Interstitial cells of Cajal are normally distributed in both ganglionated and aganglionic bowel in Hirschsprung's disease

Résumé Introduction : La chirurgie de la maladie de Hirschsprung est fréquemment compliquée d'une atteinte post-opératoire de la motilité intestinale. Des anomalies du système nerveux entérique (SNE) telles que la dysplasie neuronale intestinale de type B, l'hypoganglionose ou l'aganglionose, présents dans le segment abaissé, peuvent être la cause de certaines de ces complications mais aucune information n'est disponible quant au rôle des cellules interstitielles de Cajal (CIC) sur la motilité intestinale dans la phase post-opératoire. Ces cellules sont considérées avoir un rôle de pacemaker dans le tractus gastro-intestinal. L'objectif de cette étude était de décrire la distribution des CIC dans le segment proximal du côlon réséqué lors de cures chirurgicales de maladie de Hirschsprung et de confronter ces observations à l'évolution clinique post-opératoire. Matériel et Méthodes : L'incidence des complications post-opératoires a été déterminée par une revue rétrospective des dossiers de 48 patients opérés pour maladie de Hirschspung entre 1977 et 1999 et par l'étude histologique et immuno-histochimique des pièces réséquées chez ces patients. Nous avons comparé la distribution des CIC dans le segment proximal du côlon avec celle du côlon sain de 16 enfants contrôles par microscopie optique. L'immunohistochimie au c-Kit a été utilisée pour marquer spécifiquement les CIC sur échantillons paraffinés. Ces résultats ont ensuite été corrélés avec l'étude du SNE de ces mêmes segments, déterminée par immunohistochimie au CD56 et au protein gene product 9.5. Résultats Les complications post-opératoires suivantes furent identifiées : constipation 46%, constipation avec incontinence 15%, entérocolite 8%, décès 4% (probablement sur entérocolite). La distribution des CIC dans les segments proximaux réséqués chez les enfants avec maladie de Hirschsprung était identique à celle observée dans les segments de côlon sain, et ce indépendamment de la distribution normale ou anormale du SNE. Chez les enfants opérés pour maladie de Hirschsprung les segments réséqués présentaient les anomalies d'innervation suivantes : aganglionose 10.4%, hypoganglionose 12.5%, dysplasie neuronale intestinale de type B 6.3%, autres dysganglionoses 14.6%. Aucune relation entre ces anomalies d'innervation et les complications post-opératoires n'a été mise en évidence. Conclusion : La distribution des CIC est normale chez les patient opérés pour maladie de Hirschsprung, et ne contribue donc pas aux atteintes post-opératoires de la motilité intestinale. Cela signifie aussi que le réseau de CIC se développe noinialement dans le côlon humain, même en présence d'une innervation colique anormale ou absente. Abstract: Surgery for Hirschsprung's disease is often complicated by post-operative bowel motility disorders. The impact of intestinal neural histology on the surgical outcome has been previously studied, but no information is available concerning the influence of the distribution of interstitial cells of Cajal (ICC) on these complications. These cells are considered to be pacemakers in the gastrointestinal tract. The aim of this study was to assess the distribution of ICC in the proximal segment of resected bowel in Hirschsprung's disease and confront these results with the clinical outcome. Using immunohistochemistry for light microscopy, we compared the pattern of distribution of ICC in the proximal segment of resected bowel in Hirschsprung's disease with that in normal colon. We correlated these results with the corresponding neural intestinal histology determined by CD56 and the protein gene product 9.5 immunohistochemistry. The distribution of ICC in the proximal segment of resected bowel is identical to that of normal colon, regardless of normal or abnormal colon innervation. ICC distribution does not seem to contribute to post-operative bowel motility disorders in patients operated for Hirschsprung's disease.

Hirschsprung's disease: the "Swiss roll" technique revisited.

