Background/Purpose: In RA, autoimmunity precedes clinical symptoms by a decade and persists unabated even when downstream inflammation is well controlled; exposing lymphocytes to chronic stimulation and enforcing adaptation to persistent stress. T cells undergo rapid and extensive clonal expansion with a fairly unique need to greatly enhance metabolic activities and fulfill their energy needs by up-regulating aerobic glycolysis as well as autophagy. Glycolytic flux is mainly controlled by 6-phosphofructo-1-kinase, with its allosteric activator fructose 2,6-bisphosphate as a key regulator of the glycolytic pathway. The enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoform 3 (PFKFB3) generates fructose-2,6-bisphosphate and, hence, critically regulates the glycolytic rate under normal and pathophysiological conditions.

Methods: CD4⁺CD45RO^– T cells purified from the blood of RA patients (n=77) and age-matched controls were stimulated by TCR crosslinking. Glucose consumption, intracellular ATP and lactate production were quantified. Expression of 33 glycolysis-associated genes was profiled by RT-PCR. Protein levels of PFKFB3 and the autophagy marker LC3II were measured by Western blotting. Glycolytic flux and autophagic signaling were blocked through the PFKFB3 inhibitor N-BrEt or the autophagy inhibitor 3-methyladenine.

Results: Compared to control CD4 T cells, RA CD4 T cells respond to stimulation-induced metabolic needs with impaired glucose consumption (p=0.015), diminished ATP production (p<0.001)and reduced lactate release (p<0.001); all indicative of a defect in glycolytic flux. RA T cells express normal densities of glucose receptors but consistently fail to up-regulate PFKFB3 (p=0.012) and induce insufficient amounts of LC3II (p=0.032). Deficiency in glucose metabolism and autophagy is associated with increased apoptosis (p=0.003). This phenotype is reproduced in control T cells by knockdown of PFKFB3 or blocking autophagic signaling. Overexpression of ectopic PFKFB3 in RA T cells restores intracellular ATP (p=0.009) and lactate production (p=0.024) and conveys apoptotic resistance. Restoring PFKFB3 in RA T cells also rectifies the autophagic defect (p=0.024). Energy deprivation is a stable phenotype of RA T cells, independent from disease activity and duration.

Conclusion: The inducible glycolytic rate-limiting enzyme PFKFB3 is repressed in RA T cells, resulting in deficient glucose utilization. The energy deprivation is aggravated by insufficient access to cell-internal energy resources and renders RA T cells apoptosis sensitive. Resetting the immune system in RA will require the repair of these metabolic abnormalities as they strain immune homeostasis and sustain chronic immune stress.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/metabolic-reprogramming-of-autoimmune-t-cells-in-rheumatoid-arthritis/