Bone density in healthy men after cessation of calcium supplements: 20-month follow-up of a randomized controlled trial.

UNLABELLED: Bone density has been followed up for 20 months following completion of a trial which compared calcium 1,200 mg/day with placebo, in normal older men. Following cessation of calcium supplements, there is a small residual benefit in total body bone density, but not at the hip or spine. INTRODUCTION: Calcium supplements, or supplements of calcium-rich foods, have a positive effect on bone mineral density (BMD). However, it is uncertain whether there are any residual benefits of calcium on BMD following cessation of supplementation. METHODS: In a previously published study, 323 healthy men were randomized to receive elemental calcium 600 mg/day (n = 108), calcium 1,200 mg/day (n = 108), or placebo (n = 107) over 2 years. Consenting men from the placebo and calcium 1,200 mg/day groups (85 and 87, respectively) were followed over the next 1-2 years (mean 20 months), off trial medication. RESULTS: In the core trial, BMD increased at all sites by 1.0-1.5% at 2 years in the group receiving calcium 1,200 mg/day, compared to the group receiving placebo. In post-trial follow-up, the calcium group has some residual benefit at the total body (0.41% above placebo; P = 0.04) but there was no significant between-group differences at other sites. CONCLUSION: Following cessation of calcium supplements in healthy men, there is a small residual benefit in total body BMD, but not at the hip or spine. This is unlikely to confer a clinically significant dividend in terms of ongoing fracture prevention.

The human C3a receptor is expressed on neutrophils and monocytes, but not on B or T lymphocytes.

The pathophysiological relevance of the complement split product C3a as a proinflammatory mediator is still ill defined. The expression pattern of the human C3a receptor (C3aR) can provide important clues for the role of this anaphylatoxin in inflammation. There is strong evidence for C3aR expression on basophils, and eosinophils, but additionally, only on tumor cell lines of leukemic or hepatic origin. It is unclear whether neutrophils also express the C3aR, but need a costimulus provided by eosinophils for certain biological responses, or whether neutrophils lack the C3aR and respond to C3a via a secondary stimulus generated by eosinophils, i.e., by an indirect mode. In the present study, polyclonal antiserum raised against the second extracellular loop of the C3aR was used to characterize C3aR expression on peripheral blood leukocytes. For high degree purification of neutrophils, a negative selection method was established that decreased the contamination with CD9(bright+) eosinophils down to <0.2%. Flow cytometric analyses, functional assays, and binding assays on highly purified neutrophils confirmed C3aR expression and coupling. Monocytes were identified as an additional C3aR-positive cell population of the peripheral blood. The expression of the C3aR on eosinophils could be confirmed. In contrast, the receptor could not be detected on unchallenged B or T lymphocytes (or lymphocyte-derived Raji cells).

Inhibition of DNA synthesis by a small-cell lung carcinoma-derived protein.

A tumor-derived factor that inhibits cellular DNA synthesis was identified. The factor was extractable from a small-cell lung carcinoma cell line grown in either chemically defined medium or nu/nu mice and inhibited tritiated thymidine ([3H]dThd) incorporation by tumor cell lines of autologous, allogeneic, and xenogeneic origins. The viability of nonproliferating cells from normal tissue was not affected. Tumor extract inhibitory activity was trypsin labile but was resistant to other proteases, neuraminidase, lipase, DNase, RNase, glucosidase, extremes of pH-temperature, and reducing conditions. Inhibitory activity was reversibly bound to helix pomatia lectin but not to lentil, wheat germ, or concanavalin A lectins. Purification by size-exclusion high-performance liquid chromatography yielded a bioactive unimodal 12-kilodalton (kd) peak. The bioactive 12-kd moiety could be eluted from sodium dodecyl sulfate-polyacrylamide gels. Redosing of populations of the T-lymphoblastoid cell line CEM achieved an early (24 hr) sustained depression of pulse [3H]dThd incorporation and ultimately led to decreased population density of factor-treated populations. DNA histogram analysis demonstrated no change in cell cycle phase distribution after factor treatment. 5-Bromo-2'-deoxyuridine (BrdUrd) vs. propidium iodide with the two-parameter Fluorescence-Activated Cell Sorter analysis showed relative inhibition of non-S-phase BrdUrd uptake at 24 hours. A cell-free DNA polymerase assay demonstrated significant inhibition of non-alpha-polymerase-associated DNA synthesis in factor-treated cells. These studies suggest that this tumor-derived inhibitor of DNA synthesis represents a class of cellular products involved in the autoregulation of growth by regulation of DNA synthetic activity.

