Differential metabolism of 25-hydroxyvitamin D3 (25(OH)D3) has been shown for macrophages and fibroblast-like cells (possibly synoviocytes) cultured for two to 50 days after isolation from the synovial fluid of 12 patients with various forms of inflammatory arthritis. Macrophages synthesised the active metabolite of vitamin D3, 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), the synthesis of which was increased by bacterial lipopolysaccharide, a known macrophage activating factor. In contrast, fibroblast-like cells formed 24, 25-dihydroxyvitamin D3 (24,25(OH)2D3), synthesis of which was stimulated by 1,25(OH)2D3 and inhibited by lipopolysaccharide. The synthesis of 1,25(OH)2D3 and 24,25(OH)2D3 by macrophages and fibroblast-like cells respectively was inhibited by ketoconazole, indicating that both hydroxylases are dependent on cytochrome P-450. Mean (SEM) synovial fluid and serum 25(OH)D3 concentrations were 16.7 (1.7) and 22.2 (2.6) ng/ml and those of 1,25(OH)2D3 were 29.4 (4.8) and 43.3 (4.0) pg/ml respectively. In most cases concentrations were lower in synovial fluid than in paired serum samples, but in two patients 1,25(OH)2D3 concentrations were greater in synovial fluid than in serum, suggesting local synthesis within the affected joints.