Contribution of immunofluorescence to identification and characterization of antineutrophil cytoplasmic antibodies in inflammatory bowel disease

We evaluated the combined use of different fixatives for the identification of atypical perinuclear anti-neutrophil cytoplasmic antibodies (p-ANCA) in patients with inflammatory bowel diseases (IBD) by indirect immunofluorescence (IIF). Sera from 59 ulcerative colitis (UC) and 37 Crohn’s disease (CD) patients, and from 64 healthy controls were studied. The IIF on ethanol-, formalin-, and methanol-fixed neutrophils was used for the detection of ANCA. Enzyme linked immunosorbant assay (ELISA) was performed to identify the antigens recognized by ANCA. ANCAs were present in 35 of 59 (59.3%) UC patients and in 10 of 37 (27.02%) CD patients. Atypical p-ANCA positivity was strongly associated with UC disease (44.1% in UC vs. 8.1% in CD; p = 0.0002). The combined application of different fixatives contributed to make easy the differentiation between typical p-ANCA and atypical p-ANCA. Atypical p-ANCA determination appears to be a useful parameter for the distinction between UC and CD.     

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.     

Significance of antineutrophil cytoplasmic antibody in adult patients with Henoch-Schönlein purpura presenting mainly with gastrointestinal symptoms

AIM： To test the clinical significance of antineutrophil cytoplasmic antibody （ANCA） in evaluation of adult Henoch-Schonlein purpura （HSP） patients presenting mainly with abdominal symptoms. METHODS： Twenty-eight consecutive HSP patients who presented predominantly with abdominal symptoms were enrolled in this study. Control subjects included 27 ageand sex-matched patients with peptic ulcer disease, colon cancer, acute gastroenteritis, irritable bowel syndrome and colonic polyps. ANCA was measured by indirect immunofluorescence （IIF） in all patients, and follow-up ELISA was performed in patients with positive IIF tests. RESULTS： ANCA was detected in 9 HSP patients by IIF （2 were positive for c-ANCA and 7 were positive for p-ANCA）. No ANCA was found in the control group. The sensitivity and specificity of a positive ANCA test （either c- or p-ANCA） were 32.1% and 100% respectively. Only one out of the 9 patients with positive ANCA by IIF had positive ANCA by ELISA and the antigen was myeloperoxidase （MPO）. The patients positive for ANCA had higher HSP clinical scores, and were more likely to have renal function impairment. Patients with late purpura development were also associated with more severe clinical manifestations. CONCLUSION： A positive ANCA test is associated with more severe symptoms in HSP. After inflammatory bowel disease is excluded, a positive ANCA test provides a clue to the diagnosis of HSP presenting predominantly with abdominal symptoms.     

Antibodies to neutrophil cytoplasma in patients with ulcerative colitis and their first-degree relatives in Thailand

BACKGROUND: The prevalence of perinuclear antineutrophil cytoplasmic antibodies (p-ANCA) does not significantly vary in ethnically diverse populations. The prevalence of p-ANCA is high in ulcerative colitis and primary sclerosing cholangitis. While the prevalence of ulcerative colitis in Asian populations is low, it is interesting to know the prevalence of p-ANCA in such a population. METHODS: Sera from 33 cases of ulcerative colitis diagnosed during the last 10 years at the diarrhea clinic, Division of Gastroenterology, Siriraj Hospital, were prospectively compared with case controls consisting of 15 cases of diarrhea from non-inflammatory bowel diseases and 25 non-diarrheic patients. Indirect immunofluorescence assay was used to detect p-ANCA in all the sera. RESULTS: Positive p-ANCA tests were found in 13 of the 33 patients with ulcerative colitis and in one of the 40 controls. Sensitivity of the test was 39.4% and the specificity was 97.5%. The one patient with positive p-ANCA in the control group was the patient with irritable bowel syndrome. Of the 13 p-ANCA-positive ulcerative colitis patients, two cases were found to have proctosigmoiditis, seven cases had left-sided colitis, and four cases had pancolitis. Perinuclear antineutrophil cytoplasmic antibodies was one of the 22 cases of first-degree relatives of ulcerative colitis patients (22 relatives from 12 index ulcerative colitis cases). There was no correlation between the positivity of p-ANCA and disease activity, and extent of the disease. CONCLUSION: The prevalence of p-ANCA in Thai patients with ulcerative colitis (39.4%) is lower than that in the Western population. Although the prevalence of p-ANCA is low in the Thai population, it should serve as a useful tool in diagnosing ulcerative colitis in this part of the world where the disease is uncommon and difficult to diagnose. The negativity of p-ANCA in almost all first-degree relatives of Thai ulcerative colitis patients should be further elucidated.     