Indirect leukocyte migration inhibition in breast cancer and benign breast disease patients by mouse mammary tumor virus grown in feline kidney cells.

Indirect migration inhibition assays were performed with normal and mammary tumor-bearing C3H/HeN mice and patients with breast disease to assess cellular immunity against three different mouse mammary tumor virus (MTV) preparations grown in feline kidney cell cultures and against a mouse-derived MTV preparation. MTV obtained after passage through feline kidney cells and the mouse-derived MTV were capable of eliciting macrophage migration inhibitory factor production by mouse spleen cells obtained from normal or mammary tumor-bearing C3H/HeN mice, thus demonstrating a similar degree of antigenicity of these preparations. In experiments with human breast cancer patients' leukocytes, leukocyte inhibitory factor (LIF) was produced by 32-50% of these patients in response to the mouse-derived MTV or to three different MTV preparations obtained after passage through feline kidney cells. A significant proportion (31-54%) of benign breast disease patients also reacted with both the mouse-derived and feline-derived MTV preparations. Patients with both malignant and benign breast disease, however, had a significantly different (P less than .05) pattern of reactivity to mouse- and feline-derived MTV preparations from that observed with normal donors. Finally, some LIF activity was also observed (but not statistically significant with the use of nonparametric analysis methods) when feline leukemia virus was used as antigen with these patients. The data suggest that both breast cancer and benign breast disease patients were reactive against antigens largely specific for MTV in the feline cells and, presumably, were not reactive against feline cellular components, although the second possibility cannot be completely ruled out.

In vitro studies of cell-mediated immunity to Moloney murine leukemia virus and Moloney leukemia-associated surface antigens.

Cell-mediated immunity to Moloney murine leukemia virus (M-MuLV) and to tumor-associated surface antigens of leukemia cells induced by the virus was studied with an in vitro migration inhibition factor assay. Spleen cells of C57BL/6N mice at Day 14 following inoculation with Moloney murine sarcoma virus, produced migration inhibition factor in response to M-MuLV. The Moloney murine sarcoma virus-immune spleen cells, however, did not respond to other murine type C viruses, to AKR and Rauscher viruses, or to murine mammary tumor virus. The immune spleen cells also responded specifically to purified glycoprotein with molecular weights of 69,000 and 71,000 and proteins with molecular weights of 30,000 and 12,000, but not to protein with a molecular weight of 10,000, of the homologous M-MuLV. Migration inhibition factor production was also observed in response to soluble 3 M KCl extracts of leukemia cells, MBL-2, induced by M-MuLV. Similarly, the immune spleen cells responded to membrane fractions purified from the MBL-2 cells. Comparable membrane fractions prepared from a Gross virus-induced leukemia, E male G2, and a radiation-induced leukemia, RL male 1, were not active. The tumor-associated surface antigens of MBL-2 membranes could be solubilized by the detergent, Nonident P-40. Thus, C57BL/6N mice inoculated with Moloney murine sarcoma virus developed cell-mediated immunity to envelope and some internal antigens of M-MuLV and also to tumor-associated surface antigens of a tumor induced by this leukemia virus.

Mediation of reduction of spontaneous and experimental pulmonary metastases by ricin A-chain immunotoxin 45-2D9-RTA with potentiation by systemic monensin in mice.

