Yamba Carla Lara Pereira

Bone Repair Investigation Using rhBMP-2 and Angiogenic Protein Extracted From Latex

Background: The aim of this work was to study the new bone tissue formation after bone morphogenetic protein type 2 (rhBMP‐2) and P‐1 application, using 5 and 10 μg of each, combined to a material carrier, in critical bone defects. Methods: It was used 70 Wistar rats (male, ∼250 g) that were divided in 10 groups with seven animals on each. Groups are the following: critical bone defect only, pure monoolein gel, 5 μg of pure P‐1, 5 μg of pure rhBMP‐2, 5 μg of P‐1/monoolein gel, 5 μg of rhBMP‐2/monoolein gel, 10 μg of pure P‐1, 10 μg of pure rhBMP‐2, 10 μg of P‐1/monoolein gel, 10 μg of rhBMP‐2/monoolein gel. Animals were sacrificed after 4 weeks of the surgical procedure and the bone samples were submitted to histological, histomorphometrical, and immunohistochemical evaluations. Results: Animals treated with pure P‐1 protein, in both situations with 5 μg and 10 μg, had no significant difference (P > 0.05) for new bone formation; other groups treated with 10 μg were statistically significant (P < 0.05) among themselves and when compared with groups in which it was inserted the monoolein gel or critical bone defect only (P < 0.05). In the group involving the 10 μg rhBMP‐2/monoolein gel association, it was observed an extensive bone formation, even when compared with the same treatment without the gel carrier. Conclusion: Using this experimental animal model, more new bone tissue was found when it was inserted the rhBMP‐2, especially when this protein was combined to the vehicle, and this process seems to be dose dependent. Microsc. Res. Tech., 2011.© 2011 Wiley Periodicals, Inc.

Influence of low-level laser associated with osteogenic proteins recombinant human BMP-2 and Hevea brasiliensis on bone repair in Wistar rats

This study analyzed the newly formed bone tissue after application of recombinant human BMP‐2 (rhBMP‐2) and P‐1 (extracted from Hevea brasiliensis) proteins, 2 weeks after the creation of a critical bone defect in male Wistar rats treated or not with a low‐intensity laser (GaAlAs 780 nm, 60 mW of power, and energy density dose of 30 J/cm2). The animals were divided into two major groups: (1) bone defect plus low‐intensity laser treatment and (2) bone defect without laser irradiation. The following subgroups were also analyzed: (a) 5 μg of pure rhBMP‐2; (b) 5 μg of pure P‐1 fraction; (c) 5 μg of rhBMP‐2/monoolein gel; (d) 5 μg of P‐1 fraction/monoolein gel; (e) pure monoolein gel. Comparisons of the groups receiving laser treatment with those that did not receive laser irradiation show differences in the areas of new bone tissue. The group treated with 5 μg of rhBMP‐2 and laser irradiation was not significantly different (P >0.05) than the nonirradiated group that received the same treatment. The irradiated, rhBMP‐2/monoolein gel treatment group showed a lower area of bone formation than the nonirradiated, rhBMP‐2/gel monoolein treatment group (P < 0.001). The area of new bone tissue in the other nonirradiated and irradiated groups was not significantly different (P > 0.05). Furthermore, the group that received the 5 μg of rhBMP‐2 application showed the greatest bone formation. We conclude that the laser treatment did not interfere with the area of new bone tissue growth and that the greatest stimulus for bone formation involved application of the rhBMP‐2 protein. Microsc. Res. Tech. 2011.

Histological, Histochemical, and Protein Changes after Induced Malocclusion by Occlusion Alteration of Wistar Rats

Although disorders of the stomatognathic system are common, the mechanisms involved are unknown. Our objective was to study the changes in the masseter muscles after unilateral exodontia. Molar extraction was performed on Wistar rats (left side), and the animals were sacrificed after either 14 or 26 days. The masseter muscle was processed for histological analysis, conventional and in situ zymography, and immunohistochemistry. The morphological analysis showed unique and specific characteristics for the experimental group. By conventional zymography no significant values of 72 kDa MMP-2 (P < 0.05) were found in both of the sides of masseter muscle after 14 and 26 days of unilateral extraction. The in situ zymography showed gelatinolytic activity on all deep masseter muscles, with significant increase on the contralateral side after 14 and 26 days (P < 0.05). The immunohistochemistry demonstrated greater expression of MMP-2 than MMP-9 and MMP-14 in all masseter muscles and there were few differences in the staining of 4 TIMPs. This knowledge about morphology and molecular masticatory muscle remodeling following environmental interventions can be used to develop clinically successful treatments.

