Toll-like receptor (TLR) ligands and phytochemical feed additives (PFAs) were evaluated in this study to determine the effects of immune stimulation on gut immunity in the zebrafish, Danio rerio, and the channel catfish, Ictalurus punctatus. Rag1-/- (MT) zebrafish were used to study how the TLR ligands β-glucan and resiquimod (R848) affect the innate immune system in the gut of MT zebrafish. Enhanced expression of marker genes (NITR9, NCCRP-1 and MPEG-1) indicated stimulation of Natural Killer (NK) cells, non-specific cytotoxic cells (NCCs) and macrophages. After challenge with Edwardsiella ictaluri, MT zebrafish stimulated with β-glucan demonstrated higher survival and the presence of more macrophages/monocytes in the gut than control MT zebrafish. A PFA test diet containing a blend of prebiotic fiber, oregano, thyme, cinnamon essential oils, and Yucca schidigera (ONE Current™, OC) was fed to channel catfish for 3 months in ponds to determine the effect on channel catfish fingerling growth. Fish were fed in ponds and a tank bacterial challenge followed to test the efficacy of the product. Catfish fed OC demonstrated greater weight gain and feed conversion ratios, higher survival after challenge with E. ictaluri, greater phagocytosis or binding by macrophages and cytotoxic cells. Catfish fed OC also demonstrated greater gut surface area after 2 months feeding OC. To elucidate the effect(s) of each of the compounds in the OC diet on gut immune responses and to determine if PFAs can decrease bacterial colonization and replication within gut tissues, WT and MT zebrafish were fed diets containing different compounds included in OC. Quantification of live bacteria from gut and kidney tissue was determined after challenge with E. ictaluri. Expression levels of immune response genes were evaluated after ingestion of PFAs. Actifibe, Essential oil 25 ppm (EO 25) and Actifibe + EO demonstrated the lowest infection and colonization rate, upregulation of immune response genes, and significantly higher survival when challenged with E. ictaluri. This study demonstrates the potential for application of TLR ligand and feed administered PFAs to improve fish health. Our findings provide a more comprehensive understanding of host gut/pathogen interactions as well as suggestions for novel disease control measures.

The aim of submited Thesis was to contribute to illustration on effect of TLR and fagocytate receptors in melanoma B16-F10 treatment with mouse model. Currently, a consequence of different ligands of fagocyte receptors and their combination with liposacharide (LPS) was tested. Ligands of fagocytate receptors was used in form enabled inside tumor cells anchoring. It deals with these compounds: f-MLFKK-BAM4000, laminarin-BAM4000, mannan-BAM4000. Gained results confirmed that previously observed huge synergy of LPS (activation of TLR4) and activation of phagocytic receptors by ligands anchored to tumor cells found the clarification in the form of extensive necrotization of tumor mass at the histological level.

The prevention and treatment of cancer and infectious diseases requires vaccines that can mediate cytotoxic T lymphocyte-based immunity. A promising strategy is protein subunit vaccines composed of purified protein antigens and immunostimulatory adjuvants, such as Toll-like receptor (TLR) agonists. In this research, we developed two new biodegradable polymeric delivery vehicles for protein antigens and TLR agonists, as model vaccine delivery systems. This work was guided by the central hypothesis that an effective vaccine delivery system would have stimulus-responsive degradation and release, biodegradability into excretable non-acidic degradation products, and the ability to incorporate various TLR-inducing adjuvants. The first vaccine delivery system is a cross-linked polyion complex micelle which efficiently encapsulates proteins, DNA, and RNA. The micelle-based delivery system consists of a block copolymer of poly(ethylene glycol) (PEG) and poly(L-lysine), cross-linked by dithiopyridyl side groups to provide transport stability and intracellular release. The second delivery system consists of solid biodegradable microparticles encapsulating proteins, nucleic acids, and hydrophobic compounds. The microparticles are composed of pH-sensitive polyketals, which are a new family of hydrophobic, linear polymers containing backbone ketal linkages. Polyketals are synthesized via a new polymerization method based on the acetal exchange reaction and degrade into non-acidic, excretable degradation products. In addition, the technique of hydrophobic ion pairing was utilized to enhance the encapsulation of ovalbumin, DNA, and RNA in polyketal microparticles via a single emulsion method. Using in vitro and in vivo immunological models, we demonstrated that the micelle- and polyketal-based vaccine delivery systems enhanced the cross-priming of cytotoxic T lymphocytes. The model vaccines were composed of ovalbumin antigen and various TLR-inducing adjuvants including CpG-DNA, monophosphoryl lipid A, and dsRNA. The results demonstrate that the cross-linked micelles and polyketal microparticles have considerable potential as delivery systems for protein-based vaccines.