The Journal of Clinical Endocrinology & Metabolism Vol. 85, No. 3 1215-1219
Copyright © 2000 by The Endocrine Society
Close Correlation between Estrogen Treatment and Renal Phosphate Reabsorption Capacity
Hirokazu Uemura,
Minoru Irahara,
Naoto Yoneda,
Toshiyuki Yasui,
Kaori Genjida,
Ken-ichi Miyamoto,
Toshihiro Aono and
Eiji Takeda
Departments of Obstetrics and Gynecology (H.U., M.I., N.Y., T.Y.,
T.A.) and Clinical Nutrition (K.G., K.-i.M., E.T.), The University of
Tokushima, School of Medicine, Tokushima, 770-8503 Japan
Address correspondence and requests for reprints to: Hirokazu Uemura, M.D., Department of Obstetrics and Gynecology, The University of Tokushima, School of Medicine, 3-18-15. Kuramoto-cho, Tokushima, 770-8503 Japan. E-mail: uemura{at}clin.med
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Abstract
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To determine the influence of estrogen on the activity of renal
proximal tubular reabsorption of inorganic phosphate (Pi) in women, we
examined the changes of the renal threshold phosphate concentration
(also denoted as TmP/GFR), as well as the changes in the concentrations
of mineral components in the circulation in two groups of women—one
receiving hormone replacement therapy (HRT) and one receiving
gonadotropin-releasing hormone agonists (GnRH-a) therapy. We also
examined the changes in the concentrations of serum PTH in the
GnRH-a group. The patients in the HRT group were continuously treated
with 0.625 mg conjugated equine estrogens plus 2.5 mg
medroxyprogesterone acetate per day. The patients in the GnRH-a group
were treated with a monthly injection of 3.75 mg leuprolide acetate
depot for 6 months. The values of TmP/GFR decreased in all of the
patients who received HRT. The mean percentage change in TmP/GFR was
-14.5% (range, -24.3% to -9.6%). In contrast, in all of the
patients treated with GnRH-a, the values of TmP/GFR increased after 6
months of treatment (the mean percentage change was 28.5%; range,
18.2–78.3%) and returned to the preadministration level at 12 weeks
after stopping therapy. In these patients, both the values of TmP/GFR
and the concentrations of serum Pi correlated significantly with
circulating estradiol levels (r = -0.767, P
< 0.01 and r = -0.797, P < 0.01,
respectively), but the concentrations of serum corrected calcium did
not correlate. Moreover, in the same patients, the levels of serum
intact PTH decreased significantly (P < 0.05)
after 6 months of treatment, but at 12 weeks after stopping therapy the
trends of these levels varied among individual patients. These results
suggest that estrogen could act directly to suppress sodium-dependent
Pi reabsorption in the renal proximal tubules.
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Introduction
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IT IS WELL KNOWN that the decrease in
circulating estradiol (E2) in women at menopause
is a major trigger of the changes in bone metabolism and causes the
loss of bone density (1, 2). Administration of estrogen to
postmenopausal women produces many health benefits, such as reduction
of climacteric symptoms and prevention of arteriosclerosis (3, 4) and
osteoporosis (5, 6). On the other hand, gonadotropin-releasing hormone
agonists (GnRH-a) have been recognized to be effective against
endometriosis by inducing the decrease in circulating
E2, leading to atrophy of both the endometrium
and the endometriotic tissues. However, such long-term estrogen
deficiency is likely to have a negative influence on bone density.
Bone turnover is high at menopause. Estrogen reduces bone turnover and
especially reduces bone resorption (7). By this action, the levels of
circulating calcium (Ca) and inorganic phosphate (Pi) decrease in women
who receive estrogen replacement (8, 9, 10). Renal proximal tubular
reabsorption of Pi is a major determinant of the circulating level of
Pi and contributes to the maintenance of Pi homeostasis. Pi
reabsorption at the apical membrane is carried out by sodium-dependent
Pi (Na/Pi) cotransporters, which are classified into types I and II by
molecular characterization (11). Several hormonal and nonhormonal
factors that control serum Pi homeostasis affect the rate of Pi
reabsorption by changing the activities of type II Na/Pi cotransporter
(12, 13).
