Chitosan/2H,3H-decafluoropentane nanodroplets as anti-bacterial and skin-friendly devices for ultrasound-triggered transdermal delivery.

Hypoxia, inflammation, and impaired skin tissue remodeling represent three typical features of chronic wounds (CWs). In addition, CWs are often worsened by bacterial infections. Intriguingly, ultrasound (US)-responsive oxygen-loaded nanodroplets (OLNDs) [1-2] have proven effective in delivering oxygen to hypoxic tissues, restoring normoxia-like levels of inflammatory and matrix-remodeling molecules [3-6]. On the other hand, chitosan - a natural polysaccharyde suitable for nanodroplet’s shell manufacturing - has displayed anti-bacterial properties [7]. Based on these preconditions, in the present work chitosan-shelled/2H,3H-decafluoropentane-cored OLNDs and their oxygen-free counterpart (oxygen-free nanodroplets, OFNDs) were challenged for their anti-bacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) and Streptococcus pyogenes. Additionally, nanodroplet biocompatibility with human skin cells and suitability for US-mediated transdermal delivery was assessed. MRSA or S. pyogenes growth was monitored upon incubation with/without OLNDs and OFNDs for increasing times (2, 3, 4, 6, and 24 h). According to cell counts, both OLNDs and OFNDs significantly decreased the growth of either bacterial species up to 6 h, whereas slight bacteriostatic activity was observed at 24 h for S. pyogenes, thus confirming short-term anti-bacterial properties of chitosan in the nanodroplet’s outer shell. Notably, similar results were obtained when complementary US was applied to bacteria. Further analysis by confocal microscopy displayed OLND and OFND adhesion to the bacterial wall with no evidence of cellular internalization after 3 or 24 h of incubation. On the other hand, human keratinocytes (HaCaT cell line) were incubated with/without OLNDs or OFNDs for 24 h either in normoxia (20% O2) or hypoxia (1% O2). As assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, OLNDs did not affect cell viability, whereas OFNDs slightly did, thereby suggesting a protective role for oxygen in the nanodroplet’s inner core. Finally, US-mediated OLND transdermal delivery was studied by employing a sonophoresis assay, displaying that OLNDs effectively trespassed a porcine skin layer upon sonication. Based on these data, OLNDs appear as innovative and promising anti-bacterial/skin-friendly devices to treat infected CWs.