Hemoglobin and hematocrit levels in the prediction of complicated Crohn's disease behavior--a cohort study.

BACKGROUND: Markers that predict the occurrence of a complicated disease behavior in patients with Crohn's disease (CD) can permit a more aggressive therapeutic regimen for patients at risk. The aim of this cohort study was to test the blood levels of hemoglobin (Hgb) and hematocrit (Hct) for the prediction of complicated CD behavior and CD related surgery in an adult patient population. METHODS: Blood samples of 62 CD patients of the German Inflammatory Bowel Disease-network "Kompetenznetz CED" were tested for the levels of Hgb and Hct prior to the occurrence of complicated disease behavior or CD related surgery. The relation of these markers and clinical events was studied using Kaplan-Meier survival analysis and adjusted COX-proportional hazard regression models. RESULTS: The median follow-up time was 55.8 months. Of the 62 CD patients without any previous complication or surgery 34% developed a complication and/or underwent CD related surgery. Low Hgb or Hct levels were independent predictors of a shorter time to occurrence of the first complication or CD related surgery. This was true for early as well as late occurring complications. Stable low Hgb or Hct during serial follow-up measurements had a higher frequency of complications compared to patients with a stable normal Hgb or Hct, respectively. CONCLUSIONS: Determination of Hgb or Hct in complication and surgery naïve CD patients might serve as an additional tool for the prediction of complicated disease behavior.

Interferon-γ alters downstream signaling originating from epidermal growth factor receptor in intestinal epithelial cells: functional consequences for ion transport.

The epidermal growth factor receptor (EGFr) regulates many cellular functions, such as proliferation, apoptosis, and ion transport. Our aim was to investigate whether long term treatment with interferon-γ (IFN-γ) modulates EGF activation of downstream signaling pathways in intestinal epithelial cells and if this contributes to dysregulation of epithelial ion transport in inflammation. Polarized monolayers of T(84) and HT29/cl.19A colonocytes were preincubated with IFN-γ prior to stimulation with EGF. Basolateral potassium transport was studied in Ussing chambers. We also studied inflamed colonic mucosae from C57BL/6 mice treated with dextran sulfate sodium or mdr1a knock-out mice and controls. IFN-γ increased intestinal epithelial EGFr expression without increasing its phosphorylation. Conversely, IFN-γ caused a significant decrease in EGF-stimulated phosphorylation of specific EGFr tyrosine residues and activation of ERK but not Akt-1. In IFNγ-pretreated cells, the inhibitory effect of EGF on carbachol-stimulated K(+) channel activity was lost. In inflamed colonic tissues, EGFr expression was significantly increased, whereas ERK phosphorylation was reduced. Thus, although it up-regulates EGFr expression, IFN-γ causes defective EGFr activation in colonic epithelial cells via reduced phosphorylation of specific EGFr tyrosine residues. This probably accounts for altered downstream signaling consequences. These observations were corroborated in the setting of colitis. IFN-γ also abrogates the ability of EGF to inhibit carbachol-stimulated basolateral K(+) currents. Our data suggest that, in the setting of inflammation, the biological effect of EGF, including the inhibitory effect of EGF on Ca(2+)-dependent ion transport, is altered, perhaps contributing to diarrheal and other symptoms in vivo.

C1q/TNF-related protein-3 (CTRP-3) is secreted by visceral adipose tissue and exerts antiinflammatory and antifibrotic effects in primary human colonic fibroblasts.

BACKGROUND: The adipokine CTRP-3 (C1q/TNF-related protein-3) belongs to the C1q/TNF-related protein family which antagonizes the effects of lipopolysaccharide (LPS). The aim was to investigate the antiinflammatory and antifibrotic role of CTRP-3 in Crohn's disease (CD). METHODS: Mesenteric adipose tissue (MAT) of patients with CD or colonic cancer (CC) was resected. Human primary colonic lamina propria fibroblasts (CLPF) were isolated from controls and CD patients. Concentrations of chemokines and cytokines in the supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Expression of connective tissue growth factor (CTGF), collagen I, and collagen III was analyzed by real-time polymerase chain reaction (PCR). Recombinant CTRP-3 expressed in insect cells was used for stimulation experiments. RESULTS: CTRP-3 is synthesized and secreted by MAT resected from patients with CD, ulcerative colitis (UC), CC, and sigma diverticulitis as well as by murine and human mature adipocytes. CTRP-3 had no effect on the basal secretion of MCSF, MIF, or RANTES in MAT of CD and control patients. LPS-stimulation (10 ng/mL) significantly increased IL-8 release in CLPF of CD patients and, to a lesser extent, in cells of controls and of fibrotic CD tissue. CTRP-3 significantly and dose-dependently reduced LPS-induced IL-8 secretion in CLPF within 8 hours after LPS exposure, whereas LPS-induced IL-6 and TNF release was not affected. CTRP-3 inhibited TGF-ß production and the expression of CTGF and collagen I in CLPF, whereas collagen III expression remained unchanged. CONCLUSIONS: CTRP-3 exerts potent antiinflammatory and antifibrotic effects in CLPF by antagonizing the LPS pathway and by targeting the TGF-ß-CTGF-collagen I pathway.

