A Multicenter Study to Evaluate Harmonization of Assays for C-Terminal Telopeptides of Type I Collagen (ß-CTX): A Report from the IFCC-IOF Committee for Bone Metabolism (C-BM).

BACKGROUND: Biochemical bone turnover markers are useful tools to assess bone remodeling. C-terminal telopeptide of type I collagen (ß-CTX) has been recommended as a reference marker for bone resorption in research studies. METHODS: We describe the results of a multicenter study for routine clinical laboratory assays for ß-CTX in serum and plasma. Four centers (Athens GR, Copenhagen DK, Liege BE and Sheffield UK) collected serum and plasma (EDTA) samples from 796 patients presenting to osteoporosis clinics. Specimens were analyzed in duplicate with each of the available routine clinical laboratory methods according to the manufacturers' instructions. Passing-Bablok regressions, Bland-Altman plots, V-shape evaluation method, and Concordance correlation coefficient for ß-CTX values between serum and plasma specimens and between methods were used to determine the agreement between results. A generalized linear model was employed to identify possible variables that affected the relationship between the methods. Two pools of serum were finally prepared and sent to the four centers to be measured in 5-plicates on 5 consecutive days with the different methods. RESULTS: We identified significant variations between methods and between centers although comparison results were generally more consistent in plasma compared to serum. We developed univariate linear regression equations to predict Roche Elecsys®, IDS-iSYS, or IDS ELISA ß-CTX results from any other assay and a multivariable model including the site of analysis, the age, and weight of the patient. The coefficients of determination (R2) increased from approximately 0.80 in the univariate model to approximately 0.90 in the multivariable one, with the site of analysis being the major contributing factor. Results observed on the pools also suggest that long-term storage could explain the difference observed with the different methods on serum. CONCLUSION: Our results show large within- and between-assay variation for ß-CTX measurement, particularly in serum. Stability of the analyte could be one of the explanations. More studies should be undertaken to overcome this problem. Until harmonization is achieved, we recommend measuring ß-CTX by the same assay on EDTA plasma, especially for research purposes in large pharmacological trials where samples can be stored for long periods before they are assayed.

Health effects associated with serum calcium concentrations: evidence from MR-PheWAS analysis in UK Biobank.

We conducted a phenome-wide Mendelian randomization analysis (MR-PheWAS) to survey health effects associated with high normal serum calcium. We found causal evidence for conditions related to renal function, bone and joint health, and cardiovascular risk. These conditions collectively suggest that tissue calcification may be a key mechanism through which serum calcium influences health. INTRODUCTION: Calcium is essential for the normal functioning of the cardiovascular system, muscles, and nerves. In this MR-PheWAS study, we sought to capture the totality of health effects associated with high normal serum calcium. METHODS: We used data from up to 337,535 UK Biobank participants, and tested for associations between calcium genetic score (calcium-GS) and 925 disease outcomes, with follow-up analyses using complementary MR methods. RESULTS: Calcium-GS was robustly associated with serum calcium concentration (F statistics = 349). After multiple testing correction (P < 1.62E-4), we saw genetic evidence for an association between high serum calcium and urinary calculus (OR per 1 mg/dl 3.5, 95%CI 1.3-9.2), renal colic (9.1, 95%CI 2.5-33.5), and allergy/adverse effect of penicillin (2.2, 95%CI 1.5-3.3). Secondary analyses with independent replication from consortia meta-analyses suggested further effects on myocardial infarction and osteoarthrosis. CONCLUSION: We found causal evidence for effects of high normal serum calcium with conditions related to renal function, bone and joint health, and cardiovascular risk, which may collectively reflect influences on tissue calcification and immune function.

Clinical usefulness of bone turnover marker concentrations in osteoporosis.

Current evidence continues to support the potential for bone turnover markers (BTM) to provide clinically useful information particularly for monitoring the efficacy of osteoporosis treatment. Many of the limitations identified earlier remain, principally in regard to the relationship between BTM and incident fractures. Important data are now available on reference interval values for CTX and PINP across a range of geographic regions and for individual clinical assays. An apparent lack of comparability between current clinical assays for CTX has become evident indicating the possible limitations of combining such data for meta-analyses. Harmonization of units for reporting serum/plasma CTX (ng/L) and PINP (µg/L) is recommended. The development of international collaborations continues with an important initiative to combine BTM results from clinical trials in osteoporosis in a meta-analysis and an assay harmonization program are likely to be beneficial. It is possible that knowledge derived from clinical studies can further enhance fracture risk estimation tools with inclusion of BTM together with other independent risk factors. Further data of the relationships between the clinical assays for CTX and PINP as well as physiological and pre-analytical factors contributing to variability in BTM concentrations are required.

