Continued maturation of thymic emigrants in the periphery

Developing thymocytes are selected for recognition of molecules encoded by the major histocompatibility complex, purged of self-reactive cells and committed to either the CD4 or CD8 lineage. The 1% of thymocytes that complete these tasks emigrate and join the population of peripheral lymphocytes. Whether T cell maturation is complete at the time of thymic exit has been a subject of debate. Using mice transgenic for green fluorescent protein driven by the recombination activating gene 2 promoter to identify recent thymic emigrants, we now show that T cell differentiation continues post-thymically, with progressive maturation of both surface phenotype and immune function. In addition, the relative contribution of CD4 and CD8 recent thymic emigrants was modulated as they entered the peripheral T cell pool. Thus, T cell maturation and subset contribution are both finalized in the lymphoid periphery.     

Mutagenicity of bisphenol A (4,4'-isopropylidenediphenol) in vitro: effects of nitrosylation

Bisphenol A (4,4'-isopropylidenediphenol) is a common component of polycarbonate plastics and epoxy resins. Since bisphenol A-containing plastics and resins have found uses in food-contact items, its potential migration into foodstuffs and possible health consequences have been the focus of many recent studies. However, the potential mutagenic activation of bisphenol A by nitrosylation has received little attention. Incubation of bisphenol A with sodium nitrite under acidic conditions produced a yellow-brown product. When nitrosylated bisphenol A was tested in the Ames Salmonella/microsome assay at 100 ng to 1 mg/plate, dose-dependent increases in mutagenicity were found in both TA98 and TA100 Salmonella strains. These results indicated the presence of a direct-acting mutagenic activity causing both frameshift and base pair mutations, respectively. When compared to colony formation in untreated controls, the addition of rat liver S9 for metabolic activation had little influence on revertant colony formation. Unreacted bisphenol A dissolved in DMSO, acidic buffer, or inactivated nitrosylation solution showed negligible mutagenicity. When the nature of the mutagenic changes was examined using the Ames II trade mark Assay, a variety of base pair changes was found including T:A to A:T - S9, G:C to A:T +/- S9,C:G to A:T +/- S9 and C:G to G:C +/- S9. Bisphenol A also induced frameshift mutations at G:C sites. In addition, the presence of electrophiles was shown by the production of an intensely coloured orange-red product upon incubation of nitrosylated bisphenol A with the nucleophile 4-(4'-nitrobenzyl)pyridine. These findings suggest that migration of bisphenol A into nitrite containing foodstuffs, or its ingestion in the presence of nitrite, could lead to the formation of mutagenic compounds.     

Identification of MEN1 gene mutations in sporadic carcinoid tumors of the lung

Lung carcinoids occur sporadically and rarely in association with multiple endocrine neoplasia type 1 (MEN1). There are no well defined genetic abnormalities known to occur in these tumors. We studied 11 sporadic lung carcinoids for loss of heterozygosity (LOH) at the locus of the MEN1 gene on chromosome 11q13, and for mutations of the MEN1 gene using dideoxy fingerprinting. Additionally, a lung carcinoid from a MEN1 patient was studied. In four of 11 (36%) sporadic tumors, both copies of the MEN1 gene were inactivated. All four tumors showed the presence of a MEN1 gene mutation and loss of the other allele. Observed mutations included a 1 bp insertion, a 1 bp deletion, a 13 bp deletion and a single nucleotide substitution affecting a donor splice site. Each mutation predicts truncation or potentially complete loss of menin. The remaining seven tumors showed neither the presence of a MEN1 gene mutation nor 11q13 LOH. The tumor from the MEN1 patient showed LOH at chromosome 11q13 and a complex germline MEN1 gene mutation. The data implicate the MEN1 gene in the pathogenesis of sporadic lung carcinoids, representing the first defined genetic alteration in these tumors.      

Previous abdominal colectomy affects functional results after ileal pouch-anal anastomosis

We assessed the effect of previous abdominal colectomy on functional results after ileal J pouch-anal anastomosis (IPAA) in patients with ulcerative colitis. Twenty-five patients with colectomy prior to IPAA were compared with 22 patients who underwent noncolonic abdominal operations prior to IPAA. No differences were observed in pre- or postoperative resting anal sphincter pressure, squeeze pressure, or rectal inhibitory reflex. Previous colectomy was associated with a greater incidence of postoperative small bowel obstruction. Mean ± SEM daily stool frequency at 1 and 12 months postoperatively, respectively, was 8.9±0.8 and 5.7±0.3 for patients who had undergone previous colectomy, and 8.2±0.7 and 6.0±0.5 for the no previous colectomy group (p=not significant). At the same postoperative intervals, nocturnal stool frequency was 1.9±0.3 and 1.1±0.2 for the colectomy group and 1.5±0.3 and 0.6±0.1 for the no colectomy group (p=0.05 at 1 year). More patients in the previous colectomy group had greater than or equal to 1 nocturnal stool after 1 year (71% versus 33%,p=0.03). Although pouch capacity at 1 year was not different in the 2 groups, pouch capacity was directly related to stool frequency in the no colectomy group (r²=0.48,p=0.01), but not in the previous colectomy group (r²= 0.08,p=not significant). We conclude that previous abdominal colectomy may be associated with a higher overall incidence of small bowel obstruction. Moreover, previous colectomy is a determinant of postoperative nocturnal stool frequency after IPAA, most likely due to altered ileal pouch function. When possible, single-stage colectomy, mucosal proctectomy, and endorectal ileal pouch-anal anastomosis should be performed in patients requiring colectomy for ulcerative colitis.

