Insulin modulates the stimulation of renal 1,25-dihydroxyvitamin D3 production by parathyroid hormone

Previous studies have shown that there is an impairment in renal production of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), the major biologically active metabolite of vitamin D3, in diabetes. This impairment is not due to a deficiency in the parathyroid hormone (PTH), a major stimulator of renal 1,25(OH)2D3 production. Therefore, we have investigated the capacity of PTH to stimulate 1,25(OH)2D3 production in insulin deficiency and with insulin replacement. Experiments were performed in rats fed a 0.6% calcium, vitamin D sufficient diet for 2 weeks. Thyroparathyroidectomy was performed on all rats. Rats to be rendered diabetic were injected with streptozotocin immediately after surgery. In non-diabetic rats, PTH administration significantly increased renal 1,25(OH)2D3 production (11 +/- 2 vs 46 +/- 5 pg/min/g; P less than 0.05). In diabetic rats, however, PTH caused only a modest increase in 1,25(OH)2D3 production (11 +/- 1 vs 19 +/- 4 pg/min/g; P less than 0.05). With insulin replacement, PTH stimulation of 1,25(OH)2D3 production was markedly increased over that seen in diabetic rats (48 +/- 12 vs 19 +/- 4 pg/min/g; P less than 0.05). PTH was equally effective in raising serum calcium, depressing serum phosphorus and tubular reabsorption of phosphate in non-diabetic as well as in diabetic rats. These results demonstrate that insulin is necessary for the maximal stimulation of renal 1,25(OH)2D3 production by PTH. However, insulin is not necessary for PTH action in terms of renal handling of phosphate and inducing hypercalcaemia. These results suggest multiple pathways for the action of PTH, only some of which are insulin requiring.     

Insulin permits parathyroid hormone stimulation of 1,25-dihydroxyvitamin D3 production in cultured kidney cells

Primary cultures of chick kidney cells in serum free medium respond to PTH with increased production of 1,25(OH)2D3 only when exposed to insulin. The response of 1,25(OH)2D3 is maximal at 5 ng bPTH (1-34) per ml and decreases at higher hormone concentrations. Increased 1,25(OH)2D3 synthesis is not evident after 30 minutes exposure to bPTH and is maximal at 4-6 hours of treatment. Insulin does not increase the cyclic AMP response to PTH suggesting that whatever permissive role it is playing occurs beyond the generation of cyclic AMP.     

Infusions of parathyroid hormone in ruminants: hypercalcemia and reduced plasma 1,25-dihydroxyvitamin D concentrations

The relationship between infused synthetic bovine PTH-(1-34) and plasma concentrations of minerals and vitamin D metabolites was studied in eight calves (150-230 kg) and two thyroparathyroidectomized goats. Calves were infused iv with saline for 15-20 h. Then, calves were infused with one of three types of solution for an additional 35-h period. Three of the eight calves received 3 ng/kg X min (group H), three received 0.75 ng/kg X min (group L), and the remaining two calves received control saline over a 33-h period (group C). Blood samples were taken every 4-6 h. Plasma calcium, phosphorus, hydroxyproline, and 1,25-dihydroxyvitamin D [1,25-(OH)2D] remained relatively constant in control calves. PTH infusions into calves in group H resulted in an increase in plasma calcium from 2.4 to a plateau of 3.0 mmol/liter. PTH infusion caused no change in plasma phosphorus, but increased urinary excretion of phosphorus. Infusion of PTH caused a moderate increase in urinary calcium excretion, followed by pronounced calciuria after PTH withdrawal. Plasma concentrations of 1,25-(OH)2D decreased from about 30 pg/ml at the start of infusion to undetectable levels (less than 5 pg/ml) at the end of the infusion and for 30 h thereafter. Similar, but less pronounced, changes in plasma calcium and 1,25-(OH)2D concentration were observed in group L. Hypocalcemia and hypophosphatemia developed in the two lactating goats after thyroparathyroidectomy, and plasma 1,25-(OH)2D concentrations were decreased. PTH infusion (3 ng/kg X min) corrected the hypocalcemia and hypophosphatemia and markedly raised plasma 1,25-(OH)2D concentrations. When calcium chloride was infused in addition to PTH, the resulting hypercalcemia (3 mmol/liter) was associated with a marked reduction in plasma 1,25-(OH)2D. We conclude that the concentration of calcium in plasma has the major regulatory role on plasma 1,25-(OH)2D concentrations in ruminant species when potentially conflicting signals, such as hypercalcemia and high PTH concentrations, are present simultaneously.     

