Peripheral blood based discrimination of ulcerative colitis and Crohn's disease from non-IBD colitis by genome-wide gene expression profiling.

A molecular diagnostic assay using easily accessible peripheral blood would greatly assist in the screening and diagnosis of ulcerative colitis (UC) and Crohn's disease (CD). Transcriptional profiles in blood/biopsy samples from 12 UC (6/12), 9 CD (5/9), 6 non-inflammatory bowel disease (non-IBD) colitis (6/0), and 11 healthy (11/11) patients were assessed by Affymetrix HGU133Plus2.0 microarrays. Prediction analysis of microarrays, discriminant and ROC analyses were performed, the results were validated by RT-PCR and immunohistochemistry using also an independent set of samples (15 blood samples, 45 biopsies). A set of 13 transcripts was differentially expressed in IBD, non-IBD controls and healthy blood samples (100% specificity and sensitivity). Validated difference was found in 16 transcripts between UC, non-IBD and normal blood, and 4 transcripts between CD, non-IBD and normal samples. UC and CD blood cases could be also distinguished by 5 genes with 100% specificity and sensitivity. Some disease associated alterations in blood transcripts were also detected in colonic tissue. IBD subtypes may be discriminated from non-IBD (diverticulitis, infective and ischemic colitis) in vitro from peripheral blood by screening for differential gene expression revealed in this study. Transcriptional profile alterations in peripheral blood can be located in diseased colon.

Elevated insulin-like growth factor 1 receptor, hepatocyte growth factor receptor and telomerase protein expression in mild ulcerative colitis.

OBJECTIVE: The risk of development of colorectal carcinoma is elevated in chronic, long-standing ulcerative colitis (UC). The changes in regenerative and immortalizing pathways caused by the inflammatory process, and that have been proved to be carcinogenic in other human tissues, have not been fully and uniformly described. We assayed the expression alterations of regenerative signal receptors and cell-aging inhibitory systems within colonic crypts by considering the histological activity of the disease. METHODS: I-type insulin-like growth factor receptor (IGF1R), hepatocyte growth factor receptor (HGFR), telomerase reverse transcriptase (TERT) and telomerase associated protein (TP-1) expression were evaluated immunohistochemically on biopsy specimens from 11 mild, 11 moderate and 12 severe active inflammation of UC cases and from 10 normal colonic tissue cases. Independent colonic biopsies from 5 healthy and 7 severe UC cases were used for TaqMan real-time RT-PCR validation. RESULTS: In mild inflammation, all observed parameters showed significantly elevated epithelial protein expression (IGF1R: 22.3 +/- 9.46%; HGFR: 35.3 +/- 22.8%; TERT/TP-1: 2.1 +/- 1.87%/2 +/- 1.32%) compared to normal (p < 0.005). In moderately active inflammation, only IGF1R expression was significantly higher (50.2 +/- 8.6%) compared to normal and mild inflammation (p < 0.005). In severe inflammation, all parameters showed decreased epithelial expression; IGF1R showed decreased mRNA expression, while HGFR was overexpressed and TERT showed a decreased tendency. CONCLUSIONS: The epithelial expression of IGF1R, HGFR and TERT/TP-1 is elevated in mildly active UC. This phenomenon may allow the epithelial cells that collected genetic defects during severe inflammatory episodes pathologically to survive and proliferate.

[Endoscopy in pregnancy]. / Endoszkópos vizsgálatok terhességben.

Assessing the need of endoscopy and performing the examination is a difficult task which requires an experienced investigator. Though it is prudent to postpone the investigation to the third trimester or rather to the postpartum period; in certain clinical situations like upper gastrointestinal bleeding or biliary pancreatitis it is not possible; endoscopic intervention has to be performed in these cases just like in non-pregnant patients. There is a paucity of data in medical literature to rely on and the retrospective nature of these data makes it even more problematic. There is not more than one such a case per year per endoscopist in Hungary which requires decision in these situations; we have to think of the factors endangering the fetus but we have to keep in mind the factors that ensure the well-being of the mother by all means.