Significance of antineutrophil cytoplasmic antibody in adult patients with Henoch-Sch(o)nlein purpura presenting mainly with gastrointestinal symptoms

AIM: To test the clinical significance of antineutrophil cytoplasmic antibody (ANCA) in evaluation of adult Henoch-Schonlein purpura (HSP) patients presenting mainly with abdominal symptoms.METHODS: Twenty-eight consecutive HSP patients who presented predominantly with abdominal symptoms were enrolled in this study. Control subjects included 27 age-and sex-matched patients with peptic ulcer disease, colon cancer, acute gastroenteritis, irritable bowel syndrome and colonic polyps. ANCA was measured by indirect immunofluorescence (IIF) in all patients, and follow-up ELJSA was performed in patients with positive IIF tests.RESULTS: ANCA was detected in 9 HSP patients by IIF (2 were positive for c-ANCA and 7 were positive for p-ANCA). No ANCA was found in the control group. The sensitivity and specificity of a positive ANCA test (either c- or p-ANCA) were 32.1% and 100% respectively. Only one out of the 9 patients with positive ANCA by IIF had positive ANCA by ELISA and the antigen was myeloperoxidase (MPO). The patients positive for ANCA had higher HSP clinical scores, and were more likely to have renal function impairment. Patients with late purpura development were also associated with more severe clinical manifestations.CONCLUSION: A positive ANCA test is associated with more severe symptoms in HSP. After inflammatory bowel disease is excluded, a positive ANCA test provides a clue to the diagnosis of HSP presenting predominantly with abdominal symptoms.     

Perinuclear anti-neutrophil cytoplasmic antibody and refractory pouchitis

Refractory pouchitis (RP) is a debilitating complication of ileal pouch reservoirs that affects hypothesized that it reflects underlying Crohn's disease. Since perinuclear anti-neutrophil cytoplasmic antibody (pANCA) is found in approximately 70% of ulcerative colitis patients but only rarely in Crohn's disease patients, it may help distinguish Crohn's disease from ulcerative colitis. Therefore, to test whether RP reflects missed Crohn's disease, we determined the ANCA status of 26 patients with RP. The pANCA was positive in 42% of cases [50% of Kock pouch cases and 33% of ileoanal pull-through (IAPT) cases] and 57% of matched control subjects without pouchitis (N=42,P=NS). Moreover, 3/6 (50%) of IAPT RP subjects whose signs and symptoms most suggested Crohn's disease tested positive for pANCA. When compared to controls, IAPT cases exhibited significantly more preoperative extraintestinal manifestations (EIMs) of inflammatory bowel disease (P<0.05). The presence of preoperative EIMs was 100% predictive of postoperative EIMs (P<0.05). Review of pouch biopsies from cases of RP revealed no pathognomonic histologic features of Crohn's disease. These data confirm our previous suggestion that RP does not reflect underlying Crohn's disease but may be associated with the EIMs of inflammatory bowel disease.     

Genetic heterogeneity within ulcerative colitis determined by an interleukin-1 receptor antagonist gene polymorphism and antineutrophil cytoplasmic antibodies.

