Gut microbiome composition and metabolic activity in women with diverticulitis.

The etiopathogenesis of diverticulitis, among the most common gastrointestinal diagnoses, remains largely unknown. By leveraging stool collected within a large prospective cohort, we performed shotgun metagenomic sequencing and untargeted metabolomics profiling among 121 women diagnosed with diverticulitis requiring antibiotics or hospitalizations (cases), matched to 121 women without diverticulitis (controls) according to age and race. Overall microbial community structure and metabolomic profiles differed in diverticulitis cases compared to controls, including enrichment of pro-inflammatory Ruminococcus gnavus, 1,7-dimethyluric acid, and histidine-related metabolites, and depletion of butyrate-producing bacteria and anti-inflammatory ceramides. Through integrated multi-omic analysis, we detected covarying microbial and metabolic features, such as Bilophila wadsworthia and bile acids, specific to diverticulitis. Additionally, we observed that microbial composition modulated the protective association between a prudent fiber-rich diet and diverticulitis. Our findings offer insights into the perturbations in inflammation-related microbial and metabolic signatures associated with diverticulitis, supporting the potential of microbial-based diagnostics and therapeutic targets.

Protective effects of amoxicillin and probiotics on colon disorders in an experimental model of acute diverticulitis disease.

Acute diverticulitis disease is associated with inflammation and infection in the colon diverticula and may lead to severe morbidity. This study aimed to evaluate and compare the protective effects of amoxicillin antibiotic, either alone or in combination with probiotics (Lactobacillus acidophilus and Bifidobacterium lactis), in a rat model of acute diverticulitis disease. Acute diverticulitis was induced, in albino rats, by adding 3% weight/volume of dextran sulfate sodium (DSS) to the rats' drinking water; daily for 7 days, in addition to injecting lipopolysaccharide (LPS) enema (4 mg/kg). The impact of treatments was assessed by measuring the physiological and immunological parameters and evaluating colon macroscopic and microscopic lesions. The results showed that both treatments (especially probiotics with amoxicillin) alleviated the adverse effects of DSS and LPS. This was obvious through the modulation of the rats' body weight and the colon weight-to-length ratio. Also, there was a significant (p < 0.001) decrease in the colon macroscopic lesion score. The pro-inflammatory cytokines [(TNF)-α, (IL)-1ß, (IFN)-γ, and (IL)-18]; in the colon tissue; were significantly (p < 0.001) decreased. Also, both treatments significantly ameliorated the elevation of myeloperoxidase activity and C-reactive protein levels, in addition to improving the histopathological alterations in the colon tissue. In conclusion, amoxicillin and probiotics-amoxicillin were effective in preventing the development of experimentally induced acute diverticulitis, through their anti-inflammatory and immunomodulatory effects. Furthermore, this study has explored the role of probiotics in preventing DSS/LPS-induced acute diverticulitis, so it can be applied as a promising treatment option for acute diverticulitis disease.

COLQ and ARHGAP15 are Associated with Diverticular Disease and are Expressed in the Colon.

BACKGROUND: Diverticular disease is a common but poorly understood disease of the gastrointestinal tract. Recent studies have identified several single nucleotide polymorphisms (SNPs) that are associated with diverticular disease. MATERIALS AND METHODS: The genotypes of three SNPs (rs4662344 in ARHGAP15, rs7609897 in COLQ, and rs67153654 in FAM155A) were identified by Taqman assay in 204 patients with diverticular disease. Clinical characteristics were obtained from the medical record to study association with genotype. To evaluate gene expression in colon tissue, qPCR was performed on 24 patients with diverticulitis, and COLQ was localized using immunohistochemistry. RESULTS: The ARHGAP15 and COLQ SNPs were significantly associated with both diverticular disease and specifically diverticulitis, while the FAM155A was not associated with either. No association was found with clinical disease characteristics. Heterozygous genotypes at the ARHGAP15 SNP was associated with lower ARHGAP15 expression in colon tissues. COLQ protein localized to the myenteric plexus in the colon. CONCLUSIONS: This study confirmed association of the ARHGAP15 and COLQ SNPs with diverticular disease in our patients but could not confirm FAM155A SNP association. Neither of these SNPs appeared to associate with more severe disease, but genotype at the ARHGAP15 SNP did impact expression of ARHGAP15 in the colon. Additionally, this study is the first to localize COLQ in the colon. Its presence in the myenteric nervous system suggests COLQ SNP variants may contribute to diverticular disease by altering motility.