Protection of epithelial barrier function by the Crohn's disease associated gene protein tyrosine phosphatase n2.

BACKGROUND & AIMS: Protein tyrosine phosphatase N2 (PTPN2) has been identified as a Crohn's disease (CD) candidate gene. However, a role for PTPN2 in the pathogenesis of CD has not been identified. Increased permeability of the intestinal epithelium is believed to contribute prominently to CD. The aim of this study was to determine a possible role for PTPN2 in CD pathogenesis. METHODS: Intestinal epithelial cell (IEC) lines T(84) and HT29cl.19a were used in all studies. Protein analysis was performed by Western blotting, and protein knockdown was induced by small interfering RNA. Primary samples were from control and CD patients. RESULTS: Here, we demonstrate increased PTPN2 expression in CD intestinal biopsy specimens and that the proinflammatory cytokine interferon (IFN)-gamma increases PTPN2 expression and activity in IEC. Moreover, IFN-gamma-induced STAT1 and STAT3 phosphorylation in IEC is enhanced by PTPN2 knockdown. The cellular energy sensor adenosine monophosphate-activated protein kinase partially regulates the IFN-gamma-induced effects on PTPN2. Additionally, PTPN2 knockdown potentiates IFN-gamma-induced increases in epithelial permeability, accompanied by elevated expression of the pore-forming protein claudin-2. CONCLUSIONS: PTPN2 is activated by IFN-gamma and limits IFN-gamma-induced signalling and consequent barrier defects. These data suggest a functional role for PTPN2 in maintaining the intestinal epithelial barrier and in the pathophysiology of CD.

AMP-activated protein kinase mediates the interferon-gamma-induced decrease in intestinal epithelial barrier function.

Impaired epithelial barrier function plays a crucial role in the pathogenesis of inflammatory bowel disease. Elevated levels of the pro-inflammatory cytokine, interferon-gamma (IFNgamma), are believed to be prominently involved in the pathogenesis of Crohn disease. Treatment of T(84) intestinal epithelial cells with IFNgamma severely impairs their barrier properties measured as transepithelial electrical resistance (TER) or permeability and reduces the expression of tight junction proteins such as occludin and zonula occludens-1 (ZO-1). However, little is known about the signaling events that are involved. The cellular energy sensor, AMP-activated protein kinase (AMPK), is activated in response to cellular stress, as occurs during inflammation. The aim of this study was to investigate a possible role for AMPK in mediating IFNgamma-induced effects on the intestinal epithelial barrier. We found that IFNgamma activates AMPK by phosphorylation, independent of intracellular energy levels. Inhibition of AMPK prevents, at least in part, the IFNgamma-induced decrease in TER. Furthermore, AMPK knockdown prevented the increased epithelial permeability, the decreased TER, and the decrease in occludin and ZO-1 caused by IFNgamma treatment of T(84) cells. However, AMPK activity alone was not sufficient to cause alterations in epithelial barrier function. These data show a novel role for AMPK, in concert with other signals induced by IFNgamma, in mediating reduced epithelial barrier function in a cell model of chronic intestinal inflammation. These findings may implicate AMPK in the pathogenesis of chronic intestinal inflammatory conditions, such as inflammatory bowel disease.

Secretion of RANTES (CCL5) and interleukin-10 from mesenteric adipose tissue and from creeping fat in Crohn's disease: regulation by steroid treatment.

BACKGROUND: Creeping fat represents a characteristic feature of Crohn's disease (CD) and adipose tissue is currently being recognized as a complex compartment secreting highly active molecules. Pro- or anti-inflammatory adipose tissue-derived secretory products (adipocytokines) might play a role in the pathogenesis of CD. METHODS: Adipose tissue specimens were obtained from creeping fat contiguous to the involved intestine of 10 patients with CD. Mesenteric adipose tissue specimens resected in 13 patients with colon cancer (CC) and in seven patients with diverticulitis (DIV) served as controls. Three fat tissue specimens per well and n = 6-8 wells per patient were incubated ex vivo for 24 h. The release of regulated on activation, T-cell expressed and secreted (RANTES) and interleukin (IL)-10 into the supernatant was measured by ELISA. RESULTS: Both RANTES and IL-10 secretion could be demonstrated from total adipose tissue explants. The RANTES secretion is increased from creeping fat in CD (3691 +/- 597 pg/g fat per 24 h) when compared to mesenteric adipose tissue from patients with CC (1690 +/- 191 pg/g fat per 24 h; P < 0.0001) or DIV (1672 +/- 336 pg/g fat per 24 h; P < 0.0001). In contrast, IL-10 secretion is downregulated significantly only in patients with DIV (1418 +/- 180 pg/g fat per 24 h; P = 0.016) when compared to CC patients (2368 +/- 259 pg/g fat per 24 h). Crohn's disease patients receiving steroids had a higher secretion rate of RANTES and IL-10. CONCLUSIONS: Both RANTES and IL-10 secretion can be detected from mesenteric adipose tissue and from creeping fat. The elevated RANTES and IL-10 secretion from creeping fat in CD is not due to a CD-specific effect but caused by steroid treatment.

