Abstract
The anti-inflammatory effect of mammalian heparin analogues, named dermatan sulfate and heparin, isolated from the ascidian Styela plicata was accessed in a TNBS-induced colitis model in rats. Subcutaneous administration of the invertebrate compounds during a 7-day period drastically reduced inflammation as observed by the normalization of the macroscopic and histological characteristics of the colon. At the molecular level, a decrease in the production of TNF-alpha, TGF-beta, and VEGF was observed, as well as a reduction of NF-kappaB and MAPK kinase activation. At the cellular level, the heparin analogues attenuated lymphocyte and macrophage recruitment and epithelial cell apoptosis. A drastic reduction in collagen-mediated fibrosis was also observed. No hemorrhagic events were observed after glycan treatment. These results strongly indicate the potential therapeutic use of these compounds for the treatment of colonic inflammation with a lower risk of hemorrhage when compared with mammalian heparin.
Highlights
EXPERIMENTAL PROCEDURESAnimals—Male Wistar rats (each weighing between 250 and 300 g) obtained from a local supplier were maintained on a 12-h/12-h light and dark cycle in a temperature-controlled room (24 °C)
Th1-type response is characterized by the production of tumor necrosis factor (TNF)-␣, interleukin (IL)-12, IL-18, interferon (IFN)-␥, among other proinflammatory cytokines, and involves the production of growth factors such as transforming growth factor (TGF)-, and vascular endothelial growth factor (VEGF) (4 – 6)
The suggested efficacy observed in several open clinical studies [15,16,17], heparin has been proposed as an alternative for the treatment of Inflammatory bowel diseases (IBD)
Summary
Animals—Male Wistar rats (each weighing between 250 and 300 g) obtained from a local supplier were maintained on a 12-h/12-h light and dark cycle in a temperature-controlled room (24 °C). After rinsing in phosphate-buffered saline (PBS) containing 0.5% Tween 20 for 10 min, tissue sections were incubated with nonimmune horse serum for 30 min and, subsequently, with the respective monoclonal antibody in a humidified chamber overnight, at room temperature. After rinsing three times in PBS for 5 min each, tissue sections were incubated for 1 h with a FITC-conjugated Fab fraction of goat antimouse IgG antibody (Dako A/S). Slides were counterstained with Evan blue-diluted 0.01% in phosphate-buffered saline (PBS) and incubated at 37 °C for 15 min, mounted in an antifading medium containing buffered glycerol and p-phenylenediamine (Sigma), and observed with a Zeiss LSM 510 confocal laser scanning microscope. Two sections from each sample were incubated with either PBS alone or FITC-conjugated anti-mouse IgG antibody and served as negative controls.
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