Altered lipid metabolism in synovial fibroblasts of individuals at risk of developing rheumatoid arthritis.

OBJECTIVE: Fibroblast-like synoviocytes (FLS) can augment the inflammatory process observed in synovium of patients with rheumatoid arthritis (RA). A recent transcriptomic study in synovial biopsies revealed changes in metabolic pathways before disease onset in absence of synovial tissue inflammation. This raises the question whether alterations in cellular metabolism in tissue resident FLS underlie disease pathogenesis. MATERIALS AND METHODS: To study this, we compared the metabolic profile of FLS isolated from synovial biopsies from individuals with arthralgia who were autoantibody positive but without any evidence of arthritis (RA-risk individuals, n = 6) with FLS from patients with RA (n = 6), osteoarthritis (OA, n = 6) and seronegative controls (n = 6). After synovial digestion, FLS were cultured in vitro and cellular metabolism was assessed using quantitative PCR, flow cytometry, XFe96 Seahorse Analyzer and tritium-labelled oleate oxidation assays. RESULTS: Real-time metabolic profiling revealed that basal (p < 0.0001) and maximum mitochondrial respiration (p = 0.0024) were significantly lower in RA FLS compared with control FLS. In all donors, basal respiration was largely dependent on fatty acid oxidation while glucose was only highly used by FLS from RA patients. Moreover, we showed that RA-risk and RA FLS are less metabolically flexible. Strikingly, mitochondrial fatty acid ß-oxidation was significantly impaired in RA-risk (p = 0.001) and RA FLS (p < 0.0001) compared with control FLS. CONCLUSION: Overall, this study showed several metabolic alterations in FLS even in absence of synovial inflammation, suggesting that these alterations already start before clinical manifestation of disease and may drive disease pathogenesis.

Studies on the substrate specificity of the inducible and non-inducible acyl-CoA oxidases from rat kidney peroxisomes.

We have studied the substrate specificity of the inducible (acyl-CoA oxidase I) and non-inducible (acyl-CoA oxidase II) oxidases in peroxisome-enriched fractions from rat kidney. The two oxidases were separated by means of ion-exchange chromatography and shown to accept a variety of acyl-CoA esters as substrates, including lignoceroyl-CoA, palmitoyl-CoA, lauroyl-CoA, caproyl-CoA, and trimethyltridecanoyl-CoA. Glutaryl-CoA was found to react exclusively with the inducible enzyme, and pristanoyl-CoA exclusively with the non-inducible enzyme. We conclude that under normal non-induced conditions both acyl-CoA oxidase I and II contribute to the oxidation of the various acyl-CoA esters with the exception of pristanoyl-CoA and glutaryl-CoA, although the extent to which each enzyme contributes to the oxidation was found to differ between the various acyl-CoA esters.

The activity of one soluble component of the cell-free NADPH:O2 oxidoreductase of human neutrophils depends on guanosine 5'-O-(3-thio)triphosphate.

Neutrophil NADPH:O2 oxidoreductase activity, essential in the killing of bacteria by neutrophils, can be elicited in a cell-free system that requires plasma membranes, cytosol and sodium dodecyl sulfate. In addition, GTP or its nonhydrolyzable analog guanosine 5'-3-O-(thio)triphosphate (GTP gamma S) enhances NADPH oxidase activity. We investigated the mechanism of this effect of GTP gamma S in the cell-free system. Cytosol from human neutrophils was separated in three different soluble oxidase components (SOC I, SOC II, and SOC III). Previously we (Bolscher, B. G. J. M., Van Zwieten, R., Kramer, I. J. M., Weening, R. S., Verhoeven, A. J., and Roos, D. (1989) J. Clin. Invest. 83, 757-763) reported that the cytosol contains two components which act synergistically. We now report that one component (previously labeled SOC II) contains two different components that can be separated by ion exchange chromatography. Immunoblotting with antiserum B-1 (Volpp, B. D., Nauseef, W. M., and Clark, R. A. (1988) Science 242, 1295-1297), directed against a cytosolic complex capable of activating latent membranes in the cell-free system, showed a 47-kDa protein in SOC II and a 67-kDa protein in SOC III. SOC II also contains the 47-kDa phosphoprotein, which indicates that this phosphoprotein and the protein recognized by the antiserum are identical. Low rates of NADPH-dependent O2 consumption can be elicited by SOC II and SOC III in the absence of SOC I. This activity is independent of GTP gamma S. Addition of SOC I increases this activity 3-4-fold, only when GTP gamma S is present. Plasma membranes, incubated with SOC I plus GTP gamma S and re-isolated, showed a similar 3-4-fold enhanced O2 consumption with SOC II and SOC III. The GTP gamma S effect is exerted primarily at the level of the plasma membrane. The concentration of GTP gamma S that causes a half-maximal stimulation was 0.4 mu M. It is concluded that SOC I is a functional component of the NADPH oxidase.