A Rapid Screenable Assay for Compounds That Protect Against Intestinal Injury in Zebrafish Larva.

This chapter describes a method to assay compounds modulating NSAID-induced intestinal injury in zebrafish larvae. The assay employs the NSAID glafenine, which causes intestinal epithelial cell damage and death by inducing organelle stress responses (endoplasmic reticulum and mitochondrial) and blocking the unfolded protein response pathway. This epithelial damage includes sloughing of intestinal cells into the lumen and out the cloaca of the zebrafish larvae. Exposing larvae to acridine orange highlights this injury when visualized under fluorescence microscope; injured fish develop intensely red-staining intestines, as well as a "tube" or cord of red color extending through the intestine and out the cloaca. Using this rapid visually screenable method, various candidate compounds were successfully tested for their ability to prevent glafenine-induced intestinal injury. Because this assay involves examination of larval zebrafish intestinal pathology, we have also included our protocol for preparation and analysis of zebrafish histology. The protocol includes numerous steps to generate high-quality zebrafish histology slides, as well as protocols to establish accurate anatomic localization of any given tissue cross-section-processes that are made technically difficult by the small size of zebrafish larvae.

Vitamin D as an Immunomodulator: Risks with Deficiencies and Benefits of Supplementation.

Vitamin D refers to a class of fat-soluble secosteroids often associated with their role in absorption and metabolism of minerals such as calcium and phosphate. In recent years, our understanding of vitamin D has expanded to include its role in modulating the immune system. Of particular focus are the effects of vitamin D deficiency and supplementation on patients suffering from disorders due to dysregulation of the immune system. In patients with multiple sclerosis, rheumatoid arthritis, and inflammatory bowel disease, deficiencies in vitamin D have been associated with an increased risk of disease activity. In this review, we will look at the current state of research in regards to the relationship between vitamin D and immune-dysregulation. We will focus on both the risks associated with vitamin D deficiency as well as the benefits of vitamin D supplementation.

Oxymatrine prevents NF-κB nuclear translocation and ameliorates acute intestinal inflammation.

Oxymatrine is a traditional Chinese herbal product that exhibits anti-inflammatory effects in models of heart, brain and liver injury. We investigated the impact of oxymatrine in an acute model of intestinal injury and inflammation. Oxymatrine significantly decreased LPS-induced NF-κB-driven luciferase activity, correlating with diminished induction of Cxcl2, Tnfα and Il6 mRNA expression in rat IEC-6 and murine BMDC. Although oxymatrine decreased LPS-induced p65 nuclear translocation and binding to the Cxcl2 gene promoter, this effect was independent of IκBα degradation/phosphorylation. DSS-induced weight loss and histological damage were ameliorated in oxymatrine-treated C57BL/6-WT-mice. While this effect correlated with reduced colonic Il6 and Il1ß mRNA accumulation, global NF-κB activity as measured in NF-κB(EGFP) mice was unaffected. Our data demonstrate that oxymatrine reduces LPS-induced NF-κB nuclear translocation and activity independently of IκBα status, prevents intestinal inflammation through blockade of inflammatory signaling and ameliorates overall intestinal inflammation in vivo.