[Insulin-like growth factor receptor, hepatocyte-derived growth factor receptor and telomerase expression in ulcerative colitis]. / Inzulinszeru növekedési faktor receptor, hepatocytaeredetu növekedési faktor receptor és telomeráz fehérje expresszió colitis ulcerosában.

BACKGROUND: The risk of colorectal carcinoma development is elevated in chronic, longstanding ulcerative colitis. The changes of such regenerative and immortalizing pathways caused by the inflammatory process that are proved to be carcinogenic in other human tissues have not been fully and uniformly described. Aim of the study was to describe the expression alterations of regenerative signal receptors and cell aging inhibitory systems within colonic crypts considering the histological activity of the disease. MATERIALS AND METHODS: I-type insulin-like growth factor receptor (IGF1R), hepatocyte derived growth factor receptor (HGFR), telomerase reverse transcriptase (TERT) and telomerase associated protein (TP-1) expression were evaluated immunohistochemically on formalin fixed paraffin embedded biopsy specimen from 10 mild, 10 moderate, and 10 severe active inflammation of ulcerative colitis and from 10 normal colonic tissue. RESULTS: In mild inflammation all observed parameter showed significantly elevated protein expression in protein level compared to normal (p <0.005). In moderately active inflammation only IGF1R expression was significantly higher compared to normal and to mild inflammation (p <0.005). There were no regenerative signal expression alteration in severe inflammation compared to normal, and epithelial telomerase expression was not detectable in these cases. CONCLUSION: The expression of regenerative signal receptors and immortalizing factors is elevated in mildly (and moderately) inflamed ulcerative colitis. This phenomenon let the genetically defected epithelial cells to pathologically survive and proliferate, so thus favours the development of tumors.

[Association of coeliac disease and myasthenia gravis]. / Coeliakia es myasthenia gravis együttes elofordulása.

Coeliac disease is a malabsorption syndrome induced by the gluten-containing cereals leading to the damage of small intestinal mucosa. A case of a young woman is presented whose coeliac disease became overt in the puerperium. Despite strict adherence to gluten-free diet, her muscle weakness affecting several muscle regions persisted. Following a diagnostic work-up covering numerous co-disciplines, the diagnosis of myasthenia gravis was set up underlying her muscle symptoms. After the thorough review of the corresponding literature, the authors could not find any case report on the association of coeliac disease with myasthenia gravis.

Infliximab therapy improves the bone metabolism in fistulizing Crohn's disease.

BACKGROUND: Tumor necrosis factor-alpha (TNF-alpha) plays a pivotal role in the pathogenesis of inflammatory bowel diseases and bone resorption as well. Limited data exist about the effect of anti-TNF-alpha infliximab on bone metabolism in inflammatory-type Crohn's disease (CD). AIM: Our aim was to evaluate the effect of infliximab treatment on rapid changes of bone metabolism in fistulizing CD patients. METHODS: 27 patients with fistulizing CD were treated with three series of infliximab. Serum osteocalcin (OC) and beta-CrossLaps (bCL) were measured before administration of each infliximab infusion. 54 patients with inactive CD were controls. RESULTS: In treated patients, there were significant differences in bCL concentrations on days 0 and 14 (p < 0.01) and days 0 and 42 (p < 0.05). OC levels increased significantly between day 0 and 42 (p < 0.05). The values of bCL and OC of control groups differed from serum levels in active patients before the treatment, but not on day 42. Bone markers improved significantly in responder patients, but not in non-responders. CONCLUSION: The beneficial effect of infliximab to the bone metabolism is more expressive in patients whose fistulizing disease improves with this therapy. Our results suggest that TNF-alpha has an important role in the alteration of bone metabolism in fistulizing CD patients.

The half-and-half nail: a new sign of Crohn's disease? Report of four cases.

