An ethanol extract of the stem of Opuntia ficus-indica var. saboten (OFS) was assessed to determine the mechanism(s) of its antioxidant activity. The ethanol extract exhibited a concentration-dependent inhibition of linoleic acid oxidation in a thiocyanate assay system. In addition, the OFS extract showed dose-dependent free-radical scavenging activity, including DPPH radicals, superoxide anions (O(2)(*-)), and hydroxyl radicals (*OH), using different assay systems. The OFS ethanol extract was also found to be effective in protecting plasmid DNA against the strand breakage induced by hydroxyl radicals in a Fenton's reaction mixture. Furthermore, the extract showed significant (p < 0.01) dose-dependent protection of mouse splenocytes against glucose oxidase-mediated cytotoxicity. Finally, the OFS extract was characterized as containing a high amount of phenolics (180.3 mg/g), which might be the active compounds responsible for the antioxidant properties of the OFS extract.
BackgroundAntimicrobial peptides (AMPs) are primarily known for their innate immune defense against invading microorganisms, including viruses. In addition, recent research has suggested their modulatory activity in immune induction. Given that most subunit vaccines require an adjuvant to achieve effective immune induction through the activation of innate immunity, AMPs are plausible candidate molecules for stimulating not only innate immune but also adaptive immune responses.ResultsIn this study, we investigated the ability of human β-defensin (HBD) 2 to promote antiviral immunity in vitro and in vivo using a receptor-binding domain (RBD) of Middle East respiratory syndrome-coronavirus (MERS-CoV) spike protein (S RBD) as a model antigen (Ag). When HBD 2-conjugated S RBD was used to treat THP-1 human monocytic cells, the expression levels of antiviral (IFN-β, IFN-γ, MxA, PKR, and RNaseL) and primary immune-inducing (NOD2, TNF-α, IL-1β, and IL-6) molecules were enhanced compared to those expressed after treatment with S RBD only. The expression of chemokines capable of recruiting leukocytes, including monocytes/macrophages, natural killer cells, granulocytes, T cells, and dendritic cells, was also increased following HBD 2-conjugated S RBD treatment. More important, immunization of mice with HBD 2-conjugated S RBD enhanced the immunogenicity of the S RBD and elicited a higher S RBD-specific neutralizing antibody response than S RBD alone.ConclusionsWe conclude that HBD 2 activates the primary antiviral innate immune response and may also mediate the induction of an effective adaptive immune response against a conjugated Ag.
Oral mucosal immunization can induce protective immunity in both systemic compartments and the mucosa. Successful mucosal immunization depends on Ag delivery to the mucosal immune induction site. The high transcytotic activity of M cells within the mucosa makes these cells attractive targets for mucosal Ag delivery, although it remains unclear whether delivery of Ag to M cells only can guarantee the induction of effective immune responses. In this study, we evaluated the ability of an M cell-targeting ligand with adjuvant activity to induce immunity against ligand-fused Ag. We selected M cell-targeting ligands through biopanning of a phage display library against differentiated in vitro M-like cells and produced the recombinant Ags fused to the selected ligands using the model Ag. One of the selected peptide ligands, Co1, promoted the binding of ligand-fused Ag to mouse Peyer’s patch M cells and human M-like cells that had been defined by binding with the M cell-specific and anti-GP2 Abs. In addition, Co1 ligand enhanced the uptake of fused Ag by immunogenic tissue in an ex vivo loop assay and in vivo oral administration experiments. After oral administration, the ligand-fused Ag enhanced immune responses against the fused Ag compared with those of the control Ag without ligand. In addition, this use of the ligand supported a skewed Th2-type immune response against the fused Ag. Collectively, these results suggest that the ligand selected through biopanning against cultured M-like cells could be used as an adjuvant for targeted Ag delivery into the mucosal immune system to enhance immune induction.
In the mucosal immune system, M cells are known as specialized epithelial cells that take up luminal antigens, although the receptors on M cells and the mechanism of antigen uptake into M cells are not well-understood. Here, we report the expression of the complement C5a receptor (C5aR) on the apical surface of M cells. C5ar mRNA expression in co-cultured Caco-2 human M-like cells was six-fold higher than in mono-cultured cells. C5aR expression was detected together with glycoprotein 2, an M-cell-specific protein, on the apical surface of M-like cells and mouse Peyer's patch M cells. Interestingly, after oral administration of Yersinia enterocolitica which expresses outer membrane protein H (OmpH) that is homologous to the Skp a1 domain of Escherichia coli, a ligand of C5aR, dense clustering and phosphorylation of C5aR were detected in M cells. Finally, targeted antigen delivery to M cells using C5aR as a receptor was achieved using the OmpH a1 of Y. enterocolitica such that the induction of ligand-conjugated antigen-specific immune responses was confirmed in mice after oral immunization of the OmpH b1a1-conjugated antigen. Collectively, we identified C5aR expression on M cells and suggest that C5aR could be used as a target receptor for mucosal antigen delivery. IntroductionThe mucosa are a primary target for infection and, at the same time, the first line of defense against many pathogens in which secretory IgA is a main protective component of the immune system [1]. In order to achieve an effective induction of antigenspecific immune responses using the oral route, obstacles such as oral tolerance induction and inefficient antigen delivery to the mucosal immune induction site should be resolved [2]. One of the most effective ways to overcome these obstacles is to target antigens to M cells, which are a primary gateway for the delivery of antigens from the intestinal lumen to the mucosal lymphoid organs. In addition, M cells have high transcytotic capacity and have highly distributed APCs in their basolateral pockets [3]. [5][6][7]. Therefore, identification of target receptors on M cells that could be used for successful mucosal vaccine delivery is one of the most interesting subjects in mucosal immunology. The complement and TLR systems are central components of the innate immune system, necessary for rapid responses to external dangers. However, in comparison to the well-recorded expression of TLR1, 2, and 4 and C-C chemokine receptor 5 in the M cell, expression of the molecules associated with the complement system has not yet been reported, despite possible roles for these proteins in activities such as homeostatic regulation of the mucosa by the induction of chemotactic and pro-inflammatory cytokines or antigen uptake through crosstalk with TLRs [8]. Among the molecules associated with the complement system, the complement C5a receptor (C5aR/CD88) was traditionally thought to exist only on myeloid cells. However, C5aR expression was recently verified on non-myeloid cells, including bronchial epithelial...
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