Granulocyte-macrophage colony stimulating factor activates proteoglycan, type II collagen, and cAMP production by rat articular chondrocytes through specific binding sites

Objective. To evaluate the effects of granulocyte-macrophage colony stimula ting factor (GM-CSF) on rat articular chondrocyte (AC) with respect to DNA synthesis, collagen type II and proteoglycan (PG) synthesis and expression, and cAMP production; to examine these cells for the presence of GM-CSF-spe cific binding sites, and to study their regulation by growth factors and cy tokines. Methods. First passage monolayers of rat AC were incubated with various con centrations of recombinant human GM-CSF, and then [H-3]-thymidine, [H-3]-pr oline, and [S-35]SO4 incorporation and cAMP production were measured. The d ensity of GM-CSF-specific binding sites, the effects of growth factors and cytokines on receptor density, and the activation of certain post-receptor signaling pathways were also examined by labeling the cell monolayers with [I-125]-GM-CSF. Results. GM-CSF (6-100 U/ml) inhibited (30%) [H-3]-thymidine incorporation into DNA, and, in contrast, stimulated up to 3.6- and 2-fold [S-35]SO4 and [H-3]-proline incorporation into glycosaminoglycan side chains and collagen molecules, respectively. GM-CSF also increased aggrecan and type It collag en (Coll II) transcripts by 2- to 3-fold, respectively. These effects were associated with a concentration-dependent increase in cAMP production. A si ngle class of high affinity (K-d = 98 pM; B-max = 7.08 pM/mug DNA) binding sites of about 220 kDa were found. The [I-125]-GM-CSF binding to the cells was slightly increased with phorbol 12-myristate 13-acetate (PMA), insulin- like growth factor-I, platelet derived growth factor, basic fibroblast grow th factor, and tumor necrosis factor-alpha, and decreased with pertussis to xin, cholera toxin, and interleukin-1B. Conclusion. These results suggest that GM-CSF may play a role in the regula tion of chondrocyte metabolism as an anabolic agent and may stimulate carti lage healing under pathological conditions.