A new ex vivo model of the bone tissue response to the hyperglycemic environment - the embryonic chicken femur organotypic culture in high glucose conditions

Abstract

Diabetes mellitus (DM) embrace a group of chronic metabolic conditions with a high morbidity, causing deleterious effects in different tissues and organs, including bone. Hyperglycemia seems to be one of the most contributing etiological factors of bone-related alterations, altering metabolic functionality and inducing morphological adaptations. Despite the established models for the assessment of bone functionality in hyperglycemic conditions, in vitro studies present a limited representativeness given the imperfect cell-cell and cell-matrix interactions, and restricted three-dimensional spatial arrangement; while in vivo studies raise ethical issues and offer limited mechanistic characterization, given the modulatory influence of many systemic factors and/or regulatory systems. Accordingly, the aim of this study is to establish and characterize an innovative ex vivo model of the bone tissue response to hyperglycemia, reaching hand of the organotypic culture of embryonic chicken femurs in high glucose conditions, showcasing the integrative responsiveness of the model regarding hyperglycemia-induced alterations. A thorough assessment of the cellular and tissue functionality was further conducted. Results show that, in high glucose conditions, femurs presented an increased cell proliferation and enhanced collagen production, despite the altered protein synthesis, substantiated by the increased carbonyl content. Gene expression analysis evidenced that high glucose levels induced the expression of pro-inflammatory and early osteogenic markers, further impairing the expression of late osteogenic markers. Furthermore, the tissue morphological organization and matrix mineralization were significantly altered by high glucose levels, as evidenced by histological, histochemical and microtomographic evaluations. Attained data is coherent with acknowledged hyperglycemia-induced bone tissue alterations, validating the models' effectiveness, and evidencing its integrative responsiveness regarding cell proliferation, gene and protein expression, and tissue morpho-functional organization. The assessed ex vivo model conjoins the capability to access both cellular and tissue outcomes in the absence of a systemic modulatory influence, outreaching the functionality of current experimental in vitro and in vivo models of the diabetic bone condition.