ABSTRACT
INTRODUCTION AND OBJECTIVES: Antimicrobial photodynamic therapy combines the use of a photosensitizing drug with light and oxygen to eradicate pathogens. Trichophyton mentagrophytes is a dermatophytic fungus able to invade the skin and keratinized tissues. We have investigated the use of new methylene blue as the photosensitizing agent for antimicrobial photodynamic therapy to produce the in vitro inactivation of T mentagrophytes. MATERIAL AND METHODS: A full factorial design was employed to optimize the parameters for photoinactivation of the dermatophyte. The parameters studied were new methylene blue concentration, contact time between the photosensitizing agent and the fungus prior to light treatment, and the fluence of red light (wavelength, 620-645nm) applied. RESULTS: The minimum concentration of new methylene blue necessary to induce the death of all T. mentagrophytes cells in the initial suspension (approximate concentration, 106 colony forming units per milliliter) was 50µM for a fluence of 81J/cm2 after a contact time of 10minutes with the photosensitizing-agent. Increasing the concentration to 100µM allowed the fluence to be decreased to 9J/cm2. CONCLUSIONS: Comparison of our data with other published data shows that the susceptibility of T. mentagrophytes to antimicrobial photodynamic therapy with new methylene blue is strain-dependent. New methylene blue is a photosensitizing agent that should be considered for the treatment of fungal skin infections caused by this dermatophyte.
Subject(s)
Methylene Blue/analogs & derivatives , Photochemotherapy , Photosensitizing Agents/pharmacology , Trichophyton/drug effects , Drug Evaluation, Preclinical , Humans , In Vitro Techniques , Methylene Blue/pharmacology , Microbial Sensitivity Tests , Photochemistry , Suspensions , Trichophyton/radiation effectsABSTRACT
The 'injury hypothesis' in organ transplantation suggests that ischemia-reperfusion injury is involved in the adaptative alloimmune response. We previously found that a strong immune/inflammatory response was induced by ischemia during kidney transplantation in rats. We show here that immature dendritic cells (DCs) undergo hypoxia-mediated differentiation comparable to allogeneic stimulation. Hypoxia-differentiated DCs overexpress hypoxia inducible factor-1alpha (HIF-1alpha) and its downstream target genes, such as vascular endothelial growth factor or glucose transporter-1. Rapamycin attenuated DC differentiation, HIF-1alpha expression, and its target gene expression in a dose-dependent manner along with downregulated interleukin-10 secretion. Coculture of hypoxia-differentiated DCs with CD3 lymphocytes induced proliferation of lymphocytes, a process also neutralized by rapamycin. Furthermore, in vivo examination of ischemia-reperfusion-injured mouse kidneys showed a clear maturation of resident DCs that was blunted by rapamycin pretreatment. Our results suggest that hypoxia is a central part of the 'injury hypothesis' triggering DC differentiation under hypoxic conditions. Rapamycin attenuates the hypoxic immune-inflammatory response through inhibition of the HIF-1alpha pathway.
Subject(s)
Antibody Formation , Cell Hypoxia/physiology , Dendritic Cells/immunology , Antibody Formation/drug effects , Cell Differentiation , Cells, Cultured , Dendritic Cells/drug effects , Dendritic Cells/physiology , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/biosynthesis , Sirolimus/pharmacologyABSTRACT
The aim of the study was to characterize the role of cold ischemia in the process of acute rejection using an experimental renal transplant model. Syngeneic renal transplants were performed between Wistar Agouti rats and allogeneic grafts using Wistar-Agouti rats as recipients of Brown-Norway kidneys. For cold ischemia (CI), kidneys were preserved in Euro-Collins (4 degrees C/ 2.5 hours). Rats were bilaterally nephrectomized at the moment of renal transplant and did not receive any immunosuppressant. The groups were NoAR (n = 6): immediate syngeneic transplant; CI-NoAR (n = 6): syngeneic transplant with CI; AR (n = 13): immediate allogeneic graft; CI-AR (n = 6): allogeneic graft with CI. Allogeneic rats were followed for the survival study. Syngeneic rats, with mean survival time beyond 6 months, were sacrificed on the day 7 to compare grafts with those in the allogeneic groups. H&E- and PAS-stained grafts were evaluated using the Banff criteria. Tissue INF-gamma and TNF-alpha were quantified by RT-real time-PCR on the kidney grafts. Renal insufficiency did not appear in the NoAR group, but it did from the posttransplant day 5 in both acute rejection groups. While NoAR kidneys showed well-conserved renal architecture, then AR group displayed variable degrees of tubular necrosis with scarce cellular infiltration, interstitial hemorrhage, vascular damage with fibrinoid necrosis, perivascular edema, and nuclear disruption. Cold ischemia in rejecting animals increased the mortality rate due to renal insufficiency and accelerated acute rejection. Independently of CI, the proinflammatory cytokines TNF-alpha and INF-gamma were increased in both rejection groups. In conclusion, addition of CI overactivates the acute rejection process via a humoral component.
