IADR Abstract Archives
Anti-Inflammatory Effect of Different Irradiation Modes of Azulene-Mediated Photodynamic Therapy
Objectives: This study aimed to study the effects of different irradiation modes in Azulene-mediated photodynamic therapy on the amount of inflammatory cytokine PGE2.
Methods: Azulene was dissolved in ethanol/distilled water to obtain 1, 10 and 100 µM and added 10 µM dimethyl anthracene, a singlet oxygen probe, in 96-black well plate then irradiated in arbitrary laser unit (wavelength 638 nm, 1.515 mW/cm2, light bulb-to-well bottom distance = 2 cm) to obtain energy densities 4 or 40 J/cm2 by either of following irradiation modes; 1) continuous, 2) fractionation (1/4 of final energy density x4; resting 15 minutes between each session), 3) pulse (by on 300 msec/ off 700 msec). Subsequently, singlet oxygen was relatively quantified by Varioscan® (Thermo Fisher Scientific, USA) fluorescence microplate reader (excitation/emission wavelength = 375/473 nm) in term of optical density. For anti-inflammatory assays, 1 × 105 cells of human peripheral blood mononuclear cell (PBMCs) were cultured in RPMI-1640+10% Fetal bovine serum (37 °C, ≥ 90% humidity, 5% CO2) for 24 hours and 6-hour preincubated with 10 ng/ml of TNF-α to activate inflammation. Azulene concentrations were incubated for 30 minutes prior to irradiate in aforementioned conditions. 100 µl of supernatant was tranfered for measuring PGE2 using Human PGE2 ELISA kit with above fluorescence microplate reader at absorbance 420 nm. Kruskal-Wallis test with post-hoc test was used for comparisons of mean absorbance at significant level p < 0.05.
Results: The continuous mode with 40 J/cm2+10 µM Azulene could produce the highest amount of singlet oxygen (p = 0.001). The fractionation mode with 40 J/cm2+ 1 µM Azulene could reduce amount of PGE2 the most.
Conclusions: In conclusion, fractional mode with relatively low Azulene concentration in photodynamic therapy tended to reduce PGE2.
Methods: Azulene was dissolved in ethanol/distilled water to obtain 1, 10 and 100 µM and added 10 µM dimethyl anthracene, a singlet oxygen probe, in 96-black well plate then irradiated in arbitrary laser unit (wavelength 638 nm, 1.515 mW/cm2, light bulb-to-well bottom distance = 2 cm) to obtain energy densities 4 or 40 J/cm2 by either of following irradiation modes; 1) continuous, 2) fractionation (1/4 of final energy density x4; resting 15 minutes between each session), 3) pulse (by on 300 msec/ off 700 msec). Subsequently, singlet oxygen was relatively quantified by Varioscan® (Thermo Fisher Scientific, USA) fluorescence microplate reader (excitation/emission wavelength = 375/473 nm) in term of optical density. For anti-inflammatory assays, 1 × 105 cells of human peripheral blood mononuclear cell (PBMCs) were cultured in RPMI-1640+10% Fetal bovine serum (37 °C, ≥ 90% humidity, 5% CO2) for 24 hours and 6-hour preincubated with 10 ng/ml of TNF-α to activate inflammation. Azulene concentrations were incubated for 30 minutes prior to irradiate in aforementioned conditions. 100 µl of supernatant was tranfered for measuring PGE2 using Human PGE2 ELISA kit with above fluorescence microplate reader at absorbance 420 nm. Kruskal-Wallis test with post-hoc test was used for comparisons of mean absorbance at significant level p < 0.05.
Results: The continuous mode with 40 J/cm2+10 µM Azulene could produce the highest amount of singlet oxygen (p = 0.001). The fractionation mode with 40 J/cm2+ 1 µM Azulene could reduce amount of PGE2 the most.
Conclusions: In conclusion, fractional mode with relatively low Azulene concentration in photodynamic therapy tended to reduce PGE2.