Differential effects of 1,25-dihydroxyvitamin D on mineralisation and differentiation in two different types of osteoblast-like cultures.

In osteoblast cultures, 1,25-dihydroxyvitamin D (1,25D) has been shown to play either catabolic or anabolic roles on differentiation and mineralisation. We have employed osteoblast-like cells extracted from neonatal mouse calvariae and cells derived from juvenile mouse long bones to compare the biological effects of 1,25D on differentiation and mineralisation in vitro. 1,25D exerts differential effects on osteoblast-like cells depending on their stage of maturation and possibly their skeletal origin. This article is part of a Special Issue entitled 'Vitamin D Workshop'.

Determinants of insulin responsiveness in young women: Impact of polycystic ovarian syndrome, nitric oxide, and vitamin D.

BACKGROUND: Polycystic ovary syndrome (PCOS) is associated with incremental risk of atherosclerosis and possibly of cardiovascular events. Insulin resistance (IR) occurs frequently in PCOS subjects, which might be one of the mechanisms involved in engendering such risk. We sought to evaluate whether the impact of other factors potentially associated both with PCOS and with IR might differentially modulate degree of IR in women with and without PCOS. METHODS AND RESULTS: We measured body mass index (BMI), hs-CRP, plasma concentrations of asymmetric dimethylarginine (ADMA), vitamin D (25(OH)D3) levels and platelet responsiveness to nitric oxide donor sodium nitroprusside (NO responsiveness) in 47 young women (n=27 with PCOS and n=20 weight-matched controls) without metabolic syndrome, hypertension or overt cardiovascular disease. We performed univariate and multivariate regression analyses to establish correlates of the quantitative insulin-sensitivity check index (QUICKI), as a marker of IR. On univariate analysis, plasma 25(OH)D3 levels and low NO responsiveness tended to be direct correlates with QUICKI in the entire subject group. BMI, hs-CRP, and ADMA levels were significant inverse correlates of QUICKI in PCOS subjects, but not in subjects without PCOS. On multivariate analysis, NO responsiveness, and 25(OH)D3 levels, but not PCOS per se were significant correlates of QUICKI. CONCLUSIONS: In the entire cohort of young women, low NO responsiveness and vitamin D deficiency are associated with low QUICKI, while elevated ADMA, inflammatory activation and obesity are selectively associated with low QUICKI in PCOS subjects; this may contribute to the increased cardiovascular risk associated with this syndrome.

Circulating sex hormones and breast cancer risk factors in postmenopausal women: reanalysis of 13 studies.

BACKGROUND: Breast cancer risk for postmenopausal women is positively associated with circulating concentrations of oestrogens and androgens, but the determinants of these hormones are not well understood. METHODS: Cross-sectional analyses of breast cancer risk factors and circulating hormone concentrations in more than 6000 postmenopausal women controls in 13 prospective studies. RESULTS: Concentrations of all hormones were lower in older than younger women, with the largest difference for dehydroepiandrosterone sulphate (DHEAS), whereas sex hormone-binding globulin (SHBG) was higher in the older women. Androgens were lower in women with bilateral ovariectomy than in naturally postmenopausal women, with the largest difference for free testosterone. All hormones were higher in obese than lean women, with the largest difference for free oestradiol, whereas SHBG was lower in obese women. Smokers of 15+ cigarettes per day had higher levels of all hormones than non-smokers, with the largest difference for testosterone. Drinkers of 20+ g alcohol per day had higher levels of all hormones, but lower SHBG, than non-drinkers, with the largest difference for DHEAS. Hormone concentrations were not strongly related to age at menarche, parity, age at first full-term pregnancy or family history of breast cancer. CONCLUSION: Sex hormone concentrations were strongly associated with several established or suspected risk factors for breast cancer, and may mediate the effects of these factors on breast cancer risk.

Markers of bone turnover for the prediction of fracture risk and monitoring of osteoporosis treatment: a need for international reference standards.

