Person:

Dai, Tianhong

Opportunistic fungal pathogens may cause superficial or serious invasive infections, especially in immunocompromised and debilitated patients. Invasive mycoses represent an exponentially growing threat for human health due to a combination of slow diagnosis and the existence of relatively few classes of available and effective antifungal drugs. Therefore systemic fungal infections result in high attributable mortality. There is an urgent need to pursue and deploy novel and effective alternative antifungal countermeasures. Photodynamic therapy (PDT) was established as a successful modality for malignancies and age-related macular degeneration but photodynamic inactivation has only recently been intensively investigated as an alternative antimicrobial discovery and development platform. The concept of photodynamic inactivation requires microbial exposure to either exogenous or endogenous photosensitizer molecules, followed by visible light energy, typically wavelengths in the red/near infrared region that cause the excitation of the photosensitizers resulting in the production of singlet oxygen and other reactive oxygen species that react with intracellular components, and consequently produce cell inactivation and death. Antifungal PDT is an area of increasing interest, as research is advancing (i) to identify the photochemical and photophysical mechanisms involved in photoinactivation; (ii) to develop potent and clinically compatible photosensitizers; (iii) to understand how photoinactivation is affected by key microbial phenotypic elements multidrug resistance and efflux, virulence and pathogenesis determinants, and formation of biofilms; (iv) to explore novel photosensitizer delivery platforms; and (v) to identify photoinactivation applications beyond the clinical setting such as environmental disinfectants.

Background: Local microbial infections induced by multiple-drug-resistant bacteria in the orthopedic field can be intractable, therefore development of new therapeutic modalities is needed. Photodynamic therapy (PDT) is a promising alternative modality to antibiotics for intractable microbial infections, and we recently reported that PDT has the potential to accumulate neutrophils into the infected site which leads to resolution of the infection. PDT for cancer has long been known to be able to stimulate the innate and adaptive arms of the immune system. Methodology/Principal Findings: In the present study, a murine methicillin-resistant Staphylococcus aureus (MRSA) arthritis model using bioluminescent MRSA and polystyrene microparticles was established, and both the therapeutic (Th-PDT) and preventive (Pre-PDT) effects of PDT using methylene blue as photosensitizer were examined. Although Th-PDT could not demonstrate direct bacterial killing, neutrophils were accumulated into the infectious joint space after PDT and MRSA arthritis was reduced. With the preconditioning Pre-PDT regimen, neutrophils were quickly accumulated into the joint immediately after bacterial inoculation and bacterial growth was suppressed and the establishment of infection was inhibited. Conclusions/Significance: This is the first demonstration of a protective innate immune response against a bacterial pathogen produced by PDT.

Low-level laser (light) therapy (LLLT) has been clinically applied around the world for a spectrum of disorders requiring healing, regeneration and prevention of tissue death. One area that is attracting growing interest in this scope is the use of transcranial LLLT to treat stroke and traumatic brain injury (TBI). We developed a mouse model of severe TBI induced by controlled cortical impact and explored the effect of different treatment schedules. Adult male BALB/c mice were divided into 3 broad groups (a) sham-TBI sham-treatment, (b) real-TBI sham-treatment, and (c) real-TBI active-treatment. Mice received active-treatment (transcranial LLLT by continuous wave 810 nm laser, (25 mW/cm^2), (18 J/cm^2), spot diameter 1 cm) while sham-treatment was immobilization only, delivered either as a single treatment at 4 hours post TBI, as 3 daily treatments commencing at 4 hours post TBI or as 14 daily treatments. Mice were sacrificed at 0, 4, 7, 14 and 28 days post-TBI for histology or histomorphometry, and injected with bromodeoxyuridine (BrdU) at days 21–27 to allow identification of proliferating cells. Mice with severe TBI treated with 1-laser Tx (and to a greater extent 3-laser Tx) had significant improvements in neurological severity score (NSS), and wire-grip and motion test (WGMT). However 14-laser Tx provided no benefit over TBI-sham control. Mice receiving 1- and 3-laser Tx had smaller lesion size at 28-days (although the size increased over 4 weeks in all TBI-groups) and less Fluoro-Jade staining for degenerating neurons (at 14 days) than in TBI control and 14-laser Tx groups. There were more BrdU-positive cells in the lesion in 1- and 3-laser groups suggesting LLLT may increase neurogenesis. Transcranial NIR laser may provide benefit in cases of acute TBI provided the optimum treatment regimen is employed.

Purpose To investigate the effectiveness of antimicrobial blue light (aBL) as an alternative or adjunctive therapeutic for infectious keratitis. Methods: We developed an ex vivo rabbit model and an in vivo mouse model of infectious keratitis. A bioluminescent strain of Pseudomonas aeruginosa was used as the causative pathogen, allowing noninvasive monitoring of the extent of infection in real time via bioluminescence imaging. Quantitation of bacterial luminescence was correlated to colony-forming units (CFU). Using the ex vivo and in vivo models, the effectiveness of aBL (415 nm) for the treatment of keratitis was evaluated as a function of radiant exposure when aBL was delivered at 6 or 24 hours after bacterial inoculation. The aBL exposures calculated to reach the retina were compared to the American National Standards Institute standards to estimate aBL retinal safety. Results: Pseudomonas aeruginosa keratitis fully developed in both the ex vivo and in vivo models at 24 hours post inoculation. Bacterial luminescence in the infected corneas correlated linearly to CFU (R2 = 0.921). Bacterial burden in the infected corneas was rapidly and significantly reduced (>2-log10) both ex vivo and in vivo after a single exposure of aBL. Recurrence of infection was observed in the aBL-treated mice at 24 hours after aBL exposure. The aBL toxicity to the retina is largely dependent on the aBL transmission of the cornea. Conclusions: Antimicrobial blue light is a potential alternative or adjunctive therapeutic for infectious keratitis. Further studies of corneal and retinal safety using large animal models, in which the ocular anatomies are similar to that of humans, are warranted.