内神经病理学改变

慢传输型便秘（STC）是一类以胃肠动力减弱为主要特点的顽固性便秘。由于临床上难以对便秘患者进行前瞻性研究，因而对其病因及发病机制缺乏深入的认识。目前国内外STC的病理学研究对象主要为手术切除的人体标本，缺乏前瞻性的研究资料，而临床便秘患者就诊前大多有长期服药史，关于结肠的病理改变是否为继发性争议较多。本研究应用复方苯乙哌啶建立大鼠STC模型，对结肠神经系统内多种神经病理改变进行检测，并从神经病理学角度初步探讨STC的发病机制。     

便秘小鼠结肠肌的电生理变化

目的观察便秘小鼠结肠肌的电生理变化。方法用复方地芬诺酯混悬液对小鼠进行灌胃建立便秘模型,然后测定其小肠结肠段的肌电,并分析其电生理活动变化。结果与对照组比较,便秘组慢波肌电频率明显增快(P<0.01);结肠肌电慢波频率的变异系数明显增大(P<0.01);结肠肌电慢波振幅明显减少(P<0.01);结肠肌电慢波振幅变异系数明显增大(P<0.01)。结论便秘小鼠结肠肌电慢波频率及振幅的异常改变可能是导致结肠传输减慢的重要原因。     

钙敏感受体在肠神经系统中的表达

目的观察与探讨G-蛋白耦联受体(CASR)在豚鼠肠神经系统中的表达。方法以豚鼠为试验对象,先进行RT-PCR设计,然后采用Western blotting来检测豚鼠肠神经系统中CASR蛋白,并且应用间接法双重免疫荧光标记对肠神经系统整体、肠神经系统中的肌间神经丛黏膜下神经丛中的CASR分布及数量等进行测定,荧光标记物采用FITC和Cy3。结果无论是肠神经系统中的肌间神经丛还是黏膜下神经丛,不同的肠神经细胞中含有不同的神经元标记物,即均存在CASR。结论豚鼠肠神经系统存在钙敏感受体的表达,因此,关于CASR在肠神经系统中的相关研究具有重要的临床意义,可能为将来研究CASR在肠道神经系统中的功能及肠炎症中的作用奠定基础。     

大鼠泻剂结肠模型的建立

目的应用大黄和酚酞建立一种大鼠泻剂结肠的动物模型．方法健康成年Ｗｉｓｔａｒ大鼠８４只，分２期，第１期３６只，第２期４８只，均分为对照组（饲以普通饲料）、大黄组（饲料中添加有大黄粉）和酚酞组（饲料中添加酚酞）．第１期：大黄及酚酞开始剂量为１００ｍｇ／（ｋｇ·ｄ），逐渐递增，３０ｄ后，大黄剂量为３２００ｍｇ／（ｋｇ·ｄ），酚酞为４０００ｍｇ／（ｋｇ·ｄ）．第２期：大黄及酚酞起始剂量为２００ｍｇ／（ｋｇ·ｄ），３ｍｏ后，大黄最终用量为２６００ｍｇ／（ｋｇ·ｄ），酚酞为３６００ｍｇ／（ｋｇ·ｄ）．活性炭悬液推进法测定肠道传输功能，并行结肠标本ＨＥ染色及肌间神经丛嗜银染色．结果与对照组相比，两期试验中大黄组和酚酞组肠道传输均有明显减慢；肌间神经丛嗜银染色大黄组和酚酞组有肌间神经丛嗜银性减弱甚至消失．结论本泻剂结肠大鼠模型具有慢传输性便秘的肠道传输延迟和肌间神经丛嗜银性明显降低等肠道功能及病理变化，简单经济，可重复性强．     

慢传输型便秘发病机制的研究

慢传输型便秘(STC)的发病机制主要与肠神经系统(ENS)、Cajal间质细胞(ICC)、平滑肌、神经递质等有关。研究发现STC结肠组织中ENS出现退行性变化,肌间神经丛空泡变性,ICC数量减少,形态改变,平滑肌退行性变,多种神经递质发生改变。本文就STC发病机制的研究作一综述。     

