Treatment of experimental periodontal disease by laser therapy in simvastatin-modified rats

Abstract Low intensity laser can be used as a promising alternative in the treatment of periodontal disease. Objective The aim of this study was to evaluate low-level laser therapy (LLLT) as an adjuvant treatment for scaling and root planing (SRP) for the treatment of induced periodontitis in simvastatin-modified rats. Material and Methods A total of 180 rats were evenly divided into two groups: Veh – receiving oral administration of polyethylene glycol (vehicle); S – receiving oral administration of Simvastatin. Periodontal disease was induced in both groups at the first mandibular molar. After seven days, the ligature was removed and the animals were divided into subgroups according to the following local treatments: NT – no treatment; SRP – scaling and root planing and irrigation with saline solution; and LLLT ¬– SRP and laser irradiation (660 nm; 0.03 W; 4 J). Ten animals in each subgroup/local treatment were euthanized at 7, 15 and 30 days. Samples of gingival tissue were processed to analyze the tissue oxidative damage and radiographic analysis. Levels of oxidative stress were analyzed by the expressions of Tripeptideglutathione (TG), Malondialdehyde (MDA) and Carbonylated Proteins (CP). Results The animals in S group had higher levels of TG and lower levels of MDA and CP compared with Veh group (p<0.05). Radiographically, in the intragroup analysis Veh and S, LLLT showed lower bone loss (BL) compared with NT and SRP, in all experimental periods (p<0.01). In addition, a lower BL was observed for the animals of Veh group treated with LLLT compared with treatment SRP in the S group, in all experimental periods. Conclusion Within the limits of this study, we can conclude that LLLT was effective as adjuvant treatment for SRP protecting against the occurrence of oxidative tissue damages as well as for reducing alveolar bone loss in experimentally induced periodontitis simvastatin-modified rats.


Introduction
In inflammation, neutrophils are the first cells to be activated to defend the body together with macrophages 27 . Those are chemotaxically attracted by secretory cells, bacteria and other foreign bodies to inflammation areas 27 . On this site, neutrophils phagocyte the microorganisms, covered or not with complement or specific antibodies, which are killed by cytotoxic proteins derived from cytoplasmic granules and by oxygen and nitrogen reactive species such as superoxide anion 9 , hydrogen peroxide 28 , hydroxyl radical 12 and peroxynitrite 21 .
There is evidence of a more aggressive destruction of tooth support tissues with elevated levels of oxidative stress markers during the development of periodontal disease (PD) 24 . Among the oxidants, the superoxide anion 9 , in periodontal tissues, can be involved in signaling of induction of bone resorption; the hydroxyl radical 12 is extremely reactive and can damage important biomolecules, such as proteins, lipids and nucleic acids, whereas hydrogen peroxide is able to cross membranes, damaging adjacent cells and increasing the oxidative cascade 28 . Thus, most studies demonstrate that periodontitis is associated with increased lipid peroxidation 29 and increased protein carbonyls 9 as well as decreased antioxidants, such as reduced glutathione (GSH) 11,29. In addition, there are evidences of decreased oxidative injuries and changes in antioxidant system following periodontal treatment 19 .
The treatment of PD is based on the elimination of pathogenic subgingival microbiota by scaling and root planing (SRP) 24 . However, mechanical therapy used alone may be defective in the elimination of pathogenic bacteria, since they are located within soft and hard tissues or in areas that are inaccessible to periodontal instruments 7 . In addition, an important component of individuals themselves can lead to tissue destruction observed in the periodontitis. Therefore, therapeutic strategies performing the pharmacological modulation of host response have emerged as a new therapeutic approach 13 .
Simvastatin is an inhibitor of the 3-hydroxy-3methyl-glutaryl-coenzyme A (HMG -CoA reductase) enzyme, which is responsible for the synthesis of cholesterol and therefore it is widely used for the systemic treatment of diseases related to hypercholesterolemia. This drug also has antiinflammatory, immunomodulator, antioxidant, and angiogenic effects 6,25 , and it also promotes increased osteoblast formation 10,20 . Such properties offer great potential for statins to modify the course of chronic inflammatory diseases such as periodontitis 26 .
In addition to drug therapy, the use of low intensity lasers associated with scaling and root planing for the local treatment of periodontal disease has been   In the comparison between the groups (Veh and S), among the same local treatments, MDA levels in S group were significantly lower than those in Veh group (p<0.05), for NT, SRP and LLLT at 7 days ( Table 2). #Difference among same groups and local treatments (ANOVA and p<0.05) *Difference between local treatments, same group and period (ANOVA and Tukey, p<0.05) Difference between groups, same local treatment and period (ANOVA and Tukey, p<0.05)°D ifference between groups and local treatments, same period (ANOVA and Tukey, p<0.05) #Difference among same groups and local treatments (ANOVA and p<0.05) *Difference between local treatments, same group and period (ANOVA and Tukey, p<0.05) Difference between groups, same local treatment and period (ANOVA and Tukey, p<0.05)°D ifference between groups and local treatments, same period (ANOVA and Tukey, p<0.05)

Radiographic analysis
In the intragroup analysis between Veh and S, LLLT showed lower BL compared with NT and SRP, in all experimental periods (p<0.01).
In the intergroup analysis between the same local treatments, the SRP presented a lower BL (p<0.01) in      Figure 1 and Figure 2).

Conclusion
Within the limits of this study, we can conclude that LLLT was effective as adjuvant treatment for SRP protecting against the occurrence of oxidative tissue damages as well as reducing alveolar bone loss in experimentally induced periodontitis simvastatinmodified rats.