Identification of a rare LAMB4 variant associated with familial diverticulitis through exome sequencing.

Diverticulitis is a chronic disease of the colon in which diverticuli, or outpouching through the colonic wall, become inflamed. Although recent observations suggest that genetic factors may play a significant role in diverticulitis, few genes have yet been implicated in disease pathogenesis and familial cases are uncommon. Here, we report results of whole exome sequencing performed on members from a single multi-generational family with early onset diverticulitis in order to identify a genetic component of the disease. We identified a rare single nucleotide variant in the laminin ß 4 gene (LAMB4) that segregated with disease in a dominant pattern and causes a damaging missense substitution (D435N). Targeted sequencing of LAMB4 in 148 non-familial and unrelated sporadic diverticulitis patients identified two additional rare variants in the gene. Immunohistochemistry indicated that LAMB4 localizes to the myenteric plexus of colonic tissue and patients harboring LAMB4 variants exhibited reduced LAMB4 protein levels relative to controls. Laminins are constituents of the extracellular matrix and play a major role in regulating the development and function of the enteric nervous system. Reduced LAMB4 levels may therefore alter innervation and morphology of the enteric nervous system, which may contribute to colonic dysmotility associated with diverticulitis.

Tratamiento quirúrgico de la diverticulitis aguda. Estudio retrospectivo multicéntrico / Surgical treatment of acute diverticulitis. A retrospective multicentre study

INTRODUCCIÓN: Se pretende analizar los resultados a corto y medio plazo de diferentes técnicas quirúrgicas en el tratamiento de la diverticulitis aguda complicada (DAC). MÉTODOS: Estudio retrospectivo y multicéntrico de pacientes operados de urgencia o de urgencia diferida por DAC. RESULTADOS: Estudiamos a 385 pacientes: 218 hombres y 167 mujeres, de edad media 64,4 ± 15,6 años, intervenidos en 10 hospitales. La mediana (25-758 percentiles) de evolución desde el inicio de los síntomas hasta la cirugía fue de 48 h(24-72), y su indicación más frecuente, un cuadro peritonítico (66%). El abordaje fue generalmente abierto (95,1%) y los hallazgos más comunes, peritonitis purulenta (34,8%) o absceso pericólico (28,6%). La técnica más habitual fue el procedimiento de Hartmann (PHT) en 278 (72,2%), seguida de resección y anastomosis primaria (RAP) en 69 (17,9%). Se complicaron 205 pacientes (53,2%) y fallecieron 50 (13%). Edad avanzada, inmunodepresión, factores de riesgo quirúrgico y peritonitis fecal se asociaron a mayor mortalidad. El lavado peritoneal laparoscópico (LPL) tuvo elevada tasa de reintervenciones, implicando frecuentemente un estoma, y la RAP se complicó con dehiscencia de sutura en el 13,7% de pacientes, sin diferencias en la morbimortalidad al compararla con el PHT. La mediana de estancia postoperatoria fue de 12 días; su mayor duración se relacionó con la mayor edad, riesgo quirúrgico ASA, hospital y complicaciones postoperatorias. CONCLUSIONES: La cirugía por DAC tiene importante morbimortalidad y se asocia frecuentemente a un estoma terminal. Además, el LPL presenta alta tasa de reintervenciones. LA RAP, aun asociando un estoma de protección, parece de elección en muchos casos