Combined effects of dexamethasone and 1,25-dihydroxyvitamin D3 on parathyroid hormone secretion in cultured bovine parathyroid cells

The present studies investigate the effects of glucocorticoids on the function of the parathyroid glands using primary cultures of bovine parathyroid cells. Treatment of parathyroid cell cultures with dexamethasone for 48 h caused a dose-dependent stimulation of PTH secretion. The minimal concentration of dexamethasone required for a significant stimulation of PTH secretion was 0.1 nM. The stimulatory effect of dexamethasone on the secretion of PTH was found within 12 h of treatment with 100 nM dexamethasone. The steroids deoxycorticosterone and cortexolone, which do not have glucocorticoid activity were without effect of PTH secretion. Since glucocorticoids may modulate the effects of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] in other tissues, additional studies were performed to evaluate the interactions of glucocorticoids and 1,25-(OH)2D3. Addition of 1,25-(OH)2D3 to parathyroid cell cultures for 48 h significantly suppressed PTH secretion. In the presence of dexamethasone, however, 1,25-(OH)2D3 also significantly decreased PTH secretion, although it did not reduce PTH secretion to control levels. The treatment of parathyroid cell cultures with 100 nM dexamethasone did not affect the parathyroid cell content of 1,25-(OH)2D3 receptors. In summary, these studies indicate that glucocorticoids significantly increase the secretion of PTH in vitro. This stimulatory effect can be inhibited by 1,25-(OH)2D3. The parathyroid gland is an additional site of physiological antagonism of glucocorticoids and 1,25-(OH)2D3.     

Effect of experimental human magnesium depletion on parathyroid hormone secretion and 1,25-dihydroxyvitamin D metabolism

Magnesium (Mg) deficiency in man may result in hypocalcemia, impaired PTH secretion, and low serum concentrations of 1,25-dihydroxyvitamin D [1,25-(OH)2D]. To determine whether these changes are due to selective Mg depletion, we studied 26 normal subjects before and after a 3-week low Mg (less than 1 meq/day) diet. This diet induced Mg deficiency, as demonstrated by a fall in pre- to postdiet serum Mg levels from 0.80 +/- 0.01 to 0.61 +/- 0.02 mmol/L (P less than 0.001), an increase in Mg retention from 11 +/- 4% to 62 +/- 4% (P less than 0.001), and a fall in red blood cell free Mg2+ from 205 +/- 10 to 162 +/- 7 microM (P less than 0.001). Serum calcium (Ca) fell significantly from 2.36 +/- 0.02 to 2.31 +/- 0.03 mmol/L (P less than 0.05), and serum 1,25-(OH)2D fell from 55 +/- 4 to 43 +/- 3 pmol/L (P less than 0.05). PTH secretion was impaired, as demonstrated by a fall or no change in serum PTH in 20 of 26 subjects despite a fall in the serum Ca and Mg. In addition, an iv injection of Mg in eight subjects after the diet resulted in a significant rise in PTH from 15 +/- 2 to 19 +/- 2 ng/L (P less than 0.01), whereas a similar injection given to six of the subjects before the diet resulted in a significant fall from 28 +/- 5 to 13 +/- 3 ng/L (P less than 0.001). The fall in serum 1,25-(OH)2D may be due to both the decrease in PTH secretion and a renal resistance to PTH. PTH resistance was suggested, as no increase in serum 1,25-(OH)2D was observed in the six subjects in which the PTH concentration rose by mean of 68% after the diet. Also, the rise in serum 1,25-(OH)2D after a 6-h human PTH-(1-34) infusion was significantly less after Mg deprivation. The results demonstrate that mild Mg depletion can impair mineral homeostasis and may be implicated as risk factor for osteoporosis in disorders such as chronic alcoholism and diabetes mellitus, in which Mg deficiency and osteoporosis are both common.     

