THU0016 Transmitochondrial cybrids show that oxphos via, but no glycolysis via, is involved in the atp reduction of oa human chondrocytes
Authors: M. Fernández-Moreno, A. Dalmao-Fernandez, T. Hermida-Gomez, A. Rustan, J. Lund, M.E. Vazquez-Mosquera, S. Relano-Fernandez, I. Rego-Perez, F. Blanco-Garcia
Journal: Annals of the Rheumatic Diseases (2018)

<h3>Background</h3> Mitochondrial dysfunction is well documented in OA and has the capacity to alter chondrocyte function and viability, contributing to cartilage degeneration. It is important to evaluate the influence of mitochondria in the pathogenesis of OA using an <i>in vitro</i> model to explain the functional consequences of this association and help us to identify potential diagnostic biomarkers and/or therapeutic targets Transmitochondrial cybrids are a useful cellular model to study the mitochondrial biology and function implications in the cellular behaviour, since they carry different mitochondrial variants with the same nuclear background, therefore, excluding the variations because of nuclear genome. <h3>Objectives</h3> The aim of this work is test mitochondrial activity in the OA chondrocytes using transmitochondrial cybrids with mtDNA from healthy donors (without OA) and from patients with OA. <h3>Methods</h3> Cybrids were developed using 143B. TK<sup>-</sup> Rho-0 cell line (nuclear donor) and platelets (mitochondrial donors) from healthy and OA donors. Human articular chondrocytes were obtained from patients with hip replacement. The mtDNA copy number was measured by real-time PCR method. The ROS production was evaluated using flow cytometry. The metabolic status was evaluated by glucose consumption and glucose oxidation. The glycolytic activity was measure after addition of glucose, oligomycin and 2-dioxyglucose using Seahorse XFp (ECAR). The OXPHOS function was evaluated by SeaHorse XFp (OCR) after addition of oligomycin, FCCP and Rotenone/Antimycin. Appropriate statistical analyses were performed with GraphPad Prism v6. <h3>Results</h3> The analysis of mtDNA copy number showed that the OA have higher levels than N in cybrids and human chondrocytes. The analysis of ROS production showed that OA had higher levels than N in both types of cells. The metabolic status analysing glucose consumption, glucose oxidation and total glucose cellular uptake reflected higher values in OA cybrids than N cybrids. But the analysis of glycolysis data showed lower values in OA than N cybrids. The analysis of ATP obtained through glycolysis did not show any difference between cybrids. The analysis of OXPHOS function showed that OA had lower basal respiration and maximal respiratory capacity than N in both types of cells. The ATP obtained via OXPHOS was lower in OA than in N. <h3>Conclusions</h3> The analysis of OXPHOS function supports the participation of mitochondria in cybrids and human chondrocytes metabolism. Both types of cells use the mitochondria to obtain ATP and OXPHOS via, but no glycolysis, is involved in the reduction of ATP synthesis by OA cells. All these data support that N cybrids and chondrocytes use mitochondria with more efficiency. <h3>Disclosure of Interest</h3> None declared