Macrophage-Mediated In Vivo Intracellular Crystallization of Aluminum Oxyhydroxide Adjuvant in Vaccine-Induced Granulomas

Pérez, Estela ; Navarro, Marta (Universidad de Zaragoza) ; Ibarra, Alfonso (Universidad de Zaragoza) ; de Blas, Ignacio (Universidad de Zaragoza) ; Pérez, Marta (Universidad de Zaragoza) ; Luján, Lluís (Universidad de Zaragoza) ; Sebastián, Víctor (Universidad de Zaragoza)

Resumen: Aluminum (Al) in the form of γ-aluminum oxyhydroxide (γ-AlOOH) nanoparticles has been used as a vaccine adjuvant for nearly a century. At injection sites, animals and humans develop granulomatous inflammation, in which macrophages may contain intracellular crystalline-like structures, so-called crystalloid bodies (CBs), morphologically distinct from the injected adjuvant. This raised the hypothesis that CBs may originate from the in vivo crystallization of γ-AlOOH within macrophages. To test this, six sheep received two subcutaneous doses of a γ-AlOOH-adjuvanted viral vaccine, while six controls received the same antigen without an adjuvant. All injection sites (AlOOH: n = 12; controls: n = 12) were examined by hematoxylin and eosin, modified aluminum hematoxylin, and lumogallion fluorescence. Al and CBs occurred exclusively in AlOOH-vaccinated tissues. The adjuvant stock and vaccine formulations were characterized ex situ using a combination of structural and imaging techniques. Four granulomas containing CBs, collected 133 days postinjection, were further examined using in situ imaging and crystallographic approaches. No crystalline structures resembling CBs were identified in the injected products; only γ-AlOOH nanoparticles were present. In post-mortem granulomas, macrophages contained Al-positive vacuoles filled with nanoparticle aggregates compatible with γ-AlOOH, often adjacent to CBs. Crystallographic analyses demonstrated that CBs were highly ordered γ-Al2O3 microcrystals, and advanced SEM-EDS with elemental mapping similarly distinguished γ-AlOOH nanoparticles and CBs as separate phases. These findings support an in vivo formation of γ-Al2O3 microcrystals from γ-AlOOH nanoparticles within macrophages under physiological conditions. To our knowledge, these findings provide evidence consistent with Al crystallization within mammalian cells and suggest that macrophage phagolysosomal conditions may represent a previously unrecognized biomimetic pathway for γ-Al2O3 formation. This finding may offer a biological model for the development of green and low-energy synthesis strategies for γ-Al2O3, with potential industrial applications as well as relevance to immunotoxicology, pathology, and cell
biology in the context of vaccine adjuvants.

Idioma: Inglés

DOI: 10.1021/acsami.6c04089

Año: 2026

Publicado en: ACS applied materials & interfaces 18, 18 (2026), 25960-25978
ISSN: 1944-8244

Financiación: info:eu-repo/grantAgreement/ES/MCIU/RTI2018-096172-B-C33
Financiación: info:eu-repo/grantAgreement/ES/MICINN/PID2021-127847OB-I00
Financiación: info:eu-repo/grantAgreement/ES/MICIU/CEX2023-001286-S
Financiación: info:eu-repo/grantAgreement/ES/MICIU/PID2024–160339OB-I00

Tipo y forma: Artículo (Versión definitiva)

Área (Departamento): Área Sanidad Animal (Dpto. Patología Animal)
Área (Departamento): Área Anatom.Anatom.Patológ.Com (Dpto. Anatom.,Embri.Genét.Ani.)
Área (Departamento): Área Ingeniería Química (Dpto. Ing.Quím.Tecnol.Med.Amb.)
Área (Departamento): Área RPT Laboral INMA (Institutos Univ. de Investig.)