Isolation and characterization of a novel short peptide associated with Crohn's disease.

Phage display technology has been utilized to select target molecules against circulating antibodies. The aims of this study were to isolate a peptide that binds with serum from Crohn's disease (CD) patients and to examine its diagnostic and pathogenic significance. A phage display library was constructed using cDNA from Caco-2 cells. Affinity selection using this cDNA library and serum samples from patients with CD was then performed. Phage clones that specifically reacted with the CD sera were then selected using a phage enzyme-linked immunosorbent assay (ELISA). After the DNA sequences of the selected phages were determined and converted to amino acid sequences, the synthesized peptides were examined using an ELISA. The effect of the synthesized peptides on cytokine release from cultured blood mononuclear cells was investigated. An ELISA analysis for TCP-353 demonstrated that while 61·7% of the samples from CD patients were seroreactive, seroreactivity was less common among patients with ulcerative colitis (7·3%), acute colitis (0%) or colon cancer (11·4%) and among normal subjects (2·8%). The induction of interleukin (IL)-1ß, IL-6 and tumour necrosis factor (TNF)-α release, but not IL-10 release, in response to TCP-353 peptide was enhanced in CD mononuclear cells only. We isolated a novel peptide that specifically binds to CD sera and stimulates the proinflammatory responses of CD mononuclear cells. TCP-353 may have diagnostic, pathogenic and therapeutic significance with regard to the treatment of CD.

Pneumomediastinum is a frequent but minor complication during esophageal endoscopic submucosal dissection.

BACKGROUND AND STUDY AIM: Esophageal perforation caused by endoscopic submucosal dissection (ESD) induces serious pneumomediastinum. In the absence of endoscopically detected perforation, postprocedural pneumomediastinum may occur. The aim of this study was to evaluate the association between the clinical factors/courses and pneumomediastinum revealed by chest computed tomography (CT) with special reference to an exposed muscle layer during esophageal ESD. PATIENTS AND METHODS: A total of 58 patients undergoing ESD for esophageal neoplasms between February 2003 and June 2007 also underwent both chest radiography and chest CT within 1 hour after ESD. We studied the association between findings on CT scan and tumor-related and technical factors of esophageal ESD by uni- and multivariate analyses. We also analyzed the clinical factors/courses experienced by all patients. RESULTS: Pneumomediastinum was detected in 18 / 58 patients (31 %) by chest CT compared with only 1 / 58 patients (1.7 %) by chest radiography. ESD-induced exposure of the muscular layer (32 patients) was the only significant factor for pneumomediastinum (18 / 32; P < 0.0001). Clinical factors such as fever, white blood cell count, and C-reactive protein were significantly increased in the group positive for both endoscopically exposed muscular layer and pneumomediastinum (+/+, n = 18) compared with the (-/-) group (n = 26) in the early phase (day 1) after ESD. However, these factors did not affect the length of the fasting period or the length of hospital stay. CONCLUSIONS: In esophageal ESD, pneumomediastinum detected by chest CT only does not cause clinically significant complication. Endoscopic muscle exposure during ESD is a significant risk factor for pneumomediastinum, which causes mild inflammation in the early post-ESD phase.

Linkage of Crohn's disease-related serological phenotypes: NFKB1 haplotypes are associated with anti-CBir1 and ASCA, and show reduced NF-kappaB activation.

BACKGROUND AND AIMS: Genetics studies of the serum expression of antibodies to microbial antigens may yield important clues to the pathogenesis of Crohn's disease. Our aim was to conduct a linkage study using expression of anti-CBir1, anti-I2, anti-OmpC and ASCA as quantitative traits. METHODS: Expression of antibodies to microbial antigens was measured by enzyme-linked immunosorbant assay (ELISA) and a standard approximately 10 cM whole genome microsatellite study was conducted. Single nucleotide polymorphism genotyping was performed using either Illumina or TaqMan MGB technology. Nuclear factor Kappa B (NF-kappaB) activation in cells from Epstein-Barr virus (EBV)-transformed cell lines was assessed using an electrophoretic mobility shift assay and protein was measured using ELISA and western blotting. RESULTS: Evidence for linkage to anti-CBir1 expression was detected on human chromosome 4 (logarithm of odds (LOD) 1.82 at 91 cM). We therefore directly proceeded to test the association of haplotypes in NFKB1, a candidate gene. One haplotype, H1, was associated with anti-CBir1 (p = 0.003) and another, H3, was associated with ASCA (p = 0.023). Using cell lines from Crohn's disease patients with either H1 or H3, NF-kappaB activation and NF-kappaB p105 and p50 production were significantly lower for patients with H1 compared to patients with H3. CONCLUSIONS: These results suggest that NFKB1 haplotypes induce dysregulation of innate immune responses by altering NF-kappaB expression. The results also show the use of EBV-transformed lymphoblastoid cell lines to conduct phenotypic studies of genetic variation.

Polyarthritis nodosa with mesenteric aneurysms demonstrated by angiography: report of a case and successful treatment of the patient with prednisolone and cyclophosphamide.

Polyarteritis nodosa is a necrotizing angitis that predominantly affects small and medium-sized arteries. The prognosis of untreated polyarteritis nodosa is very poor. Since symptoms are diverse and no serologic test is specific for polyarteritis nodosa, the diagnosis is difficult and often delayed. We describe a patient with polyarteritis nodosa who had gastrointestinal involvement with multiple aneurysms of the inferior mesenteric artery; only abdominal angiography provided a conclusive diagnosis. Alleviation of symptoms and regression of aneurysms were observed after combination therapy of an immunosuppressive agent, cyclophosphamide, and prednisolone. We emphasize the importance of early diagnosis by angiography and aggressive therapy in patients in whom physical signs indicating definite polyarteritis nodosa are not present.

