Pulsed low-intensity laser treatment stimulates wound healing without enhancing biofilm development in vitro

Objectives: Treating infected or chronic wounds burdened with biofilms still is a major challenge in medical care. Healing-stimulating factors lose their efficacy due to bacterial degradation, and antimicrobial substances negatively affect dermal cells. Therefore, alternative treatment approaches like the pulsed low intensity laser therapy (LILT) require consideration. Methods: The effect of pulsed LILT (904 nm, in three frequencies) on relevant human cells of the wound healing process (fibroblasts (BJ), keratinocytes (HaCaT), endothelial cells (HMEC), monocytes (THP 1)) were investigated in in-vitro and ex vivo wound models with respect to viability, proliferation and migration. Antimicrobial efficacy of the most efficient frequency in cell biological analyses of LILT (3200 Hz) was determined in a human biofilm model (lhBIOM). Quantification of bacterial load was evaluated by suspension method and qualitative visualization was performed by scanning electron microscopy (SEM). Results: Pulsed LILT at 904 nm at 3200 Hz +/- 50% showed the most positive effects on metabolic activity and proliferation of human wound cells in vitro (after 72 h - BJ: BPT 0.97 +/- 0.05 vs. 0.75 +/- 0.04 (p = 0.0283); HaCaT: BPT 0.79 +/- 0.04 vs. 0.59 +/- 0.02 (p = 0.0106); HMEC: 0.74 +/- 0.02 vs. 0.52 +/- 0.04 (p = 0.009); THP-1: 0.58 +/- 0.01 vs. 0.64 +/- 0.01 (p > 0.05) and ex vivo. Interestingly, re-epithelialization was stimulated in a frequency-independent manner. The inhibition of metabolic activity after TNF-alpha application was abolished after laser treatment. No impact of LILT on monocytes was detected. Likewise, the tested LILT regimens showed no growth rate reducing effects on three bacterial strains (after 72 h - PA: -1.03%; SA: 0.02%; EF: -1,89%) and one fungal (2.06%) biofilm producing species compared to the respective untreated control. Accordingly, no significant morphological changes of the biofilms were observed after LILT treatment in the SEM. Conclusions: Frequent application of LILT (904 nm, 3200 Hz) seems to be beneficial for the metabolism of human dermal cells during wound healing. Considering this, the lack of disturbance of the behavior of the immune cells and no growth-inducing effect on bacteria and fungi in the biofilm can be assigned as rather positive. Based on this combined mode of action, LILT may be an option for hard to heal wounds infected with persistent biofilms.