The role of insulin in the stimulation of renal 1,25-dihydroxyvitamin D synthesis by parathyroid hormone in rats

To evaluate the role of insulin in 1,25-dihydroxyvitamin D [1,25(OH)2D] production in response to PTH, 25-hydroxyvitamin D-1 alpha-hydroxylase activity in kidney homogenates as well as serum 1,25(OH)2D concentration was measured both after dietary calcium (Ca) deprivation and after PTH infusion in control and streptozotocin-diabetic rats. Although serum Ca and phosphate (Pi) levels did not change significantly after dietary Ca deprivation for 1 week, urinary cAMP excretion increased significantly, indicating that dietary Ca deprivation caused secondary hyperparathyroidism without a significant change in serum Ca level. In control rats, renal 1 alpha-hydroxylase activity increased markedly from 0.11 +/- 0.05 to 1.70 +/- 0.46 ng/300 mg tissue/20 min in parallel with the change in serum 1,25(OH)2D level from 121 +/- 8 to 360 +/- 54 pg/ml in response to Ca deprivation. In contrast, serum 1,25(OH)2D level (82 +/- 3 pg/ml) and 1 alpha-hydroxylase activity (0.07 +/- 0.02 ng/300 mg tissue.20 min) were lower in the diabetic rats on a normal Ca diet than those in control rats, and the increase in both 1,25(OH)2D level and 1 alpha-hydroxylase activity in response to Ca deprivation was suppressed in diabetic rats (136 +/- 24 pg/ml and 0.38 +/- 0.12 ng/300 mg tissue.20 min, respectively, after Ca deprivation). Insulin treatment of the diabetic rats restored the baseline levels of serum 1,25(OH)2D (125 +/- 14 pg/ml) and renal 1 alpha-hydroxylase activity (0.21 +/- 0.02 ng/300 mg tissue.20 min) as well as those after Ca deprivation (340 +/- 52 pg/ml and 2.05 +/- 0.30 ng/300 mg tissue.20 min, respectively). Furthermore, when control and diabetic rats were thyroparathyroidectomized and infused with a maximal stimulatory dose of PTH, the increase in serum 1,25(OH)2D and renal 1 alpha-hydroxylase activity in response to PTH was markedly inhibited in diabetic rats. In addition, the baseline levels of serum 1,25(OH)2D and renal 1 alpha-hydroxylase activity in thyroparathyroidectomized diabetic rats were not different from those in control rats. These results are consistent with the conclusion that insulin plays an important role in the regulation of renal 1 alpha-hydroxylase activity and serum 1,25(OH)2D levels in response to PTH.     

Correlations of serum concentrations of 1,25-dihydroxyvitamin D, phosphorus, and parathyroid hormone in tumoral calcinosis

The inherited metabolic disorder tumoral calcinosis is characterized by elevated serum phosphorus and 1,25-dihydroxyvitamin D [1,25-(OH)2D] levels and paraarticular calcific tumors. The pathogenesis of this disease is obscure, but an elevated renal phosphate reabsorption threshold and increased production of 1,25-(OH)2D are postulated as defects. We studied nine affected patients and found that both serum phosphorus and renal phosphate reabsorption threshold (TmP/GFR) were positively correlated with serum 1,25-(OH)2D levels. Since tumoral calcinosis is a disorder with abnormal renal phosphate transport, we compared the TmP/GFR and serum 1,25-(OH)2D levels to values obtained in patients with two other diseases with renal phosphate transport defects: oncogenic osteomalacia and X-linked hypophosphatemic rickets. We found a significant correlation between TmP/GFR and 1,25-(OH)2D levels in all three diseases, suggesting that in these diseases 1,25-(OH)2D production is regulated in some manner by phosphate transport. Furthermore, previous work indicated that in tumoral calcinosis broad variation exists in serum phosphorus levels. In our patients a negative correlation was found between the serum PTH concentrations and both serum phosphorus levels and TmP/GFR values, respectively. We postulate that although the basic defect in tumoral calcinosis most likely resides in the proximal renal tubular cell, the variation in serum phosphorus levels and possibly disease expression is modulated in part by PTH.     