Systemic disease can produce changes in the nails. Perhaps the best known example of this is koilonychia as a sequale of iron deficiency anemia. "Half and half nail" is a type of pseudoleuconychia that can be caused by chronic renal disease, Kawasaki's disease, cirrhosis, and zinc deficiency. It has not been described in patients with Crohn's disease yet. Four male patients with Crohn's disease were observed. None of them had extraintestinal manifestations of Crohn's disease. The average duration of the disease was 5.25 (range, 1-10) years. The nail alterations were defined with a portion of the nail being as much as 15 to 40 percent of normal color distally, whereas the rest of the nail was white. The contrast between the two zones remains sharply demarcated even after constricting venous return. Histologic examination was not performed. Every patient had zinc deficiency but not hypalbuminaemia or sideropenia. After review of relevant literature (MEDLINE, PubMed, etc.), we found that half-and-half nail had not been described in Crohn's disease. This study was designed to highlight the fact that the half-and-half nail, which was thought to be a sign of chronic renal failure, also occurs in patients with Crohn's disease. The relationship of this symptom to clinical activity cannot yet be assessed and has only been observed in four cases.

Carcinoma arising in enterocutan fistulae of Crohn's disease patients: description of two cases.

Rarely, carcinoma arises from the fistulous tract of Crohn's disease. Adequate radiological examination often produces misleading pseudonegative findings. We reported two cases of fistula cancers treated with infliximab. The short time-span between the administration of this drug and the diagnosis of cancer makes the correlation between the two unlikely.

[Improvement of bone metabolism after infliximab therapy in Crohn's disease]. / Infliximabkezelés hatása a csontanyagcserére Crohn-betegségben.

BACKGROUND: Osteoporosis has received increasing attention as a potential complication of Crohn's disease. Among cytokines tumor necrosis factor-alpha plays a pivotal role in the pathogenesis of inflammatory bowel diseases by inducing a wide variety of inflammatory responses, including bone resorption. Only few data are present about the effect of infliximab on bone metabolism. AIMS: The authors evaluated the effect of infliximab on bone metabolism in patients with Crohn's disease. PATIENTS AND METHODS: Twenty seven patients (17 females, 10 males, mean age 32.58 yrs) with refractory fistulizing Crohn's disease were treated with a series of three infusions of 5 mg infliximab per kg at weeks 0, 2, and 6. Biochemical markers of bone formation (osteocalcin) and bone resorption (beta-CrossLaps) were measured before administration of each infliximab infusion. 54 patients were studied with inactive Crohn's disease (Crohn's disease activity index < 150) as a control. RESULTS: There were significant differences in beta-CrossLaps concentrations (ng/ml) between the day 0 and 14 (0.57 +/- 0.32 vs. 0.46 +/- 0.29, p < 0.01) and the day 0 and 42 (0.57 +/- 0.32 vs. 0.45 +/- 0.26, p < 0.05). The osteocalcin levels significantly increased from day 0 to day 42 (21.31 +/- 12.14 vs. 25.64 +/- 16.97, p < 0.05). The serum beta-CrossLaps and osteocalcin levels were 0.47 +/- 0.24, 27.2 +/- 8.44 in the control group respectively. These results differed from the serum levels of active patients before the treatment, but there were no notable differences at the day 42. CONCLUSION: Infliximab therapy in Crohn's disease patients displayed a rapid influence on bone metabolism by enhancing bone formation and decreasing bone resorption. In addition to its mucosal effect affecting the bone homeostasis, indicate a further rationale usage of tumor necrosis factor-alpha blockade in the therapy of inflammatory bowel diseases.

Growth in epithelial cell proliferation and apoptosis correlates specifically to the inflammation activity of inflammatory bowel diseases: ulcerative colitis shows specific p53- and EGFR expression alterations.

