Combined Effects of Low Level Laser Therapy and Inducers on the Neural Differentiation of Mesenchymal Stem Cells

Abstract

Low level laser therapy (LLLT) improves the therapeutic effectiveness of stem cell therapy for neurological injury through its ability to enhance stem differentiation and protect against neuronal apoptosis through its antioxidation effects. However, the specific mechanisms governing these effects are poorly defined. In this study, we investigated the effects of LLLT on stem cell differentiation at the molecular level, oxidative stress balance, and inflammatory factors to provide theoretical support for its clinical application. Cell viability was assessed via MTT assays. Reactive oxygen species (ROS), total superoxide dismutase (SOD), and total antioxidant capacity (TAC) were used to evaluate oxidative stress levels. Western blot analysis was used to quantitatively investigate protein expression. The levels of secreted proteins and tumor necrosis factor- $\alpha $ (TNF- $\alpha$ ) and interleukin- $1\beta $ (IL- $1\beta$ ) expression were measured by ELISA. Compared to LLLT at 808 nm, LLLT at 635 nm enhanced the proliferation of human umbilical cord-derived mesenchymal stem cells (hUCMSCs). The rates of proliferation markedly increased at a power density 20 mW/cm2. LLLT enhanced the antioxidant capacity and caused no inflammation in normal cells. Markers of neural precursors were more highly expressed at 808 nm when combined with inducers for 3 d, compared to the more modest increases observed at 635 nm. The expression of neuN on day 7 also increased, most notably when LLLT at 808 nm was combined with cerebrospinal fluid (CSF)/injured cerebrospinal fluid (iCSF). ELISA assays showed that LLLT at 808 nm with CSF/iCSF also increased the differentiation of hUCMSCs into neurons. LLLT at 808 nm combined with inducers promoted the differentiation into neurons and increased the rate of neuronal differentiation.