However, little is known about the effects of estrogen on renal
phosphate reabsorption in women. To clarify this issue, we studied the
effects of hormone replacement therapy (HRT) in climacteric women and
of GnRH-a therapy in women with endometriosis on the renal phosphate
threshold concentration (TmP/GFR) by modulating circulating
E2 levels.
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Experimental Subjects
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Patients and clinical treatment
HRT treatment.Five Japanese women who had complained of
various climacteric symptoms and/or had undergone surgical castration,
aged 46–56 yr (mean ± SD, 52.8 ± 3.7), with
body mass indices ranging from 20.6–24.6 kg/m2 (mean
± SD, 22.9 ± 1.4) were studied after providing
informed consent. They were treated with 0.625 mg conjugated equine
estrogens (CEE) (Premarin; Wyeth-Ayerst Laboratories, Inc., Radnor, PA) plus 2.5 mg medroxyprogesterone
acetate (Provera; Upjohn Co., Kalamazoo, MI) per day. All patients had
no history of estrogen-dependent cancer, hypercortisolism,
hyperthyroidism, metabolic bone disease, or renal failure. All patients
had both a negative mammogram and negative Papanicolaou smear within 6
months prior to the study. None of them had previously received
estrogen replacement therapy or any drug treatment that might have
affected Ca or Pi metabolism.
GnRH-a treatment.Five Japanese women with endometriosis, aged
30–49 yr and with body mass indices of 17.0–28.8 kg/m2
(mean ± SD, 22.9 ± 5.2), were studied after
providing informed consent. They were treated with a monthly injection
of 3.75 mg leuprolide acetate (Leuprin; Takeda Pharmaceutical Co.,
Tokyo, Japan) depot for 6 months. All patients were in good health
except for their endometriosis. None of them had received any other
hormonal treatment for their endometriosis.
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Materials and Methods
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Mineral and hormone determination
Fasted blood and urine samples were collected from the patients
during HRT at the beginning and at the 6th month of treatment. They
were also collected from the patients treated with GnRH-a at the
beginning and end (i.e. at the 6th month) of treatment and
at 12 weeks after stopping treatment. Serum was immediately separated
after blood collection and promptly frozen at -40 C until the assay.
Urine was also frozen and kept at -40 C until it was assayed.
Serum E2 was measured by a combination of
high-performance liquid chromatography after extraction and RIA.
The high-performance liquid chromatography was performed using a
CAPCELL PAK NH2 column (4.6 250, 5 um; Shiseido Co., Tokyo, Japan). RIA
was performed using a DSL Kit (Diagnostics Systems Laboratories, Inc., Webster, TX). The assay range was 1.0–50 pg/mL.
Intra- and interassay coefficients of variation were 8.4–13.8% and
15.5%, respectively.
Serum intact PTH was measured by two-site immunoradiometric assay
(Nichols, San Juan Capistrano, CA), with a normal range of 10–65 pg/mL
and assay sensitivity of 1 pmol/L.
Pi concentration in serum and urine was analyzed by the method of
Taussky and Shorr (14). Creatinine concentration in serum and urine was
measured using a Creatinine-TEST Wako (Wako Pure Chemical Industries, Ltd., Osaka, Japan).
Determination of TmP/GFR
The activity of renal tubular reabsorption of Pi is best
expressed as the renal threshold phosphate concentration (also denoted
as TmP/GFR, or the ratio of maximum rate of renal tubular reabsorption
of phosphate to GFR), which is independent of GFR and of net inflow of
phosphate. TmP/GFR was calculated using a standard nomogram (15) and
was expressed in millimoles per liter.
Statistical analysis
Data are expressed as the mean ± SD unless
otherwise stated. Statistical significances were determined by
t test and linear regression analysis. All P
values of 0.05 or less were considered statistically significant.
Analyses were carried out using a Stat Works program (Cricket Software,
Inc., Philadelphia, PA).
