Human cartilage glycoprotein 39 (HC gp-39) has been described as a major secreted product of cultured articular chondrocytes, synovial fibroblasts, and the osteosarcoma line MG63. However, its expression in these cells types has not been directly linked to corresponding cell types in vivo. In this report, expression of HC gp-39 is demonstrated from peripheral blood-derived macrophages in association with their differentiation from monocytes to macrophages. Consistent with macrophage specificity, HC gp-39 expression is also induced upon selective stimulation of the pluripotent promyelocytic leukemia cell line HL-60 toward the monocyte/macrophage lineage with vitamin D3 or phorbol 12-myristate 13-acetate (PMA), while treatments stimulating granulocyte and eosinophilic pathways do not induce expression. Furthermore, HC gp-39 expression levels correlate with the degree of morphological differentiation induced by PMA and vitamin D3 treatments. PMA-induced mRNA expression occurs by 36 h and is a secondary transcriptional response since its synthesis is inhibited by cycloheximide. Apparently, HC gp-39 expression is tied to later events in the differentiation of monocytes into macrophages. The in vivo significance of these results is validated by the in situ detection of HC gp-39 mRNA in inflammatory macrophages associated with rheumatoid synovium. Thus, macrophages appear to be an important source of HC gp-39, which has been shown to be present at elevated levels in the blood and synovium of rheumatoid arthritis patients. The implications of this extend well beyond the previously restricted observations in cell types associated with the joint and suggest a potential involvement of macrophage-derived HC gp-39 in other aspects of inflammation, tissue remodeling, and host defense.