Carol J. Landers

A distinct subset of antineutrophil cytoplasmic antibodies is associated with inflammatory bowel disease

Antineutrophil cytoplasmic antibodies (ANCAs) have recently been demonstrated to be of importance in Wegeners granulomatosis and certain other forms of vasculitis associated with glomerulonephritis. With a fixed-cell ELISA, we demonstrated that ANCAs occur in the serum of patients with inflammatory bowel disease (IBD) involving the colon. In a blinded study, sera from 21 of 25 patients with ulcerative colitis (UC) and five of 25 patients with Crohns disease had binding in the fixed-cell ELISA. The five reactive sera from patients with Crohns disease were associated with the presence of clear colonic involvement. The presence of ANCA in patients with UC was not influenced by disease distribution or activity. Indeed, such antibodies were present in four subjects with UC more than 5 years after colectomy. The IBD-associated ANCAs were distinct from ANCAs reported in patients with Wegeners granulomatosis since the pattern of staining on indirect immunofluorescence exhibited a nongranular perinuclear distribution (P-ANCA). The P-ANCA observed in IBD did not react with myeloperoxidase and thus was distinct from the P-ANCA observed in vasculitis with cresentric glomerulonephritis. IBD and, in particular, UC, is associated with a distinct subset of P-ANCA, which may have important diagnostic and potential pathophysiologic implications.

Neutrophil cytoplasmic antibodies : a link between primary sclerosing cholangitis and ulcerative colitis

Whether serum autoantibodies to neutrophil cytoplasmic components, previously found in ulcerative colitis, are also associated with primary sclerosing cholangitis was determined. In an enzyme-linked immunosorbent assay for immunoglobulin G neutrophil antibodies, neutrophil binding by primary sclerosing cholangitis sera was significantly greater than that for primary biliary cirrhosis, chronic hepatitis B, and chronic non-A, non-B hepatitis. Similar differences were seen when sera from patients with primary sclerosing cholangitis without evidence for ulcerative colitis were compared with sera from liver disease controls. Perinuclear immunofluorescence staining of neutrophils was exhibited by the majority of ulcerative colitis, primary sclerosing cholangitis, and primary sclerosing cholangitis without ulcerative colitis sera. The combination of elevated immunoglobulin G neutrophil antibodies and a perinuclear pattern was 65% sensitive and 100% specific for primary sclerosing cholangitis compared with the liver disease control sera. It is concluded that neutrophil cytoplasmic antibodies in ulcerative colitis and primary sclerosing cholangitis may be markers of shared underlying immunopathogenic mechanisms. Identification of the target antigen(s) may facilitate understanding of the underlying immune response and development of an improved disease marker assay.

Serum Immune Responses Predict Rapid Disease Progression among Children with Crohn's Disease: Immune Responses Predict Disease Progression

BACKGROUND AND AIM:Crohns disease (CD) is a heterogeneous disorder characterized by diverse clinical phenotypes. Childhood-onset CD has been described as a more aggressive phenotype. Genetic and immune factors may influence disease phenotype and clinical course. We examined the association of immune responses to microbial antigens with disease behavior and prospectively determined the influence of immune reactivity on disease progression in pediatric CD patients.METHODS:Sera were collected from 196 pediatric CD cases and tested for immune responses: anti-I2, anti-outer membrane protein C (anti-OmpC), anti-CBir1 flagellin (anti-CBir1), and anti-Saccharomyces-cerevisiae (ASCA) using ELISA. Associations between immune responses and clinical phenotype were evaluated.RESULTS:Fifty-eight patients (28%) developed internal penetrating and/or stricturing (IP/S) disease after a median follow-up of 18 months. Both anti-OmpC (p < 0.0006) and anti-I2 (p < 0.003) were associated with IP/S disease. The frequency of IP/S disease increased with increasing number of immune responses (p trend = 0.002). The odds of developing IP/S disease were highest in patients positive for all four immune responses (OR (95% CI): 11 (1.5–80.4); p= 0.03). Pediatric CD patients positive for ≥1 immune response progressed to IP/S disease sooner after diagnosis as compared to those negative for all immune responses (p < 0.03).CONCLUSIONS:The presence and magnitude of immune responses to microbial antigens are significantly associated with more aggressive disease phenotypes among children with CD. This is the first study to prospectively demonstrate that the time to develop a disease complication in children is significantly faster in the presence of immune reactivity, thereby predicting disease progression to more aggressive disease phenotypes among pediatric CD patients.

