Influence of inflammatory bowel disease on intestinal microflora.

The microflora of the jejunum, ileum, and colon has been studied from operative samples in Crohn's disease (n = 30), ulcerative colitis (n = 15), and controls (n = 40). There was no significant difference in the flora of patients with ulcerative colitis compared with controls. In Crohn's disease there was a significant increase in E. coli (P less than 0.001) and B. fragilis (P less than 0.001) in the ileum and of E. coli (P less than 0.001) and lactobacilli (P less than 0.01) in the colon. The abnormal ileal flora in Crohn's disease was unrelated to serological evidence of disease activity (indices: ESR, serum albumin, serum seromucoids), diameter of the ileum, or excision of the ileocaecal valve. The abnormal colonic flora in Crohn's disease was not related to presence of macroscopic colitis.     

Histochemical demonstration of desialation and desulphation of normal and inflammatory bowel disease rectal mucus by faecal extracts.

Experiments were carried out to assess the susceptibility of normal and inflammatory bowel disease rectal mucus to desulphation and desialation by faecal extracts and by bacterial sialidase. The effects were assessed histochemically using a combined high iron diamine (HID) and alcian blue (AB) stain for sulphomucins and sialomucins. Rectal mucus in biopsies from controls (irritable bowel syndrome) and patients with ulcerative colitis or Crohn's disease was resistant to desialation by Clostridium perfringens sialidase, but susceptible to desialation and desulphation by bacteria-free extracts of normal faeces. Periodic acid-Schiff (PAS) staining of adjacent sections similarly treated showed retention of neutral mucus. One faecal extract selectively desulphated all 42 biopsies, causing the goblet cells to change from HID positive to AB positive, suggesting that most, or all HID positive cells also contain sialomucins. This alters the interpretation of previous histochemical studies. Faecal extracts from patients with active ulcerative colitis (n = 6) had desialating and desulphating effects similar to faecal extracts from normal subjects (n = 6). Ulcerative colitis (n = 21), Crohn's disease (n = 18), and control (irritable bowel syndrome) (n = 17) rectal biopsies all showed similar susceptibility to desulphation by a pooled normal faecal extract, but rectal biopsies from patients with Crohn's disease proved more resistant to desialation than control or ulcerative colitis biopsies (p less than 0.02). These studies imply that colonic mucus undergoes continual desulphation and desialation in vivo as a result of faecal enzyme activity that is probably mainly of bacterial origin. Altered susceptibility of colonic mucus to this may be important in the pathogenesis of colonic disease.     

Thiol methyltransferase activity in inflammatory bowel disease

BACKGROUND: Luminal anionic sulphide may contribute to epithelial damage in ulcerative colitis. Thiol methyltransferase (TMT) governs sulphide detoxification by the colonic mucosa and circulating erythrocytes. AIMS: To measure levels of TMT activity in erythrocytes of surgically treated cases of colitis or in rectal biopsies of defined groups of colitis. PATIENTS: Venepuncture blood was obtained from 37 blood donors and 27 subjects who had previously undergone a proctocolectomy for colitis: 18 for ulcerative colitis and nine for Crohn's colitis. Rectal biopsies from 122 cases were obtained: 47 without mucosal disease, 33 post-colon resection for cancer, 14 with moderate to severe ulcerative colitis, 15 with quiescent ulcerative colitis, seven with acute Crohn's colitis, and six with radiation proctitis. METHODS: TMT activity was measured by high performance liquid chromatography with radioactive detection to measure (14)C methylmercaptoethanol formation, the reaction product of cell extracts incubated with mercaptoethanol and (14)C S-adenosylmethionine. RESULTS: Erythrocyte TMT activity of surgically treated cases of colitis was significantly elevated (p<0. 001) compared with control cases. TMT activity of rectal biopsies was significantly decreased (p<0.02) in acute but not quiescent ulcerative colitis, Crohn's colitis, or radiation colitis. CONCLUSIONS: Erythrocyte TMT activity was persistently elevated after proctocolectomy for Crohn's disease and ulcerative colitis. No primary defect of TMT activity was found in any case of unoperated colitis but mucosal activity was diminished with disease progression of ulcerative colitis. Studies of genetic control of TMT activity of erythrocytes in inflammatory bowel disease appear worthwhile.     