INTRODUCTION: To analyze short and medium-term results of different surgical techniques in the treatment of complicated acute diverticulitis (CAD). METHODS: Multicentre retrospective study including patients operated on as surgical emergency or deferred-urgency with the diagnosis of CAD. RESULTS: A series of 385 patients: 218 men and 167 women, mean age 64.4 ± 15.6 years, operated on in 10 hospitals were included. The median (25th-75th percentile) time from symptoms to surgery was 48 (24-72) h, being peritonitis the main surgical indication in a 66% of cases. Surgical approach was usually open (95.1%), and the commonest findings, a purulent peritonitis (34.8%) or pericolonic abscess (28.6%). Hartmann procedure (HP) was the most used technique in 278 (72.2%) patients, followed by resection and primary anastomosis (RPA) in 69 (17.9%). The overall postoperative morbidity and mortality was 53.2% and 13% respectively. Age, immunosupression, presence of general risk factors and faecal peritonitis were associated with increased mortality. Laparoscopic peritoneal lavage (LPL) was associated with an increased reoperation rate frequently involving a stoma, and anastomotic leaks presented in 13.7 patients after RPA, without differences in morbimortality when compared with HP. Median postoperative length of stay was 12 days, and was correlated with age, surgical risk, ASA score, hospital and postoperative complications. CONCLUSIONS: Surgery for CAD has important morbidity and mortality and is frequently associated with an end-stoma. Moreover LPL presented high reoperation rates. It seems better to resect and anastomose in most cases, even with an associated protective stoma

Human resident CD34+ stromal cells/telocytes have progenitor capacity and are a source of αSMA+ cells during repair.

We studied the progenitor capacity of human resident CD34+ stromal cells/telocytes (SC/TCs) in the enteric wall affected by inflammatory/repair processes (appendicitis, diverticulitis of large bowel and Crohn's disease of the terminal ileum) at different stages of evolution (inflammatory, proliferative and remodelling). In these conditions, CD34+ SC/TCs are activated, showing changes, which include the following overlapping events: 1) separation from adjacent structures (e.g., from vascular walls) and location in oedematous spaces, 2) morphological modifications (in cell shape and size) with presence of transitional cell forms between quiescent and activated CD34+ SC/TCs, 3) rapid proliferation and 4) loss of CD34 expression and gain of αSMA expression. These events mainly occur in the inflammatory and proliferative stages. During the loss of CD34 expression, the following findings are observed: a) irregular cell labelling intensity for anti-CD34, b) co-localization of CD34 and actin, c) concurrent irregular labelling intensity for αSMA and d) αSMA expression in all stromal cells, with total loss of CD34 expression. While CD34 expression was conserved, a high proliferative capacity (Ki-67 expression) was observed and vice versa. In the segments of the ileum affected by Crohn's disease, the stromal cells around fissures were αSMA+ and, in the transitional zones with normal enteric wall, activated CD34+ SC/TCs were observed. In conclusion, human resident CD34+ SC/TCs in the enteric wall have progenitor capacity and are activated with or without differentiation into αSMA+ stromal cells during inflammatory/repair processes.

Comparison of two FFPE preparation methods using label-free shotgun proteomics: Application to tissues of diverticulitis patients.

Formalin-fixed paraffin-embedded (FFPE) specimens of patients are useful sources of materials for clinical research and have recently gained interest for use in the discovery of clinical proteomic biomarkers. However, the critical step in this field is the ability to obtain an efficient and repeatable extraction using the limited quantities of material available for research in hospital biobanks. This work describes the evaluation of the peptide/protein extraction using FFPE sections treated by the following two methods before shotgun proteomic analysis: a commercial solution (FFPE-FASP) (filter aided sample preparation) and an antigen retrieval-derived protocol (On Slice AR). Their efficiencies and repeatabilities are compared using data-independent differential quantitative label-free analysis. FFPE-FASP was shown to be globally better both qualitatively and quantitatively than On Slice AR. FFPE-FASP was tested on several samples, and differential analysis was used to compare the tissues of diverticulitis patients (healthy and inflammatory tissues). In this differential proteomic analysis using retrospective clinical FFPE material, FFPE-FASP was reproducible and provided a high number of confident protein identifications, highlighting potential protein biomarkers. BIOLOGICAL SIGNIFICANCE: In clinical proteomics, FFPE is an important resource for retrospective analysis and for the discovery of biomarkers. The challenge for FFPE shotgun proteomic analysis is preparation by an efficient and reproducible protocol, which includes protein extraction and digestion. In this study, we analyzed two different methods and evaluated their repeatabilities and efficiencies. We illustrated the reproducibility of the most efficient method, FFPE-FASP, by a pilot study on diverticulitis tissue and on FFPE samples amount accessible in hospital biobanks. These data showed that FFPE is suitable for use in clinical proteomics, especially when the FFPE-FASP method is combined with label-free shotgun proteomics as described in the workflow presented in this work.