Effects of parathyroid hormone and 1,25-dihydroxyvitamin D3 on tubular handling of phosphate in hypophosphatemic rickets

A controlled metabolic study to examine the effects of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) treatment on the renal handling of phosphate was conducted in nine patients with X-linked dominant hypophosphatemic rickets, including one with autonomous secondary hyperparathyroidism. Administration of 1,25(OH)2D3 resulted in uniform reduction in serum PTH from 63.6 +/- 14.7 (SD) to 49.3 +/- 14.8 muleq/ml (P less than 0.01), elevation of the tubular threshold for phosphate (TmP/GFR) from 1.41 +/- 0.30 to 1.90 +/- 0.31 mg/dl (P less than 0.01) and increase in serum phosphate from 2.6 +/- 0.7 to 3.4 +/- 1.1 mg/dl (P less than 0.01) in eight PTH-suppressible patients. Four patients treated with phosphate before and during the study (group A) excreted significantly more phosphate than those not treated with phosphate (group B) (P less than 0.001). In the control period, group A also had depressed TmP/GFR and higher concentrations of serum phosphate and PTH. With 1,25(OH)2D3 treatment, serum phosphate in group A became remarkably higher than in group B, 4.28 +/- 0.99 vs. 2.55 +/- 0.31 mg/dl (P less than 0.02), whereas serum PTH and TmP/GFR were similar in both groups. A good inverse linear correlation was found between mean serum PTH and mean TmP/GFR of the groups before and after treatment (r = 0.947); whereas, no correlation was found between TmP/GFR and serum calcium. The patient with autonomous secondary hyperparathyroidism, who was also treated with phosphate, had the lowest TmP/GFR. Administration of 1,25(OH)2D3 had no effect on the serum PTH and phosphate concentrations or on TmP/GFR. We conclude that in patients with X-linked dominant hypophosphatemic rickets PTH modulates to some extent the tubular handling of phosphate, and that the importance of this mechanism increases with therapeutic phosphate supplementation. Simultaneous administration of 1,25(OH)2D3 suppressed PTH activity, raised serum phosphate concentrations, and elevated TmP/GFR.     

A comparison between 1,25-dihydroxy-22-oxavitamin D3 and 1,25-dihydroxyvitamin D3 regarding suppression of parathyroid hormone secretion and calcemic action

1alpha,25-dihydroxy-22-oxavitamin D(3) (maxacalcitol;OCT), a vitamin D analogue with reduced calcemic activity, showed potency 10 times greater than 1alpha,25 (OH) (2)D(3) in differentiation induction of HL-60 cells. In addition, OCT showed immunomodulating activity, anti-proliferative effect and a suppressive effect on PTH secretion in vitro. OCT suppressed the expression of PTH mRNA in both normal and nephrectomized rats, and up-regulated VDR in the parathyroid gland in uremic rats to the same degree as 1alpha,25 (OH) (2)D(3). The results of a comparison between OCT and 1alpha,25 (OH) (2)D(3) indicate that the dissociation between efficacy and side effects with OCT is greater than with 1alpha,25 (OH) (2)D(3) in uremic rats. In future, we expect that OCT would contribute a great deal to the dialysis field in clinical.     

Inhibition of interleukin-1 production by 1,25-dihydroxyvitamin D3

The hormonal form of vitamin D, 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], inhibits the proliferation of T lymphocytes and production of growth-promoting factors (including interleukin-2) (IL2) in CTLL2 murine cells. In this study, we investigated the role of monocytes in this hormone-mediated inhibitory effect, by testing the effects of 1,25-(OH)2D3 on the ability of the mitogenic lectin phytohemagglutinin (PHA) to induce T cell activation in either a monocyte-dependent or phorbol myristate acetate (PMA)-driven (monocyte-independent) system. The results indicate that proliferation of T cells and production of growth-promoting factors are inhibited by 1,25-(OH)2D3 only in the monocyte-dependent system. Preincubation of monocytes with 1,25-(OH)2D3 for various periods of time and subsequent removal of the hormone resulted in inhibition of the PHA-driven proliferation of T cells. Preincubation for 2 h resulted in 20% inhibition, while preincubation for 36 h reduced proliferation to 50% of the control value [no 1,25-(OH)2D3 exposure]. These data suggested that monocytes are important participants in 1,25-(OH)2D3-mediated events. Therefore, we tested the effects of the hormone on the production of IL1, a monocyte-derived product thought to be involved in the induction of IL2 release and the subsequent development of the T cell proliferative response. 1,25-(OH)2D3 inhibited the production of both extracellular and cell-associated immunoreactive IL1 alpha and IL1 beta. Indomethacin, a prostaglandin synthetase inhibitor, did not alter the inhibitory properties of 1,25-(OH)2D3, suggesting that prostaglandins are not responsible for the inhibitory phenomenon. We conclude that part of the ability of 1,25-(OH)2D3 to inhibit T cell proliferation may be due to direct effects on monocytes by down-regulating IL-1 production. However, it is unlikely that the immunoregulatory properties of 1,25-(OH)2D3 on T cells are mediated solely through monocytes, and it is possible that the hormone also exerts its influence directly on T cells.     