PURPOSE: Epithelial cell turnover related differences between ulcerative colitis, Crohn's colitis, and aspecific colitis are not known yet. METHODS: Totally 345 formalin-fixed, paraffin-embedded biopsy specimens from 33 ulcerative colitis, 26 Crohn's colitis, 30 aspecific colitis, and 10 healthy patients were observed with the TdT-mediated dUTP nick end labeling method and proliferating cell nuclear antigen-, p53-, and epithelial growth factor receptor immunohistochemistry. Because of epithelial growth factor receptor positivity of subepithelial cells epithelial growth factor receptor and CD45, CD68, or CD83 double fluorescence immunohistochemistry were performed on 16 freshly frozen samples from 8 severely active ulcerative colitis and 8 severely active Crohn's colitis patients to describe lamina propria's mononuclear cells, respectively. RESULTS: The epithelial growth factor receptor expression was significantly lower in each inflammatory group compared with normal (P < 0.005) and decreased significantly in mild ulcerative colitis compared with mild Crohn's colitis or aspecific colitis (P < 0.005). Numerous epithelial growth factor receptor and CD45 double-positive submucosal mononuclear cells were observed in moderate-severe inflammations. The p53-expression was significantly higher in each inflammatory group compared with normal (P < 0.05). Significant differences were found between mildly, moderately, and severely inflamed samples in ulcerative colitis (P < 0.05) compared with Crohn's colitis or aspecific colitis. Apoptotic/proliferative rates increased significantly in line with the inflammatory process (P < 0.0001/0.05), but the TdT-mediated dUTP nick end labeling and proliferating cell nuclear antigen-labeling characteristics did not show disease type specificity. CONCLUSIONS: Based on our results, the alterations of epithelial growth factor receptor and p53 expression show ulcerative colitis specificity, whereas the rate of epithelial apoptosis and proliferation are determined by the histologic activity of the inflammation. The increased epithelial growth factor receptor expression by the lamina propria's mononuclear cells in inflammation may suggest its role as an autoantigen.

Immediate DNA ploidy analysis of gastrointestinal biopsies taken by endoscopy using a mechanical dissociation device.

BACKGROUND: Quantitative DNA analysis of fresh biopsy material can contribute to a more accurate diagnosis and prognosis. The authors aimed to develop and test a mechanical, nuclear preparation protocol for quantitative DNA analysis. PATIENTS AND METHODS: Altogether 32 gastric (10 healthy, 17 gastritis, 7 adenocarcinoma) and 48 colon (21 healthy, 20 colitis ulcerosa, 7 adenocarcinoma) biopsy specimens were evaluated. The mechanical disruption was performed by Medimachine (DAKO, Denmark). The flow cytometry analysis was performed on a BD FACSStar flow cytometer. RESULTS: DNA Aneuploidy was found in gastric samples only in tumours. The S-phase fraction of the normal cases was 5.9 +/- 2.1%, 5.1% +/- 1.2% in gastritis and 10.7 +/- 1.6% in carcinomas. Seven out of 20 colitis ulcerosa and 4 out of 7 colon cancer samples were aneuploid. The S-phase fraction of normal colon cases was 5.7 +/- 3.4%, in colitis 8.1 +/- 4.2% and 15.1 +/- 5.7% in carcinomas, respectively. CONCLUSION: Mechanical nuclear isolation is a useful method for flow cytometric DNA ploidy analysis of fresh biopsy samples.

Mutant p53 expression and apoptotic activity of Helicobacter pylori positive and negative gastritis in correlation with the presence of intestinal metaplasia.