UNLABELLED: The International Osteoporosis Foundation (IOF) and the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) recommend that a marker of bone formation (serum procollagen type I N propeptide, s-PINP) and a marker of bone resorption (serum C-terminal telopeptide of type I collagen, s-CTX) are used as reference analytes for bone turnover markers in clinical studies. INTRODUCTION: Bone turnover markers (BTM) predict fracture risk, and treatment-induced changes in specific markers account for a substantial proportion of fracture risk reduction. The aims of this report were to determine their clinical potential in the prediction of fracture risk and for monitoring the treatment of osteoporosis and to set an appropriate research agenda. METHODS: Evidence from prospective studies was gathered through literature review of the PUBMED database between the years 2000 and 2010 and the systematic review of the Agency for Healthcare Research and Quality up to 2001. RESULTS: High levels of BTMs may predict fracture risk independently from bone mineral density in postmenopausal women. They have been used for this purpose in clinical practice for many years, but there is still a need for stronger evidence on which to base practice. BTMs provide pharmacodynamic information on the response to osteoporosis treatment, and as a result, they are widely used for monitoring treatment in the individual. However, their clinical value for monitoring is limited by inadequate appreciation of the sources of variability, by limited data for comparison of treatments using the same BTM and by inadequate quality control. IOF/IFCC recommend one bone formation marker (s-PINP) and one bone resorption marker (s-CTX) to be used as reference markers and measured by standardised assays in observational and intervention studies in order to compare the performance of alternatives and to enlarge the international experience of the application of markers to clinical medicine. CONCLUSION: BTM hold promise in fracture risk prediction and for monitoring treatment. Uncertainties over their clinical use can be in part resolved by adopting international reference standards.

Systematic characterisation of the rat and human CYP24A1 promoter.

The biologically active form of vitamin D, 1,25-dihydroxyvitamin D (1,25D) ligands VDR (vitamin D receptor) and binds to the vitamin D response element (VDRE) located within target genes to regulate their transcription. Previously we showed that 1,25D-mediated rat CYP24A1 induction via the two critical VDREs is dependent on a short stretch of nucleotides called vitamin D stimulating element (VSE), located approximately 30bp upstream of VDRE-1 in the rat CYP24A1 promoter. We have now undertaken systematic analysis of the human CYP24A1 and rat CYP24A1 promoters to determine if the VSE is present in the human promoter. Using electrophoretic mobility shift and dual-luciferase reporter assays, we show that the VSE is absent in the human CYP24A1 promoter. In addition, we show that 1,25D-mediated induction of human CYP24A1 is dependant upon a promoter region spanning nucleotides -470 to -392 of the human CYP24A1 promoter.

The metabolism of 25-(OH)vitamin D3 by osteoclasts and their precursors regulates the differentiation of osteoclasts.

Current evidence suggests that levels of 25-(OH)vitamin D3 (25D), rather than 1alpha,25-(OH)2vitamin D3 (1,25D), directly affect bone mineralization and that the skeleton is a site of extra-renal synthesis of 1,25D. Since cells of the monocyte lineage can also metabolise 25D, it is possible that osteoclasts participate in local production of, and the response to, 1,25D. In this study, we investigated the effects of vitamin D metabolism on osteoclastogenesis using both the murine RAW 264.7 cell line and the human peripheral blood mononuclear cell (PBMC) models. PBMC-derived osteoclasts expressed cytoplasmic cyp27b1 and nuclear vdr proteins. PBMC expressed CYP27B1 mRNA, levels of which increased during RANKL induced differentiation into osteoclasts in both cell types. While 1,25D elicited a robust CYP24 transcriptional response in PBMC, the response to 25D was approximately 100-fold less at the concentrations used. Using media devoid of pre-existing vitamin D metabolites, we found that 25D was metabolised by RAW 264.7 cells to 1,25D and resulted in significant elevation in the numbers of TRAP-positive, multinucleated osteoclasts when present in the cultures for the first 3-5 days. These results suggest that vitamin D metabolism by osteoclast lineage cells is an important regulator of osteoclast formation.

Consumption of animal products, their nutrient components and postmenopausal circulating steroid hormone concentrations.