Fusobacterium varium localized in the colonic mucosa of patients with ulcerative colitis stimulates species-specific antibody

BACKGROUND: Microbial agents are a possible cause of ulcerative colitis. We have previously reported evidence of bacteria invading the colonic mucosa of patients with ulcerative colitis. We have isolated bacteria from inflamed colonic mucosa, examined the localization of the species in the mucosa, and assayed for serum antibodies to the bacteria. METHODS: Cohorts of 31 per group were enrolled from patients with active ulcerative colitis, Crohn's disease, ischemic colitis, and colon adenomas. A group of 31 healthy controls were also studied. The presence of bacteria in biopsies of patients with ulcerative colitis was analyzed by both isolation and immunohistochemistry. Sera from patients were tested for bacterial antibodies using both Western blots and enzyme-linked immunosorbent assay (ELISA). RESULTS: Only sera from patients with ulcerative colitis gave specific reactions with Fusobacterium varium in Western blot assays. The detection rate of specific bands was higher for patients with ulcerative colitis (61%) than for subjects with either Crohn's disease (13%) or healthy controls (29%) (P < 0.001 and P = 0.021, respectively). The ELISA showed that the mean optical densities with extracts of F. varium as antigen were significantly higher for ulcerative colitis patients than for subjects with either Crohn's disease or healthy controls (P < 0.001). Immunohistochemical detection of F. varium in colonic mucosa was significantly higher in patients with ulcerative colitis (84%) than for subjects with either Crohn's disease (16%) or other controls (3-13%) (P < 0.001). CONCLUSIONS: Fusobacterium varium bacteria were present in a significant number of patients with active ulcerative colitis, and should be tested in therapeutic trials in order to confirm the causal relationship between F. varium and ulcerative colitis.     

Increased group II phospholipase A2 in colonic mucosa of patients with Crohn's disease and ulcerative colitis.

The immunochemical protein content of group II phospholipase A2 (PLA2) and PLA2 enzymatic activity were measured for colonic mucosal biopsy samples obtained from patients with either Crohn's disease of the colon or ulcerative colitis, and control patients without inflammatory bowel disease. Immunoreactive group II PLA2 (IR-PLA2 II) content and PLA2 activity in actively inflamed colonic mucosa of Crohn's disease patients were significantly higher than those in inactively inflamed mucosa of Crohn's disease patients and the colonic mucosa of controls. IR-PLA2 II content and PLA2 activity in severely inflamed mucosa of ulcerative colitis patients were significantly higher than those in the colonic mucosa of the controls. Mucosal PLA2 enzymatic activity was closely correlated with mucosal IR-PLA2 II content in patients with Crohn's disease and ulcerative colitis. These results suggest that an increase in PLA2 enzymatic activity in inflamed colonic mucosa of Crohn's disease and ulcerative colitis was mainly attributed to increased protein content of group II PLA2, and that an increase in mucosal group II PLA2 may be involved in the pathogenesis of intestinal inflammation of Crohn's disease and ulcerative colitis.     

Tumour necrosis factor alpha converting enzyme (TACE) activity in the colonic mucosa of patients with inflammatory bowel disease

BACKGROUND: Anti-tumour necrosis factor alpha (TNF-alpha) antibodies are effective in Crohn's disease and perhaps ulcerative colitis but antigenicity and the high cost have raised interest in other strategies to block TNF-alpha. These include the TNF-alpha converting enzyme (TACE) which releases soluble TNF-alpha from transmembrane pro-TNF-alpha. AIM: To investigate whether TACE activity is present in human colonic mucosa. MATERIALS AND METHODS: Detergent extracts of cell membranes from colonic biopsies were obtained from 12 controls and 28 patients with inflammatory bowel disease. Enzyme activity was measured by hydrolysis assays using pro-TNF-alpha or oligopeptide substrates spanning the known pro-TNF-alpha cleavage site at Ala(76)-Val(77). Cleavage products were identified by western blotting, high pressure liquid chromatography, or mass spectrometry. TACE protein was localised by immunohistochemistry and identified by western blotting of detergent extracts from purified lamina propria mononuclear cells (LPMNC) or epithelial cells. RESULTS: Detergent extracts released TNF-alpha from pro-TNF-alpha and cleaved a model oligopeptide as predicted. Substrate hydrolysis was sensitive to known TACE/matrix metalloproteinase (MMP) inhibitors, but not trocade which has low activity against TACE. The median TACE level was increased in active ulcerative colitis (147 arbitrary units (AU)/mg; p<0.01) but not in Crohn's disease (81 AU/mg) compared with controls (79 AU/mg). Both the full length proform and the active form of TACE protein were expressed in LPMNC cells and epithelial cells. CONCLUSIONS: Functional TACE activity is ubiquitously expressed in the human colon and increased in ulcerative colitis, raising interest in MMP inhibitors targeting TACE.     

