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Estrogen inhibits bone resorption by directly inducing apoptosis of the bone-resorbing osteoclasts.

Kameda T, Mano H, Yuasa T, Mori Y, Miyazawa K, Shiokawa M, Nakamaru Y, Hiroi E, Hiura K, Kameda A, Yang NN, Hakeda Y, Kumegawa M - J. Exp. Med. (1997)

Bottom Line: Using a recently developed technique for isolation of highly purified mammalian osteoclasts, we showed that 17 beta-estradiol (E2) was able to directly inhibit osteoclastic bone resorption.At concentrations effective for inhibiting bone resorption, E2 also directly induced osteoclast apoptosis in a dose- and time-dependent manner.ICI164,384 and tamoxifen, as pure and partial antagonists, respectively, completely or partially blocked the effect of E2 on both inhibition of osteoclastic bone resorption and induction of osteoclast apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthodontics, Nippon Dental University School of Dentistry at Niigata, Niigata 951, Japan.

ABSTRACT
Estrogen deficiency causes bone loss, which can be prevented by estrogen replacement therapy. Using a recently developed technique for isolation of highly purified mammalian osteoclasts, we showed that 17 beta-estradiol (E2) was able to directly inhibit osteoclastic bone resorption. At concentrations effective for inhibiting bone resorption, E2 also directly induced osteoclast apoptosis in a dose- and time-dependent manner. ICI164,384 and tamoxifen, as pure and partial antagonists, respectively, completely or partially blocked the effect of E2 on both inhibition of osteoclastic bone resorption and induction of osteoclast apoptosis. These data suggest that the protective effects of estrogen against postmenopausal osteoporosis are mediated in part by the direct induction of apoptosis of the bone-resorbing osteoclasts by an estrogen receptor- mediated mechanism.

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Dose- and time-dependent manner of estrogen-induced osteoclast apoptosis. (A) Purified osteoclasts were cultured on dentine slices  (150 cells/slice) for 24 h in medium (Con) or in medium containing  0.001–1 nM E2, 1–20 ng/ml TGF-β1, or 1 nM CT. Apoptotic osteoclasts  were quantified under a fluorescence microscope. (B) Time-dependent  effects of E2 on osteoclast apoptosis and osteoclast number. Under the  same culture conditions, purified osteoclasts were incubated in medium  without 0.1 nM E2 (apoptosis: ○; cell number: □) or with 0.1 nM E2  (apoptosis: •, cell number: ▪) for 6, 12, or 24 h. Apoptotic osteoclasts  are expressed as a percentage of total number of adherent osteoclasts. Values are means ± SD, n = 4. *P <0.05, **P <0.005 compared with time = 0  groups. Data are representative of those of three additional independent  experiments.
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Figure 5: Dose- and time-dependent manner of estrogen-induced osteoclast apoptosis. (A) Purified osteoclasts were cultured on dentine slices (150 cells/slice) for 24 h in medium (Con) or in medium containing 0.001–1 nM E2, 1–20 ng/ml TGF-β1, or 1 nM CT. Apoptotic osteoclasts were quantified under a fluorescence microscope. (B) Time-dependent effects of E2 on osteoclast apoptosis and osteoclast number. Under the same culture conditions, purified osteoclasts were incubated in medium without 0.1 nM E2 (apoptosis: ○; cell number: □) or with 0.1 nM E2 (apoptosis: •, cell number: ▪) for 6, 12, or 24 h. Apoptotic osteoclasts are expressed as a percentage of total number of adherent osteoclasts. Values are means ± SD, n = 4. *P <0.05, **P <0.005 compared with time = 0 groups. Data are representative of those of three additional independent experiments.

Mentions: Quantification of E2-induced osteoclast apoptosis showed a dose-dependent increase (Fig. 5 A), which correlated with the dose range of E2 for inhibition of bone resorption (Fig. 2 B). In contrast to estrogen, calcitonin, which inhibits osteoclast activity directly through its receptors on osteoclasts, did not cause osteoclasts to undergo apoptosis, suggesting that inhibition of osteoclastic bone resorption by E2 is mediated by a distinctively different mechanism than that used by calcitonin. As for detached cells, significant cellular disintegration prevented us from detecting osteoclast apoptosis in them in 24-h and older cultures. E2-induced osteoclast apoptosis was also time dependent (Fig. 5 B). E2 (0.1 nM) inhibition of formation of resorption pits by osteoclasts was correlated with a reduction in the expression of mRNAs for Ca K and CA II and with an increased induction of osteoclast apoptosis in a time course study. These findings suggest that E2 directly acts on osteoclasts and inhibits osteoclastic bone resorption by causing osteoclast inactivation partially due to apoptosis. A recent report indicated that estrogen promoted TGF-β–mediated apoptosis of in vitro–developed murine osteoclast-like cells in mixed cell cultures (21). TGF-β1 (10 ng/ml) induces apoptosis of our pure authentic osteoclasts. However, the incidence was lower than that by E2 treatment in our mature cells or that of TGF-β–induced apoptosis in murine mixed cell cultures (Fig. 5 A, reference 21). The causes of this discrepancy might be partially due to their different species; however, these results suggest that estrogen-enhanced osteoclast apoptosis mediated by TGF-β might occur in an indirect manner. The direct induction of osteoclast apoptosis by estrogen besides indirect induction might exist.


