Efeito do laser de baixa intensidade na apoptose e estresse oxidativo em células-tronco mesenquimais derivadas do tecido adiposo submetidas a toxicidade por doxorrubicina

Abstract

Background: doxorubicin is a chemotherapy agent widely used on the treatment of hematologic diseases and neoplasia. However, doxorubicin causes cell damage in healthy cells (e.g. Cardiomyocytes), a situation that is persistent even after the end of drug administration. These implications justify tools that in some way lessen the adverse effects of doxorubicin on non-diseased cells, especially in the heart. We highlight the use of stem cell therapy, in which cardioprotective effects have been reported. However, doxorubicin also imposes cytotoxicity on transplanted stem cells, reducing its therapeutic potential. Objective: to analyze whether the Low Intensity Laser (LIL) reduces apoptosis and oxidative stress in mesenchymal stem cells derived from adipose tissue (MSCA) submitted to doxorubicin toxicity. Material and Methods: MSCA at the concentration of 1×105/0.1 mL of DMEM were plated in a 96-well plate for the composition of the following experimental sets: Control - MSCA untreated with doxorubicin and/or LBI; D25 and D50 - MSCA incubated with doxorubicin at concentrations of 25 μg/mL and 50 μg/mL, respectively; D25 + LIL/D50 + LIL - MSCA irradiated with 0.2 J, 0.4 J and 0.7 J LIL (660 nm) and incubated with doxorubicin at concentrations of 25 μg/mL and 50 μg/mL, respectively. Cell viability and apoptosis were determined by flow cytometry with 7AAD and Annexin V, respectively. Cell permeabilization with DCFH was used as a marker of reactive oxygen species (ROS). Intracellular ATP content was determined by spectrophotometer luminescence assay. Cytokines IL-6, IL-10 and TNF were analyzed by ELISA. Results: irradiations of 0.4 and 0.7 J resulted in a higher concentration of viable MSCA independent of doxorubicin concentrations. Similar results were observed for the apoptosis assays in cells incubated with 50 μg of doxorubicin and irradiated with 0.2 and 0.4 J. Considering that 0.4 J was effective in increasing the viability and reducing apoptosis of the MSCA at the concentration of 50 μg, we directed to continue the experiments only with this dose of energy and concentration of the drug. Thus, 0.4 J inhibited the increase of ROS in MSCA, however, it did not result in a higher concentration of intracellular ATP. For IL-6, the irradiated group increased expression when compared to the control group. For IL-10 and TNF, there was no difference between groups. Conclusion: LIL reduced mortality, apoptosis and oxidative stress of MSCA undergoing doxorubicin toxicity.