Efeito da associação das terapias com laser de baixa intensidade (LBI) e células-tronco da polpa dentária humana em modelo experimental de doença pulmonar obstrutiva crônica (DPOC)

Abstract

Currently, COPD has a high impact on morbidity and mortality worldwide. Smoking, history of smoking, airflow rate, obstruction and pulmonary emphysema are all related to the increase of the cells CD8+/CD4+, the number of these cells CD4+, CD8+ express NF-κB, STAT4, IFN-  and perforina. A deficiency in T cells CD4+CD25+Foxp3+ (Tregs) can impair the tolerance of the immune system and thus lead to the induction of the disease. Thus, immunotherapeutic strategies, such as Low Level Laser (LBI), aim to modulate the levels of cytokines, chemokines and transcription factors and the use of mesenchymal stem cells (CTMDD), which offer the possibility of simultaneously targeting the inflammatory response and promote the regeneration of the structure are of great importance. In this sense, the objective was to analyze some inflammatory parameters in animals with COPD after LLL and MSC therapy. 80 C57BL/6 mice were divided into 8 groups: BASAL, LBI, CTMDD, CTMDD+LBI, COPD, COPD+LBI, COPD+CTMDD and COPD+LBI+CTMDD. The protocol used for the induction of COPD consisted in the application of extracts of orotracheal cigarette smoke (3x per week/7 weeks), submitted to diode laser therapy (660nm, 30mW, 180s) for 15 days and inoculation of 106 CTMDD (1x per week in the last two weeks). 24 hours after the last treatment, the bronchoalveolar lavage (LBA) and lungs were collected. We analyzed functional and structural parameters such as amount of macrophages, neutrophils and lymphocytes by flow cytometry in the BAL, levels of GM-CSF, MCP1, IL-1β, IL-6, IL-8, IL-10, TNF-a, IFN-γ and LTB4 by ELISA, mucus production, collagen deposition, alveolar enlargement and bronchoconstriction index by histology. Data were submitted to ANOVA one-way test followed by Newman-Keuls post-test and significance level adjusted to 5% (p<0,05). LBI reduced the number of inflammatory cells, the levels of IL1-β, TNF- α, IL-6, IFN-γ, MCP-1, GM-CSF (p<0,001), KC/CXCL1, LTB4 (p<0,01) and increased IL-10 (p<0,001). We observed decreased collagen (p<0,001) with less significance in the group treated with laser alone (p<0,01), reduction in the bronchoconstriction index (p<0.001) with a smaller difference in the group treated with CTMDD alone (p<0.05), decreased mucus production only in the laser treated groups (p<0.001). Attenuated alveolar enlargement (p<0.001) and lymphocytes T CD4+ (p<0,01), CD8+ is decreased in the COPD+LBI group (p<0,001) and DPOC+CTMDD (p<0,01) demonstrating an increase in the COPD+CTMDD+LBI group (p<0.001). We noticed a reduction in the number of cells CD4+STAT4+ (p<0,01), CD4+STAT4+IFN-+ (p<0,001) and increased CD4+CD25+Foxp3+ (p<0,001) with less significance in the group with associated therapies (p<0,01) and CD4+CD25+IL-10+ (p<0,001), also with minor difference in the group DPOC+CTMDD+LBI (p<0,05). The CTMDD-treated groups demonstrated a similar effect, but did not decrease the amount of neutrophils and macrophages, KC/CXCL1 levels, and mucus production. The results demonstrate that the combination of therapies reduced cell migration to the lung, cytokine and chemokine levels as well as alveolar enlargement. One possible mechanism could be to increase the population of IL-10-producing regulatory T cells (CD4+CD25+Foxp3+) in the lung. We emphasize the importance of the use of photobiomodulation, since the data demonstrate that alone the laser can regulate inflammation and treat pulmonary emphysema in individuals with COPD.