Cellular hypersensitivity to components of intestinal mucosa in ulcerative colitis and Crohn''s disease

The migration of peripheral leucocytes in vitro is examined in 36 patients with ulcerative colitis, in 34 patients with Crohn's disease, in 12 patients with ulcerative colitis or Crohn's disease, and in 31 patients with other gastrointestinal disorders. In a majority of the patients with ulcerative colitis extracts of foetal, colonic, and jejunoileal mucosa inhibit migration of leucocytes. A similar reactivity is seldom seen in Crohn's disease. Extracts of liver, kidney, and adrenal gland do not inhibit the migration. The reactivity of the ulcerative colitis group was found to be significantly different from that in controls and in the Crohn group, whereas the Crohn group did not differ significantly from the controls. The examination thus reveals a biological difference between ulcerative colitis and Crohn's disease, which are otherwise separable mainly on nosological criteria.The finding of a similar inhibition of leucocyte migration in five out of 31 patients with miscellaneous gastrointestinal disorders unrelated to ulcerative colitis and Crohn's disease was inconclusive.An antigen-induced inhibition of leucocyte migration has been shown in vitro to be a correlate of cellular hypersensitivity. The immunological mechanisms behind the present system are discussed, and it is concluded that the reactivity observed probably indicates the existence in ulcerative colitis of a state of cellular hypersensitivity to components of normal foetal, colonic, and jejunoileal mucosa.     

Virulence properties of Escherichia coli strains isolated from patients with inflammatory bowel disease.

Escherichia coli strains cultured from 74 patients with inflammatory bowel disease at different stages of disease activity (Crohn's disease (40), ulcerative colitis (34)) and 18 healthy controls were studied in relation to haemolysin and verotoxin production and enteroadherence. Disease activity was assessed by standard clinical and laboratory tests. Haemolytic E coli were isolated from 18% of patients with Crohn's disease, 24% with ulcerative colitis, and 11% of healthy controls. None of these differences was significant. No verotoxin producing strains were detected among the 216 E coli isolates examined but the extract from five strains (Crohn's (4), ulcerative colitis (1) produced a distinctive cytopathic effort on Vero cell monolayers which was later shown not to be due to verotoxin. The adhesion indices of E coli isolates cultured were: mean (SEM) 42.2 (6.4) for Crohn's disease, 43.3 (6.2) for ulcerative colitis, and 11.3 (2.0) for normal controls (p less than or equal to 0.0001). Adhesive E coli were isolated from 62% of patients with Crohn's disease and 68% with ulcerative colitis but from only 6% of normal controls (p less than or equal to 0.0002). Neither haemolysin production nor enteroadherence was dependent upon disease activity, disease location, sulphasalazine treatment, or previous intestinal resection. These results indicate that only enteroadherent E coli were frequently associated with inflammatory bowel disease; their relation to the pathogenesis of these conditions, however, remains uncertain.     

Increased production of vascular endothelial growth factor by intestinal mucosa of patients with inflammatory bowel disease.