Subject(s)
Graft Rejection/immunology , Kidney Transplantation/immunology , Acute Disease , Animals , Antibody Formation , Disease Models, Animal , Graft Survival/immunology , Ischemia , Male , Rats , Rats, Inbred BN , Rats, Wistar , Renal CirculationABSTRACT
INTRODUCCIÓN Y OBJETIVOS: La terapia fotodinámica antimicrobiana combina el uso de un fármaco fotosensibilizante, la luz y el oxígeno para erradicar microorganismos patógenos. Trichophyton mentagrophytes es un hongo dermatofito capaz de invadir la piel y tejidos queratinizados. El objetivo de este trabajo es aplicar la terapia fotodinámica antimicrobiana para la inactivación in vitro de T. mentagrophytes utilizando el nuevo azul de metileno como agente fotosensibilizador. MATERIAL Y MÉTODOS: Se aplica un diseño factorial completo para optimizar los parámetros que permiten la fotoinactivación del dermatofito. Se tiene en cuenta la concentración del nuevo azul de metileno, el tiempo de contacto entre el fotosensibilizador y el hongo antes del tratamiento con luz y la fluencia de luz roja aplicada entre 620 y 645 nm. RESULTADOS: La mínima concentración de nuevo azul de metileno que produce una mortalidad de todas las células de T. mentagrophytes de la suspensión inicial (concentración∼106 ufc/ml) es 50 M para una fluencia de 81 J/cm-2 y un tiempo previo de contacto hongo-fotosensibilizador de 10 min. Si se aumenta la concentración a 100 M la fluencia que se necesita disminuye a 9 J/cm-2. CONCLUSIONES: La comparación de nuestros datos con otros publicados muestra que la susceptibilidad de T. mentagrophytes a la terapia fotodinámica antimicrobiana con nuevo azul de metileno es cepa-dependiente. El nuevo azul de metileno es un fotosensibilizador a tener en cuenta para el tratamiento de las micosis cutáneas causadas por este dermatofito
INTRODUCTION AND OBJECTIVES: Antimicrobial photodynamic therapy combines the use of a photosensitizing drug with light and oxygen to eradicate pathogens. Trichophyton mentagrophytes is a dermatophytic fungus able to invade the skin and keratinized tissues. We have investigated the use of new methylene blue as the photosensitizing agent for antimicrobial photodynamic therapy to produce the in vitro inactivation of T mentagrophytes. MATERIAL AND METHODS: A full factorial design was employed to optimize the parameters for photoinactivation of the dermatophyte. The parameters studied were new methylene blue concentration, contact time between the photosensitizing agent and the fungus prior to light treatment, and the fluence of red light (wavelength, 620---645 nm) applied. RESULTS: The minimum concentration of new methylene blue necessary to induce the death of all T. mentagrophytes cells in the initial suspension (approximate concentration, 106 colony forming units per milliliter) was 50 M for a fluence of 81 J/cm2 after a contact time of 10 minutes with the photosensitizing-agent. Increasing the concentration to 100 M allowed the fluence to be decreased to 9 J/cm2. CONCLUSIONS: Comparison of our data with other published data shows that the susceptibility of T. mentagrophytes to antimicrobial photodynamic therapy with new methylene blue is straindependent. New methylene blue is a photosensitizing agent that should be considered for the treatment of fungal skin infections caused by this dermatophyte