PURPOSE: During pull-through for Hirschsprung's disease (HSCR), the assessment of innervation is mainly based on the presence of ganglion cells when conventional Hematoxylin and Eosin (HE) staining is used. In hypoganglionosis, the evaluation is difficult. We adapted a standardized methodology for the examination of resected bowel after HSCR surgery, using the technique described by Moolenbeek on rodent intestine and later by Meier-Ruge in children. We have analysed the entire innervation of surgically resected bowels and compared the results with the follow up of patients. METHODS: Three longitudinal strips of colon were harvested from the mesenteric, anti-mesenteric and intermediate part in the whole length of resected colon of six patients with HSCR. Each strip was divided into two parts. One of the contiguous strips was assessed with HE and Hematoxylin-Phloxin-Safran, and the other one with acetylcholinesterase (AChE) histochemistry. We analyzed the distribution of ganglion cells and nerve arrangement along the strips with both techniques and compared the results obtained in the three different regions of the bowel. RESULTS: There was no significant difference in the pattern of innervation circumferentially. There was a correlation between a progressive increase of AChE activity and nerve hypertrophy and a decrease of ganglion cells from the proximal to the distal part of the resected colon in the submucosa and the myenteric plexus. Nerve hypertrophy and AChE-positive reaction in the mucosa were found at the resection border in patients who presented postoperative complications. CONCLUSIONS: Simultaneous assessment of nerve cells, nerve fibers and AChE activity is important in the evaluation of the innervation of the bowel segment proximal to the aganglionic zone. The method described is feasible and can be adapted to older children and adults with larger bowels. These results point out the importance of assessing nerve fibers in intraoperative biopsies during pull-through procedures to prevent uncomplete surgical bowel resection.

Estudi de l'argot català a la sèrie Plats bruts : l'argot a les sèries de ficció de producció pròpia de TV3

L'interès del present projecte rau en la identificació de formes pròpies de l'argot català difoses a través de les sèries de ficció de producció pròpia de la cadena de televisió autonòmica catalana. Així, en aquest treball pretenem respondre bàsicament dues qüestions: d'una banda, si les sèries de ficció de producció pròpia de TV3, i en aquest cas Plats bruts, contribueixen a difondre l'argot català; i de l'altra, si l'existència d'un argot pròpiament català (és a dir, creat a partir de recursos de la llengua catalana) és mostra de la bona salut de la llengua.

Angiotensin II is an endogenous neurotransmitter for rat and human mesenteric resistance blood vessels

Angiotensin II (Ang II) is one of the most potent vasoconstrictors. We document here the innervation of rat and human mesenteric resistance arteries (MRA) by angiotensinergic neurons of the rat and human sympathetic coeliac ganglia. Angiotensinogen (Ang-N)-mRNA and angiotensin converting enzyme-mRNA but no renin-mRNA were detected by using quantitative real time polymerase chain reaction in total RNA extracts of rat coeliac ganglia. In the same extracts, cathepsin D-mRNA was detected: This protease also cleaves Ang I from Ang-N and could therefore account for the generation of neuronal Ang peptides in the absence of renin. In situ hybridization confirmed the presence of Ang-N-mRNA in the cytoplasm of rat coeliac ganglia. By using solid-phase extraction, high performance liquid chromatography and subsequent radioimmunoassay, Ang II and its metabolites were detected in rat and also in human coeliac ganglia. Immunoreactivity for Ang II was demonstrated in rat and human coeliac ganglia neurons and their projections innervating MRA. In addition, segmental angiotensinergic innervation of MRA was also observed. By means of confocal laser scanning microscopy we were able to demonstrate the presence of angiotensinergic synapses en passant along side of vascular smooth muscle cells. Our findings could indicate that Ang II is synthesized inside the neurons of sympathetic coeliac ganglia and may act as an endogenous neurotransmitter locally in MRA.

