Session Type: Poster Session (Tuesday)
Session Time: 9:00AM-11:00AM
Background/Purpose: Background. With the redefinition of Osteoarthritis (OA) and the understanding that the joint behaves as an organ, OA is now considered a systemic illness with a low grade of chronic inflammation. Chondroptosis, chondrosenescence and autophagy contribute to cell death and tissue damage in OA. The mitochondria are related with these three process implicated in the cartilage degeneration. Mitochondrial dysfunction is well documented in OA and has the capacity to alter chondrocyte and synoviocyte function contributing to cartilage degeneration. Transmitochondrial cybrids are a useful cellular model to study mitochondrial biology in vitro, since they carry different mitochondrial variants with the same nuclear background
Purpose: The aim of this work was to study apoptosis, senescence and autophagy using cybrids with mtDNA from healthy (N) and OA donors.
Methods: Methods Cybrids were developed using 143B.TK– Rho-0 cell line (nuclear donor) and platelets (mitochondrial donors) from healthy (N) and OA donors. The mitochondrial depolarized and morphology were evaluated incubating cells with DilC1(5) and MitoTracker Red® respectively and these parameters were analyzed using Flow Cytometer. The percentage of apoptotic cells was measured by Flow Cytometry using Annexin-V and PI. Senescence level was measured by real-time PCR method. Autophagy was evaluated through the developed of Microtubule-associated protein 1A/1B-light chain 3 (LC3) WB. The WB quantification was developed using Image J software. Appropriate statistical analyses were performed with GraphPad Prism v6.
Results: Results OA cybrids showed higher increment depolarized mitochondria under negative stimuli (2.57±1.20; 1.76±0.99; p≤0.05 respectively). Mitochondrial distribution showed that in OA cybrids mitochondria were concentrated around the nucleus whereas in N cybrids were organized in extended tubular structures. The quantification of staining reflected a decrease of fluorescence intensity in OA compared to N cybrids (14.31±3.16; 33.00±4.20 respectively, p≤0.005). When the cells were submitted for a positive stimuli and an inflammatory environment the analysis of apoptotic levels were developed and reflected that OA cybrids had an increase in positive cells for Annexin-V in comparison to N cybrids (2µM Staurosporine 15.68±6.39; 6.41±4.88 respectively, p≤0.05. 10ng/ml IL-1β 0.924±0.19; 0.47±0.24 respectively, p≤0.05). The gene expression corresponding to senescence marker protein (SMP30) showed higher levels in OA cybrids than in N (4.535±1.63; 1.21±0.42 respectively, p≤0.0005). Autophagy was analyzed studying LC3 a marker for autophagosome formation and the results showed that LC3 activation was reduced in OA cybrids (1.19±0.24; 1.41±0.21 respectively, p≤0.05).
Conclusion: Conclusion: Mitochondria from OA donors was involved in three relevant processes related with OA as cellular apoptosis, senescence and autophagy.
To cite this abstract in AMA style:Fernandez-Moreno M, Dalmao-Fernández A, Nogueira-Recalde U, Hermida-Gómez T, Vazquez-Mosquera M, Relaño S, Rego-Pérez I, Blanco F. Role of Mitochondrial DNA from OA Patients in Cellular Apoptosis, Senescence and Autophagy [abstract]. Arthritis Rheumatol. 2019; 71 (suppl 10). https://acrabstracts.org/abstract/role-of-mitochondrial-dna-from-oa-patients-in-cellular-apoptosis-senescence-and-autophagy/. Accessed April 13, 2021.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/role-of-mitochondrial-dna-from-oa-patients-in-cellular-apoptosis-senescence-and-autophagy/