Estrogen inhibits bone resorption by directly inducing apoptosis of the bone-resorbing osteoclasts.

Kameda T, Mano H, Yuasa T, Mori Y, Miyazawa K, Shiokawa M, Nakamaru Y, Hiroi E, Hiura K, Kameda A, Yang NN, Hakeda Y, Kumegawa M - J. Exp. Med. (1997)

Dose- and time-dependent manner of estrogen-induced osteoclast apoptosis. (A) Purified osteoclasts were cultured on dentine slices  (150 cells/slice) for 24 h in medium (Con) or in medium containing  0.001–1 nM E2, 1–20 ng/ml TGF-β1, or 1 nM CT. Apoptotic osteoclasts  were quantified under a fluorescence microscope. (B) Time-dependent  effects of E2 on osteoclast apoptosis and osteoclast number. Under the  same culture conditions, purified osteoclasts were incubated in medium  without 0.1 nM E2 (apoptosis: ○; cell number: □) or with 0.1 nM E2  (apoptosis: •, cell number: ▪) for 6, 12, or 24 h. Apoptotic osteoclasts  are expressed as a percentage of total number of adherent osteoclasts. Values are means ± SD, n = 4. *P <0.05, **P <0.005 compared with time = 0  groups. Data are representative of those of three additional independent  experiments.
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Related In: Results  -  Collection

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Figure 5: Dose- and time-dependent manner of estrogen-induced osteoclast apoptosis. (A) Purified osteoclasts were cultured on dentine slices (150 cells/slice) for 24 h in medium (Con) or in medium containing 0.001–1 nM E2, 1–20 ng/ml TGF-β1, or 1 nM CT. Apoptotic osteoclasts were quantified under a fluorescence microscope. (B) Time-dependent effects of E2 on osteoclast apoptosis and osteoclast number. Under the same culture conditions, purified osteoclasts were incubated in medium without 0.1 nM E2 (apoptosis: ○; cell number: □) or with 0.1 nM E2 (apoptosis: •, cell number: ▪) for 6, 12, or 24 h. Apoptotic osteoclasts are expressed as a percentage of total number of adherent osteoclasts. Values are means ± SD, n = 4. *P <0.05, **P <0.005 compared with time = 0 groups. Data are representative of those of three additional independent experiments.
Mentions: Quantification of E2-induced osteoclast apoptosis showed a dose-dependent increase (Fig. 5 A), which correlated with the dose range of E2 for inhibition of bone resorption (Fig. 2 B). In contrast to estrogen, calcitonin, which inhibits osteoclast activity directly through its receptors on osteoclasts, did not cause osteoclasts to undergo apoptosis, suggesting that inhibition of osteoclastic bone resorption by E2 is mediated by a distinctively different mechanism than that used by calcitonin. As for detached cells, significant cellular disintegration prevented us from detecting osteoclast apoptosis in them in 24-h and older cultures. E2-induced osteoclast apoptosis was also time dependent (Fig. 5 B). E2 (0.1 nM) inhibition of formation of resorption pits by osteoclasts was correlated with a reduction in the expression of mRNAs for Ca K and CA II and with an increased induction of osteoclast apoptosis in a time course study. These findings suggest that E2 directly acts on osteoclasts and inhibits osteoclastic bone resorption by causing osteoclast inactivation partially due to apoptosis. A recent report indicated that estrogen promoted TGF-β–mediated apoptosis of in vitro–developed murine osteoclast-like cells in mixed cell cultures (21). TGF-β1 (10 ng/ml) induces apoptosis of our pure authentic osteoclasts. However, the incidence was lower than that by E2 treatment in our mature cells or that of TGF-β–induced apoptosis in murine mixed cell cultures (Fig. 5 A, reference 21). The causes of this discrepancy might be partially due to their different species; however, these results suggest that estrogen-enhanced osteoclast apoptosis mediated by TGF-β might occur in an indirect manner. The direct induction of osteoclast apoptosis by estrogen besides indirect induction might exist.

Bottom Line: Using a recently developed technique for isolation of highly purified mammalian osteoclasts, we showed that 17 beta-estradiol (E2) was able to directly inhibit osteoclastic bone resorption.At concentrations effective for inhibiting bone resorption, E2 also directly induced osteoclast apoptosis in a dose- and time-dependent manner.ICI164,384 and tamoxifen, as pure and partial antagonists, respectively, completely or partially blocked the effect of E2 on both inhibition of osteoclastic bone resorption and induction of osteoclast apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthodontics, Nippon Dental University School of Dentistry at Niigata, Niigata 951, Japan.

ABSTRACT
Estrogen deficiency causes bone loss, which can be prevented by estrogen replacement therapy. Using a recently developed technique for isolation of highly purified mammalian osteoclasts, we showed that 17 beta-estradiol (E2) was able to directly inhibit osteoclastic bone resorption. At concentrations effective for inhibiting bone resorption, E2 also directly induced osteoclast apoptosis in a dose- and time-dependent manner. ICI164,384 and tamoxifen, as pure and partial antagonists, respectively, completely or partially blocked the effect of E2 on both inhibition of osteoclastic bone resorption and induction of osteoclast apoptosis. These data suggest that the protective effects of estrogen against postmenopausal osteoporosis are mediated in part by the direct induction of apoptosis of the bone-resorbing osteoclasts by an estrogen receptor- mediated mechanism.

Show MeSH
Related in: MedlinePlus