(1/580) Protein kinase C mediates experimental colitis in the rat.
Protein kinase C (PKC) plays an important role in the cell signal transduction of many physiological processes. In contrast to these physiological responses, increases in PKC activity have also been associated with inflammatory disease states, including ulcerative colitis. The objective of this study was to examine the role of PKC as a causative mediator in initiation of experimentally induced colitis in the rat. Colitis was induced in rats by intrarectal (0.6 ml) instillation of 2,4,6-trinitrobenzenesulfonic acid (TNBS; 75 mg/kg in 50% ethanol) or the PKC activator phorbol 12-myristate 13-acetate (PMA; 1.5-3.0 mg/kg in 20% ethanol). Gross and histological mucosal damage, mucosal neutrophil infiltration, mucosal PKC activity, and PKC protein content for PKC isoforms alpha, beta, delta, and epsilon were assessed 2 h to 14 days after an inflammatory challenge. Both PKC activity and mucosal injury increased significantly within 4 h of TNBS treatment. PKC activity was maximal at 7 days and declined at 14 days, whereas mucosal damage became maximal at 1 day and declined after 7 days. In contrast, neutrophil infiltration as assessed by myeloperoxidase activity only increased 12 h after TNBS treatment, became maximal 1 day after TNBS administration, and declined thereafter. PKCbeta, -delta, and -epsilon were increased in response to TNBS, whereas PKCalpha protein content was decreased. The PKC antagonists staurosporine and GF-109203X (25 ng/kg iv) reduced TNBS-induced changes in mucosal PKC activity and the degree of mucosal damage. In contrast, neutropenia induced by antineutrophil serum treatment did not significantly affect the degree of injury or mucosal PKC activity. Furthermore, activation of mucosal PKC activity with PMA also induced mucosal damage, which was also inhibited by pretreatment with a PKC antagonist. In conclusion, these results suggest that increases in PKC activity play a causative role in TNBS-induced colitis. The PKC-mediated response to TNBS does not appear to involve neutrophil infiltration. (+info)
(2/580) Susceptibility of Lewis and Fischer rats to stress-induced worsening of TNB-colitis: protective role of brain CRF.
We assessed the role of central corticotropin-releasing factor (CRF) in stress-induced worsening of colitis in inbred rat strains with hypo (Lewis/N) and hyper (Fischer344/N) CRF responses to stress. Intracolonic administration of 2,4,6-trinitrobenzenesulfonic acid (TNB) induced colitis of similar severity in both strains as assessed on day 7 by macroscopic scoring, histological evaluation, tissue myeloperoxidase (MPO) activity, and decrease in food intake and body weight. Colitis was inhibited by daily intracerebroventricular injections of CRF in both strains. Chronic stress (3 h/day, water avoidance or wrap restraint on alternate days for 6 days) aggravated colitis more in Lewis than Fischer rats (71 and 22% further increase in MPO activity, respectively). The CRF antagonist astressin injected intracerebroventricularly enhanced the colitis response to stress and caused mortality in both strains. Fischer rats had higher plasma corticosterone levels 20 min after stress alone on day 1 and after TNB plus stress on days 1 and 3 compared with Lewis. These data show that central CRF restrains the proinflammatory action of stress in experimental colitis. (+info)
(3/580) Hapten-induced colitis is associated with colonic patch hypertrophy and T helper cell 2-type responses.