BACKGROUND: Although there is strong evidence implicating genetic predisposition in the pathogenesis of the chronic inflammatory bowel diseases, the number and identity of susceptibility genes remain uncertain. Cytokine genes are tentative candidate loci, but data regarding association studies in different populations are conflicting. AIMS: To determine potential associations of interleukin-1 receptor antagonist (IL-1ra), tumour necrosis factor alpha (TNF alpha), and tumour necrosis factor beta (TNF beta) gene polymorphisms with ulcerative colitis or subsets of ulcerative colitis in a Spanish population. METHODS: Genotyping for IL-1ra, TNF alpha and TNF beta gene polymorphisms was performed by the polymerase chain reaction in 95 patients with ulcerative colitis and 74 healthy controls. A variable number of tandem repeats (VNTR) in the IL-1ra gene, and a single base pair polymorphism in the TNF alpha gene promoter region (-308) and in the first intron of the TNF beta gene were analysed. Anti-neutrophil cytoplasmic antibodies (ANCA) were detected using an indirect immunofluorescence assay. RESULTS: There were no significant differences between ulcerative colitis patients and controls in either polymorphism analysed, nor between ulcerative colitis subgroups as a function of the clinical disease pattern. However, when stratified by their ANCA status, perinuclear ANCA (p-ANCA) ulcerative colitis showed an increased frequency of the genotype 1,2 of the IL-1ra gene compared with ANCA-negative ulcerative colitis (52% versus 28%; P = 0.02, Pcorr = 0.1). Furthermore, p-ANCA ulcerative colitis had a statistically significant increase of this genotype compared with cytoplasmic ANCA (c-ANCA)/ANCA-negative ulcerative colitis (52% versus 26.5%; P = 0.01, Pcorr = 0.05). CONCLUSIONS: In the Spanish population studied, the polymorphisms analysed in the IL-1ra, TNF alpha and TNF beta genes are unlikely to be important in the overall susceptibility to ulcerative colitis. However, the combination of a subclinical (p-ANCA) and a genetic (IL-1ra gene) marker identified a distinct ulcerative colitis subgroup (p-ANCA; IL-1ra genotype 1,2). These findings provide further evidence of genetic heterogeneity within ulcerative colitis, and support the concept that ANCA may represent a subclinical marker of genetic heterogeneity.     

Identification of a prodromal period in Crohn's disease but not ulcerative colitis

OBJECTIVES: Irritable bowel syndrome, a common gastrointestinal diagnosis, has not been clearly studied in inflammatory bowel disease. Some of the residual symptoms in subjects treated with Crohn's disease and ulcerative colitis are thought to be related to irritable bowel syndrome. The aims of this study were 1) to describe the duration and nature of complaints before the diagnosis of Crohn's disease and ulcerative colitis (prodromal period), and 2) to determine the role of IBS in this prodromal period. METHODS: A total of 66 patients with confirmed inflammatory bowel disease were enrolled in the study. The subjects received a questionnaire to ascertain the nature and duration of symptoms preceding the diagnosis of Crohn's disease or ulcerative colitis, including features described under the Rome criteria for irritable bowel syndrome. RESULTS: Of the 66 subjects analyzed, 45 had Crohn's disease and 21 had ulcerative colitis. The prodromal period was 7.7 +/- 10.7 yr for Crohn's disease and 1.2 +/- 1.8 yr for ulcerative colitis (p < 0.05). Once patients meeting the Rome criteria for irritable bowel syndrome during the prodrome were excluded, the duration of the prodromal period (non-IBS) for ulcerative colitis dropped to 0.8 +/- 1.3 yr compared to 6.9 +/- 9.8 yr in the Crohn's disease group (p < 0.05). The symptoms of the non-IBS prodrome in subjects with Crohn's disease were bloating, diarrhea, stomach pain, heartburn, fever, weight loss, and fatigue. Further analysis demonstrated that subjects whose Crohn's disease initially began as colonic disease had a longer prodrome than with small bowel. In the non-IBS Crohn's group, there was also a correlation between the age at the time of diagnosis and the duration of prodrome (r = 0.67, p < 0.0001). CONCLUSIONS: There is a significant prodromal period before the time of diagnosis of Crohn's disease that is not found in ulcerative colitis even after exclusion of subjects with IBS.     

Antineutrophil cytoplasmic antibodies (ANCA) and small vessel vasculitis

We report on one patient with Wegener's granulomatosis (WG) and two patients with microscopic polyangiitis (MPA). The patient with WG had signs of a respiratory infection and showed a c-ANCA pattern with proteinase 3 (PR3) specificity. The patients with MPA presented with pulmonary haemorrhage and signs of renal damage and showed a p-ANCA pattern with myeloperoxidase (MPO) specificity. In the three patients histopathological findings confirmed the diagnosis. We discuss the clinical indications of ANCA testing and the current terminology for reporting ANCA results (c-ANCA, p-ANCA, c-ANCA (atypical) and atypical ANCA). The target antigens and diseases associated with these different patterns are considered. Finally we focus on the value of ANCA and more specific PR3-ANCA and MPO-ANCA in the diagnosis of WG and MPA. The new application domain of ANCA in Crohn's disease and ulcerative colitis is also discussed.     