Enhanced production rate of 1,25-dihydroxyvitamin D in sarcoidosis

We determined the metabolic clearance and production rates of 1,25-dihydroxyvitamin D [1,25-(OH)2D] in 5 patients with sarcoidosis who had either hypercalciuria or hypercalcemia to examine whether abnormalities in the metabolism of this hormone existed. The mean MCR in the 5 patients with sarcoidosis [40 +/- 9 (+/- SD) mL/min] was similar to that in 13 normal subjects (37 +/- 6 mL/min) and that in 9 patients with absorptive hypercalciuria and renal stones (35 +/- 4 mL/min). However, the mean serum 1,25-(OH)2D concentration was significantly higher in the patients with sarcoidosis (211 +/- 60 pmol/L) than in either of the other 2 groups. The mean 1,25-(OH)2D production rate was markedly elevated in the patients with sarcoidosis (12.4 +/- 5.3 mumol/day), being more than 2-fold greater than the normal mean value (5.4 +/- 1.2 mumol/day). The highest production rates were found in patients with hypercalcemia, whereas subjects with hypercalciuria had production rates comparable to those in the patients with absorptive hypercalciuria. These data indicate that there is no impairment in the clearance of 1,25-(OH)2D in patients with sarcoidosis and that the elevated serum 1,25-(OH)2D levels are due to an increase in its production rate.     

Effects of gonadal suppression on the regulation of parathyroid hormone and 1,25-dihydroxyvitamin D secretion in women.

Although a causal association between estrogen deficiency and bone loss has been established for many years, the mechanism by which estrogen deficiency leads to bone loss is unclear. Estrogen deficiency could induce bone loss either by a direct effect on bone cells to modify the production of bone-resorbing cytokines or by altering the production or response to calcium regulatory hormones such as PTH and 1,25-dihydroxyvitamin D. To assess the effects of ovarian hormones on calcium regulatory hormones, we evaluated the ability of calcium to suppress PTH secretion and the ability of PTH to increase serum 1,25-dihydroxyvitamin D and whole blood ionic calcium levels in women before and after GnRH analog-induced ovarian suppression. Sixteen women with endometriosis underwent i.v. infusion of calcium (1.1 mg calcium gluconate/cc in 5% dextrose) at a rate of 4 cc/kg x h (n = 7) or human PTH-(1-34) (Parathar) at a dose of 0.55 U/kg x h (n = 9) before and after 6 months of suppression of ovarian function with the GnRH analog nafarelin acetate (200 microg, intranasally, twice daily). Initial infusions were performed between days 6-10 of the menstrual cycle. Serum PTH and whole blood ionic calcium levels were measured at -20, -10, and 0 min and then every 10 min for 2 h during i.v. calcium infusions. Whole blood ionic calcium and 1,25-dihydroxyvitamin D levels were measured every 6 h for 24 h during i.v. human PTH-(1-34) infusions. Serum estradiol levels were markedly suppressed by nafarelin therapy in both groups of women. The relationship between whole blood ionic calcium and serum PTH levels was similar before and during nafarelin-induced ovarian suppression. The net change and rate of rise in serum 1,25-dihydroxyvitamin D levels in response to PTH infusion were similar before and during nafarelin therapy. Peak whole blood ionic calcium and incremental increases in ionic calcium in response to PTH were similar before and during nafarelin therapy. Our data suggest that ovarian suppression does not alter the regulation of PTH secretion in response to calcium, the ability of PTH to stimulate 1,25-dihydroxyvitamin D formation, or the skeletal sensitivity to PTH. These findings suggest that alterations in calcium regulatory hormones by estrogen deficiency are unlikely to play a major role in the pathogenesis of estrogen deficiency bone loss.     