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Resumen Hemos valorado el efecto de una colectomía abdominal previa sobre los resultados funcionales después de anastomosis ileoanal de bolsa en J (AIAB) en pacientes con colitis ulcerativa. Veinticinco pacientes con colectomía previa a la AIAB fueron comparados con 22 pacientes sometidos a operaciones abdominales no colónicas antes de la AIAB. No se hallaron diferencias en cuanto a la presión en reposo del esfínter anal (preoperatoria o postoperatoria), a la presión de compresión, o al reflejo rectal inhibitorio. La colectomía previa apareció asociada con una mayor incidencia de obstrucción del intestino delgado. La frecuencia de defecación diaria a 1 y a 12 meses postoperatorios, respectivamente, fue 8.9±0.8 y 5.7±0.3 para los pacientes que habían sido sometidos a colectomía previa, y 8.2 ±0.7 y 6.0±0.5 para el grupo sin colectomía previa (p=NS). En los mismos períodos postoperatorios la frecuencia de defecación nocturna fue 1.9±0.3 y 1.1±0.2 para el grupo con colectomía previa y 1.5±0.3 y 0.6 ±0.1 para el grupo sin colectomía (p=0.05 a 1 año). Más pacientes en el grupo de colectomía previa presentó más de una o una deposición nocturna después de un año (71% versus 33%, p=0.03). Aunque la capacidad de la boisa a un ano no apareció diferente en los 2 grupos, la capacidad de la bolsa apareció directamente relacionada con la frecuencia de la deposición en el grupo sin colectomía (r²=0.48,p=0.01), pero no en el grupo con colectomía previa (r²=0.08,p=NS). Nuestra conclusión es que una colectomía abdominal previa puede estar asociada con una mayor incidencia de obstrucción del intestino delgado. Además, la colectomía previa aparece como un factor determinante de la frecuencia de defecación nocturna después de AIAB, muy probablemente por alteración de la función de la bolsa ileal. En cuanto sea posible, se debe realizar la colectomía, proctectomía mucosal, y anastomosis ileal endorrectal en una sola etapa en los pacientes que requieran colectomía por colitis ulcerativa.

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Résumé Nous avons chercher à savoir si le fait d'avoir déjà effectué une colectomie retentissait sur les résultats de fonctionnement de l'anastomose iléo-anale avec réservoir en J (AIAR) chez le patient avec rectocolite hémorragique. Vingt cinq patients ayant eu une colectomie avant d'être opérés de leur AIAR ont été comparés à 22 patients ayant une intervention abdominale sans colectomie avant d'être opérés de leur AIAR. Aucune différence dans la pression sphinctérienne au repos pré ou post-opératoire, dans la pression de contraction ou dans le réflexe inhibiteur rectal n'a été observée. La colectomie préalable était associée à une incidence élevée d'occlusion intestinale post-opératoire. Le nombre de selles à 1 et à 12 mois post-opératoires était de 8.9±0.8 et 5.7±0.3, respectivement, chez le patient sans chirurgie colique antérieure (NS). Aux mêmes intervalles, la fréquence de selles nocturnes était de 1.9 ±0.3 et de 1.1±0.2 pour le groupe à colectomie préalable et de 1.5±0.3 et 0.6±0.1 pour le groupe sans chirurgie colique préalable (p=0.05 à un an). Dans le groupe à colectomie préalable, il y avait plus de patients qui avaient une ou plusieurs selles nocturnes après la première année (71% versus 33%;p= 0.03). Bien que la capacité du réservoir ne différait pas à 1 an entre les 2 groupes, la capacité était directement en rapport avec la fréquence des selles dans le groupe sans chirurgie colique préalable (r²=0.48; p=0.01), mais sans rapport dans le groupe avec chirurgie colique préalable (r²=0.08, p=NS). Nous concluons que la colectomie préalable est asscoiée à une incidence d'occlusion post-opératoire supérieure. De plus, la colectomie préalable est associée à une fréquence plus élevée de selles nocturnes après AIAR, probablement liée à un dysfonctionnement du réservoir. Lorsque la colectomie totale avec mucosectomie rectale, avec anastomose iléo-anale et réservoir est indiquée chez le patient ayant une rectocolite hémorragique, il vaut mieux la faire en un seul temps.