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Results
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Changes in clinical values for serum and urine
The changes in clinical chemical and hormonal values for serum and
urine in patients during HRT are shown in Table 1
. In all the patients, serum
E2 levels increased after 6 months of treatment,
whereas the concentrations of both serum corrected Ca and serum Pi
decreased. The changes in clinical chemical and hormonal values for
serum and urine in patients who received GnRH-a therapy are shown in
Table 2. In all patients treated with
GnRH-a, the concentrations of serum E2 decreased
after 6 months of treatment and returned to the preadministration level
at 12 weeks after stopping therapy, and the concentrations of both
serum corrected Ca and serum Pi changed inversely to those of serum
E2.
Changes in TmP/GFR
The changes in the values of TmP/GFR in patients during HRT are
shown in Fig. 1. In all of the patients,
the values of TmP/GFR decreased after 6 months of treatment. The mean
percentage changes in TmP/GFR was -14.5% (range, -24.3% to
-9.6%). In contrast, in all of the patients treated with GnRH-a, the
values of TmP/GFR increased after 6 months of treatment (the mean
percentage change was 28.5%; range, 18.2–78.3%) and returned to the
preadministration level at 12 weeks after stopping therapy (Fig. 2).
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Figure 1. Changes in the values of TmP/GFR in the
patients during HRT. In all patients, the values of TmP/GFR decreased
after 6 months of treatment. The mean percentage change in TmP/GFR was
-14.5% (range, -24.3% to -9.6%).
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Figure 2. Changes in the values of TmP/GFR in patients
receiving GnRH-a therapy. In all patients, the values of TmP/GFR
increased after 6 months of treatment when therapy was completed (mean
percentage change, 28.5%; range, 18.2–78.3%) and returned to the
preadministration levels at 12 weeks after stopping therapy.
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Changes in levels of serum intact PTH
The concentrations of serum intact PTH in the patients who
received GnRH-a therapy decreased significantly (P <
0.05) after 6 months of GnRH-a treatment, but at 12 weeks after
stopping therapy the trends of these levels varied among individual
patients (Fig. 3).
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Figure 3. Changes in the values of serum intact PTH in
patients receiving GnRH-a therapy. The values of serum immunoreactive
PTH decreased significantly (P < 0.05) after 6
months of treatment when therapy was completed, but at 12 weeks after
stopping therapy the trends of these levels varied among individual
patients.
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Relationship between circulating E2 and chemical
values
Because administration of CEE leads mainly to a rise in
circulating estrone levels, we analyzed the correlations between
circulating E2 levels and other indices in the
patients treated with GnRH-a. As shown in Fig. 4, the values of TmP/GFR correlated
significantly with circulating E2 levels (r
= -0.767, P < 0.01). In these patients, the
concentrations of serum Pi also correlated with circulating
E2 levels (r = -0.797, P <
0.01), but those of serum corrected Ca did not correlate with
circulating E2 levels.
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Figure 4. Correlations between the concentrations of
circulating E2 and the following indices in five patients
treated with GnRH-a: TmP/GFR (A), concentration of serum Pi (B), and
concentration of serum corrected Ca (C). The values of TmP/GFR
correlated significantly with circulating E2 levels (r
= -0.767, P < 0.01). The concentrations of serum
Pi also correlated with circulating E2 levels (r =
-0.794, P < 0.01), but those of serum corrected
Ca did not correlate with circulating E2 levels.
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Discussion
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Pi is essentially necessary to body functions because it is a
constituent of the skeleton, membrane phospholipids, nucleic acids, and
nucleotides. Moreover, it is necessary in many kinds of metabolic
reactions and regulatory processes, including protein
phosphorylation.
Pi is widely recognized to be one of the most important constituent
minerals of bone. The content of phosphorus in the body increases from
4–5 g/kg at birth to 10–12 g/kg in adulthood (16). And although
levels of Pi are influenced by several factors, such as diet,
intestinal absorption, bone metabolism, and renal reabsorption, Pi
homeostasis in the body is mainly maintained by the mechanism of renal
proximal tubular reabsorption. Previous studies in vivo and
in vitro have reported that physiological regulation of
proximal tubular Pi reabsorption is most likely related to the capacity
of apical Na/Pi cotransport (13, 17, 19).
