17 beta-estradiol and parathyroid hormone potentiate each other's stimulatory effects on alkaline phosphatase activity in SaOS-2 cells in a differentiation-dependent manner.

Rao, L.G.; Wylie, John N.; Sutherland, May Kung; Murray, Timothy M.

doi:10.1210/endo.134.2.8299560

Cited by 27 publications

(29 citation statements)

References 9 publications

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“…The stimulatory effect of intermittently added E 2 on mineralized bone nodule formation in these more differentiated cells provide stronger evidence for our previous hypothesis [21][22][23][24] that the effect of E 2 is differentiation-stage dependent. The mechanisms involved in the differential response of the cells to continuous vs. intermittent treatment with E 2 are not clear at the present time.…”

Section: )supporting

confidence: 69%

“…[21][22][23][24] We later demonstrated that the more differentiated SaOSϩDex cells had greater immunohistochemically detectable ERa. 21) In this study we have further established and characterized a third and more differentiated model of cells that mineralizes in culture, by culturing SaOSϩ Dex cells in the presence of ascorbic acid and b-glycerophosphate.…”

Section: Discussionmentioning

confidence: 99%

“…Bellows and Aubin 41) showed that each nodule formed in rat calvarial osteoblast culture is derived from a single osteoprogenitor cell that proliferated and differentiated into mature bone forming osteoblasts. Although SaOS-2 cells are a clonal cell line, we [21][22][23][24] and others 42,43) have shown that SaOS-2 cells have the potential and capacity to differentiate into various stages in culture. Another clonal cell line that was shown to differentiate and mineralize in culture was the mouse MC3T3-E1 cells.…”

Section: )mentioning

confidence: 99%

“…19,20) The other way to increase estrogen receptor is by allowing the cells to differentiate in culture as we have done. [21][22][23][24] Thus, we have shown that the immunohistochemically detectable classical estrogen receptors (ERa), were detected in greater number in the more differentiated SaOSϩDex cells grown in the presence of dexamethasone compare with the less differentiatiated SaOSϪDex cells grown in its absence. 21) Furthermore, we have shown that the effect of estrogen on alkaline phosphatase activity, in combination with other hormones, is enhanced in the more differentiated cells, [22][23][24] confirming that high level of ERa increased the response of the cells to estrogen.…”

mentioning

confidence: 99%

“…[21][22][23][24] Thus, we have shown that the immunohistochemically detectable classical estrogen receptors (ERa), were detected in greater number in the more differentiated SaOSϩDex cells grown in the presence of dexamethasone compare with the less differentiatiated SaOSϪDex cells grown in its absence. 21) Furthermore, we have shown that the effect of estrogen on alkaline phosphatase activity, in combination with other hormones, is enhanced in the more differentiated cells, [22][23][24] confirming that high level of ERa increased the response of the cells to estrogen. By culturing SaOSϩDex cells in the presence of ascorbic acid, and b-glycerophosphate in order to obtain even more differentiated osteoblastic cells that have the property of mineralizing in culture, we were able to study the effects of hormones including parathyroid hormone (PTH) 25) and estrogen 26) as well as the effect of extracellular calcium 27) on mineralized bone nodule formation.…”

mentioning

confidence: 99%

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Estrogen Added Intermittently, but Not Continuously, Stimulates Differentiation and Bone Formation in SaOS-2 Cells

Rao

Liu

Murray

et al. 2003

Biological & Pharmaceutical Bulletin

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Estrogen deficiency has been established as the major cause of postmenopausal osteoporosis, a condition resulting from a disturbance in bone remodelling. Osteoporosis is characterized by low bone mass and deterioration of the microarchitecture of bone tissue with a consequent risk of fracture and debilitation.1,2) Treatment with estrogen (E 2 ) can prevent the loss of bone among osteoporotic women, 3,4) and is the treatment of choice among postmenopausal women for the prevention of osteoporosis. 5) In spite of the accepted use of estrogen in the prevention of osteoporosis, the mechanisms involved in its action are still unclear. It is thought that the major effect of estrogen in vivo is inhibition of resorption, 6) but effects on bone formation have been reported in lower species. [7][8][9][10][11] More recently, studies 12,13) showing a sustained stimulation of osteoblast function in postmenopausal women who were exposed for prolonged period to relatively high doses of estrogen, may suggest that estrogen also has an effect on bone formation in humans. 6,14,15) Others still view the anabolic effect of estrogen as being controversial.16) If indeed estrogen has anabolic effect in vivo, its mechanism of action requires elucidation.Although a direct action of estrogen on cells of the osteoblast lineage has now been demonstrated the cellular and molecular mechanisms of its action are still not fully delineated. 17,18) Because of the notion that presence of low number of estrogen receptors in osteoblasts cause the negligible or often absence of responses to estrogen, manipulations have been carried out to increase the low abundance of estrogen receptor number in human osteoblasts, and the resultant cell models have been used to study the effects of estrogen. 19,20) The other way to increase estrogen receptor is by allowing the cells to differentiate in culture as we have done. 21-24)Thus, we have shown that the immunohistochemically detectable classical estrogen receptors (ERa), were detected in greater number in the more differentiated SaOSϩDex cells grown in the presence of dexamethasone compare with the less differentiatiated SaOSϪDex cells grown in its absence. 21)Furthermore, we have shown that the effect of estrogen on alkaline phosphatase activity, in combination with other hormones, is enhanced in the more differentiated cells, [22][23][24] confirming that high level of ERa increased the response of the cells to estrogen. By culturing SaOSϩDex cells in the presence of ascorbic acid, and b-glycerophosphate in order to obtain even more differentiated osteoblastic cells that have the property of mineralizing in culture, we were able to study the effects of hormones including parathyroid hormone (PTH) 25) and estrogen 26) as well as the effect of extracellular calcium 27) on mineralized bone nodule formation. Thus, as we have recently reviewed 17) and reported in a rapid communication, 28) we found that estrogen has a direct effect on bone formation in SaOS-2 cells when added intermittently. In this study, we repo...