We developed a model to assess the therapeutic effects of the 45-2D9-ricin A-chain immunotoxin (RTA) on pulmonary metastases. The 45-2D9 mouse monoclonal antibody recognizes a Mr 74,000 glycoprotein highly expressed by rat fibroblasts transformed with the Kirsten sarcoma virus (transformed rat fibroblasts). These cells metastasize spontaneously and form lung colonies in nu/nu and irradiated BALB/c mice. Injection i.v. of 45-2D9-RTA specifically reduced formation of spontaneous pulmonary metastases and lung colonies originating from freshly disaggregated tumor cells or cultured cells. Antibody alone or mixed with unconjugated ricin A chain and an immunotoxin that recognizes a melanoma-associated antigen were ineffective. Unconjugated 45-2D9 antibody specifically blocked the 45-2D9-RTA activity in vivo. Administration of the lysosomotrophic agents ammonium chloride and chloroquine in vivo did not potentiate immunotoxin-mediated reduction in lung colonies although they were effective in vitro. Monensin potentiated 45-2D9-RTA activity in vitro and in vivo.

Analysis of expression of cell surface antigen Mr 74,000 phosphoglycoprotein in normal, oncogene-transformed, and neoplastic rat cell lines.

A Mr 74,000 phosphoglycoprotein (gp74) present on the surface of oncogene-transformed murine cells but not untransformed NIH 3T3 cells was previously identified with mouse monoclonal antibody 45-2D9. The original cell population used as the immunogen was found to consist of two cell populations. The purpose of this study was to characterize these cell populations; determine the distribution of gp74 on normal, transformed, and neoplastic cells; and to characterize the gp74 molecule. Southern hybridization studies of cloned cell populations demonstrated that the immunizing cell population consisted of c-Ha-ras-transfected NIH 3T3 cells and Kirsten sarcoma virus-transformed rat cells (TRF cells). TRF cells showed a high level of gp74 expression. We observed that the expression of gp74 was increased on chemically and spontaneously transformed rat cells compared to untransformed rat cells. No binding of monoclonal antibody 45-2D9 was detected to rat adult and fetal tissue. Immunoperoxidase staining, immunofluorescence flow cytometry, and immunoprecipitation analysis of dimethylbenz[a]anthracene-induced metastatic 13762NF rat mammary adenocarcinoma clonal sublines demonstrated an inverse relationship between gp74 expression and metastatic phenotype. gp74 was immunoprecipitated from two low and medium metastatic clonal sublines (MTC and MTF7), but not from highly metastatic clone MTLn3 cells. Biosynthetic labeling and immunoprecipitation studies demonstrated that gp74 was phosphorylated on serine residues and was not secreted from transformed cells. No detectable protein kinase activity in an immune complex assay was associated with this molecule. We conclude that increased gp74 expression by rat cells is associated with transformed and neoplastic cells.

Serum cholesterol during treatment of hypertension with diuretic drugs.

Twenty-three men had an increase in serum levels of total cholesterol and triglycerides after starting a diet to lower their serum cholesterol. They had simultaneously started therapy with, or increased the dosage of, chlorthalidone or hydrochlorothiazide for the treatment of hypertension. To evaluate a possible role of the diuretics in the increase in serum lipid concentrations, 11 of the men were randomly allocated to spironolactone therapy for two to four months. After receiving spironolactone, cholesterol levels decreased by 24 mg/dL, whereas cholesterol levels decreased by only 3 mg/dL in the 12 men still receiving chlorthalidone (P less than .05). The triglyceride level decreased by 58 mg/dL after the change to spironolactone therapy, whereas it decreased by 10 mg/dL during continued chlorthalidone treatment (P less than .05). When the 11 men again received chlorthalidone, their cholesterol levels increased 16 mg/dL, whereas cholesterol levels decreased by 11 mg/dL in men receiving uninterrupted chlorthalidone treatment (P less than .01). These observations suggest that spironolactone may be a preferable alternative to thiazide-type agents as first-line therapy for hypertension because of the more favorable influence on serum lipid concentrations.

The influence of osteophytes and aortic calcification on spinal mineral density in postmenopausal women.

