To elucidate histamine receptor–mediated signaling pathways, transcriptional events, and target gene expression in human cartilage.

Histamine modulation of cartilage destruction was assessed by Safranin O staining and proteoglycan release. H₁, H₂, H₃, and H₄ histamine receptor–dependent regulation of transcription factors (nuclear receptor 4A1 [NR4A1], NR4A2, and NR4A3), RANKL, and osteoprotegerin (OPG) messenger RNA (mRNA) levels were measured in primary and SW‐1353 chondrocyte cells using quantitative polymerase chain reaction and selective histamine receptor antagonists. Soluble RANKL and OPG protein levels were determined using enzyme‐linked immunosorbent assays. NR4A protein levels and transactivity were evaluated by Western blot analysis, immunocytochemistry, and luciferase reporter assays. Stable depletion of NR4A1–3 was achieved by lentiviral transduction of NR4A short hairpin RNA.

Primary human chondrocyte cells expressed differential steady‐state levels of H₁–H₄ histamine receptor mRNA. In combination with tumor necrosis factor α, histamine significantly promoted cartilage proteoglycan depletion and release. Histamine modulated the expression of NR4A1–3 orphan receptors in primary and immortalized human chondrocyte cells in a time‐ and concentration‐dependent manner. Histamine selectively signaled through H₁ and H₂ histamine receptors in chondrocytes to modulate RANKL and NR4A2 expression. The temporal effects of histamine on NR4A2 gene transcription were reduced in cells pretreated with inhibitors directed against protein kinase A, MAPK, and NF‐κB signaling pathways. Histamine modulated the expression of RANKL with modest effects on OPG levels, leading to increased RANKL:OPG mRNA and protein ratios. Stable knockdown of NR4A1–3 expression resulted in reduced endogenous OPG levels and the loss of histamine‐dependent regulation of RANKL expression.

Our findings indicate that histamine, via H₁ and H₂ histamine receptors, contributes to joint disease by enhancing the ratio of RANKL to OPG expression through altered NR4A activity in human chondrocyte cells.