β-glucan microparticles as mucosal delivery system in oral vaccine development

Several enteric pathogens infect the body following oral uptake and cause life-threatening diseases such as cholera, dysentery and typhoid fever. Oral vaccination is essential to generate protective local immunity against intestinal pathogens. However, antigen delivery to the inductive sites for mucosal immunity (the small intestine Peyer’s patches, PP) has proven to be particularly challenging. Recently, there has been a lot of interest in the use of microparticles as antigen delivery systems in the development of more efficient mucosal vaccines. We evaluated the potential of β-glucan microcapsules to deliver antigen transmucosally in the PP regions of the murine small intestine.β-glucan microparticles (2-4 µm) were prepared from Saccharomyces cerevisiae and loaded with FITC or Alexa Fluor 488-conjugated bovine serum albumin. Firstly, the appropriateness of β-glucan particles as antigen delivery systems for oral and intestinal applications was assessed in stability tests. Fluorimetric analysis demonstrated that the BSA concentration within particles is stable during the first 12 hours of simulated gastric or simulated intestinal fluid treatment. Secondly, β-glucan particles were administered to male C57BL/6 mice (8-10 weeks old) via intestinal loops at a particle concentration of 100*106/ml. After one hour of incubation, transmucosal particle transport and uptake in the PP was evaluated by flow cytometry, confocal microscopy and transmission electron microscopy. Using flow cytometry, we could not observe any particle uptake in the main antigen presenting cell population, the dendritic cells, however, a modest particle uptake was repeatedly detected in the B-cell population. Confocal microscopic images showed localization of β-glucan particles in the follicle associated epithelium and B-cell internalization. Moreover, transmission electron microscopy demonstrated transcellular transport of yeast particles in M-cells.In conclusion, stability tests show that antigen concentration of β-glucan particles remains stable in gastric and intestinal environment. This means that the β-glucan particles can be administered orally without prior enteric coating and are suitable for mucosal delivery of antigen in the murine gastro-intestinal tract. Our data suggest that M-cells, but not subepithelial dendritic cells, are crucial for the transmucosal transport of β-glucan particles from the intestinal lumen to the PP and transcytosis of antigen to underlying antigen presenting cells and immune cells.Several enteric pathogens infect the body following oral uptake and cause life-threatening diseases such as cholera, dysentery and typhoid fever. Oral vaccination is essential to generate protective local immunity against intestinal pathogens. However, antigen delivery to the inductive sites for mucosal immunity (the small intestine Peyer’s patches, PP) has proven to be particularly challenging. Recently, there has been a lot of interest in the use of microparticles as antigen delivery systems in the development of more efficient mucosal vaccines. We evaluated the potential of β-glucan microcapsules to deliver antigen transmucosally in the PP regions of the murine small intestine.β-glucan microparticles (2-4 µm) were prepared from Saccharomyces cerevisiae and loaded with FITC or Alexa Fluor 488-conjugated bovine serum albumin. Firstly, the appropriateness of β-glucan particles as antigen delivery systems for oral and intestinal applications was assessed in stability tests. Fluorimetric analysis demonstrated that the BSA concentration within particles is stable during the first 12 hours of simulated gastric or simulated intestinal fluid treatment. Secondly, β-glucan particles were administered to male C57BL/6 mice (8-10 weeks old) via intestinal loops at a particle concentration of 100*106/ml. After one hour of incubation, transmucosal particle transport and uptake in the PP was evaluated by flow cytometry, confocal microscopy and transmission electron microscopy. Using flow cytometry, we could not observe any particle uptake in the main antigen presenting cell population, the dendritic cells, however, a modest particle uptake was repeatedly detected in the B-cell population. Confocal microscopic images showed localization of β-glucan particles in the follicle associated epithelium and B-cell internalization. Moreover, transmission electron microscopy demonstrated transcellular transport of yeast particles in M-cells.In conclusion, stability tests show that antigen concentration of β-glucan particles remains stable in gastric and intestinal environment. This means that the β-glucan particles can be administered orally without prior enteric coating and are suitable for mucosal delivery of antigen in the murine gastro-intestinal tract. Our data suggest that M-cells, but not subepithelial dendritic cells, are crucial for the transmucosal transport of β-glucan particles from the intestinal lumen to the PP and transcytosis of antigen to underlying antigen presenting cells and immune cells.C