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Analysis of the dynamics of the mandibular bone tissue, femur neck and spine of zoledronic acid-treated rats with osteoporosis and systemic administration of ozone

Grant number: 19/19445-9
Support Opportunities:Scholarships in Brazil - Scientific Initiation
Effective date (Start): March 01, 2020
Effective date (End): February 28, 2021
Field of knowledge:Health Sciences - Dentistry - Oral and Maxillofacial Surgery
Principal Investigator:Leonardo Perez Faverani
Grantee:Maria Eloise de Sá Simon
Host Institution: Faculdade de Odontologia (FOA). Universidade Estadual Paulista (UNESP). Campus de Araçatuba. Araçatuba , SP, Brazil


This paper will aim to analyze the potential of ozone therapy in bone tissue dynamics of osteoporotic rats treated with zolendronic acid. For this, 60 female Wistar rats, 6 months old will be submitted to bilateral ovariectomy (Ovx). After three months, four animals will be euthanized to characterize the bone tissue architecture in microtomography (micro-ct), soon after, the animals will be randomly divided into four groups (SAL, SAL + ZOL, SAL + OZN, ZOL + OZN) containing 6 animals in each. The drug protocol will be initiated in the animals of the groups (ZOL + SAL, ZOL + OZN), with zoledronate 100 µg / kg / 28 days, which will be diluted in 0.45 ml 0.9% sodium chloride solution, The protocol will be started after the third month of ovariectomy. The SAL group will receive 0.45 ml saline using the same protocol as ZOL and these therapies will continue until the end of the experiment. After three months of drug therapy (sixth month), four animals from the SAL group and four animals from the SAL + ZOL group will be euthanized to characterize the bone architecture (Micro ct). After this procedure, ozone therapy at a concentration of 0.7 mg / kg will be started every two days until the end of the experiment (SAL + OZN and ZOL + OZN groups). 30 and 60 days after the initiation of ozone therapy, six animals from each group will be euthanized for bone structural analysis and characterization of the mandible, femoral neck and spine regions. The obtained samples will be destined for analysis of calcified and decalcified bone tissue. In the first group of samples they will be dehydrated and included in photopolymerizable resin to obtain slides and laser confocal microscope analysis. This will allow the visualization of the fluorochromes calcein (which will represent old bone) and alizarin (which will represent neoformed bone) for the analysis of bone tissue dynamics. Both fluorochromes will be administered in all groups at a dose of 20 mg / kg. Another part of the calcified parts will be destined to biomechanical tests in order to evaluate the bone resistance. For histological analysis, the collected samples will be decalcified, embedded in paraffin and microtomized to obtain slides. These will be stained with hematoxylin and eosin for histological analysis of the biological events of bone dynamism ("bone turnover"). All quantitative data will be submitted to the normality curve to establish the best statistical test (parametric or nonparametric), considering p <0.05 the level of significance.

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