help button home button Endocrine Society Endocrinology
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Liel, Y.
Right arrow Articles by Schwartz, B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Liel, Y.
Right arrow Articles by Schwartz, B.
Endocrinology Vol. 140, No. 1 280-285
Copyright © 1999 by The Endocrine Society

ARTICLES

Estrogen Increases 1,25-Dihydroxyvitamin D Receptors Expression and Bioresponse in the Rat Duodenal Mucosa1

Yair Liel, Shraga Shany, Patricia Smirnoff and Betty Schwartz

Department of Medicine and the Endocrine Unit (Y.L.), Clinical Biochemistry Unit (S.S.), Soroka University Hospital of Kupat-Holim and the Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84101, Israel; and Institute of Biochemistry (P.S., B.S.), Food Science and Nutrition, Faculty of Agricultural, Food and Environmental Quality Sciences, The Hebrew University of Jerusalem, Rehovot 96100, Israel

Address all correspondence and requests for reprints to: Betty Schwartz, Institute of Biochemistry, Food Science and Nutrition, Faculty of Agriculture, The Hebrew University, Rehovot 96100, Israel. E-mail: bschwart{at}agri.huji.ac.il


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Menopause and estrogen deficiency are associated with apparent intestinal resistance to vitamin D, which can be reversed by estrogen replacement. The in vivo influence of estrogens on duodenal vitamin D receptor (VDR) was studied in three groups of rats: ovariectomized (OVX), sham-operated, and ovariectomized rats treated daily with estrogen (40 µg/kg BW) for 2 weeks (OVX + E). Estrogen administration to OVX rats resulted in a 2-fold increase in VDR messenger RNA transcripts. 1,25(OH)2D3 was shown to bind specifically to one class of receptors in duodenal mucosal extracts, with a dissociation constant of 0.03 nM. Binding was significantly increased in duodenal extracts from OVX + E rats, compared with OVX rats (735 ± 81 vs. 295 ± 26 fmol/mg protein; P < 0.001); a comparable, 1.5- to 2-fold increase in VDR protein expression was observed in Western blot analyzes of the duodenal mucosa. Markers of VDR activity were increased in estrogen-exposed rats: calbindin-9k messenger RNA transcript content was 1.4- to 1.6-fold higher, and alkaline phosphatase activity was 1.4- to 3-fold higher in sham-operated and OVX + E, respectively, compared with OVX. 25(OH)D, 1,25(OH)2D, or PTH levels were not altered by estrogen treatment. Cumulatively, these findings suggest that estrogen up-regulates VDR expression in the duodenal mucosa and concurrently increases the responsiveness to endogenous 1,25(OH)2D. Modulation of intestinal VDR activity by estrogen, and subsequent influence on intestinal calcium absorption, could be one of the major protective mechanisms of estrogen against osteoporosis.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
INTESTINAL calcium absorption takes place via two main mechanisms: passive diffusion, which occurs when luminal calcium is high; and active absorption, a complex and not fully understood process mediated by 1,25(OH)2D3, which predominates when luminal calcium is low (1). Under physiological circumstances, 1,25(OH)2D3 is primarily produced in the kidneys under the influence of PTH stimulation. PTH secretion, in turn, is dependent on extracellular free calcium concentration, as sensed by calcium-sensing-receptors located in parathyroid cells’ membranes (2). Overall, the rate of active intestinal calcium absorption is determined by physiologic interactions between various components of the PTH-vitamin D-endocrine system organized in a multilevel negative feed-back loop structure.

Intestinal calcium absorption declines with age, both in humans (3, 4) and in rats (5). A widely held hypothesis suggests that the decrease in intestinal calcium absorption results from a sequence of events initiated by low estrogen levels, causing increased bone resorption; released calcium increases extracellular space calcium concentration, which suppresses PTH secretion, followed by a subsequent decrease in 1,25(OH)2D3 production and in 1,25(OH)2D3 plasma concentration, and finally results in decreased intestinal calcium absorption (6).

Nevertheless, there is evidence that estrogen may be more directly involved in determining intestinal calcium absorption. Estrogen receptors (7, 8, 9, 10), as well as estrogen-receptor-associated proteins, pS2 antigen (11, 12, 13) and ER-D5 (14), have been consistently demonstrated in the mucosa along the alimentary tract, suggesting a specific physiological role for estrogen in the intestine. Menopause, postmenopausal osteoporosis (6), and the postovariectomy state (5) are associated with decreased circulating estrogen and a concomitant decrease in calcium absorption. Moreover, available data indicates that the decrease in the basal levels of 1,25(OH)2D3 could not solely account for the decrease in calcium absorption, suggesting that the intestine of elderly or ovariectomized women is resistant to 1,25(OH)2D3 (15, 16). In addition, estrogen administration was shown to effectively restore the normal responsiveness of the intestine to 1,25(OH)2D3 in ovariectomized premenopausal women (16) and in postmenopausal women (17, 18).

Previous studies indicated an age-related decrease in intestinal vitamin D receptor (VDR) (19, 20). The number of VDRs is a primary determinant of the biological response to 1,25(OH)2D3, as previously shown in osteoblastic cell lines (21, 22) and in association with VDR-gene polymorphism in human populations (23, 24, 25). We have previously shown that estrogen increases the number of VDRs in the osteoblast-like cell line ROS 17/2.8, and that the increase in VDR number is associated with an increased responsiveness of the cells to 1,25(OH)2D3 (22). Others have shown that estrogen increases VDR messenger RNA (mRNA) transcript in osteoblasts (26, 27, 28). Estrogen-mediated increase in VDR expression was also noted in other tissues and cell types, such as uterus (29, 30) liver (31), and human breast cancer cells (32).

