We previously showed that antimicrobial photodynamic inactivation (aPDI) of Gram-positive and Gram-negative bacteria mediated by the phenothiazinium dye, methylene blue (MB) was potentiated by addition of potassium thiocyanate (10mM). The mechanism was suggested to involve a singlet oxygen mediated reaction with SCN- to form sulfite and cyanide and then to produce sulfur trioxide radical anion. We now report that potassium selenocyanate (KSeCN) (concentrations up to 100 mM) can also potentiate (up to 6 logs of killing) aPDI mediated by a number of different photosensitizers: MB, Rose Bengal, and TPPS4 (as low as 200 nM). When a mixture of selenocyanate with these PS in solution was illuminated and then bacteria were added after the light, there was up to 6 logs killing (Gram-negative > Gram-positive) but the antibacterial species decayed rapidly (by 20 min). Our hypothesis to explain this antibacterial activity is the formation of selenocyanogen (SeCN)2 by reaction with singlet oxygen (1O2) as shown by quenching of 1O2 by SeCN- and increased photoconsumption of oxygen. The fact that lead tetra-acetate reacted with SeCN- (literature preparation of (SeCN)2) also produced a short-lived antibacterial species supports this hypothesis.
We report a novel class of highly water-soluble decacationic methano[60]fullerene decaiodides C60[>M(C3N6+C3)2]-(I−)10 [1-(I−)10] capable of co-producing singlet oxygen (Type-II) and highly reactive hydroxyl radicals, formed from superoxide radicals in Type-I photosensitizing reactions, upon illumination at both UVA and white light wavelengths. The O2‒·-production efficiency of 1-(I−)10 was confirmed by using a O2‒·-reactive bis(2,4-dinitrobenzenesulfonyl)tetrafluorofluorescein probe and correlated to the photoinduced electron-transfer event going from iodide anions to 3C60*[>M(C3N6+C3)2] leading to C60‒·[>M(C3N6+C3)2]. Incorporation of a defined number (ten) of quaternary ammonium cationic charges per C60 in 1 was aimed to enhance its ability to target pathogenic Gram-positive and Gram-negative bacterial cells. We used the well-characterized malonato[60]fullerene diester monoadduct C60[>M(t-Bu)2] as the starting fullerene derivative to provide a better synthetic route to C60[>M(C3N6+C3)2] via transesterification reaction under trifluoroacetic acid catalyzed conditions. These compounds may be used as effective photosensitizers and nano-PDT drugs for photoinactivation of pathogens.
Potassium iodide can potentiate antimicrobial photodynamic inactivation (aPDI) of a broad-spectrum of microorganisms, producing many extra logs of killing. We compared two charged porphyrins, TPPS4 (thought to be anionic and not able to bind to Gram-negative bacteria) and TMPyP4 (considered cationic and well able to bind to bacteria). As expected TPPS4 + light did not kill Gram-negative Escherichia coli, but surprisingly when 100 mM KI was added, it was highly effective at mediating aPDI (eradication at 200 nM + 10 J/cm2 of 415 nm light). TPPS4 was more effective than TMPyP4 in eradicating the Gram-positive bacteria, methicillin-resistant Staphylococcus aureus and the fungal yeast Candida albicans (regardless of KI). TPPS4 was also highly active against E. coli after a centrifugation step when KI was added, suggesting that the supposedly anionic porphyrin bound to bacteria and Candida. We conclude that TPPS4 behaves as if it has some cationic character in the presence of bacteria, which may be related to its supply from vendors in the form of a dihydrochloride salt.
Increasing concern is evident over the epidemic of traumatic brain injury in both civilian and military medicine, and the
lack of approved treatments. Transcranial low level laser therapy tLLLT) is a new approach in which near infrared laser
is delivered to the head, penetrates the scalp and skull to reach the brain. We asked whether tLLLT at 810-nm could
improve memory and learning in mice with controlled cortical impact traumatic brain injury. We investigated the
mechanism of action by immunofluorescence studies in sections from brains of mice sacrificed at different times. Mice
with TBI treated with 1 or 3 daily laser applications performed better on Morris Water Maze test at 28 days. Laser
treated mice had increased BrdU incorporation into NeuN positive cells in the dentate gyrus and subventricular zone
indicating formation of neuroprogenitor cells at 7 days and less at 28 days. Markers of neuron migration (DCX and Tuj1)
were also increased, as was the neurotrophin, brain derived neurotrophic factor (BDNF) at 7 days. Markers of
synaptogenesis (formation of new connections between existing neurons) were increased in the perilesional cortex at 28
days. tLLLT is proposed to be able to induce the brain to repair itself after injury. However its ability to induce
neurogenesis and synaptogenesis suggests that tLLLT may have much wider applications to neurodegenerative and
psychiatric disorders.