BACKGROUND/OBJECTIVES: Little is known about nutritional factors that influence circulating concentrations of steroid hormones, which are consistently associated with risk of breast cancer for postmenopausal women. We aimed to investigate the association between consumption of animal products and the plasma concentrations of steroid hormones and sex hormone-binding globulin (SHBG). SUBJECTS/METHODS: Cross-sectional analysis was conducted on plasma from 766 naturally postmenopausal women. We measured plasma concentrations of steroid hormones and SHBG, and estimated dietary intakes using a 121-item food frequency questionnaire. Log-transformed values of hormone concentrations were regressed on quartiles of intake of meat and dairy products among food items, and fats, proteins and cholesterol among nutrient intake. RESULTS: Total red and fresh red meat consumption was negatively associated with SHBG levels (P for trend=0.04 and <0.01, respectively). Mean SHBG concentrations were approximately 8% and 13% lower for women in the highest quartile compared with the lowest quartile of total red and fresh red meat consumption, respectively. Positive associations were observed between dairy product consumption and total and free estradiol concentrations (P for trend=0.02 and 0.03, respectively). Mean concentrations of total and free estradiol were 15 and 14% higher for women in the highest quartile of dairy product consumption than for those in the lowest quartile, respectively. No associations were observed with consumption of processed meat, chicken, fish, eggs, cholesterol, fats or protein. CONCLUSIONS: Our study suggests that greater consumption of total red and fresh red meat and dairy products might influence circulating concentrations of SHBG and estradiol, respectively. Confirmation and further investigation is required.

Traceability and standardization of immunoassays: a major challenge.

The clinical utility of immunoassay results is dependent on precision and trueness for the application of internationally agreed clinical protocols and common reference intervals and decision limits. The application of metrological principles to achieve traceability and standardization for these assays is being pursued to this end. Comprehensive measurement systems are available for a number of the total serum hapten assays routinely measured by immunoassays while current research is investigating the appropriateness of developing defined systems for the measurement of "free" hormone levels. The standardization or harmonization of assays for heterogeneous polypeptide hormones is also another area of research for these assays.

Regulation of the 5'-flanking region of the human CYP27B1 gene in osteoblast cells.

Synthesis of 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) is catalysed by the enzyme 25-hydroxyvitamin D(3)-1alpha-hydroxylase (CYP27B1). Regulation of CYP27B1 gene expression is poorly understood, particularly in non-renal tissues including bone where 1,25(OH)(2)D(3) is hypothesised to serve autocrine/paracrine roles. Transient transfection of ROS 17/2.8 osteoblast-like cells with reporter gene constructs containing deletions of the 5'-flanking region of the human CYP27B1 gene revealed a proximal promoter, enhancer region and strong upstream repressive region. Putative CCAAT and GC boxes, as well as Ets protein binding sites were shown to contribute to promoter and enhancer activities respectively in common with kidney and prostate cells. Inhibition of basal expression was largely attributed to a palindrome 5'-GTCTCAGAC-3' (-1015/-1007bp) that contains two putative canonical Smad binding elements. We conclude that repression of CYP27B1 gene expression may be a common event but the novel inhibitory elements we have identified may be unique to osteoblasts.

Relationship between calcium absorption and plasma dehydroepiandrosterone sulphate (DHEAS) in healthy males.

CONTEXT: Impaired gut sensitivity to 1,25-dihydroxyvitamin D (1,25(OH)(2)D), leading to reduced intestinal calcium absorption, has been reported in older men and women. While this phenomenon in postmenopausal women has been attributed to oestrogen deficiency, it is unclear whether the same observation in older men correlates with the age-related decline in androgen concentrations. OBJECTIVE: To examine the relationship between androgens and intestinal calcium absorption in older men. DESIGN: Cross-sectional study on 55 healthy male volunteers, divided into younger (n = 27) and older (n = 28) groups separated according to the median age of 59 years. MAIN OUTCOME MEASURES: Calcium absorption, total and free (calculated) testosterone, dehydroepiandrosterone sulphate (DHEAS), SHBG, and 1,25(OH)(2)D, among others, were measured. RESULTS: Calcium absorption, free testosterone and DHEAS, but not 1,25(OH)(2)D, declined significantly with age. After adjusting for age and body mass index, stepwise regression showed that 1,25(OH)(2)D and serum albumin were the only significant determinants of calcium absorption in younger men, while the sole determinant in older men was DHEAS, not testosterone. Residual deviations from the regression of calcium absorption on 1,25(OH)(2)D, reflecting the efficiency of 1,25(OH)(2)D-induced calcium absorption, was positively correlated with DHEAS (r = 0.27, P = 0.027). CONCLUSIONS: DHEAS is an independent determinant of calcium absorption in older men, although its manner of influence is, as yet, undefined. The age-related decline of DHEAS may, partly, account for the observed 'intestinal resistance to 1,25(OH)(2)D' in older men.