Expression of PROKR1 and PROKR2 in Human Enteric Neural Precursor Cells and Identification of Sequence Variants Suggest a Role in HSCR

Background. The enteric nervous system (ENS) is entirely derived from neural crest and its normal development is regulated by specific molecular pathways. Failure in complete ENS formation results in aganglionic gut conditions such as Hirschsprung's disease (HSCR). Recently, PROKR1 expression has been demonstrated in mouse enteric neural crest derived cells and Prok-1 was shown to work coordinately with GDNF in the development of the ENS. Principal Findings. In the present report, ENS progenitors were isolated and characterized from the ganglionic gut from children diagnosed with and without HSCR, and the expression of prokineticin receptors was examined. Immunocytochemical analysis of neurosphere-forming cells demonstrated that both PROKR1 and PROKR2 were present in human enteric neural crest cells. In addition, we also performed a mutational analysis of PROKR1, PROKR2, PROK1 and PROK2 genes in a cohort of HSCR patients, evaluating them for the first time as susceptibility genes for the disease. Several missense variants were detected, most of them affecting highly conserved amino acid residues of the protein and located in functional domains of both receptors, which suggests a possible deleterious effect in their biological function. Conclusions. Our results suggest that not only PROKR1, but also PROKR2 might mediate a complementary signalling to the RET/GFRα1/GDNF pathway supporting proliferation/survival and differentiation of precursor cells during ENS development. These findings, together with the detection of sequence variants in PROKR1, PROK1 and PROKR2 genes associated to HSCR and, in some cases in combination with RET or GDNF mutations, provide the first evidence to consider them as susceptibility genes for HSCR.

Is the RET proto-oncogene involved in the pathogenesis of intestinal neuronal dysplasia type B?

Hirschsprung disease (HSCR) is defined by the absence of intramural ganglia of Meissner and Auerbach along variable lengths of the gastrointestinal tract. Intestinal neuronal dysplasia (IND) type B is characterized by the malformation of the parasympathetic submucous plexus of the gut. A connection appears to exist between these two enteric nervous system abnormalities. Due to the major role played by the RET proto-oncogene in HSCR, we sought to determine whether this gene was also related to INDB. dHPLC techniques were employed to screen the RET coding region in 23 patients presenting with INDB and 30 patients with a combined HSCR+INDB phenotype. In addition, eight RET single nucleotide polymorphisms (SNPs) were strategically selected and genotyped by TaqMan technology. The distribution of SNPs and haplotypes was compared among the different groups of patients (INDB, HSCR+INDB, HSCR) and the controls. We found several RET mutations in our patients and some differences in the distribution of the RET SNPs among the groups of study. Our results suggest an involvement of RET in the pathogenesis of intestinal INDB, although by different molecular mechanisms than those leading to HSCR. Further investigation is warranted to elucidate these precise mechanisms and to clarify the genetic nature of INDB.

Navegant entre narracions: recuperant silencis i subjectivitats. Recursos psicosocials en l’atenció a víctimes de violència de gènere en l’àmbit penal (jutjats i cos policial)

El treball pretén aportar un coneixement de caràcter psicosocial sobre els contextos on els professionals del sistema jurídic-penal ofereixen atenció a les víctimes de violència de gènere, concretament, les unitats i oficines habilitades per recollir la denúncia o atestat dins de les comissaries dels Mossos d’esquadra i els jutjats d´instrucció de la dona. L´àmbit territorial va ser Barcelona i Girona. Es va obtenir informació sobre judicis realitzats a l´Audiència provincial, així com documents legals en relació al tema. S’ha analitzat les necessitats i dificultats juntament amb la identificació dels recursos psicosocials que aporten tant aquests professionals com les dones que han patit o estan patit un procés de maltractament per part de la parella. La pretensió es oferir una mirada particular per suggerir transformacions que facilitin el procés d´atenció en aquests contextos, sovint difícil i, que comporta conseqüències emocionals per part de les víctimes i dels mateixos agents que les atenen. S’ha partit de les narracions de les víctimes i dels agents jurídics, com pràctiques socials a partir de les quals podem detectar valors, posicions, normes, emocions i efectes de gènere que permetrà dirigir la intervenció. Com a eines d´anàlisi s’ha fet ús els repertoris interpretatius els quals han permès analitzar les construccions socials i els seus efectes al voltant del maltracte, de les identitats de les víctimes i els victimaris.