[Significance of CRP in the assessment of severity of acute diverticulitis]. / CRP:n merkitys akuutin divertikuliitin vakavuuden arvioinnissa.

Diverticulitis is suspected when the patient has pain and tenderness on palpation on the lower left abdomen, fever and an elevated CRP level. After a confirmed diagnosis, outpatient therapy of uncomplicated diverticulitis is possible, according to the latest research results even without antimicrobial drugs. CT scanning is usually unnecessary when the CRP level remains below 50 mg/l in the absence of signs of peritoneal irritation, but should always be performed when the value exceeds 150 mg/l. Unenhanced CT scan without contrast agent is well suited for the exclusion of complicated diverticulitis.

Regulated expression of leukocyte-specific transcript (LST) 1 in human intestinal inflammation.

INTRODUCTION: Leukocyte-specific transcript 1 (LST1) encoded peptides are involved in immunomodulation and nanotube-mediated cell-cell communication. The aim of this study was to assess the expression of LST1 in colonic epithelium and endothelium during intestinal inflammation. METHODS: LST1 expression was evaluated by RT-PCR, FACS, western blot analysis, and immunohistochemistry in intestinal epithelial Caco-2 cells, human intestinal microvascular endothelial cells and in human histological specimens from inflammatory bowel disease (IBD) patients and non-IBD colitis patients. RESULTS: LST1 expression was significantly increased upon proinflammatory stimulation in intestinal epithelial and endothelial cells. Furthermore, LST1 tissue expression was significantly enhanced in macroscopically inflamed colonic mucosal biopsies as compared to non-affected mucosal areas. CONCLUSIONS: This is the first report demonstrating regulated LST1 expression in human intestinal epithelial and microvascular endothelial cells and in inflamed colonic tissue from IBD patients. Proinflammatory expression of LST1 occurs in the setting of human IBD and is not restricted to immune cell populations. Future studies are needed to further elucidate the role of soluble and membrane-expressed LST1 in the regulation of mucosal intestinal immunity and inflammation as well as to reveal possible therapeutic implications.

Comparative analysis of the expression of metalloproteases and their inhibitors in resected crohn's disease and complicated diverticular disease.

BACKGROUND: Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs), are expressed in the gastrointestinal tract by different cellular types. Nevertheless, the imbalance between MMPs and TIMPs plays an important role in the physiopathology of diverse intestinal inflammatory processes. METHODS: An immunohistochemical study was performed using tissue arrays and specific antibodies against MMPs -1, -2, -7, -9, -11, -13, -14, and TIMPs -1, -2 and -3. Immunohistochemical staining of intestinal samples from surgical interventions from 30 patients with complicated Crohn's disease (CD) and 25 patients with diverticulitis were performed at the inflamed mucosa and in adjacent noninflamed mucosa. A reverse-transcription polymerase chain reaction (RT-PCR) analysis was performed to confirm the results obtained by immunohistochemistry. In addition, western blot experiments were carried out. RESULTS: CD inflamed mucosa showed higher global expression of MMP-2, MMP-9, and MMP-13 than diverticulitis inflamed mucosa. However, inflamed and noninflamed diverticulitis mucosal samples showed higher global expression of MMP-1, TIMP-1, and 3 than the CD samples. Epithelial cells of inflamed mucosa showed higher expression of MMP-2, 9, and 13 in CD than diverticulitis. However, the latter showed higher expression of TIMP-1. Similar differences for fibroblast-like cells and mononuclear inflammatory cells were found. CD samples presented an increased expression of MMPs and a decreased expression of TIMPs compared to diverticulitis. CONCLUSIONS: These results indicate a differential pattern of expression of MMPs and TIMPs in CD and diverticulitis and the necessity to study the potential role of MMP inhibitors as new protective agents in both diseases.