Anti-Neutrophil Cytoplasmic Antibodies in Inflammatory Bowel Disease with Special Attention for IgA-Class Antibodies

Perinuclear anti-neutrophil cytoplasmicantibodies (P-ANCA) of the IgG class have been reportedin inflammatory bowel disease, mainly in ulcerativecolitis. Since this disease affects the gastrointestinal tract, we determined whether IgA class ANCAwere present in inflammatory bowel disease. We used anindirect immunofluorescence assay for IgG and IgA ANCAtesting. Sera from 34 patients with Crohn's disease and 29 patients with ulcerative colitis werecollected together with clinical and laboratory data. Wefound IgA class ANCA of a perinuclear type in 52% ofpatients with ulcerative colitis and in 9% of Crohn's disease patients. There was a significantassociation between the presence of IgA ANCA and theoccurrence of blood in the feces in the ulcerativecolitis group (P = 0.03). IgG ANCA was found in 56% ofpatients with ulcerative colitis and in 7% of patientswith Crohn's disease. Because of partial overlap betweenIgG and IgA ANCA positivity, the sensitivity of ANCAtesting in ulcerative colitis increased from 56% up to 78% by combining IgG and IgA assays. Inconclusion, IgA ANCA occurs with a high prevalence inulcerative colitis. Moreover there is a possiblerelationship between IgA ANCA and disease activity in ulcerative colitis.     

Antineutrophil cytoplasmic antibodies in Japanese patients with inflammatory bowel disease: prevalence and recognition of putative antigens

OBJECTIVE: Our aim was to investigate the prevalence of antineutrophil cytoplasmic antibodies (ANCA) in Japanese patients with ulcerative colitis (UC) and Crohn's disease (CD), and the putative antigens recognized by perinuclear staining pattern ANCA (p-ANCA)-positive sera. METHODS: Sera from UC (n = 52) and CD (n = 43) patients, and from healthy controls (n = 74) were studied. The indirect immunofluorescence (IIF) method was used for the detection of ANCA and its binding pattern. p-ANCA-positive sera were studied further for putative antigens. ELISAs using lactoferrin (Lf), myeloperoxidase (MPO), and cathepsin G (Cat G) as antigens were performed. RESULTS: ANCA was positive in 40 of the 52 (76.9%) UC (p-ANCA in 33) and in 32 of the 43 (74.4%) CD (p-ANCA in 31) patients. UC and CD patients showed significantly higher titers of p-ANCA than controls; however, no significant difference was observed between UC and CD. In UC, 23, 17, and nine of the 33 patients with p-ANCA-positive sera showed reactivity with Lf, MPO, and Cat-G, respectively. In CD, 21, 20, and 11 of the 31 patients with p-ANCA-positive sera showed reactivity with Lf, MPO, and Cat-G, respectively. Fourteen of the UC and six of the CD patients showed reactivity with two different antigens, and seven of the UC and 11 of the CD patients showed reactivity with all three antigens. The presence of anti-Lf and anti-MPO antibodies was further confirmed by Western blotting. CONCLUSIONS: ANCA is useful in distinguishing patients with IBD from normal subjects but is not sufficient for the differential diagnosis of CD and UC. p-ANCA reactivity might be derived from the recognition of heterogeneous neutrophil-associated antigens.     

