Estrogen directly and specifically downregulates NaPi-IIa through the activation of both estrogen receptor isoforms (ERα and ERβ) in rat kidney proximal tubule

Abstract

We have previously demonstrated that estrogen (E2) downregulates phosphate transporter NaPi-IIa and causes phosphaturia and hypophosphatemia in ovariectomized rats. In the present study, we examined whether E2 directly targets NaPi-IIa in the proximal tubule (PT) and studied the respective roles of estrogen receptor isoforms (ERα and ERβ) in the downregulation of NaPi-IIa using both in vivo and an in vitro expression systems. We found that estrogen specifically downregulates NaPi-IIa but not NaPi-IIc or Pit2 in the kidney cortex. Proximal tubules incubated in a "shake" suspension with E2 for 24 h exhibited a dose-dependent decrease in NaPi-IIa protein abundance. Results from OVX rats treated with specific agonists for either ERα [4,4',4″;-(4-propyl-[1H]-pyrazole-1,3,5-triyl) trisphenol, PPT] or ERβ [4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl) trisphenol, DPN] or both (PPT + DPN), indicated that only the latter caused a sharp downregulation of NaPi-IIa, along with significant phosphaturia and hypophosphatemia. Lastly, heterologous expression studies demonstrated that estrogen downregulated NaPi-IIa only in U20S cells expressing both ERα and ERβ, but not in cells expressing either receptor alone. In conclusion, these studies demonstrate that rat PT cells express both ERα and ERβ and that E2 induces phosphaturia by directly and specifically targeting NaPi-IIa in the PT cells. This effect is mediated via a mechanism involving coactivation of both ERα and ERβ, which likely form a functional heterodimer complex in the rat kidney proximal tubule.

Klotho/fibroblast growth factor 23- and PTH-independent estrogen receptor-α-mediated direct downregulation of NaPi-IIa by estrogen in the mouse kidney

“…We used human U20S osteosarcoma cells stably expressing estrogen receptors ER␣ and/or ER␤ under the control of doxycycline as previously described (10). U20S cells were cultured in DMEM-F-12 medium containing 10% fetal bovine serum, 1ϫ antibiotic/antimycotic, 5 mg/l blasticidin, and 500 mg/l zeocin and maintained at 37°C in a 5% CO 2-95% O2 air incubator.…”

“…In contrast, estrogen depletion is associated with increased plasma levels of P i and increased rate of proximal tubule P i reabsorption (2). Recent studies in our laboratory demonstrated that estrogen treatment of ovariectomized rats is associated with significant hypophosphatemia, as a result of increased renal P i wasting, which is secondary to the downregulation of NaPi-IIa in the kidney proximal tubule cells (10,23). This effect is independent of parathyroid hormone (PTH) (23) and is not associated with alteration in phosphorus intake (10,23).…”

“…Ligand-binding studies showed that estrogen receptors are expressed in both S1 and S2 segments of the proximal tubule (18). Recently, using pharmacological agonists of ER␣ and ER␤, we demonstrated that estrogen requires the activation of both receptors to induce the downregulation of NaPi-IIa and phosphaturia in rat (10). Moreover, while our recent in vitro studies demonstrated that estrogen directly downregulates NaPi-IIa in rats (10), others have shown that estrogen increases the circulating levels of fibroblast growth factor 23 (fgf23) in rat (11).…”

“…Recent studies in our laboratory demonstrated that estrogen treatment of ovariectomized rats is associated with significant hypophosphatemia, as a result of increased renal P i wasting, which is secondary to the downregulation of NaPi-IIa in the kidney proximal tubule cells (10,23). This effect is independent of parathyroid hormone (PTH) (23) and is not associated with alteration in phosphorus intake (10,23). Thus, it is clear that estrogen treatment is associated with hypophosphatemia and increased renal P i wasting in both humans and rats.…”

“…Recently, using pharmacological agonists of ER␣ and ER␤, we demonstrated that estrogen requires the activation of both receptors to induce the downregulation of NaPi-IIa and phosphaturia in rat (10). Moreover, while our recent in vitro studies demonstrated that estrogen directly downregulates NaPi-IIa in rats (10), others have shown that estrogen increases the circulating levels of fibroblast growth factor 23 (fgf23) in rat (11). However, whether fgf23 plays a role in estrogen-induced downregulation of NaPi-IIa and phosphaturia in animals remains elusive.…”