Recurrencia del tumor ovárico de las células de la granulosa / Recurrence of ovarian granulosa cell tumor

Presentamos el caso de una mujer de 79 años, con antecedentes de un supuesto fibroma ovárico, que comienza con un cuadro de dolor en fosa ilíaca izquierda y leucocitosis, siendo diagnosticada de diverticulitis. Durante el estudio, incidentalmente se descubre una masa en la pared abdominal anterior que resultó ser una recidiva de un tumor de células de la granulosa. Realizamos una revisión bibliográfica de esta patología centrándonos, fundamentalmente, en sus aspectos clínicos y terapéuticos. Se trata de una neoplasia muy poco frecuente, sobre la que hay pocos estudios, que se caracteriza por su larga historia natural y su tendencia a recurrir años después del diagnóstico (AU)

We present the case of a 79-year-old woman who was misdiagnosed with an ovarian fibroma 5 years previously. The patient presented with pain in the right iliac fossa with leukocytosis and was diagnosed with diverticulitis. A mass was incidentally found in the anterior abdominal wall, which was later confirmed as metastasis from a granulosa cell tumor of the ovary. We provide a literature review, focussing on the clinical findings and treatment of this tumor. Few reports have been published of this uncommon entity, characterized by a long natural history and tendency to recur years after the initial diagnosis (AU)

Abnormal expression of M1/MUC5AC mucin in distal colon of patients with diverticulitis, ulcerative colitis and cancer.

The abnormal expression of gastric M1/MUC5AC mucin in precancerous lesions and colon cancer evidenced by immunohistochemistry led us to check for its presence in the mucus obtained directly from patients undergoing surgery for cancerous (adenocarcinoma) or inflammatory (diverticulitis or ulcerative colitis) diseases. In parallel, the authors quantified aberrant crypt foci (ACF) and their immunolabelling by M1/MUC5AC in mucosae of cancer and diverticulitis patients. Immuno-Radio-Metric Assay of M1/MUC5AC mucin developed by the authors was used to detect M1/MUC5AC mucin in the colonic mucus scraped from surgical specimens. M1/MUC5AC mucin was detected in the mucus of 51/69 (74%) patients with colon adenocarcinoma, versus 7/27 (26%) patients with diverticulitis (threshold: 30 units of M1 mucin per mg protein, area under ROC curve: 0.80). M1/MUC5AC was present in significantly (p < 0.001) larger amounts in the mucus of cancer versus diverticulitis patients. All (10/10) patients with ulcerative colitis tested showed levels above the threshold and their mucosae were strongly labelled with the anti-M1/MUC5AC antibody by immunohistochemistry. Patients with cancer exhibited 3 fold more ACF than those with diverticulitis, but no significant difference was observed in the mean size and M1/MUC5AC expression pattern of ACF between these two groups. The expression of M1/MUC5AC was in correlation with their size. In macroscopically normal mucosa, ACF were the most important source of M1/MUC5AC mucin. Testing of M1/MUC5AC can enhance the detection of precancerous lesions and colon cancer.

Expression of voltage-gated potassium channels in human and mouse colonic carcinoma.

PURPOSE: Voltage-gated Kv potassium channels, like ether a go-go (EAG) channels, have been recognized for their oncogenic potential in breast cancer and other malignant tumors. EXPERIMENTAL DESIGN: We examined the molecular and functional expression of Kv channels in human colonic cancers and colon of mice treated with the chemical carcinogens dimethylhydrazine and N-methyl-N-nitrosourea. The data were compared with results from control mice and animals with chemically induced DSS colitis. RESULTS: Electrogenic salt transport by amiloride-sensitive Na+ channels and cyclic AMP-activated cystic fibrosis transmembrane conductance regulator Cl- channels were attenuated during tumor development and colitis, whereas Ca2+-dependent transport remained unchanged. Kv channels, in particular Eag-1, were enhanced during carcinogenesis. Multiplex reverse transcription-PCR showed increased mRNA expression for Kv1.3, Kv1.5, Kv3.1, and members of the Eag channel family, after dimethylhydrazine and N-methyl-N-nitrosourea treatment. Eag-1 protein was detected in the malignant mouse colon and human colonic cancers. Genomic amplification of Eag-1 was found in 3.4% of all human colorectal adenocarcinoma and was an independent marker of adverse prognosis. CONCLUSIONS: The study predicts an oncogenic role of Kv and Eag channels for the development of colonic cancer. These channels may represent an important target for a novel pharmacotherapy of colonic cancer.

Secretion of RANTES (CCL5) and interleukin-10 from mesenteric adipose tissue and from creeping fat in Crohn's disease: regulation by steroid treatment.