Spectrum of Autoantibodies in Celiac Patients and Relatives

The coexistence of celiac disease together with a range of autoimmune disorders has already been reported. The aims of this study were to perform a broad spectrum of autoantibodies in celiac patients (N = 56), their first-degree relatives (N = 118), and compare the data with healthy controls (N = 101) and patients with inflammatory bowel disease (N = 42; Crohn's disease, N = 18 and ulcerative colitis, N = 24). All serum samples were tested by indirect immunofluorescence to the anti-endomysium antibodies (EmA), anti-neutrophil cytoplasmic (ANCA), anti-smooth-muscle (SMA), anti-mitochondrial (AMA), anti-nuclear (ANA), anti-liver–kidney microsomal (LKM), anti-gastric parietal cells (GPCA), and anti-thyroid microsome (TMA). EmA were detected in 100% of celiac patients ingesting gluten and in 16.1% of the first-degree relatives, while ANCA were positive only in patients with ulcerative colitis (45.6%) and Crohn's disease (16.5%). Fourteen CD patients (25%) were positive for at least one of the other autoantibodies, with significant prevalence of TMA, ANA, and GPCA, while the relatives showed 17.8% of positivity, with an increased prevalence of ANA and TMA. These results emphasize the value of screening for different autoantibodies in celiac patients and their relatives and corroborate the need for evaluation and follow-up of these individuals.     

Association of Susceptibility Locus for Inflammatory Bowel Disease on Chromosome 16 with Both Ulcerative Colitis and Crohn's Disease

A susceptibility locus for inflammatory bowel disease (IBD) on chromosome 16 (IBD1) has been linked to Crohn's disease in genome-wide linkage studies. We performed a case–control study with two markers for this locus using leukocyte DNA from 127 Crohn's patients, 83 ulcerative colitis patients, and 74 control patients. Allele, genotype, and haplotype frequencies of the polymerase chain reaction products were determined using autoradiography. Haplotype frequencies differed for ulcerative colitis and Crohn's disease, particularly for haplotype CC (22% ulcerative colitis vs 10% Crohn's disease, P = 0.002 ² = 10.0) and haplotype CD (18% Crohn's disease vs 9% ulcerative colitis, P = 0.025 ² = 5.02). These data demonstrate the association of the IBD1 locus with both ulcerative colitis and Crohn's disease in a group of unrelated IBD patients. The use of such microsatellite markers when combined with others, might help distinguish ulcerative colitis from Crohn's disease in patients with ambiguous clinical and histological features.     

Fecal Lysozyme in Assessment of Disease Activity in Inflammatory Bowel Disease

This study was undertaken to determine whethermeasurement of fecal lysozyme is helpful in determiningdisease activity in inflammatory bowel disease. In 112patients with Crohn's disease, 46 patients with ulcerative colitis, and 40 controls, fecallysozyme concentration was measured. Results werecorrelated with CDAI and AI in Crohn's disease and withTruelove and Witts' grading in ulcerative colitis. Fecal lysozyme concentration (mean ± SEM) wassignificantly (P < 0.001) higher in Crohn's disease(75 ± 14 mug/g) and ulcerative colitis (238± 33 mug/g) than in controls (6 ± 1g/g). There was only a weak correlation between fecal lysozymeconcentration and CDAI (r = 0.32; P = 0.001) and AI (r= 0.38; P < 0.0005) in patients with Crohn's diseaseand with Truelove and Witts' grading (r = 0.47; P =0.001) in ulcerative colitis. When CDAI 150 or AI 100were used as the standard for active disease, fecallysozyme concentration was elevated in 78% of patientswith active colonic Crohn's disease. In ulcerative colitis fecal lysozyme concentration wasincreased in active disease (95% in grade II and 94% ingrade III) as compared 33% in grade I. Measurement offecal lysozyme is of little help in diagnosing and determining disease activity of inflammatorybowel disease as whole, but it may be of help fordiagnosis and assessment of activity of colonicIBD.     