Modulation of VDR expression in the small intestine by estrogen could account for the relative resistance to 1,25(OH)2D3 in the senescent and estrogen-deprived intestine and its correction during estrogen replacement. It could also account, at least in part, for the protective role of estrogen replacement against osteoporosis in estrogen-deficiency states. To this end, the present study was designed to investigate the interaction between estrogen, VDRs, and markers of vitamin D bioactivity in the rat duodenal mucosa.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Chemicals and reagents were purchased from Sigma Chemical Co. (St. Louis, MO), unless otherwise specified. 1,25-dihydroxyvitamin D3 was kindly provided as a gift by Hoffmann-La Roche (Basel, Switzerland). 1{alpha} 25-Dihydroxy [26,27-methyl-[3H]-vitamin D3 (180 Ci/mmol) was obtained from Amersham Laboratories, (Amersham, England). AMV reverse transcriptase, ribonuclease inhibitor, and random hexamer were obtained from Boehringer Mannheim (Mannheim, Germany). Taq polymerase was from Beit-Ha’emek Laboratories (Beit-Ha’emek, Israel). Oligonucleotides used for PCR primers were synthesized in the Biotechnology Weizmann Institute of Sciences (Rehovot, Israel). Monoclonal rat anti-VDR antibody for immunodetection by Western blot was purchased from Chemicon International Inc. (Temecula, CA). Monoclonal rat anti-VDR antibody has extensive cross-reactivity with all avian and mammalian VDR. The enhanced chemiluminescence kit was from Amersham, Buckinghamshire, UK.

Experimental protocol
Charles-River outbred female rats (120–150 g) were divided randomly into three groups, were maintained in separate plastic cages at 12-h light, 12-h dark cycles and had free access to Purina chow. Rats were ovariectomized (OVX) or sham-operated (Sham) under light ether anesthesia. A subgroup of ovariectomized rats received 17ß-estradiol (40 µg/kg BW daily, sc) for 14 days, starting 24 h after ovariectomy (OVX + E). Estrogen was dissolved in absolute alcohol and maintained at -20 C. Immediately before administration, aliquots from the batch solution were diluted to 10% ethanol in normal saline. OVX rats received 10% ethanol vehicle only.

At the end of the treatment period, all animals were killed by decapitation; duodeni and uteri were removed. The duodenal mucosa was washed with ice-cold saline solution, laid open, and gently scraped. Uteri were removed, cleaned, and weighed to confirm estrogenic effect.

Serum collection
Blood was collected in chilled tubes for 25-hydroxyvitamin D [25-(OH)D], 1,25-dihydroxyvitamin D [1,25-(OH)2D)], and PTH determinations. The serum was subsequently separated, in a refrigerated centrifuge, and kept at -20 C until further processing.

Serum 25(OH)D and 1,25(OH)2D determination
Two-milliliter serum samples were submitted to lipid extraction by equal volumes of acetonitrile. 25-(OH)D and 1,25-(OH)2D fractions were separated by a Sep-Pak (Waters, Milford, MA) separation procedure using C-18 and silica columns (33). The 25-(OH)D fraction was submitted to a competitive protein-binding assay using diluted rachitic rat serum as the binding protein (34). Quantification of the 1,25-(OH)2D fraction was achieved by a VDR-binding assay (35) using calf thymus VDR (Nichols Institute Diagnostics, San Juan Capistrano, CA) as the ligand binder. All determinations were carried out in triplicate. Results are mean ± SE and are expressed as ng/ml serum of 25-(OH)D, or as pg/ml serum of 1,25-(OH)2D.

Serum PTH determination
Serum PTH was measured with a rat-specific PTH RIA kit from Nichols Institute Diagnostics. Two hundred microliters of serum sample was mixed with 100 microliters of 125I-labeled rat PTH. A single PTH-antibody-coated bead was added and incubated at 4 C for 24 h. At the end of the incubation, the solution was discarded, and the beads were washed in phosphate-buffered solution (0.01 M), pH 7.4. The radioactivity was assessed in a {gamma} counter. PTH concentrations were determined with a standard curve from appropriate rat-PTH standards.

RNA isolation
Total RNA was extracted from scraped duodenal mucosae, according to a protocol for single-step RNA isolation based on acid guanidinium-thiocyanate-phenol-chloroform extraction, using Tri-reagent solution (36). Aliquots of total RNA were separated in sterile tubes and quantified.

Northern blot procedures
After denaturation, 35-µg total RNA samples were submitted to electrophoresis in 1% agarose, 2.2 M formaldehyde gel, transferred to nylon membranes (Hybond N+, Amersham, Buckinghamshire, UK), and hybridized with 32P-labeled VDR, calbindin-9k, and ß-actin probes. The probes were labeled with {alpha} 32P-CTP by a random-primed DNA labeling procedure with Klenow polymerase (Random Primed-DNA Labeling Kit, Boehringer Mannheim Biochemica, Mannheim, Germany).