A novel neuro-muscular marker for the heavy subunit of neurofilament proteins, NF-H, and striated muscle myosin of Xenopus laevis.

A novel monoclonal antibody, M7, is described, that reacts on Western blots with the large subunit of the neurofilament triplet proteins (NF-H) and with striated muscle myosin of Xenopus laevis. Enzymatically digested neurofilament and myosin proteins revealed different immunoreactive peptide fragments on Western blots. Therefore, the antibody must react with immunologically related epitopes common to both proteins. Immunohistochemistry showed staining of large and small axons in CNS and PNS, and nerves could be followed into endplate regions of skeletal muscles. These muscles were characterized by a striated immunostaining of the M-lines. Despite the crossreactivity of M7 with NF-H and muscle myosin, this antibody may be a tool to study innervation of muscle fibers, and to define changes in the neuromuscular organization during early development and metamorphosis of tadpoles.

Cholinergic regulation of bone.

Bone remodeling is regulated by the two branches of the autonomic nervous system: the adrenergic and the cholinergic branches. Adrenergic activity favors bone loss, whereas cholinergic activity has been recently shown to favor bone mass accrual. In vitro studies have reported that cholinergic activity induces proliferation and differentiation of bone cells. In vivo studies have shown that the inhibition of cholinergic activity favors bone loss, whereas its stimulation favors bone mass accrual. Clinical studies have shown that bone density is associated with the function of many cholinergic-regulated tissues such as the hypothalamus, salivary glands, lacrimal glands and langerhans cells, suggesting a common mechanism of control. Altogether, these observations and linked findings are of great significance since they improve our understanding of bone physiology. These discoveries have been successfully used recently to investigate new promising therapies for bone diseases based on cholinergic stimulation. Here, we review the current understanding of the cholinergic activity and its association with bone health.

Detection of neuronal activity and metabolism in a model of dehydration-induced anorexia in rats at 14.1 T using manganese-enhanced MRI and 1H MRS.

In this study, hypothalamic activation was performed by dehydration-induced anorexia (DIA) and overnight food suppression (OFS) in female rats. The assessment of the hypothalamic response to these challenges by manganese-enhanced MRI showed increased neuronal activity in the paraventricular nuclei (PVN) and lateral hypothalamus (LH), both known to be areas involved in the regulation of food intake. The effects of DIA and OFS were compared by generating T-score maps. Increased neuronal activation was detected in the PVN and LH of DIA rats relative to OFS rats. In addition, the neurochemical profile of the PVN and LH were measured by (1) H MRS at 14.1T. Significant increases in metabolite levels were measured in DIA and OFS relative to control rats. Statistically significant increases in γ-aminobutyric acid were found in DIA (p=0.0007) and OFS (p<0.001) relative to control rats. Lactate increased significantly in DIA (p=0.03), but not in OFS, rats. This work shows that manganese-enhanced MRI coupled to (1) H MRS at high field is a promising noninvasive method for the investigation of the neural pathways and mechanisms involved in the control of food intake, in the autonomic and endocrine control of energy metabolism and in the regulation of body weight.

Sympathetic glial cells and macrophages develop different responses to Trypanosoma cruzi infection or lipopolysaccharide stimulation

Nitric oxide (NO) participates in neuronal lesions in the digestive form of Chagas disease and the proximity of parasitised glial cells and neurons in damaged myenteric ganglia is a frequent finding. Glial cells have crucial roles in many neuropathological situations and are potential sources of NO. Here, we investigate peripheral glial cell response to Trypanosoma cruzi infection to clarify the role of these cells in the neuronal lesion pathogenesis of Chagas disease. We used primary glial cell cultures from superior cervical ganglion to investigate cell activation and NO production after T. cruzi infection or lipopolysaccharide (LPS) exposure in comparison to peritoneal macrophages. T. cruzi infection was greater in glial cells, despite similar levels of NO production in both cell types. Glial cells responded similarly to T. cruzi and LPS, but were less responsive to LPS than macrophages were. Our observations contribute to the understanding of Chagas disease pathogenesis, as based on the high susceptibility of autonomic glial cells to T. cruzi infection with subsequent NO production. Moreover, our findings will facilitate future research into the immune responses and activation mechanisms of peripheral glial cells, which are important for understanding the paradoxical responses of this cell type in neuronal lesions and neuroprotection.