PMN-elastase in assessment of patients with inflammatory bowel disease

PMN-elastase is a proteinase released by activated neutrophils. PMN-elastase was determined in two independent populations with inflammatory bowel disease. In an unselected population of 70 consecutive patients with Crohn's disease and 24 patients with ulcerative colitis with different degrees of disease activity plasma PMN-elastase levels were statistically significantly higher in patients with active than in patients with inactive disease [Crohn's disease: 80.5±33.2 ng/ml vs 60.1±24.6 ng/ml (means±sd),P=0.0017; ulcerative colitis: 98.2±54.9 ng/ml vs 59.2±16.8 ng/ml,P=0.026]. PMN-elastase levels in feces were also higher in patients with active Crohn's disease (23.6±15.3 ng/g vs 13.6±12.5 ng/g,P=0.0021) and active ulcerative colitis (46.5±60.5 ng/g vs 20.2±25.0 ng/g,P=0.46), but the difference reached significance only in Crohn's disease. Correlation of disease activity and PMN-elastase in individual patients showed a statistically significant correlation between plasma and fecal elastase concentrations and disease activity in ulcerative colitis (plasma:r=0.72,P<0.001; feces:r=0.423,P<0.001) but not fecal elastase concentrations (r=0.0083,P=0.485) correlated significantly with disease activity. Plasma PMN-elastase correlated weakly with fecal PMN-elastase levels in Crohn's disease (r=0.431,P<0.01) and in ulcerative colitis (r=0.515,P=0.05). In 28 patients with highly active Crohn's disease [median severity activity index (SAI) 203] and 11 patients with highly active ulcerative colitis [median Rachmilewitz index (RI) 14] studied before and four weeks after steroid therapy, treatment lowered the median SAI to 140 and the median RI to 4.5. Mean plasma elastase concentrations decreased concomitantly from 83±44.9 ng/ml to 61.8±25.8 (P=0.0035) in patients with Crohn's disease and from 110±49.5 to 71.6±28.8 ng/ml (P=0.0069) in patients with ulcerative colitis. In conclusion, there is a release of PMN-elastase in active IBD, which can be detected in plasma as well as in feces. Plasma elastase levels reflect disease activity in patients with IBD. The variation of the data and the large overlap between different groups, however, strongly reduce the clinical value.This research was supported by the SFB 154 of the Deutsche Forschungsgemeinschaft. V. Gross is supported by a Heisenberg-Stipendium of the Deutsche Forschungsgemeinschaft.     

Autoantibodies against the bactericidal/permeability-increasing protein from inflammatory bowel disease patients can impair the antibiotic activity of bactericidal/permeability-increasing protein

Bactericidal/permeability-increasing protein (BPI) is an antineutrophil cytoplasmic autoantibody (ANCA) target antigen in inflammatory bowel disease (IBD). The aim of this study was to characterize binding regions of BPI-autoantibodies and to analyze their ability to block the antibiotic effect of BPI. Sera of 24 ulcerative colitis and Crohn's disease patients were examined in indirect immuno-fluorescence, ANCA enzyme-linked immunosorbent assay (ELISA), and by epitope mapping with 13mer peptides and Western blot for presence of BPI-autoantibodies. IgG preparations were used to determine inhibition of BPI's antimicrobial function by BPI-autoantibodies in a bacterial growth inhibition assay. BPI-autoantibodies were detected by ELISA in 18/24 patients. Epitope mapping and western blotting revealed an additional 3 patients with BPI-autoantibodies. IgG preparations of all patients with Crohn's disease and 9 of 12 ulcerative colitis patients could inhibit the antibiotic function of BPI in vitro as compared with healthy control subjects. Inhibiting BPI-autoantibodies correlated with extraintestinal manifestations, peripheral blood leukocyte counts, and anemia. BPI-autoantibodies recognizing the N-terminal portion were associated with greater mucosal damage and intestinal extent of disease. BPI is a frequent target antigen of autoantibodies in ulcerative colitis and Crohn's disease. Inhibition of the antibiotic function mediated by the N-terminal region of BPI by these autoantibodies may contribute to a proinflammatory environment in IBD patients.     

IBS-Like Symptoms in Patients with Inflammatory Bowel Disease in Remission; Relationships with Quality of Life and Coping Behavior

The aim of this study was to assess the prevalence of irritable bowel syndrome-like symptoms in healthy controls and inflammatory bowel disease patients in remission using the Rome II criteria. Furthermore, the possible relation of irritable bowel syndrome-like symptoms with the quality of life and coping behavior was studied. Seventy-three ulcerative colitis patients in remission, 34 Crohn's disease patients in remission, and 66 healthy controls completed questionnaires on irritable bowel syndrome, quality of life, and coping. Using the Rome II criteria, irritable bowel syndrome-like symptoms were found in one-third of ulcerative colitis patients and in 42% of Crohn's disease patients in remission. The presence of irritable bowel syndrome-like symptoms impaired the quality of life of patients, while no relation was found between the presence of symptoms and coping strategies.     