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Presented at the Société Internationale de Chirurgie, Toronto, Ontario, Canada, September, 1989.     

Effect of ileal oleate on interdigestive intestinal motility of the dog

To determine the effect of ileal oleate on fasting intestinal motility, pairs of duodenal and ileal catheters and bipolar duodenal and jejunal seromuscular electrodes were surgically implanted in six dogs. The ileum was perfused with either normal saline (154 mM NaCl) or oleic acid emulsion (152 mM), while intestinal myoelectric activity was continuously monitored. For transit studies, a bolus of [³H]PEG was injected into the duodenum, and jejunal and ileal alliquots were collected every 15 min for a 6-hr study period. Plasma samples were collected for radioimmunoassays of peptide YY and enteroglucagon. Ileal oleate infusion increased the MMC cycle length and decreased the number of MMCs (P<0.001) and the myoelectric spike-burst frequency/10 min in the duodenum (P<0.05). Both duodenal-jejunal (P<0.05) and duodenal-ileal transit (P<0.01) were delayed markedly by ileal perfusion with oleic acid emulsion as compared to control studies. Ileal oleate increased plasma levels of peptide YY (P<0.01) and enteroglucagon (P<0.01). Ileal perfusion with oleate therefore activated the so-called ileal brake, diminishing duodenal myoelectric spike bursts and slowing intestinal transit while concurrently increasing plasma levels of peptide YY and enteroglucagon.Dr. Dreznik is currently at Tel Aviv University, Tel Aviv, Israel, at the Sackler Faculty of Medicine.An abstract of this work was presented at the Forum of Fundamental Problems at the American College of Surgeons meeting in October 1991.Supported by the V.A. Research Advisory Group, NIH Biomedical Research Support Grant.     

Role of calcium in thromboxane B2-mediated injury to rabbit gastric mucosal cells

A sustained increase in cytosolic Ca²⁺ can damage gastric mucosal cells. The present study has examined the role of Ca²⁺ in thromboxane B₂ (TXB₂)-mediated damage of rabbit isolated gastric mucosal cells. Cells were isolated from rabbit oxyntic mucosa by collagenase-EDTA digestion. Cell metabolic activity and cell damage were estimated by alamar blue dye absorbance and trypan blue uptake, respectively. Cellular Ca²⁺ was monitored by indo-1 dye fluorescence. Addition of TXB₂ (10^–6 and 10^–8 M) to the cell suspension resulted in a decrease in metabolic activity, and this effect was reduced when Ca²⁺ was removed from the incubation Ca²⁺ and incubation of cells with the intracellular Ca²⁺ chelator, BAPTA-AM (20 M), reduced cell injury in response to TXB₂. Incubation of cells with the Ca²⁺ ionophore A23187 (1–25 M) resulted in a dose-dependent increase in trypan blue uptake and a reduction in cell metabolism. Cell unjury in response to A23187 were exacerbated by addition of TXB₂ (10^–8 M) to the cell suspension. TXB₂ treatment reduced cellular content of reduced glutathione (GSH), while exogenous GSH addition (10 mM) reduced TXB₂-mediated cell injury. These data demonstrate that TXB₂ can directly injure gastric mucosal cells. Gastric mucosal cellular damage in response to TXB₂ is mediated in part by a disruption of Ca²⁺ homeostasis as well as a reduction in cellular GSH content.This work was supported by a grant from the Medical Research Council of Canada MT6426.     

Tissue reactions to long-term electrical stimulation of the cerebellum in monkeys.

Light and electron microscopic analyses were carried out on the stimulated and unstimulated paravermal cortices of six rhesus monkeys that had electrodes implanted on their cerebella for 2 months. The electrodes and the stimulation regime (10 p.p.s.: 8 min on, 8 min off) were similar to those used to stimulate the human cerebellum for treatment of certain neurological disorders. Mere presence of the electrode array in the posterior fossa for 2 months resulted in some meningeal thickening, attenuation of the molecular layer, and loss of Purkinje cells immediately beneath the electrode array. There was no evidence of scarring. After 205 hours of stimulation (7.35 X 10(6) pulses) over 18 days, a charge of 0.5 muC/ph or estimated charge density of 7.4 muC/sq cm/ph resulted in no damage to the cerebellum attributable to electrical stimulation per se. Such a charge/phase is about five times the threshold for evocation of cerebellar efferent activity, and might be considered "safe" for stimulation of human cerebellum. Charge density/phase and charge/phase were directly related to increased cerebellar injury in the six other cerebellar cortices stimulated. Leptomeningeal thickening increased with increased charge density. Injury included severe molecular layer attenuation, ongoing destruction of Purkinje cells, gliosis, ongoing degeneration of myelinated axons, collagen intrusion, and increased levels of local polysaccharides. In all cases, even with damage that destroyed all conducting elements beneath the electrodes, there was no damage further than 1 to 2 mm from the edges of the electrode arrays.