Photobiomodulation Therapy to Promote Angiogenesis in Diabetic Mice with Hindlimb Ischemia

To assess whether photobiomodulation therapy (PBMT) induces angiogenesis in diabetic mice with hindlimb ischemia (HLI). Patients with diabetes mellitus (DM) are at high risk of developing peripheral arterial disease (PAD) in the lower extremities. PBMT has been shown to promote angiogenesis both in vitro and in vivo and could be a treatment for DM patients with PAD. Femoral artery ligation/excision in mice was performed to induce HLI as an animal model of PAD. PBMT at a dose of 660 nm and 1.91 J/cm² was delivered for 10 min on 5 consecutive days after the HLI surgery. Control mice received HLI only. Mice in the DM group were injected with streptozocin to induce diabetes before HLI surgery. Mice in the laser and DM+ laser groups received both HLI and PBMT, and the latter group had induced DM. After the laser treatment, lower limb blood flow was evaluated by laser Doppler. The capillary density and CD31 were analyzed by immunofluorescence staining, and protein levels of vascular endothelial growth factor (VEGF)-A, hypoxia-inducible factor-1α (HIF-1α), inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), and extracellular signal-regulated kinases (ERK) were measured by Western blotting of tissue samples. Compared with the control and DM mice, the laser and DM+ laser groups had more than double the capillary density and blood perfusion rate. Levels of CD31 and VEGF-A proteins in groups that received laser were increased by 1.9- to 3.2-fold compared with groups that did not undergo laser treatment. Animals treated with PBMT exhibited significantly increased HIF-1α expression and ERK phosphorylation compared with animals that did not receive this treatment, and the amount of phospho-eNOS and iNOS increased and decreased, respectively. PBMT can induce therapeutic angiogenesis, indicating that low intensity laser could be a novel treatment for PAD patients.