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Bacterial infection of pressure ulcers (PUs) are a notable source of hospitalization for individuals with diabetes. This study evaluated the safety profile and efficacy of auranofin to treat diabetic PUs infected with methicillin-resistant Staphylococcus aureus (MRSA). PUs were infected with MRSA in diabetic TALLYHO/JngJ mice and then treated with topical auranofin (2%), topical mupirocin (2%), or oral clindamycin (30 mg/kg) for four days. PUs were harvested post-treatment to enumerate bacterial burden and determine expression of cytokines/growth factors. Landrace cross pigs were exposed topically to auranofin (1%, 2%, and 3%) for 4–14 days and evaluated for signs of localized or systemic toxicity. Auranofin eradicated MRSA in PUs within four days (7.92-log10 reduction) in contrast to mupirocin (2.15-log10 reduction) and clindamycin (0.73-log10 reduction). Additionally, auranofin treatment resulted in decreased expression of pro-inflammatory cytokines and increased expression of biomarkers associated with re-epithelization of wounded tissue, confirmed with histopathologic analysis. No significant histopathologic lesions were present on porcine skin sites exposed to topical auranofin. Additionally, minimal accumulation of plasma gold and no systemic toxicity was observed in pigs exposed to topical auranofin. Auranofin appears to be a potent and safe topical agent to further investigate for treatment of mild-to-moderate MRSA-infected diabetic PUs. Published version

Pressure ulcers (PUs) are a source of morbidity in individuals with restricted mobility including individuals that are obese or diabetic. Infection of PUs with pathogens, including methicillin-resistant Staphylococcus aureus (MRSA), impairs ulcers from healing. The present study evaluated ebselen as a topical antibacterial to treat MRSA-infected PUs. Against two different S. aureus strains, including MRSA USA300, resistance to ebselen did not emerge after 14 consecutive passages. Resistance to mupirocin emerged after only five passages. Additionally, ebselen was found to exert a modest postantibiotic effect of five hours against two MRSA strains. Ebselen was subsequently evaluated in MRSA-infected PUs in two models using obese and diabetic mice. In obese mice, topical ebselen (89.2% reduction) and oral linezolid (84.5% reduction) similarly reduced the burden of MRSA in infected PUs. However, in diabetic mice, topical ebselen (45.8% reduction in MRSA burden) was less effective. Histopathological evaluation of ulcers in diabetic mice determined that ebselen treatment resulted in fewer bacterial colonies deep within the dermis and that the treatment exhibited evidence of epithelial regeneration. Topical mupirocin was superior to ebselen in reducing MRSA burden in infected PUs both in obese (98.7% reduction) and diabetic (99.3% reduction) mice. Ebselen's antibacterial activity was negatively impacted as the bacterial inoculum was increased from 10(5) CFU/mL to 10(7) CFU/mL. These results suggest that a higher dose of ebselen, or a longer course of treatment, may be needed to achieve a similar effect as mupirocin in topically treating MRSA-infected pressure ulcers. National Institute of Allergy and Infectious Diseases of the National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID) [R01AI130186] This work was supported by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (URL: https://www.niaid.nih.gov/) awarded to MNS (grant number R01AI130186). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Additional file 1: Figure S1. BBB tight junctions following blast-induced traumatic brain injury. Representative immunofluorescence microscopy images of claudin-5 (green) costained with RECA-1 (red) and DAPI (blue) in sham (A-B), single blast (C-D), triple blast (E-F) and triple blast day 3 groups (G-H) (n=4). (I) There was no significant change in claudin-5 expression in the experimental groups compared to sham control. Scale bar=50 µm, * p < 0.05; ** p < 0.01; *** p < 0.001, error bars show standard deviation (SD). Figure S2. ZO-1 expression in repetitive blast-induced traumatic brain injury. Representative immunofluorescence microscopy images of ZO-1 (green) costained with RECA-1 (red) and DAPI (blue) in sham (A-B), single blast (C-D), triple blast (E-F) and triple