Altered carbachol-induced contractile responses of rat jejunal smooth muscle following local myenteric plexus ablation

Alterations in smooth muscle responsiveness and neural pathways in adjacent tissue may occur after local myenteric denervation. The in vitro contractile responses of both longitudinal and circular muscle to the mixed muscarinic and nicotinic cholinergic agonist carbachol were determined 15, 30, and 45 days after localized myenteric plexus ablation. Denervated longitudinal muscle exhibited decreased responsiveness to carbachol at all times examined. Denervated circulated muscle was initially supersensitive, but with time became subsensitive. These changes probably reflect the loss of the nicotinic (neuronal) component of the action of carbachol. Muscle orad to the site of denervation appeared subsensitive, while muscle caudad to the lesion was supersensitive (circular) or unaffected (longitudinal). These results suggest that there are changes in ascending and descending neural pathways. Alterations in the cholinergic responsiveness of intestinal smooth muscle, both at and beyond the site of myenteric plexus ablation, may result in altered intestinal motility that could lead to functional obstruction.Supported by National Institutes of Health grant AM 32594.Contribution 209, Environmental Toxicology Center, University of Wisconsin-Madison, Madison, Wisconsin 53706.     

Nitric oxide synthase and VIP distribution in enteric nervous system in idiopathic chronic constipation

Idiopathic chronic constipation has been correlated to neural abnormalities that consist of a reduced number of myenteric plexus neurons and a decreased concentration of VIP-positive nerve fibers within the circular muscle. Recent studies hypothesized the involvement of nitric oxide in motility disorders of the human gut. To date, no information is available on nitric oxide involvement in idiopathic chronic constipation. The density of VIP- and nitric oxide-producing neurons was evaluated by immunocytochemistry using anti-VIP and anti-nitric oxide synthase antibodies in five patients with idiopathic chronic constipation. A low total neuron density was found at the myenteric plexus. The density of VIP-positive neurons was low while that of nitric oxide synthase-positive neurons was high at both plexuses. Our data confirm that idiopathic slow-transit chronic constipation is due to abnormal neurogenic factors. The presence of numerous nitric oxide synthase-positive neurons, all along the colon and at both plexuses, supports the hypothesis that an excessive production of nitric oxide may cause the persistent inhibition of contractions.Supported by MURST University Funds.     

Reaggregation of Rat Dissociated Myenteric Plexus in Extracellular Matrix Gels

The aim of this study was to investigate the growth behavior of freshly dissociated myenteric plexus in a three-dimensional extracellular matrix (ECM) environment with and without stimulation of glial cell line-derived neurotrophic factor (GDNF). Therefore, cell suspensions of the dissected myenteric plexus of newborn rats were cultured in freshly prepared gels of commercially available mixtures of collagen, laminin, and hepatoglycans as a first step towards mimicking the natural environment of the myenteric plexus. The cultures were kept either in chemically defined serum-free medium alone or supplemented with GDNF. Cultures on polylysine-coated glass cover slips served as controls. Dissociated myenteric plexus grown on polylysine formed dense clusters of neurons with radially outgrowing nerve fibers, while the neurons cultured in the gel reaggregated to much smaller clusters. These contained, depending on the culture conditions, 2–10 neurons. The morphology of the network that was seen in the gels after a few days in vitro resembled very closely the in situ situation of the submucous plexus and the myenteric plexus in hypoganglionic children. Electron microscope investigations showed a high degree of organization with fiber bundles and vesicle-containing varicosities and growth cones. Independent of the method of culturing, GDNF obviously influenced the growth behavior of the dissociated plexus. The size of the ganglia was larger, and the secondary network denser when GDNF was supplemented. Moreover, the enteric neurons in the gel cultures tended to be larger in size when treated with GDNF. Three-dimensional cultures of dissociated myenteric plexus in an ECM gel might be a valuable tool towards the understanding of the formation of the enteric nervous system during development, especially considering pathological conditions such as Hirschsprungs disease or other dysganglionic diseases.     

