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Polysaccharide microparticles as an oral vaccine platform: proof-of-concept in male rats using a GnRH-targeting
Maximiliano Wilda1,2, Maria Victoria Rocca1, M Solana Pesca3
1Centro de Virología Humana y Animal (CEVHAN) - CONICET-UAI, Ciudad Autónoma de Buenos Aires, Argentina.
Background:
Immunocastration through vaccination against gonadotropin-releasing hormone (GnRH) represents a humane alternative to surgical castration in animal production. However, oral delivery of protein antigens remains challenging due to gastrointestinal degradation and poor mucosal immunogenicity. Polysaccharide microparticles (PSMs), particularly alginate-chitosan systems, offer protection against gastric degradation and enhanced mucosal uptake.
Objective:
To evaluate whether alginate-chitosan microparticles entrapping a recombinant GnRH-fusion protein (GF45) can elicit an immunocastrating response in male rats after oral administration.
Methods:
GF45, a 45 kDa fusion peptide containing four tandem GnRH repeats fused to Rickettsia heat-shock protein fragments, was expressed in E. coli BL21(DE3) and purified under denaturing conditions followed by refolding. Microparticles were prepared by ionic gelation achieving 64% encapsulation efficiency. Male Sprague-Dawley rats (4 weeks) received oral prime-boost immunizations (weeks 4 and 7) with 200 µg GF45 in microparticles (n = 10). Controls included empty microparticles (NC, n = 10) and subcutaneous GF45 + Freund's adjuvant (PC, n = 10). Serum testosterone and anti-GF45 IgG were monitored until week 16 via ELISA. Testes were weighed, volumetrically measured by Archimedes' principle, and histologically examined using Johnsen's scoring system.
Results:
Oral GF45 reduced circulating testosterone by 95% (0.7 ± 0.3 vs 14.2 ± 1.8 ng dL-1 in NC; p < 0.0001) and induced anti-GF45 antibodies reaching 88.8% of the injectable response (titres log10 4.1 ± 0.3 vs 4.4 ± 0.2; p = 0.08). Seroconversion began at week 6 post-prime, reaching 100% by week 16. Testis weight decreased by 52% and volume by 75% (both p < 0.01), accompanied by marked seminiferous-tubule atrophy (diameter reduced from 286 ± 9 µm to 124 ± 8 µm) and reduced Johnsen scores (3.2 ± 0.4 vs 9.1 ± 0.2; p < 0.001). Empty microparticles produced no effects.
Conclusion:
Alginate-chitosan microparticles provide a practical, needle-free platform for oral immunocastration via induction of anti-GnRH autoimmunity. This polysaccharide-based system represents a versatile oral antigen carrier with potential applications in veterinary medicine and livestock production.