The assessment of vertebral bone mineral density (BMD) in the anterio-posterior projection has become widely used in the management and prevention of osteoporosis. Recently, it has been demonstrated that the presence of spinal osteophytes has a major impact on measured BMD in men, thus casting doubt on the value of these BMD measurements. We have assessed the impact of osteophytic and aortic calcification on spinal and femoral BMD measurements in 130 normal postmenopausal women, aged 45-71 yr. Lateral lumbar spine radiographs were obtained in all subjects and graded separately (0-3) for osteophytes and aortic calcification. Both forms of calcification increased with age, and BMD of all sites was correlated positively with body weight and negatively with age. The correlation coefficients between BMD and calcification scores were nonsignificant. Multiple regression analysis, including weight, age, and calcification scores, demonstrated a small but significant effect of osteophyte score on lumbar BMD (partial r2 = 0.04; P = 0.012) and a similar trend for Ward's triangle and the trochanteric region (partial r2 = 0.02; P less than 0.06). The aortic calcification score remained nonsignificant. It is concluded that the influence of spinal osteophytes on lumbar BMD in postmenopausal women is substantially less than that in men and is, therefore, unlikely to interfere with BMD estimation in most subjects. The relationship between proximal femoral BMD and osteophyte score suggests a real relationship between skeletal density and degenerative joint disease, as has been demonstrated by others.

Premature hair graying and bone mineral density.

In a recent case-control study, premature hair graying was found to be associated with osteopenia, suggesting that this might be a clinically useful risk factor for osteoporosis. We report a reexamination of this possibility in 293 healthy postmenopausal women. Subjects experiencing onset of hair graying in their 20s tended to have lower bone mineral density throughout the skeleton (adjusted for age and weight) than those with onset of graying later in life. The same was true for those in whom the majority of their hair was gray by the age of 40 yr (n = 16), in whom bone density was reduced by 7% in the femoral neck, 8% in the femoral trochanter, and 4% in the total body (P < 0.05) when compared with those not prematurely gray. Bone density at the lumbar spine and Ward's triangle showed similar trends that were not significant. However, premature hair graying explained only 0.6-1.3% of the variance in bone mineral density within the population. We conclude that premature hair graying is associated with low bone density, but that its infrequency in the normal postmenopausal population leads to its accounting for only a tiny fraction of the variance of bone density.

Determinants of the rate of bone loss in normal postmenopausal women.

Despite a large number of studies assessing relationships between putative risk factors and bone density, it is not known which factors influence the rate of axial bone loss in normal postmenopausal women. We have examined the relationships between the rate of bone loss (delta BMD) and variables related to calcium metabolism, lifestyle, diet (calcium, sodium, caffeine, and protein), body composition, muscle strength, sex hormones, and spinal osteophytosis in 122 normal postmenopausal women participating in a 2-yr prospective randomized placebo-controlled trial of calcium supplementation. Univariate correlation coefficients indicated that delta BMD at most sites was inversely related to baseline BMD and positively related to rate of change in body weight (0.10 < r < 0.36) and fat mass (0.11 < r < 0.42) during the study. Lean mass and its rate of change showed no consistent relationship to delta BMD. There was no correlation between delta BMD and any of the lifestyle, muscle strength, dietary, or hormonal indices or with the severity of spinal osteophytosis. Multiple regression analysis indicated that delta BMD in the total body was directly related to fat mass (P < 0.0001), the rate of change in fat mass (P < 0.0001), the renal tubular reabsorption of calcium (P < 0.01), and calcium treatment (P < 0.01) and inversely to the initial BMD (P < 0.0001; r2 = 0.42; P < 0.0001). Similar effects were seen throughout the skeleton, although the fraction of the variance accounted for was less in the subregions, consistent with the lower precision of measurement of regional bone density. It is concluded that baseline bone density, fat mass, and renal calcium handling are important factors influencing bone loss in normal postmenopausal women.

Determinants of total body and regional bone mineral density in normal postmenopausal women--a key role for fat mass.

