Preservation of protective capacity of hyperimmune anti-Stx2 bovine colostrum against enterohemorrhagic Escherichia coli O157:H7 pathogenicity after pasteurization and spray-drying processes.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a major etiologic agent that causes bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome (HUS). Shiga toxin (Stx) is the main virulence factor of EHEC responsible for the progression to HUS. Although many laboratories have made efforts to develop an effective treatment for Stx-mediated HUS, a specific therapy has not been found yet. Human consumption of bovine colostrum is known to have therapeutic effects against several gastrointestinal infections because of the peptide and proteins (including antibodies) with direct antimicrobial and endotoxin-neutralizing effects contained in this fluid. We have previously demonstrated that colostrum from Stx type 2 (Stx2)-immunized pregnant cows effectively prevents Stx2 cytotoxicity and EHEC O157:H7 pathogenicity. In this study we evaluated the preservation of the protective properties of hyperimmune colostrum against Stx2 (HIC-Stx2) after pasteurization and spray-drying processes by performing in vitro and in vivo assays. Our results showed that reconstituted HIC-Stx2 colostrum after pasteurization at 60°C for 60 min and spray-dried under optimized conditions preserved specific IgG that successfully neutralized Stx2 cytotoxicity on Vero cells. Furthermore, this pasteurized/dehydrated and reconstituted HIC-Stx2 preserved the protective capacity against EHEC infection in a weaned mice model. The consumption of hyperimmune HIC-Stx2 bovine colostrum could be effective for HUS prevention in humans as well as in EHEC control in calves. However, further studies need to be done to consider its use for controlling EHEC infections.

Reviewing the biological activity of chitosan in the mucosa: Focus on intestinal immunity.

Chitosan is a polymer derived from the partial deacetylation of chitin with particular characteristics, such as mucoadhesiveness, tolerability, biocompatibility and biodegradability. Biomedical uses of chitosan cover a wide spectrum of applications as dietary fiber, immunoadjuvant and regulator of the intestinal microbiota or delivery agent. Chemical modification of chitosan is feasible because its reactive amino and hydroxyl groups can be modified by a diverse array of ligands, functional groups and molecules. This gives rise to numerous derivatives that allow different formulation types influencing their activity. Considering the multiple events resulting from the interaction with mucosal tissues, chitosan is a singular candidate for strategies targeting immune stimulation (i.e., tolerance induction, vaccination). Its role as a prebiotic and probiotic carrier represents an effective option to manage intestinal dysbiosis. In the intestinal scenario where the exposure of the immune system to a wide variety of antigens is permanent, chitosan increases IgA levels and favors a tolerogenic environment, thus becoming a key ally for host homeostasis.

Intramammary inoculation with lactic acid bacteria at dry-off triggers an immunomodulatory response in dairy cows.

The use of antibiotics to prevent bovine mastitis is responsible for the emergence and selection of resistant strains. Lactic acid bacteria (LAB) could be introduced into animal feed as an alternative prevention method that would bypass the risk of resistance development. In previous research, we demonstrated that two probiotic LAB strains isolated from bovine milk were capable of stimulating the production of antibodies and the host's immune cellular response in the udder. The present study aimed to elucidate whether the antibodies of animals inoculated with these strains were able to increase phagocytosis by neutrophils and inhibit the growth of different mastitis-causing pathogens. Moreover, the effect of LAB on the expression of pro-inflammatory cytokines was assessed. Ten animals were inoculated intramammarily with 106 cells of the two strains at dry-off. The blood serum was tested for its ability to opsonize bovine mastitis pathogens, the in vitro bactericidal activity of bovine blood and milk against these pathogens was determined, and cytokine mRNA expression was quantified in milk somatic cells. The inoculated animals did not show abnormal signs of sensitivity to the LAB. Their blood serum significantly enhanced the phagocytosis of Staphylococcus spp. and the LAB. Escherichia coli and Streptococcus uberis were inhibited by the milk serum but not the blood serum, whereas Staphylococcus aureus and Staphylococcus haemolyticus were inhibited by both. In regard to cytokine expression, interleukin (IL)-1ß increased markedly for up to 4 h post-inoculation, and an increase in IL-8 was observed 4, 12 and 24 h after inoculation. Tumour necrosis factor-α mRNA increased 1 and 2 h after inoculation and a significant difference was registered at 6 h for interferon-γ. This rapid immunomodulatory response shows that inoculating animals with LAB at dry-off, when they are especially susceptible, could be a useful strategy for the prevention of bovine mastitis.

The biocompatible polysaccharide chitosan enhances the oral tolerance to type II collagen.

Chitosan is a mucoadhesive polysaccharide that promotes the transmucosal absorption of peptides and proteins. At mucosal sites chitosan exhibits immunomodulatory activities and stimulates the release of regulatory cytokines. Herein we evaluated the effect of the co-administration of chitosan in the tolerance to type II collagen (CII) using an experimental model of arthritis. Rats were fed diluent (acetic acid), 1 mg CII, 1 mg chitosan or 1 mg CII + 1 mg chitosan during 5 days before immunization with CII in Freund's complete adjuvant. Systemic effects were evaluated in draining lymph nodes after antigenic challenge or during the clinical evolution of arthritis. Specific antibodies, proliferation against CII and the production of interferon (IFN)-gamma and interleukin-10 were assessed. Clinical signs were observed 13-15 days after primary immunization. The CII : chitosan group presented the lowest incidence and developed moderate arthritis, with reduced levels of immunoglobulin (Ig)G2a anti-CII, a limited proliferation in draining lymph nodes and a lower release of IFN-gamma after restimulation with CII. Our results demonstrate that chitosan enhances the tolerance to an articular antigen with a decrease in the inflammatory responses and, as a consequence, an improvement in clinical signs.

Ability of the polysaccharide chitosan to inhibit proliferation of CD4+ lymphocytes from mucosal inductive sites, in vitro and in vivo.

OBJECTIVE: After oral administration of chitosan (a copolymer of glucosamine and N-acetylglucosamine), mesenteric lymph node (MLN) lymphocytes exhibited traits of anergy, a process coupled with inability of mature T cells to proliferate. We wondered whether biological activity of chitosan could be affecting division of lymphocytes at the mucosal inductive sites. MATERIALS AND METHODS: We studied the effect of chitosan on proliferation of carboxyfluorescein diacetate-labelled MLN lymphocytes stimulated with concanavalin A in vitro. We assessed expression of CD25 and CD71 activation markers and pro-apoptotic molecule CD95L. Moreover, we studied the effect of chitosan ex vivo, in carboxyfluorescein diacetate-labelled MLN cells isolated after feeding single or repetitive doses of the polysaccharide, and we evaluated cell cycle parameters. RESULTS: Chitosan suppressed cell proliferation and down-modulated expression of CD25 in these MLN CD4+ cells isolated from normal rats. After in vivo contact, chitosan inhibited proliferation of MLN cells and reduced secretion of interferon-gamma. Furthermore, sustained feeding produced reduction in percentage of CD4+ cells in S phase of the cell cycle. CONCLUSION: Here we demonstrate the ability of chitosan to suppress proliferation of CD4+ lymphocytes from mucosal inductive sites in vivo and in vitro This effect could be relevant in modulatory activity of chitosan in the intestinal microenvironment.