Figure 7. Functional analysis of SFs from different locations.

(a) Increased constitutive and TNF-induced expression of matrix metalloproteinase 1 (MMP1) mRNA (n=11, qPCR, mean±s.d., mean ΔCT±s.d.=9.79±0.94 of MMP1 in unsilenced, untreated knee SFs) and MMP1 protein (n=8, ELISA, box and whiskers: min to max) in knee OA SFs transfected with HOTAIR siRNA versus control siRNA (scr) for 72 h (TNF (10 ng ml⁻¹) for 24 h). (b) Suppression of HOTAIR in knee SFs (n=5) by TNF (10 ng ml⁻¹, 24 h) qPCR. (c) Increased MMP1 production in response to TNF (10 ng ml⁻¹) in knee SFs. Fold change (x-fold) of MMP1 in TNF-stimulated (24 h) versus unstimulated SFs from upper extremity joints (hands: dark grey; elbows: light grey; shoulders: black) and knees, qPCR. (d) Higher expression of TNF in synovial tissues from lower extremity (ankles: dark grey; knees: light grey; hips: black) than from upper extremity joints (hands: dark grey; elbows: light grey; shoulders: black), qPCR. (e) Increased MMP13 expression in hand compared with knee synovial tissues, qPCR. (f) Increased adhesion to culture wells (xCELLigence system) of shoulder versus hand SFs. (g) Lower proliferative potential of shoulder versus hand and knee SFs (doubling time (hours), xCELLigence system). (h) Increased leukocyte migration towards supernatants of hand versus shoulder SFs. Number of leukocytes migrating towards supernatants from SFs of different locations or towards medium. Data were normalized to the number of SFs migrating towards the supernatant from one knee SF. Healthy donors are denoted with black or grey shapes. (i) Increased leukocyte infiltration in the synovium from hands versus shoulders and hips. Increased density of synovial stroma in hands versus knees. Haematoxylin/eosin-stained RA synovial tissues from different locations (see Supplementary Table 5) were scored with a synovitis score³⁸, composed of changes in stroma, synovial lining thickness and leukocyte infiltration. Statistics: paired (a (qPCR), b) or unpaired (c,d) t-test, two-tailed Wilcoxon matched-pairs signed rank test (a (ELISA)), one-way analysis of variance (ANOVA) with Bonferroni's (e–h) or Dunn's (i) post tests. *P<0.05; **P<0.01, ***P<0.001. Open shapes are OA SFs and closed shapes are RA SFs.