25-Hydroxyvitamin D requirement for maintaining skeletal health utilising a Sprague-Dawley rat model.

To study the role of vitamin D to optimise bone architecture, we have developed an animal model to investigate the effects of frank vitamin D-deficiency as well as graded depletion of circulating 25-hydroxyvitamin D(3) (25D) levels on the skeleton. Rats fed on dietary vitamin D levels from 0 to 500 ng/day achieved diet-dependent circulating levels of 25D ranging from 11 to 115 nmol/L. Levels of serum 1,25-dihydroxyvitamin D(3) (1,25D) increased as dietary vitamin D increased between 0 and 200 ng/day at which point a maximum level was achieved and retained with higher vitamin D intakes. The renal levels of 25-hydroxyvitamin D-1alpha-hydroxylase (CYP27B1) mRNA were highest in animal groups fed on vitamin D between 0 and 300 ng/day. In contrast, renal 25-hydroxyvitamin D 24-hydroxylase (CYP24) mRNA levels increased as dietary vitamin D increased achieving maximum levels in animals receiving 500 ng vitamin D/day. This animal model of vitamin D depletion is suitable to provide invaluable information on the serum levels of 25D and dietary calcium intake necessary for optimal bone structure. Such information is essential for developing nutritional recommendations to reduce the incidence of osteoporotic hip fractures.

RNAi-mediated silencing of CYP27B1 abolishes 1,25(OH)2D3 synthesis and reduces osteocalcin and CYP24 mRNA expression in human osteosarcoma (HOS) cells.

Although local synthesis of 1,25D has been postulated to regulate parameters of cell growth and differentiation in non-renal cells, the physiological role of 1,25D production in bone cells remains unclear. We used the technique of RNA interference to inhibit the mRNA encoding the enzyme responsible for 1,25D synthesis, 25-hydroxyvitamin D 1alpha-hydroxylase (CYP27B1). Human osteosarcoma (HOS) cells were transfected with siRNA for CYP27B1 or non-silencing RNA before being treated with 25D for 48h under normal growth conditions. De novo synthesis of 1,25D was measured in the media as well as mRNA levels for CYP27B1, osteocalcin (OCN) and 25-hydroxyvitamin D 24-hydroxylase (CYP24). We demonstrated that HOS cells express CYP27B1 mRNA, metabolize 25D and secrete detectable levels of de novo synthesized 1,25D. CYP27B1 mRNA silencing by RNAi, resulted in the suppression of 1,25D production and subsequent reduction of OCN and CYP24 mRNA expression. Our findings suggest that local 1,25D synthesis has paracrine effects in the bone microenvironment implying that vitamin D metabolism in human osteoblasts represents a physiologically important pathway, possibly regulating the maturation of osteoblasts.

Effects of amylin deficiency on trabecular bone in young mice are sex-dependent.

Amylin deficiency in mice results in late-onset osteopenia. Sex differences have been identified in insulin secretion in Amylin-overexpressing transgenic mice, suggesting a possible interaction of sex steroids, growth factors, or cytokines and amylin. The aim of the current study was to compare the effects of amylin deficiency on bone in young and adult male and female mice. The metaphyses of the distal femora from male and female Amylin-deficient mice at 4, 6, and 26 weeks of age were assessed by bone histomorphometry. Femoral length was increased in Amylin-deficient male mice compared to wild-type (WT) mice at 26 weeks of age (P < 0.005) but not in females. This was associated with an increase in growth plate height in Amylin-deficient males at 4 (P < 0.01) and 6 (P < 0.05) weeks of age. Furthermore, young Amylin-deficient males had decreased trabecular number at 4 weeks of age (P < 0.05) and increased trabecular thickness at 4 and 6 weeks of age (P < 0.05) compared to WT mice, with no net change in trabecular bone volume. These effects of amylin deficiency were not observed in female mice. In conclusion, this study demonstrates that amylin deficiency exerts effects on bone during growth that are sex-dependent and suggest a possible interaction between amylin and testosterone, growth factors, or cytokines to regulate bone cell metabolism.