BACKGROUND: Creeping fat represents a characteristic feature of Crohn's disease (CD) and adipose tissue is currently being recognized as a complex compartment secreting highly active molecules. Pro- or anti-inflammatory adipose tissue-derived secretory products (adipocytokines) might play a role in the pathogenesis of CD. METHODS: Adipose tissue specimens were obtained from creeping fat contiguous to the involved intestine of 10 patients with CD. Mesenteric adipose tissue specimens resected in 13 patients with colon cancer (CC) and in seven patients with diverticulitis (DIV) served as controls. Three fat tissue specimens per well and n = 6-8 wells per patient were incubated ex vivo for 24 h. The release of regulated on activation, T-cell expressed and secreted (RANTES) and interleukin (IL)-10 into the supernatant was measured by ELISA. RESULTS: Both RANTES and IL-10 secretion could be demonstrated from total adipose tissue explants. The RANTES secretion is increased from creeping fat in CD (3691 +/- 597 pg/g fat per 24 h) when compared to mesenteric adipose tissue from patients with CC (1690 +/- 191 pg/g fat per 24 h; P < 0.0001) or DIV (1672 +/- 336 pg/g fat per 24 h; P < 0.0001). In contrast, IL-10 secretion is downregulated significantly only in patients with DIV (1418 +/- 180 pg/g fat per 24 h; P = 0.016) when compared to CC patients (2368 +/- 259 pg/g fat per 24 h). Crohn's disease patients receiving steroids had a higher secretion rate of RANTES and IL-10. CONCLUSIONS: Both RANTES and IL-10 secretion can be detected from mesenteric adipose tissue and from creeping fat. The elevated RANTES and IL-10 secretion from creeping fat in CD is not due to a CD-specific effect but caused by steroid treatment.

Vascular endothelial growth factor secretion from mesenteric adipose tissue and from creeping fat in Crohn's disease.

BACKGROUND: Creeping fat represents a characteristic feature of Crohn's disease (CD), and adipose tissue secretes adipocytokines and chemokines/growth factors such as vascular endothelial growth factor (VEGF). Because VEGF serum levels and mucosal VEGF expression is elevated in CD patients, the aim of the present paper was to investigate creeping fat-derived VEGF secretion in CD. MATERIAL AND METHODS: Adipose tissue was obtained from creeping fat of 10 patients with CD. Mesenteric adipose tissue was resected from 13 patients with colon cancer (CC) and from seven patients with diverticulitis (DIV). Three fat tissue specimens per well, and several wells (6-8) per patient were incubated ex vivo for 24 h. The release of VEGF into the supernatant was measured by ELISA. RESULTS: There was stable VEGF secretion from mesenteric adipose tissue of patients with CC or DIV and from creeping fat of patients with CD. Whereas the VEGF secretion rate was not different between patients with CD (465 +/- 98 pg/g fat per 24 h) and CC (399 +/- 48 pg/g fat per 24 h), VEGF secretion was significantly reduced in patients suffering from DIV (115 +/- 41 pg/g fat per 24 h; P < 0.0001 and P = 0.001, respectively). The CD patients treated with steroids had significantly lower VEGF secretion rates (294 +/- 42 pg/g fat per 24 h) than CD patients not receiving steroids (607 +/- 105 pg/g fat per 24 h; P = 0.001). CONCLUSIONS: Creeping fat is an important source of VEGF secretion. The characteristics of the inflammatory changes in CD might be due to the lack of VEGF downregulation that is seen in DIV.

Strictures in Crohn's disease are characterised by an accumulation of mast cells colocalised with laminin but not with fibronectin or vitronectin.

BACKGROUND/AIMS: Intestinal fibrosis and stricture formation is an unresolved problem in Crohn's disease. The aim of this study was to investigate whether mast cells accumulate in these tissues and whether their localisation is associated with extracellular matrix components. METHODS: Mast cells were visualised by immunohistochemical staining of the mast cell specific proteases chymase and tryptase. Their localisation in relation to extracellular matrix components was shown by immunohistochemical double labelling. RESULTS: In strictures in Crohn's disease, a striking accumulation of mast cells was seen particularly in the hypertrophied and fibrotic muscularis propria, with a mean (SEM) mast cell number of 81.3 (14.9) v 1.5 (0.9)/mm(2) in normal bowel (p<0.0005). All mast cells in the muscularis propria were colocalised with patches of laminin. In contrast, in the submucosa, laminin was exclusively found in the basal lamina of blood vessels where many adherent mast cells were seen. No colocalisation of mast cells was found with fibronectin or vitronectin. CONCLUSIONS: The large accumulation of mast cells in the muscle layer of strictured bowel suggests a functional role for these cells in the hypertrophic and fibrotic response of the smooth muscle cells. The colocalisation with laminin indicates a mechanism of interaction between smooth muscle cells and mast cells that may be important in the role of mast cells in the process of fibrosis.

