BACKGROUND-AIM Macrophages play a pivotal role in wound healing process contrasting infections and producing cytokines and growth factors. Chronic ulcers and impairment of tissue repair are characterized by persistent inflammation, hypoxia, bacterial infections and persistence of pro-inflammatory macrophages without transitioning to the anti-inflammatory phenotype. Oxygen-loaded nanodroplets (OLNDs), made of a decafluoropentane inner core carrying and releasing oxygen, and a dextran outer shell, were proposed as potential tools to improve chronic wounds healing: the final aim was to evaluate their effects on macrophages infected with Enterococcus faecalis in a hypoxic environment. METHODS THP-1 differentiated into macrophages infected with Enterococcus faecalis and murine BMDM were treated with 10% v/v OLNDs and OFNDs (Oxygen-free nanodroplets) both in normoxia (20% O2) and hypoxia (1% O2). The production of nitric oxide (NO) and reactive oxygen species (ROS) was assessed by colorimetric Griess assay and fluorescent DCFDA, respectively. The ability of macrophages to kill E. faecalis was evaluated by Giemsa staining after modified gentamicin protection assay. Cytokines, chemokines, and growth factors production was quantified by ELISA. RESULTS ROS production was stimulated by both OLNDs and OFNDs in BMDM, while NO was significantly (p<0,05) induced in a dose-dependent manner only by OLNDs. Despite this, the capability of differentiated THP-1 to kill E.faecalis was not improved. Proinflammatory cytokines TNFα, IL-6 and IL-1ß were all induced by infection, but TNFα and IL-6 were reduced by hypoxia (p<0,05) in differentiated THP-1. Both OLNDs and OFNDs further reduced TNFα and IL-6 in hypoxic and normoxic conditions (p<0,05). CONCLUSIONS Even though OLNDs induced the key molecules (NO and ROS) used by macrophages to kill microorganisms, E.faecalis killing was not improved. OLNDs and OFNDs inhibited TNFα and IL-6 secretion induced by E.faecalis infection, showing an anti-inflammatory activity probably due to the outer dextran shell. These data suggest that OLNDs can be envisaged as a double edge tool able to modulate the inflammatory response of macrophages during hypoxia.

Effects of oxygen-laden nonodroplets on enterococcus faecalis-infected macrophages in hypoxia / F. Perego, S. D'Alessandro, R.M. Ticozzi, B. Bressan, A. Troia, M. Prato, D. Taramelli, N. Basilico. ((Intervento presentato al convegno Pathobiology: from molecular disease to clinical application tenutosi a Firenze nel 2019.

Effects of oxygen-laden nonodroplets on enterococcus faecalis-infected macrophages in hypoxia

F. Perego;S. D'alessandro;R. M. Ticozzi;D. Taramelli;N. Basilico
2019-09-13

Abstract

BACKGROUND-AIM Macrophages play a pivotal role in wound healing process contrasting infections and producing cytokines and growth factors. Chronic ulcers and impairment of tissue repair are characterized by persistent inflammation, hypoxia, bacterial infections and persistence of pro-inflammatory macrophages without transitioning to the anti-inflammatory phenotype. Oxygen-loaded nanodroplets (OLNDs), made of a decafluoropentane inner core carrying and releasing oxygen, and a dextran outer shell, were proposed as potential tools to improve chronic wounds healing: the final aim was to evaluate their effects on macrophages infected with Enterococcus faecalis in a hypoxic environment. METHODS THP-1 differentiated into macrophages infected with Enterococcus faecalis and murine BMDM were treated with 10% v/v OLNDs and OFNDs (Oxygen-free nanodroplets) both in normoxia (20% O2) and hypoxia (1% O2). The production of nitric oxide (NO) and reactive oxygen species (ROS) was assessed by colorimetric Griess assay and fluorescent DCFDA, respectively. The ability of macrophages to kill E. faecalis was evaluated by Giemsa staining after modified gentamicin protection assay. Cytokines, chemokines, and growth factors production was quantified by ELISA. RESULTS ROS production was stimulated by both OLNDs and OFNDs in BMDM, while NO was significantly (p<0,05) induced in a dose-dependent manner only by OLNDs. Despite this, the capability of differentiated THP-1 to kill E.faecalis was not improved. Proinflammatory cytokines TNFα, IL-6 and IL-1ß were all induced by infection, but TNFα and IL-6 were reduced by hypoxia (p<0,05) in differentiated THP-1. Both OLNDs and OFNDs further reduced TNFα and IL-6 in hypoxic and normoxic conditions (p<0,05). CONCLUSIONS Even though OLNDs induced the key molecules (NO and ROS) used by macrophages to kill microorganisms, E.faecalis killing was not improved. OLNDs and OFNDs inhibited TNFα and IL-6 secretion induced by E.faecalis infection, showing an anti-inflammatory activity probably due to the outer dextran shell. These data suggest that OLNDs can be envisaged as a double edge tool able to modulate the inflammatory response of macrophages during hypoxia.
Settore MED/04 - Patologia Generale
Settore MED/07 - Microbiologia e Microbiologia Clinica
Settore MED/46 - Scienze Tecniche di Medicina di Laboratorio
Effects of oxygen-laden nonodroplets on enterococcus faecalis-infected macrophages in hypoxia / F. Perego, S. D'Alessandro, R.M. Ticozzi, B. Bressan, A. Troia, M. Prato, D. Taramelli, N. Basilico. ((Intervento presentato al convegno Pathobiology: from molecular disease to clinical application tenutosi a Firenze nel 2019.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/707050
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