The use of transcranial low-level laser (light) therapy (tLLLT) to treat stroke and traumatic brain injury (TBI) is attracting increasing attention. We previously showed that LLLT using an 810-nm laser 4 h after controlled cortical impact (CCI)-TBI in mice could significantly improve the neurological severity score, decrease lesion volume, and reduce Fluoro-Jade staining for degenerating neurons. We obtained some evidence for neurogenesis in the region of the lesion. We now tested the hypothesis that tLLLT can improve performance on the Morris water maze (MWM, learning, and memory) and increase neurogenesis in the hippocampus and subventricular zone (SVZ) after CCI-TBI in mice. One and (to a greater extent) three daily laser treatments commencing 4-h post-TBI improved neurological performance as measured by wire grip and motion test especially at 3 and 4 weeks post-TBI. Improvements in visible and hidden platform latency and probe tests in MWM were seen at 4 weeks. Caspase-3 expression was lower in the lesion region at 4 days post-TBI. Double-stained BrdU-NeuN (neuroprogenitor cells) was increased in the dentate gyrus and SVZ. Increases in double-cortin (DCX) and TUJ-1 were also seen. Our study results suggest that tLLLT may improve TBI both by reducing cell death in the lesion and by stimulating neurogenesis.
Low-level laser (or light) therapy (LLLT) is attracting growing interest to treat both stroke and traumatic brain
injury (TBI). The fact that near-infrared light can penetrate into the brain allows non-invasive treatment to be carried
out with a low likelihood of treatment-related adverse events. It is proposed that red and NIR light is absorbed by
chromophores in the mitochondria of cells leading to changes in gene transcription and upregulation of proteins
involved in cell survival, antioxidant production, collagen synthesis, reduction of chronic inflammation and cell
migration and proliferation. We developed a mouse model of controlled cortical impact (CCI) TBI and examined the
effect of 0, 1, 3, and 14 daily 810-nm CW laser treatments in the CCI model as measured by neurological severity
score and wire grip and motion test. 1 laser Tx gave a significant improvement while 3 laser Tx was even better.
Surprisingly 14 laser Tx was no better than no treatment. Histological studies at necropsy suggested that the
neurodegeneration was reduced at 14 days and that the cortical lesion was repaired by BrdU+ve neural progenitor
(stem) cells at 28 days. Transcranial laser therapy is a promising treatment for acute (and chronic TBI) and the lack
of side-effects and paucity of alternative treatments encourages early clinical trials.
Multi-drug resistant Acinetobacter baumanii infections represent a growing problem, especially in traumatic wounds and
burns suffered by military personnel injured in Middle Eastern conflicts. Effective treatment using traditional antibiotics
can be extremely difficult and new antimicrobial approaches are being investigated. One of these antimicrobial
alternatives could be the combination of non-toxic photosensitizers (PS) and visible light known as photodynamic
therapy (PDT). We report on the establishment of a new mouse model of full thickness thermal burns infected with a
bioluminescent derivative of a clinical Iraqi isolate of A. baumannii and its PDT treatment by topical application of a PS
produced by covalent conjugation chlorin(e6) to polyethylenimine followed by illumination of the burn surface with red
light. Application of 108A. baumannii cells to the surface of 10-second burns made on the dorsal surface of shaved
female BALB/c mice led to chronic infections that lasted on average 22 days characterized by a remarkably stable
bacterial bioluminescence. PDT carried out on day 0 soon after applying bacteria gave over three logs of loss of bacterial
luminescence in a light exposure dependent manner, while PDT carried out on day 1 and day 2 gave approximately a
1.7-log reduction. Application of PS dissolved in 10% or 20% DMSO without light gave only modest reduction in
bacterial luminescence from mouse burns. Some bacterial regrowth in the treated burn was observed but was generally
modest. It was also found that PDT did not lead to inhibition of wound healing. The data suggest that PDT may be an
effective new treatment for multi-drug resistant localized A. baumannii infections.
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