Strong systemic and mucosal immune responses to surface-modified PLGA microspheres containing recombinant Hepatitis B antigen administered intranasally

Abstract

Surface-modified dl-lactide/glycolide copolymer (PLGA) microspheres with chitosan (CS) were developed for nasal immunization using recombinant Hepatitis B (HBsAg) surface protein for the induction of humoral, cellular and mucosal immunity. Modified PLGA microspheres were characterized in vitro for their size, shape, entrapment efficiency and zeta potential. The nasal clearance rate was evaluated by gamma scintigraphy in rabbits. The antigen integrity, in vitro release and its stability at 37 °C were also evaluated. The designed cationic microspheres possessed 27.2 mV zeta potential and an average size less than 10 μm with antigen loading efficiency of 80 ± 5%. However, zeta potential of unmodified PLGA microspheres was measured to be negative (−8.7 mV). The modified PLGA microspheres showed the lowest nasal clearance rate when compared with unmodified PLGA microspheres and lactose powder. The antigen integrity was retained intact in encapsulated form as well as on release. The immune-stimulating activity was studied by measuring anti-HBsAg titre, secretory IgA level in serum, vaginal, nasal and salivary secretions (mucosal secretions) and cytokine level (interleukin-2 (IL-2) and interferon-γ (IFN-γ)) in spleen homogenates following nasal administration of modified PLGA microspheres in Balb/c mice and compared with alum-HBsAg vaccine injected subcutaneously. The serum anti-HBsAg titre obtained after nasal administration of modified PLGA microspheres was comparable with titre recorded after alum-HBsAg was administered subcutaneously. Moreover, alum-HBsAg vaccine did not elicit sIgA in mucosal secretions as it was induced and measured in the case of nasal administration of modified PLGA microspheres. Similarly, there was no cellular response (cytokine level) in case of alum-HBsAg vaccine. Modified PLGA microspheres (cationic microspheres) thus produced humoral (both systemic and mucosal) and cellular immune responses upon nasal administration. These data demonstrate high potential of modified PLGA microspheres for their use as a carrier adjuvant for nasal subunit vaccines.