Sero-reactivity to microbial components in Crohn's disease is associated with disease severity and progression, but not NOD2/CARD15 genotype

BACKGROUND AND AIMS:Antibodies directed against the porin protein C of Escherichia coli (anti-OmpC) and Pseudomonas fluorescens (anti-I2) have recently been described in Crohns disease (CD). Those directed against Saccharomyces cerevisiae (ASCA) and the perinuclear component of neutrophils (pANCA) have been more widely studied and may be of diagnostic importance. We aimed to assess the frequency of anti-OmpC, anti-I2, ASCA, and pANCA, in an independent Scottish CD cohort, establish phenotypic associations, and compare with a U.S. cohort.METHODS:One hundred and forty-two well-characterized CD patients (76 females, median age 39 yr (17–88)) were studied. CD was classified by the Vienna classification. Sera were assayed for anti-OmpC, anti-I2, ASCA, and pANCA. Allele specific primers were used for NOD2/CARD15 genotyping.RESULTS:Anti-OmpC, anti-I2, ASCA, and pANCA were present in sera from 37%, 52%, 39%, and 14% of CD patients, respectively. Multivariate analysis demonstrated independent associations of anti-OmpC to be progression of disease type (p= 0.005) and long disease duration (p= 0.002), and those of anti-I2 to be long disease duration (p= 0.002) and the need for surgery (p= 0.033). ASCA were associated with disease progression (p < 0.001). When the presence and magnitude of all antibody responses were considered, reactivity to microbial components was associated with long disease duration (p < 0.001), progression of disease type (p < 0.001), penetrating disease (p= 0.008), small bowel disease (p < 0.02), and the need for surgery (p < 0.001). There was no association of antibody status to NOD2/CARD15 genotype.CONCLUSION:Reactivity to microbial components is associated with severe CD characterized by small bowel involvement, frequent disease progression, longer disease duration, and greater need for intestinal surgery.

TL1A (TNFSF15) Regulates the Development of Chronic Colitis by Modulating Both T-Helper 1 and T-Helper 17 Activation

BACKGROUND & AIMS TL1A is a tumor necrosis factor-like molecule that mediates a strong costimulation of T-helper (T(H)) 1 cells. Expression of TL1A is increased in the mucosa of Crohns disease patients and murine models of ileitis. The aim of this study was to determine the possible role of TL1A in chronic intestinal inflammation. METHODS We used dextran sodium sulfate (DSS)-induced chronic colitis to investigate the effects of TL1A on the development of colitis. The cytokine profile in the gut-associated lymphoid tissue (GALT) was measured. Neutralizing anti-TL1A antibodies were injected intraperitoneally into DSS-induced chronic colitis and G protein alphai2(-/-) T-cell transfer colitis models. Severity of colitis was evaluated by body weight, colon length, histology, and cytokine production. RESULTS DSS-induced chronic colitis was characterized by the infiltration of CD4(+) T cells. TL1A, death receptor 3, interferon (IFN)-gamma, and interleukin (IL)-17 were increased significantly in GALT of DSS-treated mice. TL1A up-regulated both IFN-gamma production from T(H)1 cells and IL-17 production from T(H)17 cells in GALT CD4(+) T cells. Furthermore, IFN-gamma and IL-17 production from CD4(+) T cells, induced by IL-12 and IL-23 respectively, was enhanced synergistically by combination with TL1A. Anti-TL1A antibody prevented chronic colitis and attenuated established colitis by down-regulation of both T(H)1 and T(H)17 activation. CONCLUSIONS Our results reveal that TL1A is an important modulator in the development of chronic mucosal inflammation by enhancing T(H)1 and T(H)17 effector functions. The central role of TL1A represents an attractive, novel therapeutic target for the treatment of Crohns disease patients.

