Reduced expression of glucocorticoid receptor levels in human osteoarthritic chondrocytes. Role in the suppression of metalloprotease synthesis.
Glucocorticoid occupancy of a large percentage of glucocorticoid receptor (GR) is necessary for the suppression of matrix metalloprotease synthesis by human articular chondrocytes. In this study, we evaluated the levels of GR binding, cellular GR protein, and messenger RNA expression in both normal and osteoarthritic (OA) human articular chondrocytes and compared the degree of suppression of collagenase synthesis by glucocorticoids in cultures of the two cell types in order to investigate whether or not the GR system played an important role in the pathophysiology of OA. By radioreceptor binding assay, we recorded 56,320 +/- 8,230 sites per cell (mean +/- SE, n = 9) in primary cultures of normal chondrocytes and 27,480 +/- 14,240 sites in OA cells (n = 10, P < 0.0001). Equilibrium dissociation constant (Kd) values did not vary between normal (12.4 +/- 1.4 nmol/L) and OA (13.0 +/- 1.8 nmol/L). Subculturing of primary OA chondrocytes resulted in the up-regulation of the number of GR binding sites per cell to values comparable to those obtained in normal chondrocytes. Analysis of protein-immuno dot-blots of cytosolic extracts from normal (n = 4) and OA chondrocytes (n = 4) revealed that the former cytosols contained a 1.9 +/- 0.2 (P < 0.05) higher relative density of GR protein than the latter. By comparing the optical densities of GR-polymerase chain reaction products generated from normal (n = 6) and OA (n = 9) chondrocyte total RNA (normalized using an internal standard, glyceraldehyde phosphate dehydrogenase), we established a relative ratio, normal/OA, of 1.4. Experiments comparing the biological responsiveness of normal and OA chondrocytes to glucocorticoid suppression of interleukin-1-stimulated metalloprotease synthesis showed that dexamethasone inhibited collagenase synthesis in a dose-dependent manner with an IC50 of 6.3 +/- 1.2 x 10(-10) mol/L (n = 5) in normal cells while an IC50 of 5.0 +/- 0.4 x 10(-9) mol/L (P < 0.05) was recorded using OA (n = 5) chondrocytes. The results suggest that OA chondrocytes express fewer GR than normal cells as a result of a decrease in specific gene expression. The decreased responsiveness of OA cells to circulating glucocorticoids may be among the factors responsible for an increased level of metalloprotease synthesis by chondrocytes in OA cartilage.
DiBattista
J A
JA
Rheumatic Disease Unit, Notre-Dame Hospital, Montréal, Québec, Canada.
Martel-Pelletier
J
J
Antakly
T
T
Tardif
G
G
Cloutier
J M
JM
Pelletier
J P
JP
D001483:Base Sequence; D015153:Blotting, Western; D002358:Cartilage, Articular / Q000378:metabolism*; D017364:Collagenases / Q000378:metabolism; D016207:Cytokines / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D005260:Female; D006801:Humans; D008297:Male; D008666:Metalloendopeptidases / Q000096:biosynthesis*; D008875:Middle Aged; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000235:genetics; D010003:Osteoarthritis / Q000378:metabolism* / Q000473:pathology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism*
D016207:Cytokines; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; D017364:Collagenases; D008666:Metalloendopeptidases