The effects of estrogen receptors α‐ and β‐specific agonists and antagonists on cell proliferation and energy metabolism in human bone cell line

Abstract

In cultured human osteoblasts estradiol-17β (E2) modulated DNA synthesis, the specific activity of creatine kinase BB (CK), 12 and 15 lipoxygenase (LO) mRNA expression and formation of 12- and 15-hydroxyeicosatetraenoic acid (HETE). We now investigate the response of human bone cell line (SaOS2) to phytoestrogens and estrogen receptors (ER)-specific agonists and antagonists. Treatment of SaSO2 with E2, 2,3-bis (4-hydroxyphenyl)-propionitrile (DPN; ERβ-specific agonist), 4,4',4″-[4-propyl-(1H)-pyrazol-1,3,5-triyl] tris-phenol (PPT; ERα-specific agonist), biochainin A (BA), daidzein (D), genistein (G) and raloxifene (Ral) showed increased DNA synthesis and CK. Ral inhibited completely all stimulations except DPN and to some extent D. The ERα-specific antagonist methyl-piperidino-pyrazole (MPP) and the ERβ-specific antagonist 4-[2-phenyl-5,7-bis (tri-fluoro-methyl) pyrazolo [1,5-a]pyrimidin-3-yl] phenol (PTHPP) inhibited DNA synthesis, CK and reactive oxygen species (ROS) formation induced by estrogens according to their receptors affinity. The LO inhibitor baicaleine inhibited only E2, DPN and G's effects. E2 and Ral unlike all other compounds had no effect on ERα mRNA expression, while ERβ mRNA expression was stimulated by all compounds. All compounds modulated the expression of 12LO and 15LO mRNA, except E2, PPT and Ral for 12LO, and 12- and 15-HETE productions and stimulated ROS formation which was inhibited by NADPH oxidase inhibitors diphenyleneiodonium chloride (DPI) and N-acetyl cysteine and the estrogen inhibitor ICI. DPI did not affect hormonal-induced DNA and CK. In conclusion, we provide evidence for the separation of mediation via ERα and ERβ pathways in the effects of estrogenic compounds on osteoblasts, but the role of LO/HETE/ROS is unclear.

Premature T cell senescence in Ovx mice is inhibited by repletion of estrogen and medicarpin: a possible mechanism for alleviating bone loss

“…5b). Studies by Somjen et al [42] have shown that E2 and their agonists stimulate ROS production in human osteosarcoma cell line SaOS-2 and this effect is blocked in presence of antiestrogen. Thus, we studied if pretreatment with ICI 182,780 blunts the ability of E2 and Med to inhibit (5) Ovx results in the production of more senescent T cell population that produce more TNF-α thus leading to decreased CD28 expression on CD4 + T cells and enhanced osteoclastogenesis TNF-α-induced ROS generation.…”

Sex specific response of cultured human bone cells to ERα and ERβ specific agonists by modulation of cell proliferation and creatine kinase specific activity

“…We found that vitamin D, PTH as well as other hormones including phytoestrogens stimulate DNA and CK as well as LO mRNA and HETE formation [10,11]. We also found that the ER␣ and ER␤ specific agonists have differential effects on all these parameters in the human osteoblastic cell line SaSO 2 [12]. In the present study we analyzed the response of primary human osteoblasts in culture from males or females to the ER␣ agonist PPT and the ER␤ agonist DPN at different parameters, in order to see whether the sex-specificity is retained also using the specific ERs agonist similar to E2 itself.…”

“…(a) for 3 days daily for the expression of mRNA for 12LO and 15LO type 1 expression or for estrogen receptor ␣ (ER␣) and estrogen receptor ␤ (ER␤) were determined as previously described [18,19] or; (b) for 1 h with serum-free medium, followed by the addition of vehicle or estrogenic compounds: E2 at 30 nM, DPN at 420 nM and PPT at 390 nM for 10 min and HETE were extracted and assayed as previously described [11,12,20] or; (c) for 24 h with the addition of vehicle or estrogenic compounds: E2 at 30 nM, DPN at 420 nM and PPT at 390 nM for DNA synthesis and for CK specific activity as previously described [12,20]. In these experiments raloxifene (Ral) was used at 3000 nM.…”

“…RNA was extracted and expression of 12 and 15LO enzymes was carried out by RT-PCR as described previously [11,12,20]. …”

“…We now attempt to determine whether or not the estrogens receptor specific multiple ligands affect primary cultures of osteoblast-like cells (hObs) derived from pre-or post-menopausal cells compared to male-bone derived cells and to see whether the effects are similar to what was found using SaSO 2 cells [12].…”