BACKGROUND/AIMS: Vascular endothelial growth factor (VEGF) is a heparin-binding glycoprotein with potent angiogenic, mitogenic and vascular permeability-enhancing activities specific for endothelial cells. Recent studies have shown significantly increased VEGF serum levels in patients with active Crohn's disease and ulcerative colitis. The origin of the circulating VEGF is not yet completely described. The present investigation examines the VEGF production of colonic mucosa in consideration of mucosal disease activity in patients with inflammatory bowel disease. METHODOLOGY: Fifteen patients with inflammatory bowel disease were studied, 9 patients with Crohn's disease and 6 patients with ulcerative colitis. Biopsies were taken from endoscopically inflamed and non-inflamed colonic mucosa. Therefore, an analysis of the spontaneous VEGF production of cultured biopsies without stimulus and of the histological grade of inflammation scored on a scale of 0-3 (normal mucosa--severe chronic colitis) were performed. Eight patients with irritable bowel syndrome served as controls. VEGF levels in the supernatant of cultured mucosal biopsies were measured using an enzyme linked immunosorbent assay. RESULTS: VEGF production is expressed as pg/mg wet weight of the biopsies. Inflamed mucosa of patients with active ulcerative colitis (16.27 +/- 10.39, p = 0.003, n = 6) and active Crohn's disease (9.88 +/- 5.98, p < 0.012, n = 9) showed a significantly higher spontaneous production of VEGF by colonic mucosa than normal mucosa of controls (3.16 +/- 1.63, n = 8). In addition, there was an increased unstimulated VEGF production by cultured inflamed mucosa of patients with Crohn's disease compared with non-inflamed mucosa (3.88 +/- 3.66, p < 0.015, n = 9). In both Crohn's disease and ulcerative colitis, there was no significant difference between VEGF production by non-inflamed mucosa and normal mucosa of controls. CONCLUSIONS: The present study identifies the intestinal mucosa as one of the origins of the elevated VEGF serum levels in patients with active inflammatory bowel disease and verifies the findings of recent studies about the importance of VEGF in Crohn's disease and ulcerative colitis.     

Enhanced colonic nitric oxide generation and nitric oxide synthase activity in ulcerative colitis and Crohn's disease.

Recent studies have suggested that nitric oxide (NO.), the product of nitric oxide synthase in inflammatory cells, may play a part in tissue injury and inflammation through its oxidative metabolism. In this study the colonic generation of oxides of nitrogen (NOx) and nitric oxide synthase activity was determined in ulcerative colitis and Crohn's disease. Colonic biopsy specimens were obtained from inflammatory bowel disease patients and from normal controls. Mucosal explants were cultured in vitro for 24 hours and NOx generation was determined. Nitric oxide synthase activity was monitored by the conversion of [3H]-L-arginine to citrulline. Median NOx generation by inflamed colonic mucosa of patients with active ulcerative colitis and Crohn's colitis was 4.2- and 8.1-fold respectively higher than that by normal human colonic mucosa. In ulcerative colitis and Crohn's colitis nitric oxide synthase activity was 10.0- and 3.8-fold respectively higher than in normal subjects. Colonic NOx generation is significantly decreased by methylprednisolone and ketotifen. The decrease in NOx generation by cultured colonic mucosa induced by methylprednisolone suggests that NO synthase activity is induced during the culture and the steroid effect may contribute to its therapeutic effect. Enhanced colonic NOx generation by stimulated nitric oxide synthase activity in ulcerative colitis and Crohn's disease may contribute to tissue injury.     

Detection of colonic antigen(s) in tissues from ulcerative colitis using purified colitis colon tissue-bound IgG (CCA-IgG)

In our report of a disease-specific colonic tissue-bound antibody (CCA) from patients with ulcerative colitis (Proc Natl Acad Sci USA 1978;75:4528), crude CCA was largely fragmented, and the yield was small. We have modified the extraction procedure to increase the yield of intact IgG present in CCA by sequential elution, storage of tissues in presence of a protease inhibitor, 2 mM Phenylmethylsulfonyl Fluoride, and use of Phenylmethylsulfonyl Fluoride in extraction buffers. Intact CCA-IgG was purified using protein A-Sepharose 4B affinity chromatography. 125I-CCA-IgG formed immune complexes in vitro with the aqueous extracts of colonic mucosa from 5 patients with ulcerative colitis but not from 6 patients with Crohn's disease and 6 normal colons from patients with carcinoma (p less than 0.01). We also performed reverse experiments by iodination of the colonic mucosal extracts with Bolton-Hunter reagent and incubating them with several preparations of CCA-IgG. Mucosal extracts of colon from 3 patients with ulcerative colitis bound significantly with CCA-IgG when compared with the identical extracts from 3 patients with Crohn's disease, 3 with a normal colon (p less than 0.005), and with the control human IgG (p less than 0.025). These studies demonstrate a better method of extraction and purification of intact CCA-IgG. Intact CCA-IgG binds to a specific protein(s) present in the homogenates of colonic mucosa from patients with ulcerative colitis but not from patients with Crohn's disease and normal colon.     

