Long-term culture in dexamethasone unmasks an abnormal phenotype in osteoblasts isolated from osteoporotic subjects.
We have shown that osteoblastic cells derived from trabecular bone explants of osteoporotic subjects (OP cells) exhibited an altered alkaline phosphatase (ALP) response to 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] compared to control (CON) cells. Our hypothesis that OP cells have other intrinsic abnormalities was investigated using our cell models representing two different stages of differentiation. OP and CON cells were cultured in the absence (-DEX) or presence (+DEX) of 10 nM dexamethasone (DEX) in 10% fetal calf serum (FCS) prior to exposure to serum-free medium containing 1 nM of PTH and/or 17-beta estradiol (E2). Both OP and CON cells responded to DEX with a two-fold increase in basal ALP activity. While E2 or PTH+E2 had no effect on OP cells, both treatments inhibited ALP activity in CON cells (p<0.05). OP and CON cells grown in DEX also expressed PTH-stimulated adenylate cyclase (AC) activities higher than those of (-DEX) cells. OP+DEX cells, however, exhibited activities which were 8-fold higher than those of CON+DEX cells (p<0.001). In OP+DEX cells, E2 stimulated basal AC activity (p<0.05) but did not affect PTH-stimulated activity. In contrast, in CON+DEX cells, E2 had no effect on basal activity but inhibited PTH-stimulated AC activity (p<0.001). Osteocalcin production was 4-fold lower in OP+DEX cells compared to OP-DEX and CON cells (p<0.05) while osteocalcin mRNA levels were significantly lower in OP+DEX and CON+/-DEX cells compared to OP-DEX cells (p<0.05). E2 did not affect osteocalcin protein or mRNA levels in either OP or CON cells. No differences in mRNA levels were found for estrogen receptor-alpha (ER-a) in OP+/-DEX cells whereas these levels were significantly higher in CON+DEX compared to CON-DEX cells (p<0.05). These results indicate that DEX amplified the differences between OP and CON cells and confirm the presence of intrinsic osteoblastic abnormalities in patients with osteoporosis that persist in culture.
Rao
L G
LG
Calcium Research Laboratory, suite 2022, St. Michael's Hospital, 38 Shuter Street, Toronto, Ontario, Canada M5B 1A6. leticia.rao@utoronto.ca
Murray
T M
TM
Wylie
J N
JN
McBroom
R J
RJ
Sutherland
M Kung
MK
D000262:Adenylyl Cyclases / Q000378:metabolism; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000469:Alkaline Phosphatase / Q000378:metabolism; D002117:Calcitriol / Q000494:pharmacology; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology*; D004958:Estradiol / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D010006:Osteoblasts / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology*; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D010024:Osteoporosis / Q000201:enzymology / Q000235:genetics / Q000473:pathology*; D010281:Parathyroid Hormone / Q000494:pharmacology; D010641:Phenotype; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D044168:Receptor, Parathyroid Hormone, Type 1 / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D024042:Collagen Type I; D047628:Estrogen Receptor alpha; D010281:Parathyroid Hormone; D012333:RNA, Messenger; D044168:Receptor, Parathyroid Hormone, Type 1; D015675:Osteocalcin; D004958:Estradiol; D003907:Dexamethasone; D000469:Alkaline Phosphatase; D000262:Adenylyl Cyclases; D002117:Calcitriol