Implant macrogeometry and functionalization with anti-SOST as a strategy to improve peri-implant repair in osteopenic rats

• Grant number: 23/02175-4
• Support Opportunities: Scholarships in Brazil - Doctorate
• Effective date (Start): September 01, 2023
• Effective date (End): February 28, 2027
• Field of knowledge: Health Sciences - Dentistry - Oral and Maxillofacial Surgery
• Principal Investigator: Roberta Okamoto
• Grantee: Paula Buzo Frigério
• Host Institution: Faculdade de Odontologia (FOA). Universidade Estadual Paulista (UNESP). Campus de Araçatuba. Araçatuba , SP, Brazil

Abstract

Since Branemark's discovery in the 1960s, osseointegrated titanium implants have been the best alternative for edentulous patients. The main objective of this project is to evaluate strategies related to the macrogeometric characteristics and surface treatment of implants that enable the improvement of reparative bone tissue in the presence of systemic compromise. The preliminary stage (in vitro) will be the functionalization of titanium implants with the anti-sclerostin antibody (a-SOST), using the layer-by-layer technique. Physical and in vitro tests (cell cultures) will be performed in order to evaluate the properties of the functionalized surface to improve osteogenic responses. The studies developed in cultures of undifferentiated mesenchymal cells obtained from rat femurs will allow the selection of the best concentration of the biomolecule to be used in drug delivery. The MTT (3-4,5-dimethylthiazzo bromide), cell viability, genotoxicity, alkaline phosphatase activity and mineralization nodule formation will be tested from the mesenchymal cells on titanium discs with different concentrations of the biomolecule. After selecting the concentration of the anti-sclerostin antibody that promotes the best cell response, physical tests will be performed to characterize the surface through scanning electron microscopy and the release tests of the biomolecule to characterize its acting time in situ. The second part of the project will consist of in vivo experiments to evaluate the effect of this functionalized surface during peri-implant repair. We will use 128 rats divided into two large groups, according to sham surgery or osteoporosis induction: 1 - SHAM (n = 64): rats submitted to sham surgery, without osteopenia induction, and 2 - OVX (n = 64): rats submitted to bilateral ovariectomy for osteopenia induction. Within each experimental group there will be two subgroups, according to the macrogeometric surface of the implants: 1 - SOL: implants with solid surface and 2 - PO: implants with porous surface, then, they will be subdivided according to the surface treatment: 1 - a-SOST: implants that will be functionalized with the anti-sclerostin antibody and 2 - SF: implants that will not have functionalization on their surface. After 30 days of the ovariectomy surgery, all animals will have implants installed in the right and left tibial metaphysis (n = 256 implants), being them solid (SOL) or porous (PO) with (a-SOST) or without (SF) the treated surface. On day 44 the animals will receive 20 mg/kg calcein, and after 10 days they will receive 30 mg/kg alizarin for the confocal laser microscopy analysis. The euthanasia will occur in two periods, 14 (n = 64 rats) or 28 days (n = 64 rats) after the implantation surgery. The proposed analyses aim to characterize in decalcified sections the histology and the immunostaining of proteins related to bone repair at 14 days postoperatively. In both periods, removal torque (biomechanical) and molecular biology (real-time PCR) analyses will also be performed for the expression of genes also related to bone repair. Only at 28 days, the analyses of computerized microtomography will be carried out, seeking to characterize the micro-architecture of the repair bone and through the evaluation by confocal microscopy the pattern of mineral precipitation on the collagen matrix and bone turnover will be evaluated. At 28 days, samples will also be selected to be embedded in methylmethacrylate (PMMA) to be submitted to ultrastructural analysis to be performed in the laboratory of Prof. Kathryn Grandfield, researcher associated with this project. (AU)