An Evaluation of Urease A Subunit Nanocapsules as a Vaccine in a Mouse Model of Helicobacter pylori Infection

Abstract

Using removable silica templates, protein nanocapsules comprising the A subunit of Helicobacter pylori urease (UreA) were synthesised. The templates were of two sizes, with solid core mesoporous shell (SC/MS) silica templates giving rise to nanocapsules of average diameter 510 nm and mesoporous (MS) silica templates giving rise to nanocapsules of average diameter 47 nm. Both were shown to be highly monodispersed and relatively homogenous in structure. Various combinations of the nanocapsules in formulation were assessed as vaccines in a mouse model of H. pylori infection. Immune responses were evaluated and protective efficacy assessed. It was demonstrated that vaccination of mice with the larger nanocapsules combined with an adjuvant was able to significantly reduce colonisation.

Keywords: Helicobacter pylori; nanocapsule; silica nanoparticle template; urease alpha subunit; vaccination.

Figure 1

(A) SDS-PAGE gel of recombinant UreA following endotoxin removal and final concentration. Lane 1: Precision Plus Protein™ Unstained Protein Standards; Lane 2: neat sample; Lane 3: ½ dilution of sample; Lane 4: 1/10 dilution of sample; Lane 5: estimated 1 µg of sample. (B) Immunoblot of recombinant UreA following endotoxin removal and final concentration. Lane 1: estimated 1 µg of sample; Lane 2: Precision Plus Protein™ Kaleidoscope™ Prestained Protein Standards.

Figure 2

TEM image analysis of SC/MS silica templates (A) and MS templates (B). (C,D) show SC/MS UreA-based nanocapsules and MS UreA-based nanocapsules, respectively. Imaging revealed that templates and nanocapsules were homogenous in size and morphology within each size range.

Figure 3

DLS analysis of nanocapsules. This indicates SC/MS UreA nanocapsules were well dispersed. MS UreA nanocapsules showed some evidence of aggregation. Both capsule size ranges possessed a negative zeta potential.

Figure 4

Titre of immunoglobulins in immunised mice pre-challenge (left) and post-challenge (right) with asymptomatic H. pylori. (A,B) IgG. (C,D) IgG1. (E,F) IgG2c. See Table 1 for group details. Note that the third group (PBS-Naïve) was not challenged. * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001 and **** p ≤ 0.0001.

Figure 5

qPCR detection of H. pylori burdens in stomachs of vaccinated mice. Mice vaccinated with SC/MS nanocapsules with TiterMax had significantly reduced levels of gastric H. pylori burden compared to the infected control (** p ≤ 0.01, one-tailed). No significant difference was detected between mice vaccinated with any other vaccine groups.

Figure 6

Analysis of infiltrating immune cells in stomach tissue of vaccinated mice. (A–C) Infiltration of CD4⁺, CD8⁺ and CDIIIb⁺ cells, respectively. (Note that cells values for SL3261pYZ97 vaccinated mice are pools of cells from 5 mice, SEM not possible) * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001 and **** p ≤ 0.0001.