BACKGROUND: Mutation of the p53 gene is detectable in most cases of gastric cancer, as it is the most common genetic alteration in human malignancies. It is also well documented that Helicobacter pylori infection plays an important role in gastric carcinogenesis. There is still no clarification, however, concerning how genetic instability influences the homeostasis of gastric epithelium. We have studied the effect of H. pylori infection on apoptosis of the antral epithelium in the presence/absence of intestinal metaplasia and the expression of the p53 oncoprotein. The relationship between these two processes is analysed. METHODS: Antral biopsies were taken from 36 patients who underwent routine upper endoscopy (17 men, 19 women, mean age 61.0 years). The biopsies were fixed in formalin and embedded in paraffin. Patients were classified into two histological groups: (1) as chronic gastritis without intestinal metaplasia (n = 19), and (2) chronic gastritis with intestinal metaplasia (n = 17). An immunohistochemical method was used to detect the expression of p53 oncoprotein, and the terminal transferase mediated dUTP nick end-labelling (TUNEL) method was used to detect apoptotic cells. RESULTS: In the absence of intestinal metaplasia, both the apoptotic index (0.0272 +/- 0.011 vs 0.0128 +/- 0.006) and expresssion of p53 (35.55 +/- 31.16 vs 18.33 +/- 19.65) were significantly higher in H. pylori positive cases compared to H. pylori negative cases. In the presence of intestinal metaplasia, p53 expression was further increased (P < 0.05), but apoptosis was similar to that observed in H. pylori negative gastritis without intestinal metaplasia. In the presence of intestinal metaplasia, H. pylori infection did not influence apoptosis (0.013 +/- 0.004 vs 0.011 +/- 0.004), or p53 ratio (70.16 +/- 22.54 vs 68.50 +/- 28.96). In the sequence of gastritis-intestinal metaplasia the two indices show a close negative correlation (P < 0.05). CONCLUSION: In the absence of intestinal metaplasia H. pylori infection increases both apoptotic activity and expression of p53 oncoprotein in the gastric mucosa. The lack of increased apoptosis with a higher p53 expression in the presence of intestinal metaplasia suggests an increased genetic instability and also may suggest that mutation of the p53 gene is an early step in the multistep process of gastric carcinogenesis.

[Changes in cell kinetics and clinical course in inflammatory bowel diseases]. / Sejtkinetikai változások és a klinikai kép összefüggése gyulladásos bélbetegségekben.

INTRODUCTION: The pathogenesis of inflammatory bowel diseases is still unknown, but is accessible from several ways. One possibility is the immunohistochemical analysis of cellular changes in the intestinal mucosa. The increased epithelial cell turnover is connected with false immunological pathways in the subepithelial layer. Behind these disturbances which can lead to chronic remitting inflammatory processes the imbalance of apoptosis and proliferation plays a key role. AIM: Of this study is to summarize our current knowledge of the cell kinetical alterations considering histological activity of disease. CONCLUSIONS: On the basis of current understanding it is known that the apoptosis and proliferation of epithelial cells increase in active inflammation compare to normal, although an uniform standpoint of evaluating is still missing. Some alterations of apoptosis and antigen presentation are described in the mononuclear cells of the subepithelial layer. Getting acquainted with and describing the changes of cell kinetics provide facilities to develop new and effective diagnostical methods and therapies.

[Changes in the proliferation and apoptosis of colonic epithelial cells in correlation with histologic activity of ulcerative colitis]. / A bélhámsejtek osztódásának és apoptózisának változása colitis ulcerosa szöveti aktivitásának függvényében.

BACKGROUND: The imbalance of epithelial cell kinetics in ulcerative colitis can lead to several gastrointestinal disorders such as ulcers or neoplasias. The changes of epithelial cell kinetics caused by the inflammatory process have not been fully and uniformly described. Aim of the study was to compare the rate of epithelial cell apoptosis and proliferation within colonic crypts considering the histological activity of ulcerative colitis. MATERIALS AND METHODS: Formalin fixed paraffin embedded biopsy specimens from mild, moderate, and severe active inflammation of ulcerative colitis and from normal colonic tissue were observed. Number of cases (apoptosis/proliferation): normal: 8/10; mild UC: 8/10; moderate: 8/8; severe: 12/8. For detecting apoptosis the TUNEL-POD-DAB method, and for proliferation the PCNA (PC-10)-Biotin-Streptavidin-AEC method were used, constrained with haematoxylin. Using light microscope, at least 500 crypt epithelial cells were encountered axially in whole, well-orientated colonic crypts per each specimen. RESULTS: The number of TUNEL positive epithelial cells were significantly higher in moderate and severe active inflammation compared to each other and to normal (p < 0.0001). Between mild inflammation and normal significant alteration was not found. The number of PCNA positive cells were significantly higher in each groups of inflammation compared to normal or each other. CONCLUSION: These results demonstrate that there is a strong correlation between the histological activity of ulcerative colitis and elevated crypt epithelial cell turnover, although the alterations in proliferation develop in the earlier stages of inflammation.