Vitamin D and growth hormone regulate growth hormone/insulin-like growth factor (GH-IGF) axis gene expression in human fetal epiphyseal chondrocytes.
OBJECTIVE
Cell proliferation and gene expression regulation were studied in human fetal epiphyseal chondrocytes to ascertain the involvement of GH-IGF axis components in human fetal growth regulation by 1,25-dihydroxyvitamin D(3) (VitD) and growth hormone (GH).


METHODS
Chondrocytes from primary cultures were plated in serum-free medium for 48 h and incubated for a further 48 h with VitD (10(-11) to 10(-6)M) and/or IGF-I (100 ng/ml) and/or GH (500 ng/ml). We analyzed (3)H-thymidine incorporation into DNA and IGF-I, IGFBP-3, GHR, SOX9, COL2A1, aggrecan and COMP gene expression by real-time quantitative PCR.


RESULTS
VitD dose-dependently and significantly inhibited (3)H-thymidine incorporation whereas GH had no effect on proliferation and, when combined with VitD, the same inhibition was observed as with VitD alone. IGF-I (100 ng/ml) significantly stimulated proliferation and opposed inhibition by VitD. VitD dose-dependently stimulated IGF-I (11.1+/-19.8 at VitD10(-6)M), IGFBP-3 (2.6+/-0.9), GHR (3.8+/-2.8) and COMP (1.5+/-0.6) expression whereas it inhibited SOX9 (0.7+/-0.2), COL2A1 (0.6+/-0.3) and aggrecan (0.6+/-0.2) expression and had no significant effect on IGF-II. IGF-I stimulated IGF-I, IGFBP-3, SOX9, COL2A1 and aggrecan expression and opposed COL2A1 and aggrecan gene expression inhibition by VitD. GH alone had no effect on gene expression whereas, in the presence of VitD, significantly-increased IGF-I expression stimulation was observed above values obtained with VitD alone (17.5+/-7.4).


CONCLUSIONS
Our results suggest that VitD regulation of fetal growth cartilage could have consisted of parallel enhancing of cell differentiation and conditioning to a phenotype more sensitive to regulation by other hormones such as GH as shown by increased GHR and IGF-I expression, but not by IGF-II expression which was not regulated.
Fernández-Cancio
M
M
Research Institute, Hospital Vall d'Hebron, Autonomous University of Barcelona, CIBERER (Centro de Investigación en Red de Enfermedades Raras), Instituto de Salud Carlos III, Barcelona, Spain. mfernand@ir.vhebron.net
Audi
L
L
Carrascosa
A
A
Toran
N
N
Andaluz
P
P
Esteban
C
C
Granada
M L
ML
D053674:Aggrecans / Q000235:genetics; D049109:Cell Proliferation; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004838:Epiphyses / Q000166:cytology* / Q000196:embryology; D005786:Gene Expression Regulation / Q000187:drug effects*; D019382:Human Growth Hormone / Q000235:genetics* / Q000494:pharmacology*; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000235:genetics; D007334:Insulin-Like Growth Factor I / Q000235:genetics*; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055758:SOX9 Transcription Factor / Q000235:genetics; D014807:Vitamin D / Q000031:analogs & derivatives* / Q000494:pharmacology
D053674:Aggrecans; D018972:Insulin-Like Growth Factor Binding Protein 3; D012333:RNA, Messenger; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; D019382:Human Growth Hormone; D014807:Vitamin D; C097949:1,25-dihydroxyvitamin D; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II