Irradiation at 830 nm stimulates nitric oxide production and inhibits pro-inflammatory cytokines in diabetic wounded fibroblast cells

Background and Objective: Wound healing in diabetic patients remains a chief problem in the clinical setting and there is a strong need for the development of new, safe, reliable therapies. This study aimed to establish the effect of irradiating diabetic wounded fibroblast cells (WS1) in vitro on pro-inflammatory cytokines and the production of nitric oxide (NO). Materials and Methods: Normal, wounded and diabetic wounded WS1 cells were exposed to an 830nm laser with 5 J/cm2 and incubated for a pre-determined amount of time. Changes in cellular viability, proliferation and apoptosis were evaluated by the Trypan blue assay, VisionBlue™ fluorescence assay and caspase 3/7 activity respectively. Changes in cytokines (interleukin - IL-6, IL-1b and tumour necrosis factor-alpha, TNF-α) were determined by ELISA. NO was determined spectrophotometrically and reactive oxygen species (ROS) was evaluated by immunofluorescent staining. Results: Diabetic wounded WS1 cells showed no significant change in viability, a significant increase in proliferation at 24 and 48 hours (P<0.001 and P<0.01 respectively) and a decrease in apoptosis 24 hours postirradiation (P<0.01). TNF-α levels were significantly decreased at both 1 and 24 hours (P<0.05), while IL-1β was only decreased at 24 hours (P<0.05). There was no significant change in IL-6. There was an increase in ROS and NO (P<0.01) 15 minutes post-irradiation. Conclusion: Results show that irradiation of diabetic wounded fibroblast cells at 830nm with 5 J/cm ² has a positive effect on wound healing in vitro. There was a decrease in pro-inflammatory cytokines (IL-1β and TNF-α) and irradiation stimulated the release of ROS and NO due to what appears to be direct photochemical processes.