Probe preparation by PCR
Complementary DNA sequences for the detection of mRNA transcripts of VDR, ß-actin, and calbindin-9k genes by Northern blot procedures were obtained by RT-PCR of intestinal RNA extract, according to Iida et al. (37) using appropriate primers and reverse transcriptase enzyme. The PCR buffer consisted of 10 mM Tris (pH 8.3), 50 mM KCl, 2 mM MgCl2, 0.01% gelatin, 0.2 mM deoxynucleotide triphosphates, and 1.25 U/50 µl Taq polymerase. The VDR-gene PCR primers consisted of bases 41–60 in the sense direction (5'-GTGACTTTGACCGGAACGTG-3') and bases 301–320 in the antisense direction (5'-ATCATCTCCCTCTTACGCTG-3') of the VDR gene (37), and the calculated PCR product length was 280 bp. The PCR program involved 35 cycles: 40 sec at 94 C, 60 sec at 50 C, and 90 sec at 72 C.

The calbindin-9k primers corresponded to the reported complementary DNA (cDNA) sequence of the rat intestinal vitamin D-dependent calcium binding protein (38). They consisted of bases 1–21 in the sense direction (5'-GAGACCTCACCTGTTCCTGTC-3') and bases 387–406 (5'-GTCTCGGAATTTGCTTTAT-3') in the antisense direction. The calculated PCR product length was 406 bp. The PCR program involved 30 cycles: 60 sec at 94 C, 90 sec at 55 C, and 90 sec at 72 C.

The primers used to obtain the probe to the housekeeping gene ß-actin corresponded to bases 2846–2863 in the sense direction (5'-TCCTAGCACCATGAAGATC-3') and to bases 3140–3158 in the antisense direction (5'-AAACGCAGCTCAGTAACAG-3'). The PCR program used for these sets of primers involved 30 cycles: 40 sec at 94 C, 60 sec at 60 C, and 90 sec at 72 C. The calculated PCR product length was 190 bp (37).

In ensuing probe amplification PCR was performed using cDNA templates obtained by RT-PCR and appropriate primers for VDR, ß-actin, and calbindin-9k genes, at the conditions described previously.

1,25(OH)2D receptor (VDR) assay
VDR protein expression was determined by a ligand-binding assay, as previously described (39), in 250 µl of the cells’ 30,000 x g-soluble fraction after homogenate extraction with hypertonic buffer containing 0.3 M KCl. Parallel incubations of the soluble fractions were carried out with different concentrations of [3H]-1,25(OH)2D3, between 20–160 fmol, with or without the addition of 100-fold molar concentration of the radio-inert (cold) 1,25(OH)2D3. Kd (dissociation constant) and maximal specific binding capacities (Bmax) were determined by Scatchard analysis.

Western blot analysis
Intestinal tissue lysates were separated on a 10% SDS gel and electroblotted onto nitrocellulose gels. Immunodetection used mouse monoclonal antibodies raised against VDR. After incubation with the anti-VDR antibodies, the gels were treated with goat antirat Igs conjugated to horseradish peroxidase. Visualization of the VDR protein was attained with an enhanced chemiluminescence kit, according to the manufacturer’s instructions.

Quantification of autoradiogram signals
The signals of the Northern and Western autoradiograms were quantified using a densitometric scanner with the Fujix Phosphor-Imager apparatus and the BAS-1000 Bio-Imaging Analyzer (Fuji Photo Film Co., Ltd. Co., Japan). Results of the densitometric determinations of VDR and calbindin-9-kDa mRNA were corrected according to the corresponding ß-actin gene-transcription.

Alkaline phosphatase activity in the colonic mucosa
Cytosolic enzyme activity was determined in homogenates from freshly isolated duodenal scrapings, with p-nitrophenylphosphate as substrate, as previously described (40).

Protein determination
Protein concentration in the different suspensions was determined by a Pierce kit (Pierce Chemical Co., Rockford, IL) (41), employing a microbicinchoninic acid-based protein assay and BSA as the protein standard.

Statistical analysis
Statistical evaluation for two-group analysis was carried out by the unpaired Student’s t test. Results are expressed as mean ± SE.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Throughout this study, uterine weight served as an independent marker for estrogenic activity in the different groups of rats (Table 1Go). The markedly reduced uterine weight in OVX rats confirmed least exposure to estrogen in this group, compared with the Sham and OVX + E groups. The higher uterine weights of the OVX + E group are attributed to the supraphysiological dose of estradiol and to lack of the physiological cyclicity in estrogen production.


View this table:
[in this window]
[in a new window]
  		Table 1. Effect of estrogen on rats’ uterine weight, duodenal VDR protein expression by Western-blot, and duodenal VDR binding characteristics

 
Effect of estradiol on VDR transcription
Northern blot analysis for mRNA transcripts of VDR indicated that the strongest expression was achieved after estrogen treatment (Fig. 1Go). Expression of the VDR mRNA transcripts was also notable in Sham rats.



View larger version (65K):
[in this window]
[in a new window]
  		Figure 1. Phosphor-Imager results of Northern blot analysis of RNA (35 µg/lane) from rat duodenal tissues probed with 32P-labeled rat VDR cDNA and exposed for 10 h. Lane 1, OVX; lane 2, Sham; lane 3, OVX + E. Rat VDR mRNA expression is most intense in duodenal tissue from OVX + E rats (lane 3).

 
Densitometric quantification of the signals produced by the hybridization in each treatment group, normalized according to ß-actin mRNA expression, indicated an increase of approximately 2-fold in VDR mRNA content in estrogen-exposed animals (Sham and OVX + E), compared with OVX alone (Fig. 2Go).