Function of exudative neutrophilic granulocytes in patients with Crohn's disease or ulcerative colitis

Chemotactic, phagocytic, and oxidative metabolic activity of exudative leukocytes was measured in patients with Crohn's disease (n = 20) and with ulcerative colitis (n = 20). Unstimulated and casein-stimulated migration in Boyden chambers did not differ from that of healthy controls (n = 21). Patients with Crohn's disease had reduced serum-independent phagocytosis compared with healthy controls (p less than 0.01) and patients with ulcerative colitis (p less than 0.01). Serum-dependent phagocytosis by leukocytes from patients with Crohn's disease did not differ from that in controls but was slightly increased in patients with ulcerative colitis (p less than 0.02). Unstimulated leukocytes showed increased oxidative metabolic activity in both patient groups compared with controls (p less than 0.01), which was negatively correlated with the disease activity in Crohn's disease (p less than 0.02). The study shows that mobilized leukocytes from patients with Crohn's disease differ from those mobilized in ulcerative colitis and supports the concept of an abnormal inflammatory reaction in Crohn's disease.     

Different distribution of mast cells and macrophages in colonic mucosa of patients with collagenous colitis and inflammatory bowel disease

BACKGROUND/AIMS: Chronic inflammatory cells in colonic mucosa is a histopathologic feature in patients with collagenous colitis and inflammatory bowel disease. The aim of this study was to compare the distribution of mast cells and macrophages in the colonic mucosa of patients with collagenous colitis, Crohn's disease, and ulcerative colitis. METHODOLOGY: Patients with histologically confirmed collagenous colitis (n = 13), Crohn's disease (n = 20) or ulcerative colitis (n = 20) and normal control patients (n = 20) were included in this study. Biopsy specimens were obtained from the sigmoid colon of each patient, and immunostained using antibodies to tryptase (AA1) and CD68. The number of mast cells and macrophages located in upper and lower part of the lamina propria was determined. RESULTS: The number of mast cells in the upper part of lamina propria in patients with collagenous colitis (286 +/- 89/mm2, mean +/- SD), Crohn's disease (330 +/- 84/mm2) and ulcerative colitis (355 +/- 90/mm2), was higher than normal controls (201 +/- 44/mm2). The number of mast cells in the lower part of lamina propria in patients with Crohn's disease (345 +/- 87/mm2) and ulcerative colitis (363 +/- 86/mm2) was higher than collagenous colitis (266 +/- 63/mm2) and normal controls (309 +/- 60/mm2). The number of macrophages in the lower part of lamina propria in patients with Crohn's disease (330 +/- 63/mm2) and ulcerative colitis (301 +/- 60/mm2) was higher than in collagenous colitis (247 +/- 46/mm2) and normal controls (242 +/- 52/mm2), although there were no significant differences in the number of macrophages present in the upper part of the lamina propria among the four groups. CONCLUSIONS: Our data showed the presence of a different distribution of mast cells and macrophages in collagenous colitis and inflammatory bowel disease, and these suggest that because mucosal mast cells have been implicated in the development of Th2 response collagenous colitis is more of a Th2 type reaction rather than Th1.     

Antibodies against laminaribioside and chitobioside are novel serologic markers in Crohn's disease

BACKGROUND & AIMS: New serologic markers of inflammatory bowel disease may be useful for differentiating between Crohn's disease and ulcerative colitis and for disease stratification. We profiled sugar-binding antibodies to identify novel antiglycan antibodies that may be associated with inflammatory bowel disease. METHODS: Serum samples were obtained from patients with diagnosed Crohn's disease or ulcerative colitis and from control patients. The presence of antiglycan antibodies was evaluated using either a glycan array (GlycoChip; Glycominds, Ltd, Lod, Israel) in patients with Crohn's disease (n = 72) or ulcerative colitis (n = 56) and in healthy controls (n = 41) or using an enzyme-linked immunosorbent assay in patients with Crohn's disease (n = 124), ulcerative colitis (n = 106), and in control patients (n = 101). RESULTS: Inaddition to antibodies against mannan, antibodies to laminaribioside (Glc[beta1,3]Glc[beta]) and chitobioside (GlcNAc[beta1,4]GlcNAc[beta]) had the highest discriminative capability between Crohn's disease and ulcerative colitis (P < .001 and P < .05, respectively). Importantly, 44% (12/27) of anti-Saccharomyces cerevisiae antibody-negative Crohn's disease patients were positive for antilaminaribioside or antichitobioside. In patients with inflammatory bowel disease positive for antibodies against either laminaribioside, chitobioside, or mannan, the diagnosis of Crohn's disease was suggested with a sensitivity of 77.4% and specificity of 90.6%. Having at least 2 of these antibodies increased the specificity to 99.1%. In Crohn's disease, higher levels of antibodies against laminaribioside or mannan were significantly associated with small intestinal disease (P = .03 and P < .0001, respectively). CONCLUSIONS: Antilaminaribioside and antichitobioside carbohydrate antibodies are novel serologic markers associated with Crohn's disease. These antibodies may contribute to the diagnosis and improved stratification of Crohn's disease.     