To investigate the potential involvement of T helper (Th)2-type responses in murine models of intestinal inflammation, we used trinitrobenzene sulfonic acid (TNBS)-hapten to induce inflammatory bowel disease in situations where Th1-type responses with interferon (IFN)-gamma synthesis are either diminished or do not occur. Intracolonic administration of TNBS to either normal (IFN-gamma+/+) or Th1-deficient IFN-gamma knockout (IFN-gamma-/-) BALB/c mice resulted in significant colitis. In IFN-gamma-/- mice, crypt inflammation was more severe than in IFN-gamma+/+ mice and was accompanied by hypertrophy of colonic patches with a lymphoepithelium containing M cells and distinct B and T cell zones resembling Peyer's patches. Hapten-specific, colonic patch T cells from both mouse groups exhibited a Th2 phenotype with interleukin (IL)-4 and IL-5 production. TNBS colitis in normal mice treated with anti-IL-4 antibodies or in IL-4(-/-) mice was less severe than in either IFN-gamma+/+ or IFN-gamma-/- mice. Our findings now show that the Th2-type responses in TNBS colitis are associated with colonic patch enlargement and inflammation of the mucosal layer and may represent a model for ulcerative colitis. (+info)
(4/580) Experimental colitis increases blood-brain barrier permeability in rabbits.
Extraintestinal manifestations of inflammatory bowel disease are numerous. This study examined the effects of two models of acute colitis on cerebral blood flow (CBF) and permeability of the blood-brain barrier in rabbits. CBF (measured with radiolabeled microspheres), or the extraction ratio or permeability-surface area (PS) product of the blood-brain barrier to fluorescein and FITC-dextran, was measured 48 h after colitis induction with acetic acid (HAc) or trinitrobenzene sulfonic acid (TNBS). PS product for fluorescein increased (P < 0.05) in TNBS colitis (1.33 x 10(-5) +/- 0.52 x 10(-5) ml/s and 0.48 x 10(-5) +/- 0.13 x 10(-5) ml/s (mean +/- SE) for treated (n = 14) and untreated (n = 10) animals, respectively. PS product for the larger FITC-dextran was not different in TNBS colitis (0.24 x 10(-5) +/- 0.09 x 10(-5) ml/s, n = 7) compared with untreated controls (0.19 x 10(-5) +/- 0.04 x 10(-5) ml/s, n = 8). PS product for fluorescein increased (P < 0.01) in HAc colitis compared with vehicle (2.66 x 10(-5) +/- 1.46 x 10(-5) ml/s and 0.33 x 10(-5) +/- 0.05 x 10(-5) ml/s, respectively; n = 6 in each group). The extraction of fluorescein from the blood to the brain increased by 75% during TNBS colitis when compared with vehicle (P < 0.05). CBF and cerebrovascular resistance did not change from the untreated control after TNBS colitis. Our data suggest that, irrespective of induction method, acute colitis increases the permeability of the blood-brain barrier to small molecules without changing CBF. (+info)
(5/580) Reduced oxidative and nitrosative damage in murine experimental colitis in the absence of inducible nitric oxide synthase.
BACKGROUND: Oxidative and nitrosative stress have been implicated in the pathogenesis of inflammatory bowel diseases. AIMS: To study the role of nitric oxide (NO) derived from inducible NO synthase (iNOS) in an experimental model of murine enterocolitis. METHODS: Trinitrobenzene sulphonic acid (TNBS) was instilled per rectum to induce a lethal colitis in iNOS deficient mice and in wild type controls. The distal colon was evaluated for histological evidence of inflammation, iNOS expression and activity, tyrosine nitration and malondialdehyde formation (as indexes of nitrosative and oxidative stress), myeloperoxidase activity (as index of neutrophil infiltration), and tissue localisation of intercellular adhesion molecule 1 (ICAM-1). RESULTS: TNBS administration induced a high mortality and weight loss associated with a severe colonic mucosal erosion and ulceration, increased myeloperoxidase activity, increased concentrations of malondialdehyde, and an intense staining for nitrotyrosine and ICAM-1 in wild type mice. Genetic ablation of iNOS gene conferred to mice a significant resistance to TNBS induced lethality and colonic damage, and notably reduced nitrotyrosine formation and concentrations of malondialdehyde; it did not, however, affect neutrophil infiltration and intestinal ICAM-1 expression in the injured tissue. CONCLUSION: Data show that activation of iNOS is required for nitrosative and oxidative damage in experimental colitis. (+info)
(6/580) Intestinal inflammation in adhesion molecule-deficient mice: an assessment of P-selectin alone and in combination with ICAM-1 or E-selectin.
