Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2012 Jun 8:3:193.
doi: 10.3389/fmicb.2012.00193. eCollection 2012.

Nitric oxide releasing nanoparticles for treatment of Candida albicans burn infections

Chitralekha Macherla  1 , David A Sanchez, Mohammed S Ahmadi, Ernestine M Vellozzi, Adam J Friedman, Joshua D Nosanchuk, Luis R Martinez
Affiliations

Nitric oxide releasing nanoparticles for treatment of Candida albicans burn infections

Chitralekha Macherla et al. Front Microbiol. .

Abstract

Candida albicans is a leading fungal cause of burn infections in hospital settings, and sepsis is one of the principle causes of death after a severe burn. The prevalence of invasive candidiasis in burn cases varies widely, but it accounts for 3-23% of severe infection with a mortality rate ranging from 14 to 70%. Therefore, it is imperative that we develop innovative therapeutics to which this fungus is unlikely to evolve resistance, thus curtailing the associated morbidity and mortality and ultimately improving our capacity to treat these infections. An inexpensive and stable nitric oxide (NO)-releasing nanoparticle (NO-np) platform has been recently developed. NO is known to have direct antifungal activity, modulate host immune responses and significantly regulate wound healing. In this study, we hypothesized that NO-np would be an effective therapy in the treatment of C. albicans burn infections. Using a murine burn model, NO-np demonstrated antifungal activity against C. albicans in vivo, most likely by arresting its growth and morphogenesis as demonstrated in vitro. NO-np demonstrated effective antimicrobial activity against yeast and filamentous forms of the fungus. Moreover, we showed that NO-np significantly accelerated the rate of wound healing in cutaneous burn infections when compared to controls. The histological evaluation of the affected tissue revealed that NO-np treatment modified leukocyte infiltration, minimized the fungal burden, and reduced collagen degradation, thus providing potential mechanisms for the therapeutics' biological activity. Together, these data suggest that NO-np have the potential to serve as a novel topical antifungal which can be used for the treatment of cutaneous burn infections and wounds.

Keywords: Candida albicans; burn healing; collagen; nanoparticles; nitric oxide.