blast day 3 groups (G-H) (n=4). (I) There is no significant change in ZO-1 expression compared to sham control. Scale bar=50 µm, * p < 0.05; ** p < 0.01; *** p < 0.001, error bars show standard deviation (SD). Figure S3. Basement membrane of the BBB following blast-induced traumatic brain injury. Representative immunofluorescence microscopy images of collagen IV (green) costained with RECA-1 (red) and DAPI (blue) in sham (A-B), single blast (C-D), triple blast (E-F) and triple blast day 3 groups (G-H) (n=4). (I) While not significant, collagen IV expression in single blast and triple blast injury was modestly diminished compared to sham control. Three days following repetitive injury, recovery of collagen IV expression was identified. Scale bar=50 µm, * p < 0.05; ** p < 0.01; *** p < 0.001, error bars show standard deviation (SD). Figure S4. PDGFR-β expression in blast-induced traumatic brain injury. Representative immunofluorescence microscopy images of PDGFR-β (green) costained with RECA-1 (red) and DAPI (blue) in sham (A-B), single blast (C-D), triple blast (E-F) and triple blast day 3 groups (G-H) (n=4). (I) Quantitative analysis revealed that PDGFR-β expression was slightly elevated in the triple blast group. A rescue phenotype was observed three days following bTBI. Scale bar=50 µm, * p < 0.05; ** p < 0.01; *** p < 0.001, error bars show standard deviation (SD). Figure S5. Aquaporin-4 expression in blast-induced traumatic brain injury. Representative immunofluorescence microscopy images of AQP4 (green) costained with RECA-1 (green) and DAPI (blue) in sham (A-B), single blast (C-D), triple blast (E-F) and triple blast day 3 groups (G-H) (n=4). (I) Quantitative analysis revealed a descending AQP-4 expression trend in the single blast (1.10-fold, not significant), triple blast (1.10-fold, not significant), and triple blast day 3 (1.16-fold, not significant) groups compared to sham control. Scale bar=50 µm, * p < 0.05; ** p < 0.01; *** p < 0.001, error bars show standard deviation (SD). Figure S6. Astrocyte coverage of the BBB following blast-induced traumatic brain injury. Representative immunofluorescent microscopy images of brain capillaries (red) were costained with RECA-1 (red), aquaporin-4 (green), GFAP (white) and nuclei (DAPI). Scale bar=10 µm. White arrows demonstrate astrocytic displacement. Vessels in A-H are presented in longitudinal section, and I-P are presented in cross-section. Figure S7. Microglial activation in repetitive blast injury. Representative immunohistochemistry images of IBA-1 (brown) in sham (A), single blast (B), triple blast (C), and triple blast day 3 groups. (A) In the sham control, microglia are predominantly ramified. (B-D) Hypertrophic (black arrows), bushy (blue arrows), and amoeboid (red arrows) morphology can be seen in response to single and repetitive blast injuries. Scale bar=50 µm.

Cyanide is a highly toxic industrial chemical that is widely used by manufactures. Smoke inhalation during household fires is the most common source of cyanide poisoning while additional risks to civilians include industrial accidents and terrorist attacks. Despite the risks to large numbers of individuals, an antidote capable of administration at scale adequate for a mass casualty, prehospital scenario does not yet exist. Previously, we demonstrated that intravenous cisplatin analogues accelerate recovery from cyanide poisoning in mice and rabbits. Of the dozens of platinum-based organometallic complexes tested, hexachloroplatinate (HCP) emerged as a promising lead compound, exhibiting strong affinity for cyanide and efficacy across model systems. Here, we show HCP is an antidote to lethal cyanide exposure and importantly is effective when delivered intramuscularly. The pharmacokinetic profile of HCP exhibited bioavailability in the systemic circulation 2.5 minutes post-treatment and subsequent renal clearance of HCP-cyanide. HCP restored parameters of cellular physiology including cytochrome oxidase redox state and TCA cycle metabolism. We next validated these findings in a large animal model (swine). Finally, preclinical safety studies in mice revealed minimal toxicity. Cumulatively, these findings demonstrate hexachloroplatinate is a promising lead compound for development of an intramuscular injectable cyanide antidote for mass casualty scenarios.