Increased serum levels of YKL-40 in patients with inflammatory bowel disease

BACKGROUND AND AIMS: Initiation of a fibrotic process has been suggested as part of the intestinal response to chronic inflammation in inflammatory bowel disease. YKL-40 has been proposed as a new serum marker of fibrosis. We studied compared the serum levels of YKL-40 in patients with ulcerative colitis or Crohn's disease with inflammatory and healthy controls. PATIENTS AND METHODS: YKL-40 serum levels were measured in 179 patients with inflammatory bowel disease (94 ulcerative colitis, 85 Crohn's disease), in 23 with intestinal inflammation of other causes, and 70 matched healthy controls using a commercially available enzyme-linked immunosorbent assay. YKL-40 levels were assessed in terms of disease activity, type and localization. RESULTS: Mean serum YKL-40 levels were 102.6+/-82.7 ng/ml in ulcerative colitis patients and 112.2+/-83.7 ng/ml in Crohn's disease patients, significantly higher than in healthy controls (64.1+/-21.4 ng/ml) but not significantly different from inflammatory controls (77.8+/-23.1 ng/ml). Disease activity and C-reactive protein levels were significantly correlated with YKL-40 levels in both ulcerative colitis and Crohn's disease. Crohn's disease patients with ileum localization had significantly higher YKL-40 levels than those with ileocolonic or colonic disease. Patients with stenotic disease had mean YKL-40 levels not significantly different than those with nonstenotic disease. CONCLUSION: Serum levels of YKL-40 are increased in patients with inflammatory bowel disease, and this is associated with the inflammatory process rather than with the degree of fibrosis.     

Diagnostic value of high resolution sonography in Crohn's disease and ulcerative colitis]

OBJECTIVE: To evaluate the diagnostic accuracy of high resolution sonography in patients with inflammatory bowel disease (MICI). PATIENTS AND METHODS: In patients with Crohn's disease (n = 48), ulcerative colitis (n = 23), indeterminate colitis (n = 3), inflammatory (n = 21) and non-inflammatory (n = 23) controls, high resolution sonography was performed and compared to colonoscopy (+/- retrograde ileoscopy) and/or baryum studies of the small bowel and the colon. RESULTS: Diagnosis of intestinal inflammation or not was correct in 69/74 MICI patients (sensitivity: 94.4%, specificity: 66.7%, global accuracy: 93.2%). Segment location was accurate in 58/74 (sensitivity: 80.3%, specificity: 66.7%, global accuracy: 79.7), more frequently in Crohn's disease, than in ulcerative colitis. Five out of six complications of Crohn's disease were diagnosed. In Crohn's disease, the method was more accurate in case of colonic or ileocolonic involvement. CONCLUSION: High resolution sonography is a reliable diagnostic tool for the detection of intestinal inflammation and related complications in MICI. In can be of value in the follow-up and seems particularly interesting in the case of temporary contraindication of invasive methods.     

Antineutrophil cytoplasm autoantibodies against bactericidal/permeability-increasing protein in inflammatory bowel disease.