PubMed Disclaimer

Figures

Figure 1
Figure 1
The effect of NO-np on C. albicans growth kinetics was determined using Bioscreen C analysis. C. albicans was grown in the absence (untreated) or presence of nanoparticles with NO (NO-np) or without NO (np). Each point represents the average of three measurements. *P < 0.001 in comparing the np- or NO-np-treated group with untreated group. #P < 0.001 in comparing the np-treated group with the NO-np-treated group. This experiment was performed twice, with similar results each time.
Figure 2
Figure 2
Healing effectiveness of NO-np in C. albicans induced burn infections. (A) Burn injuries of Balb/c mice untreated, treated with np, and treated with NO-np, day 0, 1, 5, 10, and 15. Bar = 5 mm. Five animals per group were used. These experiments were performed twice with similar results. (B) Burn area closure of Balb/c mice skin lesions relative to the initial 5 mm wound. Time points are the averages of the results for measurements of six different wounds, and error bars denote SDs. *P < 0.001 in comparing the np- or NO-np-treated group with untreated group. #P < 0.001 in comparing the np-treated group with the NO-np-treated group. These experiments were performed twice with similar results. (C) NO-np prevents cutaneous spread of candidiasis in mice. Three animals per group per time point were used.
Figure 3
Figure 3
Antifungal efficacy of NO-np in C. albicans induced burn infections. (A) Burn fungal burden (colony forming units, CFU) in mice infected topically with 107 yeast cells and treated with NO-np is significantly lower than untreated or np-treated mice (n = 5). Bars are the averages of the results and brackets denote standard deviations. Asterisks denote P value significance (*P < 0.05; **P < 0.001; ns, no significance) calculated by analysis of variance and adjusted by use of the Bonferroni correction. (B) Histological analysis of Balb/c mice untreated C. albicans-infected, np-treated C. albicans-infected, and C. albicans-infected treated with NO, day 7. Mice were infected with 107 C. albicans. Representative H&E-stained sections of the skin lesions are shown with the Insets showing Periodic acid schiff staining of C. albicans. Black arrows indicate fungal pseudohyphae and hyphae. Scale bars: 20 μm.
Figure 4
Figure 4
Impact of NO-np on fungal growth and morphogenesis. (A) Time-lapse (0–60 min) microscopy was used to explore the effect of NO-np on C. albicans cell division and morphological transformation. White single arrow heads indicate budding yeast and/or pseudohyphae whereas white double arrow heads indicate nanoparticle aggregates. Images were collected at 40×. Scale bar: 10 μm. (B) Time-lapse microscopy images of untreated, np, and NO-np groups at 120 min. Images collected at 40×. Scale bar: 10 μm. (C) XTT reduction assay was used to measure the metabolic activity of NO-np-treated filamentous C. albicans in vitro. The 100% activity was set as the measurements in wells with filamentous fungi without the addition of either NO-np or np. Bars represent the average of the results and error bars denote standard deviations. Asterisks denote P value significance (**P < 0.001) calculated by analysis of variance and adjusted by use of the Bonferroni correction. These experiments were performed twice with similar results.
Figure 5
Figure 5
Effect of NO-np on C. albicans collagen degradation in burn injuries. (A) Histological analysis of Balb/c mice untreated C. albicans-infected, np-treated C. albicans-infected, and C. albicans-infected treated with NO-np, day 7. Mice were infected with 107 fungal cells. The blue stain indicates collagen. Scale bar: 20 μm. (B) Quantitative measurement of collagen intensity in 20 representative squares of the same size for untreated C. albicans-infected, np-treated C. albicans-infected, and C. albicans-infected treated with NO-np. Bars are the averages of the results, and error bars denote standard deviations. Asterisks denote P value significance (*P < 0.05) calculated by analysis of variance and adjusted by use of the Bonferroni correction.
Figure 6
Figure 6
Influence of NO-np on neutrophil and macrophage-like cell infiltration into burn wounds. (A) Histological analysis of untreated, np-treated and NO-np-treated wounded Balb/c mice, day 3. The brown staining indicates neutrophil infiltration. Representative MPO-immunostained sections of the skin lesions are shown. Scale bars: 20 μm. (B) Histological analysis of untreated, np-treated and NO-np-treated wounded Balb/c mice, day 7. The brown staining indicates macrophage-like cell infiltration. Representative Iba-1-immunostained sections of the skin lesions are shown. Scale bars: 25 μm.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Alonso R., Llopis I., Flores C., Murgui A., Timoneda J. (2001). Different adhesins for type IV collagen on Candida albicans: identification of a lectin-like adhesin recognizing the 7S(IV) domain. Microbiology 147 (Pt 7), 1971–1981 - PubMed
    1. Bahn Y. S., Molenda M., Staab J. F., Lyman C. A., Gordon L. J., Sundstrom P. (2007). Genome-wide transcriptional profiling of the cyclic AMP-dependent signaling pathway during morphogenic transitions of Candida albicans. Eukaryotic Cell 6, 2376–239010.1128/EC.00318-07 - DOI - PMC - PubMed
    1. Baroli B., Ennas M. G., Loffredo F., Isola M., Pinna R., López-Quintela M. A. (2007). Penetration of metallic nanoparticles in human full-thickness skin. J. Invest. Dermatol. 127, 1701–1712 - PubMed
    1. Broughton G. I. I., Janis J. E., Attinger C. E. (2006). The basic science of wound healing. Plast. Reconstr. Surg. 117 (Suppl. 7), 12S–34S10.1097/01.prs.0000225430.42531.c2 - DOI - PubMed
    1. Cabrales P., Han G., Nacharaju P., Friedman A. J., Friedman J. M. (2011). Reversal of hemoglobin-induced vasoconstriction with sustained release of nitric oxide. Am. J. Physiol. Heart Circ. Physiol. 300, H49–H5610.1152/ajpheart.00665.2010 - DOI - PMC - PubMed