View larger version (56K):
[in this window]
[in a new window]
  		Figure 2. Histogram of the effect of different treatments on rat VDR (Fig. 1Go) and calbindin-9k (Fig. 5Go) mRNA content in dodenal tissue from OVX, Sham, and OVX + E rats. The intensities of the transcripts were quantified by pixel densitometry and normalized to the respective intensities of the rat ß-actin mRNA products obtained in the different treatment protocols.

 
Effect of estradiol on VDR protein expression
1,25(OH)2D3 binding studies. 1,25(OH)2D3 was shown to bind specifically to one class of receptors in hypertonic-salt extracts of duodenal mucosal extracts (Fig. 3Go and Table 1Go). VDR concentration (maximal bound 1,25(OH)2D3 per mg protein; Bmax) from Sham and OVX + E rats was significantly (at least 2-fold) higher than that of duodenal extracts from OVX rats.



View larger version (20K):
[in this window]
[in a new window]
  		Figure 3. Effect of estrogen on 1,25(OH)2D3 binding to duodenal mucosal extracts. Representative Scatchard plots.

 
Western-blot analysis. Western-blot analysis detected a VDR immunoreactive protein of 48 kDa (Fig. 4Go). The intensity of the corresponding bands from Sham and OVX + E were markedly stronger than those obtained from OVX rats. Quantification of VDR expression indicated a 1.6- to 2-fold increase in VDR-protein expression in the estrogen-exposed rats, compared with the estrogen-deprived rats (Table 1Go).



View larger version (21K):
[in this window]
[in a new window]
  		Figure 4. Effect of estrogen on VDR protein expression, by Western-blot analysis in colonic mucosal extracts. Extracts from all treatment groups expressed the expected 48-kDa protein. See Table 2Go for results of densitometric analysis of the corresponding bands.

 
Effect of estradiol on functional expression of VDR in vivo
Duodenal alkaline phosphatase activity. Alkaline phosphatase activity, a nonspecific vitamin D-induced marker of intestinal cell differentiation (42, 43), was determined in extracts of duodenal mucosa. The results indicate a significant increase in alkaline phosphatase activity in intestinal extracts from estrogen-exposed (Sham or OVX + E) rats, compared with estrogen-deprived (OVX) rats (Table 2Go).


View this table:
[in this window]
[in a new window]
  		Table 2. Effect of estradiol treatment on alkaline phosphatase activity in cytosolic extracts of rat duodenal mucosae

 
Northern blot analysis of calbindin-9k mRNA expression. Calbindin-9k is a vitamin D-dependent calcium-binding protein specifically modulated by 1,25(OH)2D3 at the genomic level (44, 45, 46). Using PCR and ethidium bromide staining, a single band of the expected size (406 bp) calbindin-9k product was detected (not shown). The purified cDNA fragment provided the probe for the subsequent Northern blot analysis. Northern blot band intensities of the transcribed calbindin-9k gene of duodenal extracts from Sham and OVX + E rats were markedly stronger, compared with OVX extracts (Fig. 5Go). Quantification of the signals obtained by each treatment was attained by densitometry and was corrected according to the corresponding ß-actin signal for each sample. The results indicate a 1.6-fold increase in calbindin-9k mRNA content in estrogen-exposed, compared with estrogen-deprived, rats (Fig. 2Go).



View larger version (42K):
[in this window]
[in a new window]
  		Figure 5. Phosphor-Imager result of a Northern blot analysis of RNA (35 µg/lane) from rat duodenal tissues probed with 32P-labeled rat calbindin-9k cDNA and exposed for 10 h. Lane 1, OVX; lane 2, Sham; lane 3, OVX + E2. Rat calbindin-9k mRNA expression is most intense in duodenal tissue from OVX + E rats.

 
Serum PTH and vitamin D metabolites
Estrogen treatment did not affect serum levels of either vitamin D metabolites or PTH (Table 3Go). Serum levels of PTH, 25-(OH)D and 1,25-(OH)2D were similar in rats from all treatment groups.


View this table:
[in this window]
[in a new window]
  		Table 3. Effect of estradiol treatment on PTH and vitamin D metabolites

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
These results reveal a direct relationship between estrogen exposure, VDR mRNA-transcript content, and VDR protein expression in the duodenal mucosa, suggesting that estrogen has a pivotal role in supporting VDR expression in the duodenal mucosa. Chan et al. (47) have previously documented a 30% decrease in intestinal VDR of OVX rats; however, those results could not be verified by a subsequent study (31).

Having documented that estrogen up-regulates VDR in the duodenal mucosa, we examined VDR-related bioresponses and observed a significant increase in colonic mucosal alkaline phosphatase activity and calbindin-9k steady-state mRNA content in duodenal mucosa of estrogen-exposed (compared with estrogen-deprived) rats. Alkaline phosphatase comprises a family of specific isoenzymes, the activity of which is associated with the differentiated phenotype of enterocytes and colonocytes (42, 43); calbindin-9k is one of the best documented biological markers of the hormonal action of 1,25-(OH)2D3 at the genomic level (44, 45, 46). The present results indicate that, in addition to modulation of VDR expression, in vivo exposure to estrogen markedly enhances the response of markers of VDR activity in the duodenal mucosa to endogenous 1,25-(OH)2D. Neither serum 1,25-(OH)2D nor PTH concentrations were altered by estrogen repletion; thus, we may conclude that the increase in intestinal VDR expression and activity was directly related to estrogen and is unlikely the result of an indirect effect of estrogen via PTH or 1,25-(OH)2D on VDR expression (48, 49).