In order to determine which clinical, anthropometric, dietary, and biochemical variables are independent predictors of total and regional bone mineral density (BMD) in normal postmenopausal women, a cross-sectional study of 140 normal postmenopausal women has been carried out. Subjects were white, aged 45-71 yr (mean 58 yr), and had no history of disorders or medication use likely to influence bone or calcium metabolism. Multiple regression analysis was used to derive models for total and regional BMD in terms of the other variables measured. The analysis indicated that total body BMD was positively related to fat mass (P less than 0.0001), serum estrone (P = 0.0095) and age at menopause (P = 0.0165), and negatively related to age (P less than 0.0001), 24-h urine calcium (P = 0.0002), sex hormone-binding globulin (P = 0.0003), and serum alkaline phosphatase activity (P = 0.0029) (R2 = 0.61). Similar relationships were found in the subregions of the total body scans and in the lumbar spine and proximal femur, with insulin-like growth factor-1, parity, and age at menarche also being related to BMD at at least two of these sites. Lean body mass was not an independent correlate of BMD at any site once fat mass was taken into account. Muscle strength, physical activity, alcohol intake, and dietary intakes of calcium, sodium and protein did not emerge as significant predictors of BMD in this homogeneous group of postmenopausal women. We conclude that total body fat is the most significant predictor of BMD throughout the skeleton and this relationship is not explicable in terms of either estrone production in fat tissue or the dependence of skeletal load-bearing on fat mass. The mechanism underlying this relationship is an important question to be addressed in bone biology.

In Xenopus oocytes the human C3a and C5a receptors elicit a promiscuous response to the anaphylatoxins.

The Xenopus laevis oocyte has been widely utilized for cloning and functional expression of G-protein coupled receptors (GPCR). This system was used for the functional expression and characterization of the recently identified human C3a receptor. Complementary RNA from the human C3a receptor was transcribed in vitro and microinjected into Xenopus oocytes for functional characterization. A positive response to a synthetic C3a peptide agonist and to C3a, but not to platelet activating factor or fMetLeuPhe was detected. In addition, a response of approximately one third the amplitude obtained with C3a was obtained with rC5a. Conversely, oocytes co-injected with the C5a receptor and total RNA isolated from U937 cells responded to C5a as well as to C3a and the C3a synthetic peptide. A functional response with the anaphylatoxin C3a receptor in oocytes was dependent on co-injection of a pertussis toxin sensitive complementary human factor which could be supplied by co-injection of total RNA isolated from U937 cells. Oocytes expressing the anaphylatoxin C3a and C5a receptors responded to both agonists, in each case the response to the cognate ligand was substantially more robust than the response elicited by the other anaphylatoxin.

Cloning and characterization of a rabbit ortholog of human Galpha16 and mouse G(alpha)15.

A cDNA was cloned from a rabbit spleen cDNA library which encoded a G-protein alpha subunit peptide of 374 amino acids, that at the peptide level exhibited 86% and 79% identity with human Galpha16 and mouse G(alpha)15, respectively. The rabbit G(alpha)subunit cDNA was subcloned into a mammalian expression vector and transiently co-transfected into HEK-293 cells along with cDNAs encoding the human C3a, C5a, or nociceptin/orphanin FQ receptors. In all three cases the rabbit G alpha subunit behaved similarly to G(alpha)15 or G(alpha)16 and effectively coupled the transfected receptors to intracellular calcium mobilization pathways. By nucleotide sequence homology and functional activity the rabbit G(alpha) subunit appears to be the ortholog of human G(alpha)16 and mouse G(alpha)15.

Relationships between upper-arm anthropometry and soft-tissue composition in postmenopausal women.

The value of upper-arm anthropometry as a measure of lean- and fat-tissue masses in 140 normal postmenopausal white women was assessed by studying the relationships of these measures to total and regional fat, and lean masses measured by dual-energy x-ray absorptiometry. Midarm circumference (MAC) was highly correlated with total and regional fat masses (r = 0.85-0.89, P less than 0.0001) but less so with lean-tissue masses (r = 0.26-0.34). Triceps-skinfold thickness was also correlated with fat masses (r = 0.70-0.74, P less than 0.0001) but not with lean mass (r = 0.02-0.09). The derived index, arm muscle area (AMA), was less closely correlated with fat mass than was MAC (r = 0.59-0.61), but its correlation with lean-tissue mass was comparable to that for MAC (r = 0.37-0.43). Multiple-regression analysis confirmed that the anthropometric indices were more closely correlated with fat than with lean mass. It is concluded that all these indices are useful measures of fat mass but that none, including AMA, is a specific index of lean-tissue mass in normal postmenopausal women.