Clinical trial: comparison of alendronate and alfacalcidol in glucocorticoid-associated osteoporosis in patients with ulcerative colitis.

BACKGROUND: Bone loss is often observed in patients with ulcerative colitis, particularly if they require glucocorticoids. AIM: To determine whether the bisphosphonate, alendronate, is safe and effective in preserving bone mass compared to the active vitamin D3, alfacalcidol, in ulcerative colitis patients receiving glucocorticoids. METHODS: Thirty-nine patients with ulcerative colitis and treated with glucocorticoids were randomized to receive alendronate (5 mg/day) or alfacalcidol (1 microg/day) daily for 12 months. Loss of bone mass was evaluated by bone mineral density, bone resorption by urinary N-telopeptide for type I collagen, and bone formation by serum bone alkaline phosphatase. RESULTS: Alendronate, but not alfacalcidol, significantly increased bone mineral density in the lumbar spine. Alendronate decreased serum bone alkaline phosphatase levels, but alfacalcidol did not. Urinary N-telopeptide for type I collagen levels decreased in both groups, but were significantly lower in the alendronate group. There were no significant differences in the adverse events in the two groups. CONCLUSION: Our study indicates that alendronate is a safe, well-tolerated and more effective therapy than alfacalcidol for preventing glucocorticoid-associated bone loss in patients with ulcerative colitis.

A form of circulating interleukin-6 receptor component soluble gp130 as a potential interleukin-6 inhibitor in inflammatory bowel disease.

The presence and the role of soluble gp130, the soluble form of a component of the interleukin (IL)-6 receptor complex, were investigated in inflammatory bowel disease. The serum concentrations of soluble gp130 were increased in ulcerative colitis (active disease, median, 93.5 ng/ml; interquartile range, 26-125 ng/ml; inactive disease, 81 ng/ml, 24.8-137.3 ng/ml) and to a lesser extent in Crohn's disease (active disease, 66 ng/ml, 44.4-87.6 ng/ml; inactive disease, 63 ng/ml, 43.5-82.5 ng/ml) compared to normal controls (43 ng/ml, 27-59 ng/ml). Paired analysis of serum samples showed a decrease of IL-6 and soluble IL-6 receptor concentrations in both diseases and an increase of soluble gp130 concentrations, especially in ulcerative colitis, just after the resolution of disease exacerbation. Size fractionation of the serum revealed that a part of the IL-6 co-eluted with soluble gp130 and soluble IL-6 receptor. The IL-6-induced proliferation of murine B9 hybridoma was enhanced by recombinant soluble IL-6 receptor, whereas the proliferation was inhibited by recombinant soluble gp130. These results indicate that soluble gp130 may function as a natural inhibitor of the IL-6 actions in inflammatory bowel disease.

Diminished cytokine signalling against bacterial components in mononuclear leucocytes from ulcerative colitis patients after leukocytapheresis.

Infiltration by circulating inflammatory cells is a prominent local inflammatory feature of ulcerative colitis (UC). Several trials have suggested that leukocytapheresis by filtration can benefit patients with active UC. We investigated how this therapy might modulate the inflammatory response. Patients with active UC who were beginning repeated filtration leukocytapheresis were studied. Mononuclear cell preparations were obtained from blood before and after the first treatment, and expression of cytokine signalling components and the cell-proliferative response were analysed in vitro. Leukocytapheresis reduced lipopolysaccharide-induced production of proinflammatory cytokines (interleukin-1, -6, -8 and tumour necrosis factor-alpha, P < 0.05 for all) and activation of intracellular signalling components (nuclear factor-kappaB, mitogen-activated protein kinases, and signal transducer and activator of transcription-3), as well as surface expression of toll-like receptor-4 (P < 0.05) in mononuclear cells. The therapy also reduced the cell-proliferative response by mononuclear cells stimulated with sonicated bacterial preparations from autologous intestine (P < 0.05). These results indicate that activated mononuclear cells in the peripheral blood of patients with active UC are removed by leukocytapheresis and replaced by cells with a lower activation status. This replacement may partly explain the therapeutic benefit.

Heat shock cognate protein 70 encodes antigenic epitopes recognised by HLA-B4601-restricted cytotoxic T lymphocytes from cancer patients.

Heat shock cognate protein 70 (HSC70), a highly conserved protein and a member of the family of molecular chaperones, has the ability to induce cytotoxic T lymphocyte (CTL) responses through binding and carrying antigenic peptides. We demonstrated in this study that the HSC70 gene encodes two antigenic peptides recognised by HLA-B46-restricted and tumour-reactive CTLs established from tumour-infiltrating lymphocytes of a colon cancer. These HSC70-derived peptides, at amino-acid positions 106-114 and 233-241, had the ability to induce HLA-B46-restricted and peptide-specific CTLs, which are reactive to tumour cells, from peripheral blood mononuclear cells of the majority of epithelial cancer patients tested. These results, along with those from the previous studies, indicate the two ways of HSC70 involvement in the immune responses to tumours: chaperones and antigens, and thus may provide a new insight for the development of HSC70-directed cancer-specific immunotherapy.