Drak2-deficient (Drak2−/−) mice are resistant to multiple models of autoimmunity yet effectively eliminate pathogens and tumors. Thus, DRAK2 represents a potential target to treat autoimmune diseases. However, the mechanisms by which DRAK2 contributes to autoimmunity, particularly type 1 diabetes (T1D), remain unresolved. Here, we demonstrate that resistance to T1D in non-obese diabetic (NOD) mice is due to the absence of Drak2 in T cells and requires the presence of regulatory T cells (Tregs). Contrary to previous hypotheses, we show that DRAK2 does not limit TCR signaling. Rather, DRAK2 regulates IL-2 signaling by inhibiting STAT5A phosphorylation. We further demonstrate that enhanced sensitivity to IL-2 in the absence of Drak2 augments thymic Treg development. Overall, our data indicate that DRAK2 contributes to autoimmunity in multiple ways by regulating thymic Treg development and by impacting the sensitivity of conventional T cells to Treg-mediated suppression.
Silver nanoparticles (AgNP) are widely used for their antibacterial properties. Incorporation of AgNP into food-related products and health supplements represents a potential route for oral exposure to AgNP; however, the effects of such exposure on the gastrointestinal system are mostly unknown. This study evaluated changes in the populations of intestinal-microbiota and intestinal-mucosal gene expression in Sprague-Dawley rats (both male and female) that were gavaged orally with discrete sizes of AgNP (10, 75 and 110 nm) and silver acetate. Doses of AgNP (9, 18 and 36 mg/kg body weight/day) and silver acetate (100, 200 and 400 mg/kg body weight/day) were divided and administered to rats twice daily (∼10 h apart) for 13 weeks. The results indicate that AgNP prompted size- and dose-dependent changes to ileal-mucosal microbial populations, as well as, intestinal gene expression and induced an apparent shift in the gut microbiota toward greater proportions of Gram-negative bacteria. DNA-based analyses revealed that exposure to 10 nm AgNP and low-dose silver acetate caused a decrease in populations of Firmicutes phyla, along with a decrease in the Lactobacillus genus. Analysis of host gene expression demonstrated that smaller sizes and lower doses of AgNP exposure prompted the decreased expression of important immunomodulatory genes, including MUC3, TLR2, TLR4, GPR43 and FOXP3. Gender-specific effects to AgNP exposure were more prominent for the gut-associated immune responses. These results indicate that the oral exposure to AgNP alter mucosa-associated microbiota and modulate the gut-associated immune response and the overall homeostasis of the intestinal tract.
Nanomaterial comes in several forms (metal, metal oxide, and carbon-based), sizes, coatings, and very diverse characteristics. These physical characteristics decide the outcome of their interaction with the biological system. Due to the use of nanomaterials in the items that may have direct interaction with the gastrointestinal system, it is very imperative to have a meta-analysis of what physical characteristics and concentration levels of nanomaterials are of greatest concern regarding exposure due to dietary consumption. Here, in this chapter, our focus is on the antibacterial activity of nanomaterials against bacteria found in the gastrointestinal tract, with a specific emphasis on the mucosal microbiome.
Abstract The proteins of cheese are rapidly solubilised by heating to 95 °C in buffered 2 % sodium dodecyl sulfate, 5 % 2‐mercaptoethanol. Electrophoretic analysis of the solubilised proteins by either one‐dimensional sodium dodecyl sulphate‐polyacrylamide gel electrophoresis or high resolution two‐dimensional electrophoresis yields reproducible patterns characteristic of an individual cheese and its extent of ripening. The patterns reveal (i) the residual amounts of milk casein and whey proteins, and (ii) the appearance of casein degradation products, including pink‐violet components as detected by Coomassie Blue staining.
Silver staining and high-resolution electrophoretic methods have been used to compare the protein composition of rat parotid saliva evoked in response to (i) parasympathetic stimulation (including the nonadrenergic, noncholinergic, atropine-associated secretion), (ii) sympathetic stimulation, or (iii) the infusion of neuropeptides with secretagogue activity (substance P, calcitonin gene-related peptide, neuropeptide Y, or vasoactive intestinal peptide). The different stimuli influenced the protein concentration and flow rate of the evoked secretion but had little effect upon the protein composition of the saliva. In contrast to earlier studies using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Coomassie blue staining, the combination of silver staining and two-dimensional electrophoresis (2-DE) revealed many newly detected proteins. The results indicate that the protein composition of rat parotid saliva is more complex than previously reported but is unaffected by the mode of stimulation.
The variation in cytokine production during bacterial invasion of human intestinal epithelial cells (IECs) is a contributing factor for progression of the infection. A few Salmonella enterica Heidelberg strains isolated from poultry products harbor transmissible plasmids (TPs), including those that encode a type-IV secretion system. Earlier, we showed that these TPs are responsible for increased virulence during infection. This study examines the potential role of these TPs in cytokine production in IECs. This study showed that S. Heidelberg strains containing TPs (we refer as virulent strains) caused decreased interleukin (IL)-10 production in IECs after 1 h infection. The virulent strains induced a high level of tumor necrosis factor-α production under identical conditions. The virulent strains of S. Heidelberg also altered the production of IL-2, IL-17, and granulocyte macrophage colony-stimulating factor compared to an avirulent strain. As a part of infection, bacteria cross the epithelial barrier and encounter intestinal macrophages. Hence, we examined the cytotoxic mechanism of strains of S. Heidelberg in macrophages. Scanning electron microscopy showed cell necrosis occurs during the early stage of infection. In conclusion, virulent S. Heidelberg strains were able to modify the host cytokine profile during the early stages of infection and also caused necrosis in macrophages.
Abstract The proteins of parasympathetically stimulated cat parotid saliva were analyzed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and high resolution two‐dimensional electrophoresis (2‐DE). SDS‐PAGE revealed up to 30 polypeptide bands in microliter volumes of unconcentrated saliva. The patterns were highly reproducible and characterized by prominent bands of M r 57000, M r 30000 and M r 15000. The major protein ( M r 30000) appeared as a dimer ( M r 60000) when electrophoresed under non‐reducing conditions but dissociated into its monomeric form when the SDS concentration of the denatured samples was increased from 1 to 5%. This indicates a noncovalent association. The protein patterns of saliva from different cats differed slightly but sequential samples from the same cat (collected during 90 min of stimulation) showed little change in protein pattern apart from a fall in total protein content. Following 2‐DE, the major protein ( M r 30000) appeared as a complex array of at least eight spots in two tiers (p I 5.2–6.2; M r 28000 and 32000). The characteristics of this protein are discussed with reference to allergy to cats.