Spinal cord stimulation treatment for angina pectoris: more than a placebo?

BACKGROUND: The effects of thoracolumbal spinal cord stimulation (SCS) are confined to restricted microcirculatory areas. This limitation is generally attributed to a predominantly segmental mode of action on the autonomic nervous system. The goal of this study was to determine whether SCS applied close to supraspinal autonomic centers would induce generalized hemodynamic changes that could explain its alleged antianginal properties. METHODS: Invasive hemodynamic tests were performed in 15 anesthetized Göttingen minipigs submitted to iterative cervical SCS of various duration and intensity. RESULTS: Hemodynamic changes exceeding 10% were observed in 59 of 68 SCS sessions (87%). Their extent and time to peak varied with SCS intensity. At 2, 5, and 10 V, significant (t test p < 0.05) peak changes occurred in cardiac output (+34%, +29%, and +28%, respectively), stroke volume (+19%, +16%, +15%), mean pressure (+9%, +27%, +40%), heart rate (+14%, +23%, +14%), systemic (-17%, NS, NS), and pulmonary vascular (25%, NS, NS) resistances. Strikingly, at 2 V, the increase in cardiac output (+34%) was higher than the synchronous rise in rate pressure product (+22%), indicating efficient cardiac work. At 10 V, however, the cardiac work was inefficient (rate pressure product + 53%/cardiac output + 28%). CONCLUSIONS: Low-voltage cervical neuromodulation reduces the postcharge and improves cardiac work efficiency. The resulting reduction in oxygen myocardial demand may account for decreased anginal pain.

Calbindin D-28k- and substance P-immunoreactive primary sensory neurons: peripheral projections in chick hindlimbs.

The peripheral projections of two distinct subpopulations of primary sensory neurons, expressing either calbindin D-28k or substance P, were studied in chick hindlimbs by immunodetecting calbindin D-28k with a rabbit antiserum and substance P with a mouse monoclonal antibody. Calbindin D-28k-immunoreactive axons provided an innervation restricted to specific mechanoreceptors such as muscle spindles, Herbst and Merkel corpuscles, or collars of feather follicles but were absent from Golgi tendon organs. In contrast, substance P-positive axons spread out diffusely in muscles and skin, formed loose plexuses, and extended free branches to the endomysium, arteries, superficial dermis, or dermal pulp of feather follicles. The present results show that calbindin D-28k- and substance P-immunoreactive primary sensory neurons provide distinct modes of innervation to selective targets in peripheral tissues. The results suggest a possible correlation between CaBP-expressing nerve endings and rapidly adapting mechanoreceptors.

A comparative study of two methods of surgical treatment for painful neuroma.

Painful neuromas may follow traumatic nerve injury. We carried out a double-blind controlled trial in which patients with a painful neuroma of the lower limb (n = 20) were randomly assigned to treatment by resection of the neuroma and translocation of the proximal nerve stump into either muscle tissue or an adjacent subcutaneous vein. Translocation into a vein led to reduced intensity of pain as assessed by visual analogue scale (5.8 (SD 2.7) vs 3.8 (SD 2.4); p < 0.01), and improved sensory, affective and evaluative dimensions of pain as assessed by the McGill pain score (33 (SD 18) vs 14 (SD 12); p < 0.01). This was associated with an increased level of activity (p < 0.01) and improved function (p < 0.01). Transposition of the nerve stump into an adjacent vein should be preferred to relocation into muscle.