Histopathological analysis and evaluation of macrophage infiltration in knee joints of FVIII^−/− , EPCR⁺⁺ FVIII^−/− , and EPCR^−/− FVIII^−/− mice 2 weeks after needle injury. Joint bleeding in FVIII^−/−, EPCR⁺⁺FVIII^−/−, and EPCR^−/−FVIII^−/− mice was induced by needle puncture of joint. The mice were left either untreated (0) or treated with a single dose or 3 doses of rFVIIa, as described in the Figure 1 legend. Fourteen days after injury, knee joints were excised and sectioned, and tissue sections were stained with H&E (A) or the macrophage marker F4/80 (C). As a control, knee joint tissue sections of uninjured mice were also stained. The original magnification of images shown in the top lane is ×4. The area identified in the square box was reimaged at higher magnification (×40) and shown in the bottom lane. The histology of uninjured knee joints exhibits distal femur and proximal tibia, synovium with ≤4 cell layers of the synovial lining, and subsynovial cells associated with fat cells in the synovium with the meniscus in the center. (A) Two thin black arrows pointing each other indicate width of synovial lining layer, the yellow arrowhead points out red blood cells, and the thick black arrows point out synovial villi. (C) Arrows point out macrophages. (B) The joint tissue pathology was quantified by scoring H&E-stained sections on a 0 to 6 scale for synovial hyperplasia (0, normal, less than 4 cell layers thick; 1, 4-5 layers thick 1; 6-7 layers thick; 3, >7 layers), presence of red blood cells (0, absent; 1, present), villus formation (0, absent; 1, present), and discoloration by hemosiderin (0, absent; 1, present). (D) Macrophage infiltration was quantified on a 0 to 4 scale (0, absence of macrophages; 1, scattered macrophages; 2, line of macrophages; 3, a cluster of macrophages; 4, sheets of macrophages). *P < .05; **P < .01; ***P < .001.