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.     

Substance P Enhances Neuronal Area and Epithelial Cell Proliferation After Colon Denervation in Rats

We evaluated the neurotrophic effect of the neurokinin SP in the induced megacolon and the cell proliferation of the colonic epithelium after treatment. Colon segments of Wistar rats were chemically denervated by topical application of 2 mM benzalconium chloride and animals were daily injected intraperitoneally with SP (70 g/kg body wt) for 15 days. Control rats received either SP or were denervated and treated with saline. Neuronal profiles of the myenteric plexus were studied by immunohistochemistry to motor protein myosin V and cell proliferation by PCNA immunolabeling. Denervation induced a significant reduction in the number of neurons and an enlargement of the surviving perikarya (from 263.7 m² in the control-saline group to 468.9 m² in the denervated groups). The total area occupied by neurons was maintained in the denarvated SP group but was significantly smaller in the denervated-saline group. The proliferative index was significantly higher in the denervated groups, of which the SP-treated group showed the highest index. These results suggest that SP may have a neurotrophic effect for the neurons of the myenteric plexus chemically denervated and that this denervation stimulates cell proliferation, especially after SP administration.     

Ultrastructure of rat duodenal myenteric plexus revealed by quick-freezing and deep-etching method

A quick-freezing and deep-etching (QF-DE) method was employed with whole-mount strips of rat duodenal muscle walls to exhibit the cytoskeletons of the myenteric plexus. Nerve fibers in the myenteric plexus, which contained fewer neurofilaments than other types of neurons examined, had many varicosed contours, and were bundled by enteroglial cells. Cytoskeleton arrays were rarely observed in the varicosed regions, where synaptic vesicles were often seen, although other nerve regions contained many neurofilaments running almost in parallel with the nerve fiber bundle. Enteroglial cells had short cytoskeletons predominantly across the cytoplasm, becoming thinner the around varicosed regions of the nerve bundles. Such enteroglial extruded areas were often in close association with neighboring nerve fibers, indicating intercommunications between the nerve fibers. In distal parts of enteric nerve processes, there were numerous synaptic vesicles, but few neurofilaments. Smooth muscle cells were closely associated with the enteric nerve processes. Fine network structures, responsible for the extracellular matrix, were present between the smooth muscle cells and the enteric nerve processes. These specific structures of the myenteric plexus could be important for signalling or for the transportation of neurotransmitters involved in gut motility. (Received Feb. 25, 1998; accepted July 6, 1998)     

Clinical and morphofunctional features of idiopathic myenteric ganglionitis underlying severe intestinal motor dysfunction: a study of three cases

Ganglionitis, i.e., the inflammatory neuropathy characterized by a marked lymphoplasmacellular infiltrate in the myenteric plexus, may underlie a variety of paraneoplastic, infectious, or neurological disorders, although occasional cases are idiopathic in origin. We report clinical, manometric, morphofunctional, and immunological features of three cases of idiopathic ganglionitis. All patients had megacolon and underwent surgery for repeated episodes of intestinal subocclusion. Esophageal, GI, and colonic manometry performed in one patient showed dysmotility of the whole gut. Histological examination of colonic and ileum specimens identified a prominent lymphoplasmacellular infiltrate within the myenteric plexus along with a marked decrease of a wide array of neuronal peptides/transmitters. In one patient, tissue analysis revealed progressive neuronal changes up to marked myenteric neuron damage. The inflammatory infiltrate in all patients comprised CD4+ and CD8+ T lymphocytes. Abundance of both subclasses of lymphocytes suggests that immune-mediated mechanisms were responsible for neuronal degeneration. In one patient, systemic steroid therapy brought a significant clinical improvement. The immunosuppressive approach deserves further investigation in patients with severe gut motor abnormalities attributable to idiopathic myenteric ganglionitis.