Evidence of T cell receptor beta-chain patterns in inflammatory and noninflammatory bowel disease states.

T cell activation, as defined by expression of relevant cell surface molecules, such as the interleukin-2 receptor (CD25), is increased in many chronic relapsing diseases, including inflammatory bowel disease (IBD). These T cells are generally activated through contact of their clonotypic T cell receptor (TCR) with a peptide antigen presented by a major histocompatibility complex molecule. One of the putative antigenic contact sites for the TCR is the third complementarity determining region (CDR3) of the TCR beta-chain variable region (TCRBV). Therefore, analysis of the TCRBV CDR3 provides insight into the diversity of antigens encountered by a given T cell population. This study evaluated the TCRBV CDR3 usage of the activated intestinal lymphocytes from human subjects with IBD, diverticulitis (inflammatory control), and a normal tissue control. Public patterns, as demonstrated by shared TCRBV CDR3 amino acid sequences of activated intestinal T cell subpopulations, were observed. In particular, a public pattern of TCRBV22, a conserved valine in the fifth position, and use of TCRBJ2S1 or TCRBJ2S5 was present in three of four Crohn's disease subjects while not present in the ulcerative colitis subjects. However, the private patterns of TCRBV CDR3 region amino acid sequences were far more striking and easily demonstrated in all individuals studied, including a normal noninflammatory control. Thus we conclude that selective antigenic pressures are prevalent among an individual's activated intestinal lymphocytes.

Butyrate increases colonocyte protein synthesis in ulcerative colitis.

Butyrate promotes epithelial cell healing and improves symptoms when administered rectally in patients with distal ulcerative colitis (UC). It was hypothesized that butyrate may enhance healing in patients with UC by stimulating colonocyte proliferation and/or protein production. Mucosa from the descending colon was obtained from patients with UC (n = 5), Crohn's disease (n = 8), diverticulitis (n = 6), and cancer (normal tissue 10 cm from tumor; n = 10). Epithelial cells were isolated using dispase/collagenase and differential sedimentation and incubated for 4 hr at 37 degrees C with either Na butyrate (10 mM) or NaCl (10 mM). Protein synthesis was assessed by [14C]leucine incorporation and proliferation was determined with [3H]thymidine. Mean viability and purity were >88%. Spontaneous proliferation was significantly increased in UC when compared to diverticulitis and normal controls. Butyrate significantly increased protein synthesis in UC epithelial cells when compared to saline control. The therapeutic effects of butyrate in patients with UC may be due to its use by epithelial cells as a metabolic fuel to increase protein production and promote healing.

Production of vitamin B 12 analogues in patients with small-bowel bacterial overgrowth.

We investigated the presence of vitamin B 12 analogues (cobamides) and the bacterial conversion of 57Co-B12 (vitamin B12 cyanocobalamin, [57Co]-CN-Cbl) into cobamides in the intestinal contents of four patients with bacterial overgrowth. The (57Co)-CN-Cbl bound to intrinsic factor was given orally. Jejunal contents were aspirated for 24 h and cultured aerobically and anaerobically. The CN-Cbl and cobamides were separated by electrophoresis and chromatography and identified by bioautography. Radioactivity of cobamide zones from duplicate chromatograms showed bacterial conversion of (57Co)-cn-cbl into cobamides. Cobamides ([Ade]CNCBA, [2-Me Ade] CNCba, [CN]2Cbi and factor E) were found in the intestinal contents in three of the four patients, and in two of three patients cobamides represented more than 25% of the administered CN-Cbl. Thus bacterial production of cobamides, both de novo and from ingested CN-Cbl bound to intrinsic factor, occurs in humans with bacterial overgrowth states and results in a significant loss of vitamin B12 to the host.