BACKGROUND: Bactericidal/permeability-increasing protein (BPI), a constituent of primary neutrophil granules, is a potent natural antibiotic and an antineutrophil cytoplasm antibody (ANCA) antigen in cases of vasculitis in which the target antigen is neither myeloperoxidase (MPO) nor proteinase-3 (PR3). AIM: To investigate BPI as a possible target antigen for ANCAs in inflammatory bowel disease. METHODS: ANCAs were detected by routine immunofluorescence (IIF) and solid phase enzyme linked immunosorbent assay (ELISA) performed for antibodies to the purified neutrophil granule proteins; MPO, PR3, cathepsin-G, lactoferrin, and BPI in serum samples from 88 patients with inflammatory bowel disease (36 with Crohn's disease, 52 with ulcerative colitis). Thirty patients with bacterial enteritis acted as controls. RESULTS: Significantly more patients with ulcerative colitis were ANCA positive by IIF (60%) than patients with Crohn's disease (28%) or infectious enteritis (23%) (p < 0.001). IgG anti-BPI antibodies were present in 29% of patients with ulcerative colitis, 14% of patients with Crohn's disease, and 23% of patients with infectious enteritis, occurring in 44% of those patients with inflammatory bowel disease who were ANCA positive by IIF. Antibodies to other ANCA antigens were rare. The presence of ANCAs was not related to either disease activity or extent; presence of anti-BPI antibodies was significantly related to both a lower serum albumin concentration (p = 0.001) and a higher erythrocyte sedimentation rate (p = 0.02) in patients with ulcerative colitis, and to colonic involvement in patients with Crohn's disease (p = 0.01). CONCLUSION: BPI is a significant minority target antigen for ANCAs in inflammatory bowel disease that seems related to colonic Crohn's disease and disease activity in ulcerative colitis. Anti-BPI antibodies occur in infectious enteritis.     

The role of CD40 in ulcerative colitis: histochemical analysis and clinical correlation

OBJECTIVES : CD40 co-stimulator seems to be implicated in the loss of tolerance against self-antigens in many autoimmune diseases. The evidence suggests that in the pathogenesis of ulcerative colitis there is an activity state against self-antigens of the gut wall and flora. The aim of this study was to analyse the expression of CD40 in ulcerative colitis, comparing it with Crohn's disease and nonspecific inflammation of the colon and to determine whether there is a relationship between its expression and the activity stage of the disease. METHODS : The expression of CD40 in the colonic samples of 51 patients (30 ulcerative colitis, 9 Crohn's disease and 12 nonspecific inflammation) was analysed by immunohistochemistry. Twenty-four patients with ulcerative colitis were scored according to clinical, endoscopic and histological classification. RESULTS : The mean percentage of CD40+ cells per field in the colonic mucosa was: ulcerative colitis 21 +/- 11%, Crohn's disease 24 +/- 9%, nonspecific inflammation 7 +/- 7%. The ulcerative colitis patients were statistically significantly different compared to the patients with nonspecific inflammation (P < 0.005), even when comparing the patients in remission (P < 0.05). The expression in Crohn's disease was similar to that in ulcerative colitis. The expression of CD40 in ulcerative colitis was directly proportional to the state of activity of the disease according to the clinical (P < 0.02), endoscopic (P < 0.01) and histological (P < 0.02) criteria. CONCLUSIONS : The expression of CD40 in the colonic mucosae of patients with ulcerative colitis is significantly increased and is proportional to the state of activity. The results seem to confirm the hypothesis that a loss of tolerance could be involved in the pathogenesis of this disease.     

Immunohistochemical analysis of the distribution of measles related antigen in the intestinal mucosa in inflammatory bowel disease

BACKGROUND: Measles virus is implicated in the aetiology of Crohn's disease. This measles hypothesis is mainly supported by immunohistochemical findings that the measles related antigen is present in the intestine of patients with Crohn's disease. Recently we isolated this antigen from the intestine of a patient with Crohn's disease using a molecular cloning technique and produced the monoclonal antibody against it (designated 4F12). AIM: To discover whether the measles related antigen is uniquely present in Crohn's disease. SUBJECTS/METHODS: Colonic mucosa samples from 20 patients with Crohn's disease, 20 with ulcerative colitis, 11 with non-inflammatory bowel disease (IBD) colitis, and nine controls were immunohistochemically stained with the anti-measles monoclonal antibody 4F12. The numbers of positive cells, the ratio of positive cells to nucleated cells, and the staining intensity of the positive cells were compared. Furthermore, the distribution of the measles antigen in other human organs was examined. RESULTS: Both the number of positive cells and the ratio of positive cells to nucleated cells were significantly increased in Crohn's disease, ulcerative colitis, and non-IBD colitis compared with controls (p<0.05) but were similar among the three disease groups. The staining intensity of the positive cells was also similar among the three disease groups. Small numbers of positive cells were observed in the oesophagus, stomach, duodenum, jejunum, and lung. CONCLUSIONS: The presence of the measles related antigen in the colonic mucosa was not unique to Crohn's disease. These results, together with the observation that such a measles related antigen was derived from host protein, do not support the hypothesis that measles virus causes Crohn's disease.