Conversion of murine Fabs isolated from a combinatorial phage display library to full length immunoglobulins.

The use of combinatorial Ig libraries displayed on the surface of bacteriophage has advantages over traditional hybridoma techniques for the generation of mAbs but in many instances full length Igs may be more desirable than Fab fragments. Two murine Fabs reactive with the human complement component C5a, recovered from a combinatorial library, were converted to full length IgG2a mAbs. The VH and VL domains of these antibodies were removed from the bacterial expression vector used for the combinatorial library construction, and subcloned into individual mammalian expression vectors containing the corresponding Ig heavy and light chain constant regions. The subcloning relied on 5' restriction endonuclease sites encoded by the oligonucleotide primers originally used to amplify the Ig cDNAs and 3' sites conserved in CH1 and C kappa. These vectors were co-transfected into COS cells yielding full length IgG2a versions of the anti-C5a antibodies. The mAbs, purified from the culture supernatant, retained the full activity of the Fabs, binding specifically to and neutralizing human recombinant C5a. Refined versions of the mammalian expression vectors have been constructed for single step conversion of murine recombinant Fabs, recovered from combinatorial libraries, to IgG2a mAbs.

Elevation of serum lipid levels during diuretic therapy of hypertension.

In a study attempting to improve coronary risk status, serum cholesterol and triglyceride levels were measured before and during treatment of 74 patients with mild primary hypertension. In 35 patients there was a satisfactory reduction in elevated blood pressure levels with diet therapy alone. In the remaining 39 patients a diuretic drug was required in addition to the diet. Diet therapy alone was followed by a decrease of 11 mg/100 ml in mean serum cholesterol (p less than 0.01 versus pretreatment value) and no change in serum triglyceride. The sue of diuretics was accompanied by an average increase of 11 mg/100 ml in serum cholesterol and of 34 mg/100 ml in serum triglyceride (p less than 0.01 versus pretreatment level for both). In a subgroup of 21 patients with greatest elevations in lipid levels during the administration of diuretics, little improvement in coronary risk status occurred because the increase in serum cholesterol balanced the decrease in systolic blood pressure, according to Framingham risk tables. If the level of serum lipids is a factor in the pathogenesis of coronary atherosclerosis then the observed effect of diuretic drugs to elevate serum cholesterol and triglyceride levels may explain, in part, the continuing high rate of occurrence of myocardial infarction during the treatment of hypertension.

Long-term effects of calcium supplementation on bone loss and fractures in postmenopausal women: a randomized controlled trial.

PURPOSE: To determine the long-term effects of calcium supplements or placebo on bone density in healthy women at least 3 years postmenopause. PATIENTS AND METHODS: Eighty-six women from our previously reported 2-year study agreed to continue on their double-blind treatment allocation (1 g elemental calcium or placebo) for a further 2 years, with 78 women (40 on placebo) reaching the 4-year end point. Median (interquartile range) dietary calcium intakes for the whole group were 700 mg (range 540 to 910) per day at baseline, 670 mg (range 480 to 890) per day at 2 years, and 640 mg (range 460 to 880) per day at 4 years. The bone mineral density (BMD) of the total body, lumbar spine, and proximal femur was measured every 6 months by dual-energy, x-ray absorptiometry. RESULTS: There was a sustained reduction in the rate of loss of total body BMD in the calcium group throughout the 4-year study period (P = 0.002), and bone loss was significantly less in the calcium-treated subjects in years 2 through 4 also (difference between groups 0.25% +/- 0.11% per year, P = 0.02). In the lumbar spine, bone loss was reduced in the calcium group in year 1 (P = 0.004), but not subsequently. There was, however, a significant treatment effect at this site over the whole 4-year period (P = 0.03). In the proximal femur, the benefit from calcium treatment also tended to be greater in the first year and was significant over the 4-year study period in the femoral neck (P = 0.03) and the trochanter (P = 0.01). Nine symptomatic fractures occurred in 7 subjects in the placebo group and 2 fractures in 2 subjects receiving calcium (P = 0.037). CONCLUSIONS: Calcium supplementation produces a sustained reduction in the rate of loss of total body BMD in healthy postmenopausal women.