The present study, in addition to previous observations in osteoblasts (26, 27, 28, 50), indicates that estrogen increases total VDR mRNA content. Theoretically, estrogen could exert its effect on a genomic level directly, by stimulating VDR gene transcription. However, an estrogen response element has not yet been discovered in association with the VDR gene (51, 52); it is thus possible that estrogen influences VDR gene transcription indirectly, through activation of other transcription factor(s) or through stabilization of the VDR mRNA transcript.

Available data indicates that the difference in VDR expression between young and adult rats is in the order of 1.5-fold in favor of young rats (19), comparable with the acute effect of ovariectomy on VDR expression observed in the present study, and with the effect of estrogen deprivation on intestinal calcium absorption (3, 4, 16, 17). Although some controversy still surrounds the issue of the role of VDR gene polymorphism in determining bone mineralization, recent observations indicate that the start-codon polymorphism, which is associated with differences in the VDR gene transcript level of a magnitude similar to that observed in the present study, is associated with significant differences in bone mineral density between carriers of the various polymorphic alleles (23, 24, 25). Taken together, these observations support the notion that changes in VDR expression, such as those observed in the present study, may be of physiological and pathophysiological significance. Our results suggest that estrogen deficiency directly influences VDR expression in the senescent intestinal mucosa, subsequently affecting intestinal calcium absorption. The effect of estrogen on intestinal VDR expression also provides a pertinent mechanism for the stimulatory effect of estrogen replacement on intestinal calcium absorption observed in estrogen-deficient rats and in women (53, 54).

Estrogen is one of the most effective available treatments for osteoporosis during menopause and in estrogen-deficiency states. The results of the present study, combined with previous observations from our group, indicate a modulatory effect of estrogen in osteoblastic cells (22), to suggest a unifying (although not exclusive) mechanism by which estrogen may protect the skeleton. Taken together, our previous observation in osteoblastic cells (22) and the present results, suggest that estrogen simultaneously increases VDR expression and specific 1,25-(OH)2D-related activities in osteoblasts and in the intestinal mucosa, resulting in stimulation of bone-matrix protein synthesis, along with an increase in mineral supply. These complementary consequences of estrogen activity are likely the result of a similar molecular mechanism of estrogen activity in different types of cells, and they may account, at least in part, for the remarkable effectiveness of estrogen in maintaining bone integrity and in preventing osteoporosis.


   Footnotes
 
1 This study was supported, in part, by a grant from the Chief Scientist’s Office, the Ministry of Health of the State of Israel, and by a grant from the Faculty of Health Sciences, Ben-Gurion University of the Negev. Back

