Alveolar bone healing in rats: micro-CT, immunohistochemical and molecular analysis.

Jaqueline Suemi Hassumi,Ricardo Garcia Mureb Jacob,André Luis Da Silva Fabris,Alexandre Rodrigues Freire,Gabriel Mulinari-Santos,Alaíde Gonçalves,Leonardo Pérez Faverani,Ana Cláudia Rossi,Roberta Okamoto

doi:10.1590/1678-7757-2017-0326

Abstract

Alveolar bone healing after upper incisor extraction in rats is a classical model of preclinical studies. The underlying morphometric, cellular and molecular mechanism, however, remains imprecise in a unique study. Objectives The aim of this study was therefore to characterize the alveolar bone healing after upper incisor extraction in rats by micro computed tomographic (Micro-CT), immunohistochemical and real-time polymerase chain reaction (RT-PCR) analysis.Material and Methods Thirty animals (Rattus norvegicus, Albinus Wistar) were divided into three groups after upper incisors extraction at 7, 14, and 28 days. Micro-CT was evaluated based on the morphometric parameters. Subsequently, the histological analyses and immunostaining of osteoprotegerin (OPG), receptor activator of nuclear kappa B ligand (RANKL) and tartrate resistant acid phosphate (TRAP) was performed. In addition, RT-PCR analyses of OPG, RANKL, the runt-related transcription factor 2 (RUNX2), osteocalcin (OC), osteopontin (OPN), osterix (OST) and receptor activator of nuclear kappa B (RANK) were performed to determine the expression of these proteins in the alveolar bone healing.Results Micro-CT: The morphometric parameters of bone volume and trabecular thickness progressively increased over time. Consequently, a gradual decrease in trabecular separation, trabecular space and total bone porosity was observed. Immunohistochemical: There were no differences statistically significant between the positive labeling for OPG, RANKL and TRAP in the different periods. RT-PCR: At 28 days, there was a significant increase in OPG expression, while RANKL expression and the RANKL/OPG ratio both decreased over time.Conclusion Micro-CT showed the newly formed bone had favorable morphometric characteristics of quality and quantity. Beyond the RUNX2, OC, OPN, OST, and RANK proteins expressed in the alveolar bone healing, OPG and RANKL activity showed to be essential for activation of basic multicellular units during the alveolar bone healing.

Highlights

Given the search for a favorable bone and an ideal support for dental implant placement, understanding the alveolar bone healing in preclinical studies is crucial
real-time polymerase chain reaction (RT-PCR): At 28 days, there was a significant increase in OPG expression, while receptor activator of nuclear kappa B ligand (RANKL) expression and the RANKL/OPG ratio both decreased over time
Beyond the RUNX2, OC, OPN, OST, and receptor activator of nuclear kappa B (RANK) proteins expressed in the alveolar bone healing, OPG and RANKL activity showed to be essential for activation of basic multicellular units during the alveolar bone healing

Summary

Introduction

Given the search for a favorable bone and an ideal support for dental implant placement, understanding the alveolar bone healing in preclinical studies is crucial. Bone is a dynamic tissue, where bone cells drive the molecular and cellular mechanisms involved in the bone healing. Bone is a dynamic tissue, where bone cells drive the molecular and cellular mechanisms involved in the bone healing1,12 These cells act together signaling molecules to maintain the bone turnover. The mechanisms of development and maintenance of bone occurs constantly, since local factors such as mechanical stimulation and systemic factors can interfere in this process. Bone turnover is balanced by formation and resorption, allowing the maintenance of bone mass and ensuring calcium and phosphate levels.

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