Efeito da incorporação da testosterona ao compósito de poli ácido lático-co-glicólico/policaprolactona/Fosfato de cálcio bifásico na resposta biológica in vitro e in vivo

Abstract

Testosterone is a natural androgen hormone, which plays a key role in maintaining bone mass and it was suggested that it has a topical activity in the bone tissue formation. The calcium phosphate ceramic/polymer composite are promising materials in bone tissue engineering, because it combines the bioactivity of bioceramics with the flexibility of the polymers. The aim of this research was to characterize the testosterone (T) incorporation to the composite of poly (lactic-co-glycolic acid) (PLGA), polycaprolactone (PCL) and biphasic calcium phosphate (BCP) and to evaluate the cellular response and biocompatibility in vitro and in vivo. The PLGA/PCL/BCP/T and PLGA/PCL/BCP composites were prepared and characterized by thermal analysis and SEM-EDS. The results of the thermal analysis showed that the testosterone incorporation to the composite modified the profile of thermal degradation of the PLGA/PCL/BCP composite. The morphology and the surface composition of the composites determined by SEM and EDS, respectively, showed a porous structure homogeneously distributed and prevalence of calcium and phosphate in the composition. In vitro bioactivity test has shown that after 21 days in SBF was greater the exposure of the pores and increased intensity of the peaks of calcium and phosphate, featuring a layer similar to hydroxyapatite. Primary culture osteoblasts derived from neonatal Wistar rat calvaria were grown with the composites and after 7, 14 and 21 days, in vitro assays were performed to evaluate the biological activity and cytotoxicity increased significantly (P < 0.05), the viability and proliferation of osteoblasts with respect to the composite without hormone. The alkaline phosphatase activity, the collagen production by 14 days and the concentration of osteocalcin in osteoblasts cultures seeded with the PLGA/PCL/BCP/T composite were significantly higher (P < 0.05) compared to cultures with the composite without testosterone. As well as, the matrix mineralization process in the group of composite with T was accelerated, featuring mineralization nodules from seven days. The concentration of osteoprotegerin and nitric oxide in the cultures supernatants were not influenced by the addition of hormone to the PLGA/PCL/BCP composite. The results of subcutaneous implantation in rats of the composites showed that they are biocompatible, that promote neovascularization and there was a strong tissue/biomaterial interaction, regardless of the T addition. The association of testosterone to the composite stimulates bone cells positively increasing the viability and proliferation, the extracellular matrix production and mineralization. Therefore, the PLGA/PCL/BCP/T composite is a promising biomaterial for bone tissue engineering.