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Metabolic reprogramming in chondrocytes to promote mitochondrial respiration reduces downstream features of osteoarthritis.

ABSTRACT: Metabolic dysfunction in chondrocytes drives the pro-catabolic phenotype associated with osteoarthritic cartilage. In this study, substitution of galactose for glucose in culture media was used to promote a renewed dependence on mitochondrial respiration and oxidative phosphorylation. Galactose replacement alone blocked enhanced usage of the glycolysis pathway by IL1β-activated chondrocytes as detected by real-time changes in the rates of proton acidification of the medium and changes in oxygen consumption. The change in mitochondrial activity due to galactose was visualized as a rescue of mitochondrial membrane potential but not an alteration in the number of mitochondria. Galactose-replacement reversed other markers of dysfunctional mitochondrial metabolism, including blocking the production of reactive oxygen species, nitric oxide, and the synthesis of inducible nitric oxide synthase. Of more clinical relevance, galactose-substitution blocked downstream functional features associated with osteoarthritis, including enhanced levels of MMP13 mRNA, MMP13 protein, and the degradative loss of proteoglycan from intact cartilage explants. Blocking baseline and IL1β-enhanced MMP13 by galactose-replacement in human osteoarthritic chondrocyte cultures inversely paralleled increases in markers associated with mitochondrial recovery, phospho-AMPK, and PGC1α. Comparisons were made between galactose replacement and the glycolysis inhibitor 2-deoxyglucose. Targeting intermediary metabolism may provide a novel approach to osteoarthritis care.

SUBMITTER: Ohashi Y

PROVIDER: S-EPMC8302637 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Complement component 3 fragment C3a is an anaphylatoxin involved in promoting cellular responses important in immune response and host defense. Its receptor (C3a receptor, C3aR) is distributed on the plasma membrane; however, lysosomal localization in immune cells has been reported. Oxidative stress increases intracellular reactive oxygen species (ROS), and ROS activate complement signaling in immune cells and metabolic reprogramming. Here we tested oxidative stress and intracellular complement in mitochondrial dysfunction in RPE cells using high resolution live-cell imaging, and metabolism analysis in isolated mitochondria using Seahorse technology. While C3aR levels were unaffected by oxidative stress, its cell membrane levels decreased and mitochondrial (mt) localization increased. Trafficking was dependent on endocytosis, utilizing endosomal-to-mitochondrial cargo transfer. H2O2-treatment also increased C3a-mtC3aR co-localization dose-dependently. In isolated mitochondria from H2O2-treated cells C3a increased mitochondrial Ca2+ uptake, that could be inhibited by C3aR antagonism (SB290157), mitochondrial Ca2+ uniporter blocker (Ru360), and Gαi-protein inhibition (pertussis toxin, PTX); and inhibited mitochondrial repiration in an SB290157- and PTX-dependent manner. Specifically, mtC3aR activation inhibited state III ADP-driven respiration and maximal respiratory capacity. Mitochondria from control cells did not respond to C3a. Furthermore, transmitochondrial cybrid ARPE-19 cells harboring J haplogroup mitochondria that confer risk for age-related macular degeneration, showed high levels of mtC3aR and reduced ATP production upon C3a stimulation. Our findings suggest that oxidative stress increases mtC3aR, leading to altered mitochondrial calcium uptake and ATP production. These studies will have important implication in our understanding on the balance of extra- and intracellular complement signaling in controlling cellular health and dysfunction.

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Metabolic reprogramming in chondrocytes to promote mitochondrial respiration reduces downstream features of osteoarthritis.

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