Received April 2, 1998.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Bringhurst RF 1995 Calcium and posphate distribution, turnover, and metabolic actions. In: DeGroot LI (ed) Endocrinology. Saunders Company, Philadelphia, pp 1015–1043
  2. Chattopadhyay N, Mithal A, Brown EM 1996 The calcium-sensing receptor: a window into the physiology and pathophysiology of mineral ion metabolism. Endocr Rev 17:289–307[Abstract/Free Full Text]
  3. Avioli LV, McDonald JE, Lee SIN 1965 The influence of age on the intestinal absorption of 47Ca in women and its relation to 47Ca absorption in postmenopausal osteoporosis. J Clin Invest 44:1960–1967
  4. Bullamore JR, Gallagher JC, Wilkinson R, Nordin BEC, Marshall DH 1970 Effect of age on calcium absorption. Lancet 2:535–537[CrossRef][Medline]
  5. Russell JE, Morimoto S, Brige SJ, Fausto A, Avioli LV 1986 Effects of age and estrogen on calcium absorption in the rat. J Bone Miner Res 1:185–189[Medline]
  6. Gallagher JC, Riggs BL, Eisman J, Hamstra A, Arnoud SB, DeLuca HF 1979 Intestinal calcium absorption and serum vitamin D metabolites in normal subjects and osteoporotic patients. J Clin Invest 64:729–736
  7. Fernandez E, Lavecchia C, Davanzo B, Franceschi S, Negri E, Parazzini F 1996 Oral contraceptives, hormone replacement therapy and the risk of colorectal cancer. Br J Cancer 73:1431–1435[Medline]
  8. Francavilla A, Di Leo A, Polimeno L, Conte D, Barone M, Fanitta G, Chiumarulo C, Rizzo G, Rubino M 1987 Nuclear and cytosolic estrogen receptors in human colon carcinoma and in surrounding noncancerous colonic tissue. Gastroenterology 93:1301–1306
  9. Meggouh F, Lointier P, Saez S 1991 Sex steroid and 1,25-dihydroxyvitamin D3 receptors in human colorectal adenocarcinoma and normal mucosa. Cancer Res 51:1227–1233[Abstract/Free Full Text]
  10. Hendrickse CW, Jones CE, Donovan IA, Neoptolemos JP, Baker PR 1993 Oestrogen and progesterone receptors in colorectal cancer and human colonic cancer cell lines. Br J Surg 80:636–640[Medline]
  11. Theisinger B, Guthke R, Blin N, Welter C, Seitz G 1993 Influence of steroid hormones on pS2/BCEI gene expression in xenografted colon tumors. In Vivo 7:411–414[Medline]
  12. Luqmani YA, Ryall G, Shousha S, Coombes RC 1992 An immunohistochemical survey of pS2 expression in human epithelial cancers. Int J Cancer 50:302–304[Medline]
  13. Welter C, Theisinger B, Rio MC, Seitz G, Schuder G, Blin N 1994 Expression pattern of breast-cancer-associated protein pS2/BCEI in colorectal tumors. Int J Cancer 56:52–55[Medline]
  14. Takeda H, Yamakawa M, Takahashi T, Imai Y, Ishikawa M 1992 An immunohistochemical study with an estrogen receptor-related protein (ER-D5) in human colorectal cancer. Cancer 69:907–912[CrossRef][Medline]
  15. Francis RM, Peacock M, Storer JH, Nordin BEC 1984 Calcium malabsorption in elderly women with vertebral fractures: evidence for resistance to the action of vitamin D on the bowel. Clin Sci 66:103–107[Medline]
  16. Gennari C, Agnusdei D, Nardi P, Civitelli R 1990 Estrogen preserves a normal intestinal responsiveness to 1,25-dihydroxyvitamin D3 in oophorectomized women. J Clin Endocrinol Metab 71:1288–1293[Abstract/Free Full Text]
  17. Civitelli R, Agnusdei D, Nardi P, Zacchei F, Avioli LV, Gennari C 1988 Effects of one-year treatment with estrogens on bone mass, intestinal calcium absorption, and 25-hydroxyvitamin D-1 alpha-hydroxylase reserve in postmenopausal osteoporosis. Calcif Tissue Int 42:77–86[Medline]
  18. Heaney RP, Recker RR, Saville PD 1978 Menopausal changes in calcium balance performance. J Lab Clin Med 92:953–963[Medline]
  19. Horst RL, Goff JP, Reinhardt TA 1990 Advancing age results in reduction of intestinal and bone 1,25-dihydroxyvitamin D receptor. Endocrinology 126:1053–1057[Abstract/Free Full Text]
  20. Takamoto S, Seino Y, Sacktor B, Liang CT 1990 Effect of age on 1,25-dihydroxyvitamin D3 receptors in Wistar rats. Biochim Biophys Acta 1034:22–28[Medline]
  21. Dokoh S, Donaldson CA, Haussler MR 1984 Influence of 1,25-dihydroxyvitamin D3 on cultured osteogenic sarcoma cells: correlation with the 1,25-dihydroxyvitamin D3 receptors. Cancer Res 44:2103–2109[Abstract/Free Full Text]
  22. Liel Y, Kraus S, Levy J, Shany S 1992 Evidence that estrogens modulate activity and increase the number of 1,25-dihydroxyvitamin D3 receptors in osteoblast-like cells (ROS 17/2.8). Endocrinology 130:2597–2601[Abstract/Free Full Text]
  23. Arai H, Miyamoto K, Taketani Y, Yamamoto H, Iemori Y, Morita K, Tonai T, Nishisho T, Mori S, Takeda E 1997 A vitamin D receptor gene polymorphism in the translational initiation codon: effect on protein activity and relation to bone mineral density in Japanese women. J Bone Miner Res 12:915–921
  24. Harris SS, Eccleshall TR, Gross C, Dawson-Hughes B, Feldman D 1997 The vitamin D receptor start codon polymorphism (FokI) and bone mineral density in premenopausal American black and white women. J Bone Miner Res 12:1043–1048[CrossRef][Medline]
  25. Gross C, Eccleshall TR, Malloy PJ, Villa ML, Marcus R, Feldman D 1996 The presence of a polymorphism at the translation initiation site of the vitamin D receptor gene is associated with low bone mineral density in postmenopausal Mexican-American women. J Bone Miner Res 11:1850–1855[Medline]
  26. Mahonen A, Pirskanen A, Maenpaa PH 1991 Homologous and heterologous regulation of 1,25-dihydroxyvitamin D3 receptor messenger RNA levels in human osteosarcoma cells. Biochim Biophys Acta 1088:111–118[Medline]