Vascular endothelial growth factor secretion from mesenteric adipose tissue and from creeping fat in Crohn's disease.

BACKGROUND: Creeping fat represents a characteristic feature of Crohn's disease (CD), and adipose tissue secretes adipocytokines and chemokines/growth factors such as vascular endothelial growth factor (VEGF). Because VEGF serum levels and mucosal VEGF expression is elevated in CD patients, the aim of the present paper was to investigate creeping fat-derived VEGF secretion in CD. MATERIAL AND METHODS: Adipose tissue was obtained from creeping fat of 10 patients with CD. Mesenteric adipose tissue was resected from 13 patients with colon cancer (CC) and from seven patients with diverticulitis (DIV). Three fat tissue specimens per well, and several wells (6-8) per patient were incubated ex vivo for 24 h. The release of VEGF into the supernatant was measured by ELISA. RESULTS: There was stable VEGF secretion from mesenteric adipose tissue of patients with CC or DIV and from creeping fat of patients with CD. Whereas the VEGF secretion rate was not different between patients with CD (465 +/- 98 pg/g fat per 24 h) and CC (399 +/- 48 pg/g fat per 24 h), VEGF secretion was significantly reduced in patients suffering from DIV (115 +/- 41 pg/g fat per 24 h; P < 0.0001 and P = 0.001, respectively). The CD patients treated with steroids had significantly lower VEGF secretion rates (294 +/- 42 pg/g fat per 24 h) than CD patients not receiving steroids (607 +/- 105 pg/g fat per 24 h; P = 0.001). CONCLUSIONS: Creeping fat is an important source of VEGF secretion. The characteristics of the inflammatory changes in CD might be due to the lack of VEGF downregulation that is seen in DIV.

Profiling adipocytokine secretion from creeping fat in Crohn's disease.

BACKGROUND: Adipose tissue is recognized as a compartment secreting highly active molecules. Creeping fat represents a characteristic feature of Crohn's disease (CD). Proinflammatory or anti-inflammatory adipose-derived secretory products, now generally called adipocytokines, may play a role in the pathogenesis of CD. MATERIALS AND METHODS: Adipose tissue specimens were obtained from creeping fat contiguous to the involved intestine of 10 patients with CD. Mesenteric adipose tissue specimens resected from 13 patients with colon cancer (CC) and from 7 patients with diverticulitis served as controls. Three fat tissue specimen per well and 6 to 8 wells per patient were incubated ex vivo for 24 h. The release of adiponectin, resistin, leptin, interleukin-6, macrophage colony-stimulating factor, monocyte chemotactic protein-1, and migration inhibitory factor was measured by ELISA. The expression of AdipoR1 and AdipoR2 receptors was investigated by real-time quantitative polymerase chain reaction in a subset of adipose tissues. RESULTS: The secretion of adiponectin and macrophage colony-stimulating factor, as well as leptin and migration inhibitory factor, was significantly upregulated in CD compared with CC and diverticulitis or CC only, respectively. Resistin, interleukin-6, and monocyte chemotactic protein-1 were not specifically induced in CD but were associated with unspecific inflammation. Adiponectin receptor expression was not different in CC, CD, or diverticulitis. Steroid treatment in CD patients had differential effects on the ex vivo secretion of adipocytokines. CONCLUSIONS: A specific secretion pattern of proinflammatory and anti-inflammatory adipocytokines indicates the significance of adipose tissue in intestinal inflammation in CD. Future investigations of this intestinal compartment may provide novel insights into the pathophysiological role of creeping fat and CD.

The potential of adiponectin in driving arthritis.

Articular adipose tissue is a ubiquitous component of human joints, but its local functions are largely unknown. Because recent studies revealed several links between adipose tissue, adipocytokines, and arthritis, we investigated the expression of the adipocytokine adiponectin and its functional role in articular adipose tissue and synovium of patients with different arthritides. In contrast to its protective role in endocrinological and vascular diseases, adiponectin was found to be involved in key pathways of inflammation and matrix degradation in the human joint. The effects of adiponectin in human synovial fibroblasts appear to be highly selective by inducing only two of the main mediators of rheumatoid arthritis pathophysiology, IL-6 and matrix metalloproteinase-1, via the p38 MAPK pathway. Owing to the observation that these effects could be inhibited by different TNF-alpha inhibitors, adipocytokines such as adiponectin may also be key targets for therapeutic strategies in inflammatory joint diseases. In summary, articular adipose tissue and adipocytokines cannot be regarded as innocent bystanders any more in chronic inflammatory diseases such as arthritis.