Background: In hospitalized patients, chronic wounds are often worsened by microbial complications, and tissue hypoxia is a common microenvironmental feature during inflammation associated with bacterial infection, usually associated with Staphylococcus aureus, in particular methicillin-resistant S. aureus (MRSA) strains. The limited solubility, short half-life, and variable adsorptions of drugs along with the limited efficacy of transdermal drug delivery are driving the attention towards new therapeutic approaches. The known benefits of nanotechnology combined with the emerging advantages of using natural antimicrobial polysaccharides, such as chitosan, have paved the way to develop nonconventional and innovative adjuvant therapies for infected wounds. Chitosan-shelled oxygenloaded nanobubble (OLNB) and chitosan-shelled oxygen-loaded nanodroplet (OLND) potential, as treatments for infected wounds, was here assessed by investigating their antibacterial activity against MRSA along with toxicity on human keratinocytes. Material/methods: Medium molecular weight (MW) chitosan-shelled OLNBs and OLNDs were prepared and characterized for morphology, size, and avarage diameter. For confocal microscopy analyses, MRSA bacteria were stained with propidium iodide (PI) and incubated alone or with 10% v/v fluorescein isotiocyanate (FITC)-labeled OLNBs, oxygen-free nanobubbles (OFNBs), OLNDs, and oxygen-free nanodroplets (OFNDs) up to 24 h. Cytostatic activity was evaluated by incubation of MRSA alone or with 10% v/v OLNBs, OFNBs, OLNDs, and OFNDs within 24 h. Toxicity on human skin cell was studied by lactate dehydrogenase assay after incubating HaCaT keratinocytes alone or with 10% v/v OLNBs, OFNBs, OLNDs, OFNDs either in normoxia and hypoxia for 24 h. Results: Both OLNBs and OLNDs adhered on staphylococcal cell walls (Fig. 1, A) and showed shortterm (up to 4 h) antibacterial activity against MRSA, due to chitosan presence as antimicrobial compound in the shell (Fig. 1, B). No differences between OLNB and OFNB preparation were observed, as well as between OLND and OFND formulations. OLNBs, as well as OLNDs were not toxic to keratinocytes, whereas OFNBs and OFNDs slightly affected hypoxic cell viability, thus suggesting a protective role for oxygen on human skin cells. Nanodroplets are able to release from the core more oxygen than nanobubbles, so we focused attention on this nanocarrier. A new OLND formulation was prepared using low molecular weight (LW) chitosan for the shell manufacturing. Interestingly, LW OFNDs, compared with OFNDs, were not toxic to keratinocytes and LW OLNDs (in contrast with OLNDs and OFNDs), as well LW OFNDs, are internalised by MRSA (Fig. 1, C). Conclusions: Chitosan-shelled OLNBs and OLNDs development has the potential to be a highly innovative, cost-effective and non-toxic product for the concurrent treatment of hypoxic infected tissues. Such treatment could represent an innovative therapy of infected chronic wounds improving the patient quality of life, especially the elderly. Based on these data, future studies on chitosan nanodroplets, especially manufactured with LW chitosan, are heavily encouraged.

Chitosan-shelled oxygen-loaded nanobubble and nanodroplet comparison: new insights into infected chronic wounds caused by MRSA

FINESSO, NICOLE;BANCHE, Giuliana;ALLIZOND, Valeria;PRATO, Mauro;LUGANINI, ANNA;GIRIBALDI, Giuliana;KHADJAVI, AMINA;TULLIO, Viviana Cristina;MANDRAS, Narcisa;CAVALLI, Roberta;GUIOT, Caterina;CUFFINI, Annamaria
2017

Abstract

Background: In hospitalized patients, chronic wounds are often worsened by microbial complications, and tissue hypoxia is a common microenvironmental feature during inflammation associated with bacterial infection, usually associated with Staphylococcus aureus, in particular methicillin-resistant S. aureus (MRSA) strains. The limited solubility, short half-life, and variable adsorptions of drugs along with the limited efficacy of transdermal drug delivery are driving the attention towards new therapeutic approaches. The known benefits of nanotechnology combined with the emerging advantages of using natural antimicrobial polysaccharides, such as chitosan, have paved the way to develop nonconventional and innovative adjuvant therapies for infected wounds. Chitosan-shelled oxygenloaded nanobubble (OLNB) and chitosan-shelled oxygen-loaded nanodroplet (OLND) potential, as treatments for infected wounds, was here assessed by investigating their antibacterial activity against MRSA along with toxicity on human keratinocytes. Material/methods: Medium molecular weight (MW) chitosan-shelled OLNBs and OLNDs were prepared and characterized for morphology, size, and avarage diameter. For confocal microscopy analyses, MRSA bacteria were stained with propidium iodide (PI) and incubated alone or with 10% v/v fluorescein isotiocyanate (FITC)-labeled OLNBs, oxygen-free nanobubbles (OFNBs), OLNDs, and oxygen-free nanodroplets (OFNDs) up to 24 h. Cytostatic activity was evaluated by incubation of MRSA alone or with 10% v/v OLNBs, OFNBs, OLNDs, and OFNDs within 24 h. Toxicity on human skin cell was studied by lactate dehydrogenase assay after incubating HaCaT keratinocytes alone or with 10% v/v OLNBs, OFNBs, OLNDs, OFNDs either in normoxia and hypoxia for 24 h. Results: Both OLNBs and OLNDs adhered on staphylococcal cell walls (Fig. 1, A) and showed shortterm (up to 4 h) antibacterial activity against MRSA, due to chitosan presence as antimicrobial compound in the shell (Fig. 1, B). No differences between OLNB and OFNB preparation were observed, as well as between OLND and OFND formulations. OLNBs, as well as OLNDs were not toxic to keratinocytes, whereas OFNBs and OFNDs slightly affected hypoxic cell viability, thus suggesting a protective role for oxygen on human skin cells. Nanodroplets are able to release from the core more oxygen than nanobubbles, so we focused attention on this nanocarrier. A new OLND formulation was prepared using low molecular weight (LW) chitosan for the shell manufacturing. Interestingly, LW OFNDs, compared with OFNDs, were not toxic to keratinocytes and LW OLNDs (in contrast with OLNDs and OFNDs), as well LW OFNDs, are internalised by MRSA (Fig. 1, C). Conclusions: Chitosan-shelled OLNBs and OLNDs development has the potential to be a highly innovative, cost-effective and non-toxic product for the concurrent treatment of hypoxic infected tissues. Such treatment could represent an innovative therapy of infected chronic wounds improving the patient quality of life, especially the elderly. Based on these data, future studies on chitosan nanodroplets, especially manufactured with LW chitosan, are heavily encouraged.
27th European Congress of Clinical Microbiology and Infectious Diseases
Vienna, Austria
22 - 25 April 2017
ESCMID Online Lecture Library
ESCMID
P0643
P0643
https://www.escmid.org/escmid_publications/escmid_elibrary/
Finesso, N; Banche, G; Allizond, V; Prato, M; Luganini, A; Troia, A; Giribaldi, G; Khadjavi, A; Tullio, V; Mandras, N; Cavalli, R; Guiot, C; Cuffini, AM
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2318/1633959
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