Degradable magnesium implant-associated infections by bacterial biofilms induce robust localized and systemic inflammatory reactions in a mouse model

Abstract

Biomaterial-associated Pseudomonas aeruginosa biofilm infections constitute a cascade of host immune reactions ultimately leading to implant failure. Due to the lack of relevant in vivo biofilm models, the majority of the studies report host immune responses to free-living or planktonic bacteria, while bacteria in clinical situations live more frequently as biofilm communities than as single cells. The present study investigated host immune responses to biomaterial-associated P. aeruginosa biofilms in a clinically relevant mouse model. Previously, we reported metallic magnesium, a prospective biodegradable implant, to be permissive for bacterial biofilm in vivo even though it exhibits antibacterial properties in vitro. Therefore, magnesium was employed as biomaterial to investigate in vivo biofilm formation and associated host immune responses by using two P. aeruginosa strains and two mouse strains. P. aeruginosa formed biofilm on subcutaneously implanted magnesium disks. Non-invasive in vivo imaging indicated transient inflammatory responses at control sites, whereas robust prolonged interferon-β (IFN-β) expression was observed from biofilm in a transgenic animal reporter. Furthermore, immunohistology and electron microscopic results showed that bacterial biofilms were located in 2D immediately on the implant surface and at a short distance in the adjacent tissue. These biofilms were surrounded by inflammatory cells (mainly polymorphonuclear cells) compared to the controls. Interestingly, even though the number of live bacteria in various organs remained below detectable levels, splenomegaly indicated systemic inflammatory processes. Overall, these findings confirmed the resistance of biofilm infections in vivo to potentially antibacterial properties of magnesium degradation products. In vivo imaging and histology indicated the induction of both local and systemic host inflammatory responses to P. aeruginosa biofilms. Even though the innate host immune defenses could not eliminate the local infection for up to two weeks, there was no apparent systemic bacteremia and all the animals investigated survived the infection.

Non-Invasive Luciferase Imaging of Type I Interferon Induction in a Transgenic Mouse Model of Biomaterial Associated Bacterial Infections: Microbial Specificity and Inter-Bacterial Species Interactions

“…The main difficulty in monitoring host immunity against biomaterials-related bacterial infections is that the majority of the animal models involve tissue extractions at selected time points, followed by time consuming immunological staining [ 20 ]. Therefore, non-invasive in vivo imaging of bioluminescent markers was employed to monitor inflammation within the same animal at multiple time points, thereby achieving more reproducible results while dramatically reducing the consumption of experimental animals [ 21 , 22 ]. Moreover, bioluminescent imaging significantly reduced the issues related to animal-to-animal variations and improved the biostatistical quality of data [ 23 ].…”

“…Previously, a luciferase reporter mouse model which allows tracking of interferon beta (IFN-β) gene induction in vivo was established. Recently, this mouse model was used successfully for the non-invasive imaging of IFN-β upon infections with Pseudomonas aeruginosa biofilms on degradable implants [ 22 ]. This study suggested that IFN-β luciferase reporter mouse model could be used as a highly sensitive reporter system to investigate host inflammation against biomaterial-related infections with diverse periodontal pathogens.…”

Interacción de los materiales biofuncionales y/o bioactivos con los medios biológicos

“…Un aspecto interesante relacionado con el estudio de la corrosión se refiere a la liberación de iones metálicos al medio que puede generar toxicidad en el tejido o ambiente aledaño [1]. En particular, la alta velocidad de corrosión de los materiales implantables base Mg produce cambios locales de pH, liberación de iones Mg 2+ y burbujas de H 2 induce efectos nocivos al tejido de las zonas aledañas al implante y disminuye las propiedades mecánicas del metal [19,[27][28][29][30]. Para reducir los efectos nocivos de la biodegradación de los BMB se han desarrollado diversas aleaciones con componentes adecuados y modificaciones superficiales como estrategias para controlar la taza de corrosión pero no se ha hallado una solución definitiva para eliminar dichos efectos.…”

“…Los fenómenos de la interfaz BMB/medio biológico son muy complejos. Si bien la alcalinidad producida por la corrosión del Mg y sus aleaciones puede resultar benéfica porque reduce la proliferación bacteriana, dicha degradacion también induce efectos dañinos al tejido de las zonas aledañas al implante, relacionadas con el incremento de pH, liberación de H 2 (asociados al proceso de corrosión) y disminución de las propiedades mecanicas del metal [19,[27][28][29][30]. Al respecto, se han encontrado formaciones de cavidades gaseosas subcutáneas en el sitio de implantación de materiales base Mg frecuentemente formadas en periódos iniciales postimplante, generando respuesta inflamatoria en experimentos in vivo [19].…”

“…Reportes recientes de ensayos in vitro indican como potencial ventaja que los biomateriales base Mg podrían proveer un efecto antibacteriano. Sin embargo, éste es un tema controversial que requiere mayor discusión [29,93,[264][265][266][267]. Según nuestro conocimiento no existen informes previos sobre el estudio de la distribución de bacterias asociadas con la aleación AZ31 durante su degradación, que es importante para contribuir a la elucidación de los procesos complejos que ocurren en la interfaz metal degradable / medio biológico, la cual muestra cambios continuos.…”