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Section: )supporting

confidence: 69%

Section: Discussionmentioning

confidence: 99%

Section: )mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Estrogen Added Intermittently, but Not Continuously, Stimulates Differentiation and Bone Formation in SaOS-2 Cells

Rao

Liu

Murray

et al. 2003

Biological & Pharmaceutical Bulletin

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Functional properties of a conditionally phenotypic, estrogen-responsive, human osteoblast cell line

Bodine¹,

Green

Harris³

et al. 1997

J. Cell. Biochem.

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Osteoblasts are established targets of estrogen action in bone. We screened 66 conditionally immortalized clonal human osteoblast cell lines for estrogen receptors (ERs) using reverse transcriptase-polymerase chain reaction (RT-PCR) analysis for ER alpha mRNA and transactivation of adenovirus-estrogen response element (ERE)-tk-luciferase by 17 beta-estradiol (17 beta-E2) for functional ER protein. One of these cell lines, termed HOB-03-CE6, was chosen for further characterization. The cells, which were conditionally immortalized with a temperature-sensitive SV40 large T antigen, proliferated at the permissive temperature (34 degrees C) but stopped dividing at the nonpermissive temperature (> or = 39 degrees C). Alkaline phosphatase activity and osteocalcin secretion were upregulated by 1 alpha, 25-dihydroxyvitamin D3 in a dose-dependent manner. The cells also expressed type I collagen and other bone matrix proteins, secreted a variety of growth factors and cytokines, formed mineralized nodules based on alizarin red-S and von Kossa histochemical staining, and responded to dexamethasone, all-trans retinoic acid, and transforming growth factor-beta 1. This cell line expressed 42-fold less ER message than MCF-7 human breast cancer cells, as determined by quantitative RT-PCR. However, adenovirus-ERE-tk-luciferase activity was upregulated three- to fivefold in these cells by 17 beta-E2 with an EC50 of 64 pM. Furthermore, this upregulation was suppressed by co-treatment with the anti-estrogen ICI-182, 780. Cytosolic extracts of these cells specifically bound [125I]-17 beta-E2 in a concentration-dependent manner with a Bmax of 2.7 fmoles/mg protein (approximately 1,200 ERs/cell) and a Kd of 0.2 nM. DNA gel-shift analysis using a [32P]-ERE demonstrated the presence of ERs in nuclear extracts of these cells. Moreover, binding of the extracts to this ERE was blocked by a monoclonal antibody to the human ER DNA-binding domain. We evaluated these cells for 14 of 20 reported endogenous responses to 17 beta-E2 in osteoblasts. Although most of these responses appeared to be unaffected by the steroid, 17 beta-E2 suppressed parathyroid hormone-induced cAMP production, as well as basal interleukin-6 mRNA expression; conversely, the steroid upregulated the steady-state expression of alkaline phosphatase message in these cells. In summary, we have identified a clonal, conditionally phenotypic, human osteoblast cell line that expresses functional ERs and exhibits endogenous responses to 17 beta-E2. This cell line will be a valuable in vitro model for exploring some of the molecular mechanisms of estrogen action in bone.

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Regulation of the collagenase-3 receptor and its role in intracellular ligand processing in rat osteoblastic cells

et al. 1998

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We have previously described a specific, saturable receptor for rat collagenase-3 in the rat osteosarcoma cell line, UMR 106-01. Binding of rat collagenase-3 to this receptor is coupled to the internalization and eventual degradation of the enzyme and correlates with observed extracellular levels of the enzyme. In this study we have shown that decreased binding, internalization, and degradation of 125I-rat collagenase-3 were observed in cells after 24 h of parathyroid hormone treatment; these activities returned to control values after 48 h and were increased substantially (twice control levels) after 96 h of treatment with the hormone. Subcellular fractionation studies to identify the route of uptake and degradation of collagenase-3 localized intracellular accumulation of 125I-rat collagenase-3 initially in Golgi-associated lysosomes and later in secondary lysosomes. Maximal lysosomal accumulation of the radiolabel and stimulation of general lysosomal activity occurred after 72 h of parathyroid hormone treatment. Preventing fusion of endosomes with lysosomes (by temperature shift, colchicine, or monensin) resulted in no internalized 125I-collagenase-3 in either lysosomal fraction. Treatment of UMR cells with the above agents or ammonium chloride decreased excretion of 125I-labeled degradation products of collagenase-3. These experiments demonstrated that degradation of collagenase-3 required receptor-mediated endocytosis and sequential processing by endosomes and lysosomes. Thus, parathyroid hormone regulates the expression and synthesis of collagenase-3 as well as the abundance and functioning of the collagenase-3 receptor and the intracellular degradation of its ligand. The coordinate changes in the secretion of collagenase-3 and expression of the receptor determine the net abundance of the enzyme in the extracellular space.

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17 beta-estradiol and parathyroid hormone potentiate each other's stimulatory effects on alkaline phosphatase activity in SaOS-2 cells in a differentiation-dependent manner.

Cited by 27 publications

References 9 publications

Estrogen Added Intermittently, but Not Continuously, Stimulates Differentiation and Bone Formation in SaOS-2 Cells

Estrogen Added Intermittently, but Not Continuously, Stimulates Differentiation and Bone Formation in SaOS-2 Cells

Functional properties of a conditionally phenotypic, estrogen-responsive, human osteoblast cell line

Regulation of the collagenase-3 receptor and its role in intracellular ligand processing in rat osteoblastic cells

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