Enzyme linked immunosorbent assay (ELISA) and immunoprecipitation studies on anti-goblet cell antibody using a mucin producing cell line in patients with inflammatory bowel disease.

Circulating anti-goblet cell antibody and its corresponding antigen in patients with inflammatory bowel disease were investigated. Anti-goblet cell antibody in the serum was examined by immunocytochemistry and enzyme linked immunosorbent assay (ELISA), using a colonic cancer cell line, HT29-18-N2, which differentiates into intestinal goblet cells. The frequencies of anti-goblet cell antibody detected by immunocytochemistry were 14 in 48 patients with ulcerative colitis (29%) and five in 15 patients with Crohn's disease (33%). By ELISA, the frequencies of anti-goblet cell antibody were 38% in ulcerative colitis and 33% in Crohn's disease. This antibody did not relate directly to anti-neutrophil cytoplasmic antibodies (ANCA), although the serum samples positive for anti-goblet cell antibody were commonly positive for ANCA in ulcerative colitis. Immunoprecipitation and SDS polyacrylamide gel electrophoresis (PAGE) study showed that the antibody in the ELISA positive serum samples recognised a > 200 kD goblet cell antigen, which remained unchanged after reduction, indicating that it consists of single chain polypeptides. These results suggest that there is a subgroup of inflammatory bowel disease that has circulating anti-goblet cell antibody reactive with a > 200 kD antigen. The antibody detected by newly established ELISA will be a disease marker for this group and the identification of the corresponding antigens may be important for the understanding of the underlying immune abnormalities.     

Ulcerative Colitis and Antineutrophil Cytoplasmic Antibodies in Hong Kong Chinese

Inflammatory bowel diseases are known to be rare among the Chinese. The diagnosis of ulcerative colitis has been difficult in some of the Asian countries where infective colitis is more prevalent. Twenty-three Hong Kong Chinese patients diagnosed to have ulcerative colitis were reviewed. The symptoms were relatively mild and extraintestinal manifestation had been rare. Patients responded well to steroid therapy and sulfasalazine. Three patients in this series were found to have cyst and/or trophozoites of Entamoeba histolytica in stool. In this series, 19 patients were tested for antineutrophil cytoplasmic antibody (ANCA). Fourteen patients (73.5%) were positive, of which six (31.5%) showed a perinuclear staining pattern and eight (42%) demonstrated a cytoplasmic pattern. Five patients (26.5%) were negative for any ANCA, and none was positive for both. Sera of these patients were also tested for anti-α granules, anti-myeloperoxidase, and anti-lactoferrin activities. None was positive. Control sera collected from 16 patients with irritable bowel syndrome were all negative for the tests. In conclusion, testing of ANCAs may help in making the diagnosis of idiopathic inflammatory bowel disease in difficult situations.     

NoMycobacterium paratuberculosis found in crohn's disease using the polymerase chain reaction

M. paratuberculosis has been considered as a putative causative factor of Crohn's disease. However, its detection in diseased tissue samples using the polymerase chain reaction yielded conflicting results. We validated this technique for the detection of mycobacteria (any species) andM. paratuberculosis before applying it to 72 intestinal biopsies from patients with Crohn's disease (N=36), ulcerative colitis (N=13), and control subjects (N=23). Possible polymerase chain reaction inhibitors were detected by spiking template DNA with the equivalent of twoM. paratuberculosis genomes. Mycobacteria were found in 17/36 (47%), 6/13 (46%), and 13/23 (57%) tissue samples of Crohn's disease, ulcerative colitis, and controls, respectively. NoM. paratuberculosis were detected in any sample. It is concluded that mycobacteria are present with a similar frequency in the intestinal tissues or luminal inclusions of patients with inflammatory bowel disease and of those unaffected by the disease. Our data do not support a role forM. paratuberculosis in Crohn's disease.This work was partially supported by the Damien Foundation (Belgium) and Wilson-Cook, Inc. Jean-Marc Dumonceau is a fellow of the Fondation Erasme and of the Communauté Francaise de Belgique.