Related Experiment Videos
Activation of macrophages by Photofrin II during photodynamic therapy
R W Steubing1, S Yeturu, A Tuccillo
1Department of Surgery, Beckman Laser Institute and Medical Clinic, University of California, Irvine 92715.
Abstract:
In order to obtain information about the activation of macrophages (M phi s) during photodynamic therapy (PDT), the influence of Photofrin II (Pf II) on the viability of thioglycollate-elicited murine M phi s and the subsequent generation of superoxide anion was studied. Irradiations were performed at an energy density of 5 J cm-2, a power density of 150 mW cm-2 and a wavelength of 405 nm. Viability of M phi s was assessed using the acridine orange-ethidium bromide assay. Superoxide anion generation was determined using ferricytochrome c (cyt c) and nitroblue tetrazolium (NBT) reduction. Our results indicate that the M phi s are highly susceptible to PDT as their viability is decreased to approximately 30% by 1 microgram ml-1 Pf II at the energy density indicated above. Within the first 30 min of addition of the photosensitizer, a reducing agent is generated intracellularly by the stimulation of the M phi s. An extracellular release of superoxide anion does not occur, as measured by the cyt c assay. Preincubation of the cells for 1 or 24 h with Pf II and a second challenge with phorbol myristate acetate (PMA) does not enhance the reduction of NBT. Thus, Pf II exerts an immediate effect on the M phi s which could be interpreted as a first step for subsequent reactions.
Insights
Photodynamic therapy (PDT) significantly reduces macrophage viability using Photofrin II (Pf II). Intracellular reducing agents are generated, but extracellular superoxide anion release is not detected, indicating an immediate Pf II effect on macrophages.
Area of Science:
- Photodynamic Therapy
- Immunology
- Cell Biology
Background:
- Macrophages (M phi s) play a crucial role in immune responses.
- Photodynamic therapy (PDT) is an established cancer treatment modality.
- Understanding PDT's impact on immune cells like macrophages is vital for optimizing therapeutic strategies.
Purpose of the Study:
- To investigate the effects of Photofrin II (Pf II) on macrophage viability during PDT.
- To determine if PDT influences superoxide anion generation in macrophages.
- To elucidate the immediate cellular responses of macrophages to Pf II-mediated PDT.
Main Methods:
- Thioglycollate-elicited murine macrophages were exposed to Photofrin II (Pf II) and irradiated at 405 nm.
- Macrophage viability was assessed using the acridine orange-ethidium bromide assay.
- Superoxide anion generation was measured via ferricytochrome c (cyt c) and nitroblue tetrazolium (NBT) reduction assays.
Main Results:
- PDT with 1 microgram/mL Pf II significantly decreased macrophage viability to approximately 30%.
- An intracellular reducing agent was generated within 30 minutes of Pf II addition.
- No extracellular superoxide anion release was detected using the cyt c assay.
- Preincubation with Pf II followed by phorbol myristate acetate (PMA) challenge did not enhance NBT reduction.
Conclusions:
- Macrophages are highly susceptible to PDT-induced cell death mediated by Pf II.
- Pf II triggers an immediate intracellular response in macrophages, potentially involving reducing agent generation.
- PDT with Pf II does not appear to induce extracellular superoxide anion release from macrophages.
- These findings suggest an immediate, intracellular effect of Pf II on macrophages that may initiate subsequent biological reactions.