Metabolic Reprogramming in the Inflamed Joint Inhibits Pro-Inflammatory Mechanisms
M. Biniecka, Canavan M, Gao W, Ng CT, Cregan S, Smith T, McGarry T, Veale DJ, Fearon U
Centre for Arthritis and Rheumatic Diseases, Dublin Academic Medical Centre, St. Vincent’s University Hospital, Dublin, Ireland
Hypoxia is a powerful trigger of synovial cell activation, proliferation and survival. Metabolic turnover of the inflamed synovium outpaces a dysfunctional vascular supply which may induce a cellular metabolic switch towards glycolysis.
To examine the relationship between synovial hypoxia, cellular bioenergetics and mitochondrial dysfunction with inflammation.
Primary RASFCs were cultured with hypoxia, DMOG or lactic acid. Mitochondrial respiration (reactive oxygen species [ROS], mitochondrial membrane potential [MMP], mitochondrial mass [MM]), mitochondrial genome mutations and morphology, along with cell invasion, pro-inflammatory cytokines, glucose and lactate were assessed using specific functional assays and TEM. RASFC metabolism was assessed by the XF24 Flux analyser (Seahorse), which simultaneously quantifies real-time measurements of aerobic (Oxygen Consumption Rate) and anaerobic (Extracellular Acidification) bioenergetic profiles. In vivo synovial tissue glycolysis and oxidative phosphorylation (GAPDH, PKM2, GLUT1, ATP), inflammation and angiogenesis were quantified by immunohistology in synovial tissue (ST) from 44 patients with active inflammatory arthritis and paired ST oxygen (tpO2) measurements measured at arthroscopy. Finally the effect of blocking glycolysis using a small molecule 3PO on RASFCs and endothelial cell (EC) migration, angiogenesis, pro-inflammatory cytokines and transcriptional regulation of HIF1alpha, pSTAT3 and Notch signalling was examined.
In RASFC, hypoxia increased mitochondrial mutations, MM, MMP, ROS and invasion but inhibited ATP indicating altered cellular energy metabolism.
Hypoxia alters cellular bioenergetics by downregulating mitochondrial respiration and promoting a switch to glycolysis in the inflamed joint. This enables synovial cells to generate sufficient ATP to support abnormal angiogenesis, immune responses, cellular invasion and pannus formation.