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 stimulating 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-specific binding sites; and to study their regulation by growth factors and cytokines.


METHODS
First passage monolayers of rat AC were incubated with various concentrations of recombinant human GM-CSF, and then [3H]-thymidine, [3H]-proline, and [35S]SO4 incorporation and cAMP production were measured. The density 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 [125I]-GM-CSF.


RESULTS
GM-CSF (6-100 U/ml) inhibited (30%) [3H]-thymidine incorporation into DNA, and, in contrast, stimulated up to 3.6- and 2-fold [35S]SO4 and [3H]-proline incorporation into glycosaminoglycan side chains and collagen molecules, respectively. GM-CSF also increased aggrecan and type II collagen (Coll II) transcripts by 2- to 3-fold, respectively. These effects were associated with a concentration-dependent increase in cAMP production. A single class of high affinity (Kd = 98 pM; Bmax = 7.08 pM/microg DNA) binding sites of about 220 kDa were found. The [125I]-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 growth factor, and tumor necrosis factor-alpha, and decreased with pertussis toxin, cholera toxin, and interleukin-1beta.


CONCLUSIONS
These results suggest that GM-CSF may play a role in the regulation of chondrocyte metabolism as an anabolic agent and may stimulate cartilage healing under pathological conditions.
Quintero
M
M
Department of Rheumatology, Hospital and University of Los Andes, Merida, Venezuela.
Colantuoni
G
G
Khatib
A M
AM
Panasyuk
A
A
Lomri
A
A
Mitrovic
D R
DR
D000818:Animals; D001665:Binding Sites; D002358:Cartilage, Articular / Q000166:cytology; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects* / Q000378:metabolism*; D024043:Collagen Type II / Q000032:analysis / Q000187:drug effects*; D002373:Cyclic AMP Receptor Protein / Q000032:analysis / Q000187:drug effects*; D005260:Female; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology*; D008297:Male; D023421:Models, Animal; D011509:Proteoglycans / Q000032:analysis / Q000187:drug effects*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D012016:Reference Values; D012680:Sensitivity and Specificity
D024043:Collagen Type II; D002373:Cyclic AMP Receptor Protein; D011509:Proteoglycans; D012333:RNA, Messenger; D016178:Granulocyte-Macrophage Colony-Stimulating Factor