Background/Purpose: Pathological subsets of fibroblast-like synoviocytes (FLS) have recently been identified as key players in the aggravation of both persistent joint inflammation and destruction in rheumatoid arthritis (RA). In RA, these FLS are considered responsible for the acidification of the synovial fluid through local lactate production and secretion. Lactate transporters such as monocarboxylate transporter-1 (MCT1) and -4 (MCT4) are upregulated on FLS in RA. This facilitates a local build-up of lactate that influence cells in their vicinity. Inhibiting MCT4 in CIA mice led to amelioration of arthritis. In this study, we hypothesized that pathological FLS could be directly targeted by MCT inhibitors to decrease their cytokine production. We investigated the effect of therapeutically inhibiting MCT-1 and -4 in human pathological RA FLS cultures.

Methods: Synovial fluid derived FLS (SF-FLS) were isolated from the synovial fluid of patients with RA (n=6) and evaluated at passage 4, and compared with normal healthy dermal fibroblasts (NHDF). These cells were analyzed by flow cytometry for expression of the surface proteins CD34, CD45, thymocyte differentiation antigen-1 (Thy-1) and podoplanin (PDPN) and additionally evaluated by light microscopy. Subsequently, they were cultured with or without added INFg or TNFa (both 10 ng/mL) and a specific inhibitor of MCT-1 (AZD3965; 1 µM) or MCT-4 (syrosingopine; 10 µM) for 24 hours. The supernatants were analyzed with an Iscus microdialyzer and ELISA.

Results: The homogeneous subpopulations of unstimulated L/D^neg+CD45^neg FLS, isolated from the synovial fluid, were positive for both Thy-1 and PDPN [94.6%, (79.9-97.4]). Without stimulation, RA SF-FLS showed substantial lactate secretion [2.567 mmol/L, (2.113-3.020)] with a factor of 1.2 more than healthy dermal fibroblasts. These levels did not increase significantly after the cells were stimulated with either INFγ or TNFα. Unstimulated pathological FLS also secreted substantial amounts of MCP-1 [2107 pg/mL, (98-4117)], elevated by a factor of >12 compared with healthy dermal fibroblasts (n=3) (p< 0.05). Furthermore, the levels between lactate and MCP-1 in the supernatant showed a significant positive correlation [rho = 0.89, (0.27-0,99)] (p< 0.02). Only when the MCT inhibitors were combined, a significant reduction in lactate secretion was detected after 24 hours. This was seen in unstimulated (p< 0.05) and INFγ stimulated cells (p< 0.05) but not after TNFα stimulation (ns) of FLS cultures. Concomitantly we detected a decrease in MCP-1 production in pathological FLS by a factor of 0.8 (0.6-0.9) (p< 0.05). This reduction in MCP-1 secretion was not seen if the cells were pre-stimulated with either INFγ or TNFα prior to MCT inhibition. The reduction in both lactate and MCP-1 levels were not detectable in the NHDF cultures.

Conclusion: In pathological RA FLS the production of lactate and MCP-1 were strongly correlated and blocking the transport of lactate across the cellular membrane in these cells let to decreased chemokine production. Overall, this human in vitro study supports that targeting metabolic lactate-transporters could be a useful and promising therapeutic strategy in RA, directly affecting pathological FLS.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/in-rheumatoid-arthritis-inhibition-of-the-lactate-monocarboxylate-transporters-1-and-4-in-pathological-fibroblast-like-synoviocytes-led-to-decreased-chemokineproduction/