Chronic stress effects in contralateral medial pterygoid muscle of rats with occlusion alteration.

Temporomandibular disorder (TMD) has a high prevalence in our society, characterized by a severe pain condition of the masticatory muscles and temporomandibular joint. Despite the indication of multiple factor initiators of TMD, there is still controversy about its etiology and its pathophysiology is poorly understood. Using rats as experimental animals we investigated the effect of unpredictable chronic stress with or without unilateral molar extraction on the contralateral medial pterygoid muscle. Our hypothesis is that these two factors induce changes in morphology, oxidative metabolism and oxidative stress of muscle fibers. Young adult male Wistar rats (±200g) were divided into four groups: a group with extraction and unpredictable chronic stress (E+US); with extraction and without stress (E+C); without extraction and with unpredictable chronic stress (NO+US); and a control group without either extraction or stress (NO+C). The animals were subjected to unilateral extraction of the upper left molars, under intraperitoneal anesthesia with 4% Xylazine (10mg/kg) and 10% Ketamine (80mg/kg) on day zero. The rats of groups E+US and NO+US were submitted to different protocols of stress, from the 14th day after the extraction. The protocols were different every day for five consecutive days, which were repeated from the 6th day for five days more. Contralateral medial pterygoid muscles were obtained on the 24th day after the start of the experiment for morphological, metabolic, capillary density, and oxidative stress analysis. The data from capillary density showed a decrease of capillaries in animals subjected to dental extraction, compared with those without extraction and an increase of laminin expression in the group submitted to the unpredictable chronic stress when compared to the unexposed to stress. SDH test revealed a decrease of light fibers in the group submitted to unilateral extraction of molars, compared with this area in the control group. In E+US and NO+US groups, the deeply stained fibers increased compared to NO+C.·The exodontia factor was able to increase the ROS activity in muscle, whereas the stress factor does not significantly alter ROS in this tissue. It was concluded that both unpredictable chronic stress and the extraction induce metabolic and density of capillary changes in the contralateral medial pterygoid muscle to extraction, suggesting that these factors for a longer period of this experiment could induce muscle damage related to TMD.

The Therapeutic Use of Propolis Extract in Alveolar Bone Contaminated with Bacterial Endotoxin

Propolis is a resinous substance obtained by bees, whose antibacterial, anti-inflammatory, antiviral, antifungal, immune stimulant, and local anesthetic wound healing properties have been considered for clinical practice. In particular, its anti-inflammatory and antibacterial characteristic seems to be a novel target for infectious process from dental origin. This work aimed to evaluate the propolis antibacterial potential against a bacterial endotoxin on dental alveoli. First, some properties of green propolis extract were analyzed (in vitro): 1) physicochemical profile 2) Minimum Inhibitory Concentration (MIC) against endotoxin from Gram negative Escherichia coli, and 3) its immunoregulatory activity on leukocytes from the spleen of rats. Then, an inflammatory process was induced in rats by a contamination with lipopolysaccharide (LPS) that is recognized as an endotoxin. For this purpose, rats were subjected to extractions of maxillary first molars, right and left, which immediately had the right dental socket contaminated with 0.1L of LPS (100 μg/kg). After 14 days from exodontia, these individuals were divided in groups treated with Pure Propolis Extract (EPP) and groups without therapy. The contaminated alveolar bone or the same area from animals without inflammation-induced were removed for histological and immunohistochemical processing. Our data reveal an important therapeutic action from green propolis. In vitro tests indicated low cytotoxicity for this compound. By a hematoxilin and eosin analysis, the group infected and treated with propolis presented the alveoli with more new bone tissue, characterized by bony trabeculae circling small cavities filled by a loose connective tissue containing blood vessels. Additionally, a histochemical marker of osteoclasts, tartrateresistant acid phosphatase (TRAP), was used to determine the new bone formation rate. The propolis induced more TRAP formation on alveolar bone infected by LPS. Our findings highlight the potential of propolis to be applied in dental material.