  27. Maenpaa P, Mahonen A, Pirskanen A 1991 Hormonal regulation of vitamin D receptor levels and osteocalcin synthesis in human osteosarcoma cells. Calcif Tissue Int 49:S85–S86
  28. Ishibe M, Nojima T, Ishibashi T, Koda T, Kaneda K, Rosier RN, Puzas JE 1995 17Beta-estradiol increases the receptor number and modulates the action of 1,25-dihydroxyvitamin D3 in human osteosarcoma-derived osteoblast-like cells. Calcif Tissue Int 57:430–435
  29. Walters MR 1981 An estrogen-stimulated 1,25-dihydroxyvitamin D3 receptor in rat uterus. Biophys Biochem Res Commun 103:721–726
  30. Levy J, Zuili I, Yankowitz N, Shany S 1984 Induction of cytosolic receptors for 1{alpha},25 dihydroxyvitamin D3 in the immature rat uterus by oestradiol. J Endocrinol 100:265–269[Abstract/Free Full Text]
  31. Duncan WE, Glass AR, Wray HL 1991 Estrogen regulation of the nuclear 1,25-dihydroxyvitamin D3 receptor in rat liver and kidney. Endocrinology 129:2318–2324[Abstract/Free Full Text]
  32. Escaleira MT, Sonohara S, Brentani MM 1993 Sex steroids induced up-regulation of 1,25-(OH)2 vitamin D3 receptors in T 47D breast cancer cells. J Steroid Biochem Mol Biol 45:257–263[CrossRef][Medline]
  33. Adams JS, Singer FR, Gacad MA, Sharma OP, Hayes MJ, Vouros P, Holick MF 1985 Isolation and structural identification of 1,25-dihydroxyvitamin D3 produced by cultured alveolar macrophages in sarcoidosis. J Clin Endocrinol Metab 60:969–966
  34. Shany S, Rapoport J, Zuili I, Yankowitz N, Chaimovitz C 1986 Enhancement of 24.25-dihydroxyvitamin D levels in patients treated with continuous ambulatory peritoneal dialysis. Nephron 42:141–145[Medline]
  35. Reinhardt TA, Horst RL, Orf JW, Hollis BW1984 A microassay for 1,25(OH)2D3 not requiring HPLC. Applications to clinical studies. J Clin Endocrinol Metab 58:91–98
  36. Chomczynski P, Sacchi N 1987 Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156–159[Medline]
  37. Iida K, Taniguchi S, Kurokawa K 1993 Distribution of 1,25-dihydroxyvitamin D3 receptor and 25-hydroxyvitamin D3–24-hydroxylase mRNA expression along rat nephron segments. Biochem Biophys Res Commun 194:659–664[CrossRef][Medline]
  38. Darwish HM, Krisinger J, Strom M, DeLuca HF 1987 Molecular cloning of the cDNA and chromosomal gene for vitamin D-dependent calcium-binding protein of rat intestine. Proc Natl Acad Sci USA 84:6108–6111[Abstract/Free Full Text]
  39. Belleli A, Shany S, Levy J, Guberman R, Lamprecht SA 1992 A protective role of 1,25-dihydroxyvitamin D3 in chemically induced rat colon carcinogenesis. Carcinogenesis 13:2293–2298[Abstract/Free Full Text]
  40. Schwartz B, Avivi C, Lamprecht SA 1991 Isolation and characterization of normal and neoplastic colonic epithelial cell populations. Gastroenterology 100:692–702[Medline]
  41. Kirschbaum B 1986 Use of the bicinchoninic acid assay in measuring urinary proteins. Clin Chem 32:572[Free Full Text]
  42. Gum JR, Kam WK, Byrd JC, Hicks JW, Sleisenger MH, Kim YS 1987 Effects of sodium butyrate on human colonic adenocarcinoma cells. Induction of placental-like alkaline phosphatase. J Biol Chem 262:1092–1097[Abstract/Free Full Text]
  43. Deng G, Liu G, Hu L, Gum Jr JR, Kim YS 1992 Transcriptional regulation of the human placental-like alkaline phosphatase gene and mechanisms involved in its induction by sodium butyrate. Cancer Res 52:3378–3383[Abstract/Free Full Text]
  44. Christakos S, Gabrielides C, Rhoten WB 1989 Vitamin D-dependent calcium binding proteins: chemistry, distribution, functional considerations, and molecular biology. Endocr Rev 10:3–26[Abstract/Free Full Text]
  45. Romagnolo B, Cluzeaud F, Lambert M, Colnot S, Porteu A, Molina T, Tornasset M, Vandewalle A, Kahn A, Perret C 1996 Tissue-specific and hormonal regulation of calbindin-D9K fusion genes in transgenic mice. J Biol Chem 271:16820–16826
  46. Fleet JC, Wood RJ 1994 Identification of calbindin D-9k mRNA and its regulation by 1,25-dihydroxyvitamin D3 in Caco-2 cells. Arch Biochem Biophys 308:171–174[CrossRef][Medline]
  47. Chan SDH, Chiu DKH, Atkins D 1984 Oophorectomy leads to a selective decrease in 1,25-dihydroxycholecalciferol receptors in rat jejunal villous cells. Clin Sci 66:745–748[Medline]
  48. Krishnan AV, Feldman D 1997 Regulation of vitamin D receptor abundance. In: Feldman D, Glorieux FH, Pike JW (eds) Vitamin D. Academic Press, San Diego, pp 179–200
  49. Gensure RC, Antrobus SD, Fox J, Okwueze M, Talton SY, Walters MR 1998 Homologous up-regulation of vitamin D receptors is tissue specific in the rat. J Bone Miner Res 13:454–463[CrossRef][Medline]
  50. Davoodi F, Brenner RV, Evans SR, Schumaker LM, Shabahang M, Nauta RJ, Buras RR 1995 Modulation of vitamin D receptor and estrogen receptor by 1,25(OH)2-vitamin D3 in T-47D human breast cancer cells. J Steroid Biochem Mol Biol 54:147–153
  51. Jehan F, DeLuca HF 1997 Cloning and characterization of the mouse vitamin D receptor promoter. Proc Natl Acad Sci USA 94:10138–10143[Abstract/Free Full Text]
  52. Miyamoto K, Kesterson RA, Yamamoto H, Taketani Y, Nishiwaki E, Tatsumi S, Inoue Y, Morita K, Takeda E, Pike JW 1997 Structural organization of the human vitamin D receptor chromosomal gene and its promoter. Mol Endocrinol 11:1165–1179[Abstract/Free Full Text]
  53. Arjmandi BH, Hollis BW, Kalu DN 1994 In vivo effect of 17 beta-estradiol on intestinal calcium absorption in rats. Bone Miner 26:181–189[Medline]
  54. Mauras N, Vieira NE, Yergey AL 1997 Estrogen therapy enhances calcium absorption and retention and diminishes bone turnover in young girls with Turner’s syndrome: a calcium kinetic study. Metab Clin Exp 46:908–913