Increased Immune Reactivity Predicts Aggressive Complicating Crohn’s disease in Children

BACKGROUND & AIMS The ability to identify children with CD who are at highest risk for rapid progression from uncomplicated to complicated phenotypes would be invaluable in guiding initial therapy. The aims of this study were to determine whether immune responses and/or CARD15 variants are associated with complicated disease phenotypes and predict disease progression. METHODS Sera were collected from 796 pediatric CD cases and tested for anti-Cbir1 (flagellin), anti-outer membrane protein C, anti-Saccharomyces cerevisiae, and perinuclear antineutrophil cytoplasmic antibody by using enzyme-linked immunosorbent assay. Genotyping (Taqman MGB) was performed for 3 CARD15 variants (single nucleotide polymorphisms 8, 12, and 13). Associations between immune responses (antibody sum and quartile sum score, CARD15, and clinical phenotype were evaluated. RESULTS Thirty-two percent of patients developed at least 1 disease complication within a median of 32 months, and 18% underwent surgery. The frequency of internal penetrating, stricturing, and surgery significantly increased (P trend < .0001 for all 3 outcomes) with increasing antibody sum and quartile sum score. Nine percent of seropositive groups had internal penetrating/stricturing versus 2.9% in the seronegative group (P = .01). Twelve percent of seropositive groups underwent surgery versus 2% in the seronegative group (P = .0001). The highest antibody sum group (3) and quartile sum score group (4) demonstrated the most rapid disease progression (P < .0001). Increased hazard ratio was observed for antibody sum group 3 (7.8; confidence interval, 2.2-28.7), P < .002 and quartile sum score group 4 (11.0; confidence interval, 1.5-83.0, P < .02). CONCLUSIONS The rate of complicated CD increases in children as the number and magnitude of immune reactivity increase. Disease progression is significantly faster in children expressing immune reactivity.

Neutrophil autoantibodies in ulcerative colitis: Familial aggregation and genetic heterogeneity

The possibility that the neutrophil autoantibodies associated with ulcerative colitis represent a genetic marker of susceptibility was investigated by determining their prevalence in unaffected relatives of patients. Neutrophil autoantibodies were detected using an enzyme-linked immunosorbent assay, and positive values were confirmed by indirect immunofluorescence. An increased prevalence of neutrophil antibodies was found not only in the probands (68%, 26/38) but also in their clinically unaffected family members (15.7%, 17/108) compared with controls (2.9%, 1/35) (P less than 0.0001 and P less than 0.05, respectively). These results were confirmed with sera from a second center, where 86.4% (19/22) of probands were positive and 20.9% (9/43) of their relatives were positive. The prevalence of neutrophil autoantibodies in the relatives of probands who were antibody positive (21.4%) was significantly greater than the prevalence in relatives of probands who were antibody negative (7%; P less than 0.05). The findings are consistent with these antibodies being a potential marker of genetic susceptibility to ulcerative colitis and suggest the possibility of genetic heterogeneity within this disease.