The effect of the antiestrogen tamoxifen on bone mineral density in normal late postmenopausal women.

PURPOSE: To assess the effect of the antiestrogenic agent tamoxifen on bone mineral density in normal late postmenopausal women. METHODS: A randomized, double-blind, placebo-controlled trial was performed with 57 healthy, late postmenopausal women (mean 11 +/- 7 years since menopause). Subjects were assigned to take either tamoxifen 20 mg/d or placebo for 2 years. Total body, lumbar spine, and proximal femoral (femoral neck, Ward's triangle, trochanter) bone mineral densities were measured every 6 months using dual-energy x-ray absorptiometry. Serum and urine indices of bone turnover were measured at baseline, 6 months, and 2 years. RESULTS: In the women given tamoxifen, the mean bone mineral density of the lumbar spine increased by 1.4%, while that in the women given placebo declined by 0.7% (P < 0.01 for difference between groups). Total body bone mineral density declined in both groups, but less so in the tamoxifen-treated women (P < 0.05). At both sites, the effect of tamoxifen was maximal after 1 year, with no further separation of the groups thereafter. There was no significant effect of tamoxifen on bone mineral density in the proximal femur. Tamoxifen produced significant falls in serum alkaline phosphatase (P < 0.0001), ionized calcium (P < 0.0001), and phosphate (P < 0.01), and in urinary excretion of hydroxyproline, n-telopeptides, and calcium (P < 0.05 for each). CONCLUSIONS: In normal late postmenopausal women, tamoxifen at a dose of 20 mg/d exerts a small protective effect on bone mineral density, comparable in magnitude to that of calcium supplementation and less than that of either estrogen or the bisphosphonates. Tamoxifen is unlikely to supersede any of these therapies in the management of postmenopausal osteoporosis.

Hydrochlorothiazide reduces loss of cortical bone in normal postmenopausal women: a randomized controlled trial.

PURPOSE: Thiazide diuretics reduce urine calcium excretion and might therefore reduce postmenopausal bone loss. In some, but not all, case-control studies, their use has been associated with a reduced incidence of hip fractures. We studied the effects of hydrochlorothiazide on bone loss in normal postmenopausal women. SUBJECTS AND METHODS: We performed a randomized, double-blind, 2-year trial of the effects of hydrochlorothiazide (50 mg per day) and placebo on bone mineral density in normal postmenopausal women. Participants were not required to have either low bone mineral density or hypertension. Bone mineral density was measured using dual-energy x-ray absorptiometry. RESULTS: One hundred eighty-five women entered the study, of whom 138 completed 2 years of follow-up. In an intention-to-treat analysis, hydrochlorothiazide produced significant benefits on bone mineral density of the total body (between-group difference at 2 years of 0.8%, 95% confidence interval [CI]: 0.3% to 1.3%, P <0.0001), legs (0.9%, 95% CI: 0.2% to 1.7%, P <0.0001), mid-forearm (1.2%, 95% CI: 0.2% to 2.2%, P = 0.02), and ultradistal forearm (1.7%, 95% CI: 0.1% to 3.2%, P = 0.04). There was no effect in the lumbar spine (0.5%, 95% CI: -0.5% to 1.6%) or femoral neck (0.2%, 95% CI: 1.3% to 1.7%). The between-group changes tended to be greatest during the first 6 months, except in the mid-forearm where there appeared to be a progressive divergence. An as-treated analysis produced similar results. Urine calcium excretion and indices of bone turnover decreased in the thiazide group, but parathyroid hormone concentrations did not differ between the groups. Treatment was tolerated well. CONCLUSIONS: Hydrochlorothiazide (50 mg per day) slows cortical bone loss in normal postmenopausal women. It may act directly on bone as well as on the renal tubule. The small size of the effect suggests that thiazides may have a role in the prevention of postmenopausal bone loss, but that they are not an appropriate monotherapy for treating osteoporosis.