The effect of calcitriol on fasting urine calcium loss and renal tubular reabsorption of calcium in patients with mild renal failure--actions of a permissive hormone.

AIM: Renal production of 1,25-dihydroxycholecalciferol is attenuated in early renal failure. Renal tubular reabsorption of calcium is diminished in moderate renal failure and we wished to see if this were true in the early stages and whether supplementary calcitriol would bring about correction. We were interested in the idea of 1,25-dihydroxycholecalciferol being a permissive agent, operating indirectly. METHODS: We measured calcium-related variables, including calculated ultrafiltrable serum calcium, before and after calcitriol 0.5 microg daily for six days in 34 subjects with stable mild renal failure. RESULTS: The mean serum creatinine was 0.21 (+/- 0.08) mmol/l. The mean serum Ca++ was normal (1.18 mmol/l) but nine patients had values outside the normal range and in six cases, with low-normal serum Ca++ levels, there was a diminished tubular reabsorption. In five cases, basal serum Ca++ was mildly elevated. The coefficient of variation for serum Ca++ was 4.4%. PTH (1-84) levels were mildly elevated and 1,25-dihydroxycholecalciferol levels low-normal. The urine Ca/Cr, representing net bone resorption, was elevated in six cases. After calcitriol, the mean serum Ca++ level rose slightly and the coefficient of variation decreased to 3.6%. Changes in Ca++ whether upward or downward were accounted for by minor alterations in tubular reabsorption and a tendency to less net bone resorption. The initial Ca++ predicted (negatively) the magnitude of the correction. Neither the prevailing PTH nor the 1,25-dihydroxycholecalciferol levels explained any of the observed changes. CONCLUSION: In early renal failure, there may be impaired regulation of serum Ca++. Despite elevated PTH, mild hypocalcemia may exist in the presence of increased net bone resorption relative to GFR. Hypocalcemia was accounted for by reduced renal tubular reabsorption of calcium which corrected after calcitriol. Net bone resorption tended to fall after calcitriol. Mild hypercalcemia, when present, was corrected by a reduction in tubular reabsorption. Calcitriol did not have a simple unidirectional effect but instead contributed to efficiency of the homeostatic mechanisms controlling the serum Ca++ set-point.

Response of the 5'-flanking region of the human 25-hydroxyvitamin D 1alpha-hydroxylase gene to physiological stimuli using a transgenic mouse model.

The enzyme 25-hydroxyvitamin D 1alpha-hydroxylase, or CYP27B1, is the key enzyme in the two-step activation process of vitamin D to 1,25-dihydroxyvitamin D (1,25D). While a number of regulators of the renal CYP27B1 enzyme activity have been recognized for some years, their underlying molecular mechanisms remain largely unknown, and the DNA regions involved in the in vivo regulation of gene expression by these factors have not been delineated. We have generated a transgenic mouse line that expresses 1501 bp of 5' flanking region together with 44 bp of 5' untranslated region of the human CYP27B1 gene fused to the firefly luciferase reporter gene. Animals expressing the luciferase gene demonstrated that both luciferase protein and mRNA for CYP27B1 were localized to proximal convoluted tubule cells of the kidney. In 2-week-old animals, the expression of the transgene and the endogenous CYP27B1 mRNA levels in the kidney were highest and fell with increasing age. Both reporter gene expression and CYP27B1 mRNA levels were downregulated in response to increasing amounts of dietary calcium in a dose-dependent manner. Vitamin D deficiency resulted in an increase in both the reporter gene and CYP27B1 expression. Interestingly, the increase in CYP27B1 mRNA levels was substantially higher than the increase in reporter gene expression, suggesting either that there is a post-transcriptional mechanism that increases the amount of CYP27B1 mRNA or that other regulatory elements are required to maximize the effect of vitamin D deficiency. These findings demonstrate that the 1501 bp 5' flanking region of the CYP27B1 gene directs expression to the proximal convoluted tubules of the kidney and is responsible for increasing transcriptional activity when dietary calcium and vitamin D levels are depleted. It also responds in the kidney to the physiological regulators of development and ageing.