Chitosan Microparticals for protein and antigen delivery: Perparation, Interactions with antigen presenting cells and vivo immune response

Abstract

Chitosan microparticles of different molecular weights and sources for protein and antigen delivery were prepared by spray drying. Formulation and process parameters were firstly optimized in order to obtain the microparticles with controlled properties. Bovine serum albumin was subsequently incorporated into microparticles. Physicochemical properties of the resultant microparticles were evaluated in terms of particle size and size distribution, zeta potential, particle morphology, particle surface composition, protein content and entrapment efficiency, in vitro release, and integrity of encapsulated protein. Size of chitosan microparticles was ranged between 3.185- 7.177 μm. Integrity and structural comformation of entrapped protein were retained at protein loading of 5% w/w or higher. The microparticles were further modified by cospray drying with excipients or crosslinking with tripolyphosphate anions. Incorporation of gelatin resulted in an increase in zeta potential of microparticles, while poloxamer 407 and Eudragit E had no effect. The added excipents mostly induced faster and/or higher amount of released protein. Crosslinking the particles caused an obvious reduction of drug release. For comparison, poly(lactic-co-glycolic acid) (PLGA) and poly(α-butyl cyanoacrylate) (BCA) micro-/nanoparticles were prepared by double-emulsion solvent-evaporation and anionic polymerization methods, respectively, and characterized by their properties. All micro-/nanoparticles were subjected to cytotoxicity test with dendritic cells and macrophages. The chitosan, gelatin/chitosan, poloxamer 407/chitosan and PLGA microparticles were relatively non-toxic to both cells, while the opposite results were obtained from Eudragit E/chitosan and BCA micro-/nanoparticles. The non-toxic chitosan and composite microparticles were efficiently taken up by both cells. The Japanese Encephalitis (JE) antigen was finally incorporated into the selected microparticles and administered subcutaneously into mice. The chitosan of both low and high molecular weight and the gelatin/chitosan microparticles elicited a comparable immune response to the commercial JE vaccine. At week 12 after the first immunization, the serum IgG titer, induced by the low molecular weight chitosan particles, still kept increasing, while that of the others were stable or started to decline. The chitosan microparticles of low molecular weight presented a high potential application as an efficient vaccine delivery system.