Enhanced mucosal and systemic immune responses obtained by porous silica nanoparticles used as an oral vaccine adjuvant: Effect of silica architecture on immunological properties

Abstract

Three different kinds of silica (S2, S1 and SBA-15) with different particle sizes (130, 430 nm and 1–2 μm) and different pore characteristics (i.e. pore size and shape) were developed as oral vaccine immunological adjuvants and the relationship between the silica architecture and immunological properties was investigated. The silica particles were characterized using SEM, TEM and nitrogen adsorption. Model antigen bovine serum albumin (BSA) was successfully entrapped into the silica pores to produce a sustained release vaccine delivery system. Compared with the responsiveness induced by parenteral administration of BSA emulsified in Freund's complete adjuvant (FCA), oral immunization with the silica/BSA formulation produced a stimulated humoral and mucosal (sIgA) response. The IgG and IgA titers induced by loading BSA was as follows: S1 > S2 > SBA-15. The highest IgG and IgA titers of S1 were attributed to its large honeycombed pores and the optimal particle diameter of 430 nm. The corresponding IgG1 and IgG2a titers were also investigated to confirm that BSA loaded in nanoparticles by oral immunization can induce both T-helper 1- and T-helper 2- (Th1 or Th2) mediated responses. We believe that the results of our research will open up new avenues for the formulation of oral vaccines.

Introduction

The respiratory, gastrointestinal and urinogenital mucosa are the main portals of entry for microbial pathogens (such as HIV, hepatitis and influenza). Therefore, secretory IgA induced by vaccination via the oral route can effectively prevent many infectious diseases by forming a defense network involving distant mucosal sites (Jakobsen and Jonsdottir, 2003, Mestecky et al., 1997, Patel et al., 2007, Minato et al., 2003). In addition, the immunity generated in this way could be transferred to provide a robust systemic immune response. Furthermore, the oral route allows easy administration, has a low risk of contamination, at a reduced cost and with increased patient compliance. However, oral immunization of vaccines is one of the greatest challenges in pharmaceutical technology. This is because the acidic gastric pH and proteolytic enzymes create a harmful environment that destroys the activity of vaccines, leading to an inefficient uptake by the gut and associated lymphoid tissue (GALT) and resulting in a poor immune response. In addition, it is generally believed that particle translocation mainly happens in the follicle associated epithelium (FAE) (Jani et al., 1990, Lavelle et al., 1995, O’Hagan, 1990, Mestecky et al., 1997), because M cells mainly located within the FAE are responsible for the efficient uptake of large particles via endocytosis (Des Rieux et al., 2006). So, the development of optimum formulations which can optimize particle uptake by M cells is another important factor that should be considered when designing oral vaccine adjuvants.

In recent years, there have been a number of important breakthroughs in the development of biodegradable materials such as liposomes, chitosan and polymeric nanoparticles for use as oral vaccine immunological adjuvants. However, these structurally unstable “soft” materials are prone to hydrolysis in the harsh gastric environment; some of them would even start leaking as soon as they come into contact with water (Kwon et al., 2007, Slowing et al., 2008). This premature release presents a major challenge for antigen-delivery due to the harsh gastrointestinal environment. In addition, when antigens are loaded in some traditional carriers, such as starch microcapsules, PLA/PLGA microspheres and liposomes, they are prone to denature due to the introduction of organic solvents. Therefore, the development of novel oral vaccine adjuvants and novel oral vaccine delivery systems would be very helpful.

Based on the above considerations, we have designed three kinds of porous silica nanoparticles as oral vaccine adjuvants and loaded a model antigen (BSA) into the pores. In recent years, research has led to advances in the fabrication and characterization of porous silica nanoparticles with very attractive properties, such as good chemical stability, good biocompatibility, low toxicity, and the option of modifying the particle size and pore diameter (Du and He, 2010). Here, we report their application as an oral vaccine immunological adjuvant taking advantages of their unique characteristics. It would be expected that a porous silica nanoparticle would be an excellent candidate for oral vaccine delivery. Firstly, the porous silica carriers allow high loading of antigens due to their high surface area and large pore volume. Secondly, the nonsoluble silica could provide slow release of antigens, creating a “depot” effect and its stable and rigid framework would provide suitable protection against the effects of gastric acid and proteases in the alimentary tract. Thirdly, instead of using an organic solvent, the antigens will retain their activity when they are incorporated into the silica pores using phosphate buffered solution. Finally, although the optimal size for nanoparticles to be transcytosed by M cells still remains controversial, the synthetic silica adjuvants with sizes of 130 nm, 430 nm and 1–2 μm are expected to optimize the uptake by M cells.

The relationship between the silica architecture (including pore size/geometry as well as particle size) and immunology was systematically investigated in our research following oral immunization of BALB/c mice with model antigen (BSA). The obtained BSA/silica systems were characterized using SEM, TEM and nitrogen adsorption and then were compared with regard to their efficacy in stimulating immune responses (humoral and mucosal).