Figure 2.

Activation of lining macrophages at the onset of AIA. (A) Confocal fluorescence microscopy of mouse knee demonstrates CXCL1 immunostaining localizing in the lining at 4 h after AIA induction (large panel; VSIG4, magenta; CXCL1, green; CD68, blue; nuclei, gray; scale bar = 200 µm) with representative quantification of CXCL1 expression in regards to the synovial lining of the image in A; each point is one CXCL1 “surface” rendered by Imaris software with median and interquartile range indicated. Zoomed region of the lining shows expression of CXCL1 in the cells of the lining niche (bottom left panel, scale bar = 7 µm), whereas further enlargement demonstrates colocalization of CXCL1 and VSIG4 indicated with a white arrowhead, as well as CXCL1 expression in VSIG4⁻ lining cell indicated with a red arrowhead (bottom right panel, scale bar = 2 µm). F, femur; FP, fat pad; M, meniscus; PB, patellar bursa; T, tibia. Images representative of 10 sections from five different mice and two independent experiments. (B) Heatmap of genes within the GO term “Neutrophil recruitment” expressed in synovial macrophages shows higher expression of chemokines in lining cells; CXCL1 indicated with an arrowhead. (C) Confocal image of sub-patellar synovial region demonstrating fluorescent mBSA and CXCL1 coincidence in the lining area (left panel; VSIG4, magenta; CXCL1, green; CD68, blue; mBSA, yellow; nuclei, gray; scale bar = 40 µm). Enlarged region shows lining macrophages with intracellular mBSA and CXCL1 expression (right panel, arrowheads, scale bar = 7 µm). F, femur; P, patella; PB, patellar bursa; PT, patellar tendon.