In cases where bone resorption is accelerated, such as at menopause,
phosphate enters the circulation along with Ca from bone. In contrast,
administration of estrogen to women at menopause leads to a reduction
of bone resorption (20), thereby suppressing the flux of Ca and Pi into
the circulation and ultimately decreasing the circulating level of Pi.
However, little is known about the effects of estrogen on renal
reabsorption of Pi in women.
In our studies, the values of TmP/GFR decreased with the increase of
estrogen level in all women who received HRT, and it increased with the
decrease of estrogen level in all women treated with GnRH-a.
Administration of CEE to women leads mainly to a rise in the
circulating estrone level, rather than in the E2
level, and thus we analyzed the correlations between circulating
E2 levels and other indices in the patients
treated with GnRH-a. We also studied the changes of serum intact PTH
levels in these patients. The concentrations of circulating
E2 correlated to those of serum Pi but did not
correlate to those of serum corrected Ca. These results suggest that
estrogen regulates the renal proximal tubular Pi reabsorption directly
or indirectly, but that estrogen does not regulate the renal Ca
reabsorption.
Previous studies have suggested that long-term estrogen administration
leads to an increase in circulating immunoreactive PTH (21, 22, 23), due
probably to a slight decrease in serum Ca resulting from inhibition of
bone resorption. It is recognized that PTH has an inhibitory effect on
the renal proximal tubular Na/Pi cotransporter (13, 19). Based on this
fact, we might offer another possible explanation that the effect of
estrogen on the renal Na/Pi cotransporter might be mediated by the
change of circulating PTH concentrations. However, although short-term
administration of estrogen to postmenopausal women reduces circulating
PTH, it also reduces the value of TmP/GFR (24). Therefore, the
influence of estrogen on the change of TmP/GFR does not always occur
through the change of circulating PTH. Moreover, Beers et
al. (25) recently reported that in thyroparathyroidectomized and
ovariectomized rats 17ß-E2 injection suppressed
the capacity for Na/Pi cotransport across the renal brush border
membrane. This report strongly suggests that estrogen has a direct
inhibitory effect on the renal proximal tubular Na/Pi cotransporter in
rats. In our studies, the values of TmP/GFR changed inversely to
circulating E2 levels, but the changes in TmP/GFR
were not associated with the changes of serum intact PTH levels.
Together with the results of Beers et al. (25), our present
findings suggest that estrogen may regulate renal Pi reabsorption by
acting directly on E2 receptors in renal proximal
tubular cells in women. However, a typical estrogen response element
was not observed in the sequence of 5'-flanking region of a type II
human Na/Pi transporter (NaPi-3) (26).
Pi is an activator of bone turnover. It is well known that estrogen
reduces bone turnover in postmenopausal women by its direct effect on
osteoblasts, which have been shown to have estrogen receptors (27), and
through several kinds of cytokines or growth factors (28, 29). Our
results indicate the further possibility that estrogen may reduce bone
turnover by reducing plasma Pi levels via a suppression in the capacity
for Na/Pi cotransport across the renal brush border membrane. However,
further investigation will be required to define the relationship
between the bone turnover rate and circulating Pi level.
In summary, we examined the relation between the changes of TmP/GFR and
the changes in circulating E2 levels in women
treated with HRT or GnRH-a and found that the changes of TmP/GFR and
circulating E2 were inversely related, whereas
those of TmP/GFR and serum intact PTH were not related. Our results
suggest that estrogen could act directly to suppress the capacity for
Na/Pi cotransport at the renal brush border membrane. To elucidate the
detailed mechanisms of estrogens on renal Na/Pi cotransport, more
fundamental studies will be needed.
Received September 2, 1999.
Revised November 10, 1999.
Accepted November 30, 1999.
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Abstract
Introduction