Biopsy specimens from patients with inflammatory bowel disease have demonstrated an up-regulation of P-selectin, suggesting a role for P-selectin in intestinal inflammation. We examined the role of P-selectin in experimental intestinal inflammation using mice deficient in P-selectin alone or in combination with either ICAM-1 or E-selectin. Colitis was induced using acetic acid or trinitrobenzene sulfonic acid (TNBS). Damage scores and neutrophil infiltration 24 h post acetic acid were not different between wild-type and P-selectin- or P-selectin/ICAM-1-deficient mice, whereas P/E-selectin-deficient mice had enhanced leukocyte recruitment and damage. At 72 h an attenuation in damage scores and a slight decrease in neutrophil infiltration was observed in the P- and P/ICAM-deficient animals. The P/E-selectin-deficient mice maintained enhanced leukocyte recruitment and damage. In wild-type mice P-selectin expression was elevated 48 and 72 h post acetic acid-induced inflammation. Surprisingly, P-selectin or P-selectin/ICAM-1 deficiency did not improve the inflammation induced by TNBS over 7 days. In fact, increased mortality was observed. Anti-adhesion therapy may play only a limited, beneficial role and often a detrimental role in intestinal inflammation. (+info)
(7/580) Active site characterization of RNase Rs from Rhizopus stolonifer: involvement of histidine and lysine in catalysis and carboxylate in substrate binding.
Chemical modification studies on purified RNase Rs revealed the involvement of a single histidine, lysine and carboxylate residue in the catalytic activity of the enzyme. RNA could not protect the enzyme against DEP- and TNBS-mediated inactivation whereas, substrate protection was observed in case of EDAC-mediated inactivation of the enzyme. K(m) and k(cat) values of the partially inactivated enzyme samples suggested that while histidine and lysine are involved in catalysis, carboxylate is involved in substrate binding. Active site nature of RNase Rs suggests that the inability of the enzyme to readily convert 2',3'-cyclic nucleotides to 3'-mononucleotides is probably due to the absence of catalytically active second histidine residue. (+info)
(8/580) Activation of pancreatic stellate cells in human and experimental pancreatic fibrosis.
The mechanisms of pancreatic fibrosis are poorly understood. In the liver, stellate cells play an important role in fibrogenesis. Similar cells have recently been isolated from the pancreas and are termed pancreatic stellate cells. The aim of this study was to determine whether pancreatic stellate cell activation occurs during experimental and human pancreatic fibrosis. Pancreatic fibrosis was induced in rats (n = 24) by infusion of trinitrobenzene sulfonic acid (TNBS) into the pancreatic duct. Surgical specimens were obtained from patients with chronic pancreatitis (n = 6). Pancreatic fibrosis was assessed using the Sirius Red stain and immunohistochemistry for collagen type I. Pancreatic stellate cell activation was assessed by staining for alpha-smooth muscle actin (alphaSMA), desmin, and platelet-derived growth factor receptor type beta (PDGFRbeta). The relationship of fibrosis to stellate cell activation was studied by staining of serial sections for alphaSMA, desmin, PDGFRbeta, and collagen, and by dual-staining for alphaSMA plus either Sirius Red or in situ hybridization for procollagen alpha(1) (I) mRNA. The cellular source of TGFbeta was examined by immunohistochemistry. The histological appearances in the TNBS model resembled those found in human chronic pancreatitis. Areas of pancreatic fibrosis stained positively for Sirius Red and collagen type I. Sirius Red staining was associated with alphaSMA-positive cells. alphaSMA staining colocalized with procollagen alpha(1) (I) mRNA expression. In the rat model, desmin staining was associated with PDGFRbeta in areas of fibrosis. TGFbeta was maximal in acinar cells adjacent to areas of fibrosis and spindle cells within fibrotic bands. Pancreatic stellate cell activation is associated with fibrosis in both human pancreas and in an animal model. These cells appear to play an important role in pancreatic fibrogenesis. (+info)