This article has been cited by other articles:


Home page
 Cancer Prevention ResearchHome page
P. Protiva, H. S. Cross, M. E. Hopkins, E. Kallay, G. Bises, E. Dreyhaupt, L. Augenlicht, M. Lipkin, M. Lesser, E. Livote, et al.
Chemoprevention of Colorectal Neoplasia by Estrogen: Potential Role of Vitamin D Activity
Cancer Prevention Research, January 1, 2009; 2(1): 43 - 51.
[Abstract] [Full Text] [PDF]

Home page
 J Mol EndocrinolHome page
L. A. Gilad and B. Schwartz
Association of estrogen receptor {beta} with plasma-membrane caveola components: implication in control of vitamin D receptor
J. Mol. Endocrinol., June 1, 2007; 38(6): 603 - 618.
[Abstract] [Full Text] [PDF]

Home page
 EndocrinologyHome page
Y. Song and J. C. Fleet
Intestinal Resistance to 1,25 Dihydroxyvitamin D in Mice Heterozygous for the Vitamin D Receptor Knockout Allele
Endocrinology, March 1, 2007; 148(3): 1396 - 1402.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Endocrinol. Metab.Home page
Y. Zhang, W.-P. Lai, C.-F. Wu, M. J. Favus, P.-C. Leung, and M.-S. Wong
Ovariectomy worsens secondary hyperparathyroidism in mature rats during low-Ca diet
Am J Physiol Endocrinol Metab, March 1, 2007; 292(3): E723 - E731.
[Abstract] [Full Text] [PDF]

Home page
 J EndocrinolHome page
L. A. Gilad, O. Tirosh, and B. Schwartz
Phytoestrogens regulate transcription and translation of vitamin D receptor in colon cancer cells.
J. Endocrinol., November 1, 2006; 191(2): 387 - 398.
[Abstract] [Full Text] [PDF]

Home page
 J EndocrinolHome page
L. A. Gilad, T. Bresler, J. Gnainsky, P. Smirnoff, and B. Schwartz
Regulation of vitamin D receptor expression via estrogen-induced activation of the ERK 1/2 signaling pathway in colon and breast cancer cells
J. Endocrinol., June 1, 2005; 185(3): 577 - 592.
[Abstract] [Full Text] [PDF]

Home page
 J. Nutr.Home page
Y. Song and J. C. Fleet
1,25 Dihydroxycholecalciferol-Mediated Calcium Absorption and Gene Expression Are Higher in Female than in Male Mice
J. Nutr., August 1, 2004; 134(8): 1857 - 1861.
[Abstract] [Full Text] [PDF]

Home page
 J. Nutr.Home page
M. Cifuentes, J. P. Advis, and S. A. Shapses
Estrogen Prevents the Reduction in Fractional Calcium Absorption Due to Energy Restriction in Mature Rats
J. Nutr., August 1, 2004; 134(8): 1929 - 1934.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Endocrinol. Metab.Home page
E. M. Colin, A. G. Uitterlinden, J. B. J. Meurs, A. P. Bergink, M. Van De Klift, Y. Fang, P. P. Arp, A. Hofman, J. P. T. M. van Leeuwen, and H. A. P. Pols
Interaction between Vitamin D Receptor Genotype and Estrogen Receptor {alpha} Genotype Influences Vertebral Fracture Risk
J. Clin. Endocrinol. Metab., August 1, 2003; 88(8): 3777 - 3784.
[Abstract] [Full Text] [PDF]

Home page
 J. Nutr.Home page
P. D. O'Loughlin and H. A. Morris
Oophorectomy Acutely Increases Calcium Excretion in Adult Rats
J. Nutr., July 1, 2003; 133(7): 2277 - 2280.
[Abstract] [Full Text] [PDF]

Home page
 J. Nutr.Home page
M. Cifuentes, A. B. Morano, H. A. Chowdhury, and S. A. Shapses
Energy Restriction Reduces Fractional Calcium Absorption in Mature Obese and Lean Rats
J. Nutr., September 1, 2002; 132(9): 2660 - 2666.
[Abstract] [Full Text] [PDF]

Home page
 EndocrinologyHome page
S. Peleg, M. Uskokovic, A. Ahene, B. Vickery, and Z. Avnur
Cellular and Molecular Events Associated with the Bone-Protecting Activity of the Noncalcemic Vitamin D Analog Ro-26-9228 in Osteopenic Rats
Endocrinology, May 1, 2002; 143(5): 1625 - 1636.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Endocrinol. Metab.Home page
S. Pattanaungkul, B. L. Riggs, A. L. Yergey, N. E. Vieira, W. M. O’Fallon, and S. Khosla
Relationship of Intestinal Calcium Absorption to 1,25-Dihydroxyvitamin D [1,25(OH)2D] Levels in Young Versus Elderly Women: Evidence for Age-Related Intestinal Resistance to 1,25(OH)2D Action
J. Clin. Endocrinol. Metab., November 1, 2000; 85(11): 4023 - 4027.
[Abstract] [Full Text]

Home page
 J. Nutr.Home page
R. Vieth, S. Milojevic, and V. Peltekova
Improved Cholecalciferol Nutrition in Rats Is Noncalcemic, Suppresses Parathyroid Hormone and Increases Responsiveness to 1,25-Dihydroxycholecalciferol
J. Nutr., March 1, 2000; 130(3): 578 - 584.
[Abstract] [Full Text]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Liel, Y.
Right arrow Articles by Schwartz, B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Liel, Y.
Right arrow Articles by Schwartz, B.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Endocrinology Endocrine Reviews J. Clin. End. & Metab.
Molecular Endocrinology Recent Prog. Horm. Res. All Endocrine Journals