High-Titer Antineutrophil Cytoplasmic Antibodies in Type-1 Autoimmune Hepatitis

BACKGROUND/AIMS Autoimmune hepatitis (AIH), an unresolving liver inflammation characterized by periportal hepatitis and presence of serum autoantibodies, is distinguished by smooth muscle antibody and/or antinuclear antibody seropositivity. Type-2 AIH is characterized by antibodies to liver/kidney microsome type-1. Antineutrophil cytoplasmic antibodies (ANCAs) are highly specific for ulcerative colitis, primary sclerosing cholangitis, and, in this report, type-1 AIH determined by positive enzyme-linked immunosorbent assay confirmed by perinuclear indirect immunofluorescence staining. The studies described characterize the frequency and nature of ANCA in severe type-1 AIH patients and define AIH-ANCA specificity and diagnostic significance. METHODS One hundred four patients, characterized by clinical, immunoserological, virological, histological, and antigenic criteria, were studied. ANCA expression was assayed by enzyme-linked immunosorbent assay. RESULTS High-titer ANCA is present in 96% of patients with type-1 AIH with 92% showing a perinuclear staining pattern (pANCA). Whereas many patients were seropositive for antinuclear antibodies, the titer of ANCA did not correlate with the titer of antinuclear antibodies. ANCA expression did not correlate with the presence or absence of other autoimmune disorders. Finally, 80% of patients with AIH examined expressed only immunoglobulin G1 pANCA contrasting with the 33% of patients with PSC with pANCA subclass specificity. CONCLUSIONS The presence of ANCA seems to be an independent and selective marker for type-1 AIH.

Ulcerative colitis: a genetically heterogeneous disorder defined by genetic (HLA class II) and subclinical (antineutrophil cytoplasmic antibodies) markers.

Newly described distinct associations of HLA class II genes with ulcerative colitis (UC) (DR2) and Crohns disease (CD) (DR1/DQ5) provide strong evidence for genetic heterogeneity of susceptibility between these two forms of inflammatory bowel disease. A familial distribution of antineutrophil cytoplasmic antibodies (ANCAs, a subclinical marker of UC) in UC families has further implied the existence of heterogeneity within UC. To test the hypothesis that the heterogeneity within UC indicated by ANCAs has a genetic basis that resides within the HLA region, we studied 89 UC cases and an ethnically matched control group (n = 50). Serological and molecular typing techniques were applied to define HLA class II genes (DR, DQ). ANCAs were detected using an enzyme-linked immunosorbent assay, and positive values were confirmed by indirect immunofluorescence. We observed that ANCA-positive UC patients (n = 70) had a significantly increased frequency of DR2 compared with ANCA-negative controls (n = 46) (44% vs 22%, P = 0.01). In contrast, the frequency of DR2 in ANCA-negative UC cases (21%) was virtually identical to that in controls (22%, P = 0.9). Furthermore, the ANCA-negative UC patients had an increase in the DR4 allele compared with ANCA-positive UC (P = 0.004). Thus, with the combination of a subclinical marker (ANCAs) and molecular genetic markers, genetic heterogeneity has been demonstrated within UC: ANCA-positive UC associated with DR2, and ANCA-negative UC likely associated with DR4.

Association of Antineutrophil Cytoplasmic Antibodies With Resistance to Treatment of Left-Sided Ulcerative Colitis: Results of a Pilot Study

OBJECTIVE To determine the frequency of antineutrophil cytoplasmic antibodies with perinuclear staining in patients with treatment-resistant left-sided ulcerative colitis. METHODS We studied four groups: treatment-resistant left-sided ulcerative colitis, treatment-responsive left-sided ulcerative colitis, ulcerative colitis historical controls, and healthy control subjects. Antineutrophil cytoplasmic antibodies were detected by enzyme-linked immunosorbent assay, and positive results were confirmed by demonstration of a perinuclear staining pattern by indirect immunofluorescence assay. RESULTS The frequency of perinuclear antineutrophil cytoplasmic antibodies was significantly greater in treatment-resistant left-sided ulcerative colitis (90%) than in treatment-responsive left-sided ulcerative colitis (62%) (P = 0.03) or in ulcerative colitis historical controls (60%). CONCLUSION The increased frequency of perinuclear antineutrophil cytoplasmic antibodies in treatment-resistant left-sided ulcerative colitis suggests a possible association between these antibodies and relative resistance to medical therapy in patients with ulcerative colitis.