1,25-dihydroxyvitamin D suppresses circulating levels of parathyroid hormone in a patient with primary hyperparathyroidism and coexistent sarcoidosis

CONTEXT: PTH is excessively secreted to develop hypercalcemia and accelerate bone turnover in patients with primary hyperparathyroidism. PTH stimulates the production of 1,25-dihydroxyvitamin D [1,25(OH)2D] that in turn suppresses the synthesis of PTH in parathyroid cells. OBJECTIVE: The objective of the study was to clarify whether 1,25(OH)2D indeed inhibits circulating levels of PTH and influences bone turnover, even in a patient with primary hyperparathyroidism. DESIGN, SETTING, AND PATIENT: We evaluated PTH levels in a patient with primary hyperparathyroidism and coexistent sarcoidosis whose serum 1,25(OH)2D levels were independent of PTH. INTERVENTIONS AND MAIN OUTCOME MEASURES: The present case was treated with prednisolone before and after surgical resection of parathyroid adenoma, and Ca-regulating hormones and bone markers were measured. RESULTS: Serum Ca and PTH levels significantly decreased after parathyroid surgery, whereas serum 1,25(OH)2D levels remained high. Prednisolone administration promptly decreased serum 1,25(OH)2D levels and reciprocally increased PTH levels despite consistent serum Ca levels either before or after surgery. PTH levels were negatively correlated with serum 1,25(OH)2D levels before and after surgery. Urine N-telopeptides, serum osteocalcin, and bone-type alkaline phosphatase all decreased to physiological ranges after parathyroid surgery. CONCLUSIONS: These results suggest that 1,25(OH)2D indeed inhibits the production of PTH not to exacerbate hypercalcemia in a patient with primary hyperparathyroidism. Furthermore, PTH but not 1,25(OH)2D may primarily be involved in the stimulation of bone turnover.     

Prolactin but not growth hormone stimulates 1,25-dihydroxyvitamin D3 production by chick renal preparations in vitro

We studied the effect of PRL from two species (bovine and turkey) and GH from two species (bovine and turkey) on 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] production by two whole cell preparations from vitamin D-deficient chick kidneys (slices and tubules). We observed that 8 ng/ml turkey PRL stimulated 1,25(OH)2D3 production by renal tubules and slices. Ovine PRL had a similar effect on 1,25(OH)2D3 production but at higher concentrations. In contrast, neither bovine GH nor turkey GH stimulated 1,25(OH)2D3 production appreciably at doses up to 1000 ng/ml. The effect of PRL on 1,25(OH)2D3 production by renal tubules required a 3-h preincubation, although its effect on 1,25(OH)2D3 production by renal slices was immediate. We conclude that PRL, but not GH, directly stimulates 1,25(OH)2D3 production by the chick kidney.     

Glomerular filtration rate and parathyroid hormone are associated with 1,25-dihydroxyvitamin D in men without chronic kidney disease

Abstract. Karhapää P, Pihlajamäki J, Pörsti I, Kastarinen M, Mustonen J, Niemelä O, Tuomi H, Kuusisto J (University of Eastern Finland, Kuopio; University of Tampere, Tampere; Finnish Medical Agency, Kuopio; and University of Tampere, Tampere, Finland). Glomerular filtration rate and parathyroid hormone are associated with 1,25‐dihydroxyvitamin D in men without chronic kidney disease. J Intern Med 2012; 271 : 573–580. Background and aim. Vitamin D, estimated glomerular filtration rate (eGFR) and parathyroid hormone (PTH) are related to cardiovascular disease risk. We examined the associations between the levels of 25‐hydroxyvitamin D (25‐D) and 1,25‐dihydroxyvitamin D (1,25‐D) and both eGFR and PTH. Design and setting. Cross‐sectional population‐based study in Kuopio, Eastern Finland. Subjects. A total of 909 men without known chronic kidney disease (CKD) and not receiving antidiabetic medication, aged from 45 to 73 years, were included in the study. Main outcome measures. Fasting levels of 25‐D, 1,25‐D, creatinine and PTH were measured, and an oral glucose tolerance test (OGTT) was performed. Results. High levels of 25‐D were associated with low levels of eGFR and PTH (β = ?0.17, P = 9 × 10^?7 and β = ?0.28, P = 6 × 10^?17, respectively, adjusted for age, body mass index and levels of calcium, phosphorus and glucose in a 2‐h OGTT, and also for either eGFR or PTH). By contrast, high 1,25‐D levels were associated with high levels of eGFR and PTH (β = 0.17, P = 2 × 10^?6 and β = 0.19, P = 5 × 10^?8, respectively, adjusted as mentioned earlier and additionally for 25‐D). Eighteen per cent of men in the highest 25‐D quartile were in the lowest 1,25‐D quartile and also had a lower eGFR than men with high levels of both 25‐D and 1,25‐D (P = 4 × 10^?5). Finally, 15% of men in the lowest 25‐D quartile were in the highest 1,25‐D quartile and also had higher PTH levels than men with low levels of both 25‐D and 1,25‐D (P = 2 × 10^?3). Conclusion. Our findings suggest that both eGFR and PTH are significantly associated with vitamin D metabolism in men without known CKD.     

Attenuation of thyroid hormone action by 1,25-dihydroxyvitamin D3 in pituitary cells

Interactions between thyroid hormone and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] were examined in a rat pituitary tumor cell line, GH4C1. Cells were incubated in thyroid hormone-depleted medium for 2 days, and specific nuclear binding of [125I]T3 was measured. 1,25-(OH)2D3 decreased nuclear [125I]T3 binding without changing total cellular uptake of [125I]T3. This 1,25-(OH)2D3 effect required 2-3 h to become evident and 24 h to reach a maximum (40-50% of control) and was reversible. Treatment with 1,25-(OH)2D3 for 8 h changed the maximal binding capacity for [125I]T3 from 80.2 +/- 2.9 to 50.3 +/- 6.3 fmol/10(6) cells, whereas Kd was not significantly altered. The decrease in [125I]T3 binding was dose dependent, with an IC50 for 1,25-(OH)2D3 of 1 nM in thyroid hormone-depleted medium. 1,25-(OH)2D3 caused little change in [125I]T3 binding to isolated nuclei, i.e. 1,25-(OH)2D3 does not compete directly with [125I]T3 for binding. It is unlikely that 1,25-(OH)2D3 decreased [125I]T3 binding by increasing the concentration of intracellular free calcium ([Ca2+]i), since 1,25-(OH)2D3 did not change [Ca2+]i in Indo-I-loaded GH4C1 cells. Two major species (6 and 2.6 kilobases) of mRNA for c-erb-A, which have been reported to encode nuclear thyroid hormone receptors, were found by Northern blot analysis, and both were decreased by treatment with 1,25-(OH)2D3 for 8 h. T3 (2 nM) caused a 3-fold increase in GH production over 72 h and 1,25-(OH)2D3 inhibited GH induction by T3, with an IC50 at approximately 1 nM. 1,25-(OH)2D3 stimulated PRL synthesis 5-fold when 10 nM T3 was present, but not when T3 was absent. In summary, 1,25-(OH)2D3 caused a dose-dependent down-regulation of nuclear thyroid hormone receptors at a pretranslational level and diminished GH induction by T3. These results suggest that 1,25-(OH)2D3 inhibits GH synthesis indirectly, at least partly, by attenuating endogenous thyroid hormone action.     

Constitutive expression of osteoclast-stimulating activity by normal clonal osteoblast-like cells: effects of parathyroid hormone and 1,25-dihydroxyvitamin D3.

Osteoblasts are known to produce osteoclast-stimulating activity (OSA). The aim of the current study was to relate the expression of OSA to the osteoblastic phenotype and examine its regulation by calciotropic hormones. The study was performed with the normal osteoblastic cell clone CRP 10/30 and the preosteoblastic clone CRP 4/7. OSA was determined with the well described isolated osteoclast pit assay, using sperm whale dentine as substrate. In contrast to previous studies, the assay was carried out at pH 7.36, rather than at pH 6.4 or 6.9. The results indicate that over 24 h, CRP 10/30 cells produce constitutively OSA, which compared to controls corresponds to an about 7-fold increase in resorption pits. There was considerably less activity expressed by either CRP 4/7 cells or fibroblasts. OSA proved to be heat labile, and its mol wt was estimated to be over 10 kilodaltons. While PTH-(1-34) did not influence the synthesis of OSA, the number of pits formed by osteoclasts incubated with medium conditioned by 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]-treated CRP 10/30 cells was increased 3-fold above baseline values. A similar increase was obtained with 1.25-(OH)2D3 added directly to CRP 10/30-conditioned medium. These results could not be duplicated with 1,25-(OH)2D3 added to either control medium or medium conditioned by CRP 4/7 cells or fibroblasts. The present study shows that normal clonal bone cells synthesize constitutively OSA, which is not regulated by PTH or 1,25-(OH)2D3. Furthermore, the results suggest that the synthesis of bone cell-derived OSA is limited to cells expressing the mature osteoblastic phenotype. Finally, CRP 10/30-conditioned medium appears to permit 1,25-(OH)2D3 to function on osteoclasts.