Therapeutic efficacy of 1alpha,25-dihydroxyvitamin D3 and calcium in osteopenic ovariectomized rats: evidence for a direct anabolic effect of 1alpha,25-dihydroxyvitamin D3 on bone

It is an important question for clinical therapy of osteoporosis with vitamin D metabolites whether these compounds exert their beneficial effects on the skeleton indirectly through an increase in intestinal calcium absorption or whether there is also a major direct component of action on bone. In this study, female 6-month-old Fischer rats were either ovariectomized (OVX) or sham operated. One month before surgery, all rats were placed on a diet containing 0.25% calcium and were kept on this diet throughout the study. Beginning 3 months post-OVX, groups of OVX rats orally received vehicle, a calcium supplement, low dose (0.025 microg/kg x day) or high dose (0.1 microg/kg x day) 1alpha,25-dihydroxyvitamin D3 [1,25-(OH)2D3], or combinations of low and high dose 1,25-(OH)2D3 with the calcium supplement. By 3 months postsurgery, pretreatment OVX controls had lost 74% and 37% of tibial and vertebral cancellous bone, respectively. Two-way factorial ANOVA showed that a 3-month treatment of osteopenic OVX rats with 1,25-(OH)2D3 dose dependently increased vertebral and tibial cancellous bone mass (P < 0.001 and P = 0.021, respectively) and trabecular width (P < 0.001). Furthermore, 1,25-(OH)2D3 increased serum calcium (P = 0.028) and urinary calcium excretion (P < 0.001) and reduced serum PTH levels (P < 0.001), osteoclast numbers (P < 0.001), and urinary collagen cross-links excretion (P < 0.001). Calcium supplementation alone was without therapeutic effect, and there was no significant two-way interaction between the individual treatment effects of 1,25-(OH)2D3 and calcium on bone mass. These data indicate that the anabolic effects of 1,25-(OH)2D3 in osteopenic OVX rats are mediated through a direct activity on bone.     

In vivo treatment with calcitriol (1,25(OH)2D3) reverses age-dependent alterations of intestinal calcium uptake in rat enterocytes

The vitamin D endocrine system has been involved in the impairment of intestinal calcium absorption during aging. Alterations in the nongenomic mechanism of calcitriol (1,25-dihydroxy-vitamin D3; [1, 25(OH)2D3] have been recently evidenced. In enterocytes isolated from aged rats, 1,25(OH)2D3 stimulation of Ca2+ channels through the cAMP/PKA pathway is blunted. We have now investigated whether in vivo administration of calcitriol to senescent rats reverses the absence of hormonal effects in isolated intestinal cells. In enterocytes from 20-24-month-old rats given 1,25(OH)2D3 for 3 days (30 ng/100 g bw/day), calcitriol (10(-10) M, 3-5 minutes) stimulated Ca2&plus uptake and intracellular cAMP to the same degree and protein quinase A (PKA) activity to a lesser degree than in enterocytes from young animals. Significantly higher basal levels of cAMP and PKA detected in enterocytes from old rats were not affected by prior injection of animals with 1,25(OH)2D3. When the aged rats were injected with 25(OH)D3, similar Ca2+ influx, cAMP, and PKA responses to in vitro stimulation with calcitriol were obtained. 1, 25(OH)2D3-dependent changes in Ca2+ uptake by enterocytes from both young and old rats treated with calcitriol were totally suppressed by the cAMP antagonist Rp-cAMPS, whereas the response to the agonist Sp-cAMPS was markedly depressed in aged animals. These results suggest that intestinal resistance to nongenomic 1,25(OH)2D3 stimulation of duodenal cell Ca2+ uptake develops in rats upon aging and show that in vivo administration of 1,25(OH)2D3 or its precursor to senescent rats restores the ability of the hormone to stimulate duodenal cell calcium influx through the cAMP messenger system.     

Homologous up-regulation of vitamin D receptors is tissue specific in the rat

1,25-dihydroxyvitamin D3 (1,25(OH)2D3) receptors (VDR) are expressed in multiple tissues within the body. VDR levels are increased by 1,25(OH)2D3 in intestine and kidney and in numerous cell models. The ability of 1,25(OH)2D3 to affect VDR levels in other target tissues in vivo was studied by assessing VDR levels by the 3H-1,25(OH)2D3 binding assay under varied physiological conditions in the rat. When compared with vitamin D-deficient (-D) controls, rats raised on a normal vitamin D-sufficient (+D) diet showed elevated VDR levels in kidney (391 +/- 53 vs. 913 +/- 76 fmol/g of tissue;p < 0.05), but not in testis, heart, or lung. Up-regulation of the VDR also occurred in kidney of +D rats 1 day after a single 100-ng dose of 1,25(OH)2D3 (454 +/- 43 vs. 746 +/- 113 fmol/mg of DNA; p < 0.05), but no changes were seen in intestine, testis, or lung. Because 1,25(OH)2D3-induced hypercalcemia may independently affect VDR regulation, 1,25(OH)2D3 was infused into -D rats, and normocalcemia was maintained by reduced dietary calcium intake. In this model, the renal VDR was again up-regulated (446 +/- 115 vs. 778 +/- 58 fmol/mg of DNA; p < 0.05), but VDR levels in testis and lung were unaffected. Scatchard analysis and tests of 1,25(OH)2D3 dose (1-100 ng/day for 7 days) and temporal (100 ng/day for 1-7 days) responsiveness further supported the tissue-specific nature of the homologous VDR regulation. Assay of VDR levels by L-1-tosylamido-2-phenylethyl chloromethyl ketone-3H-1,25(OH)2D3 exchange assay ruled out differences in endogenous 1,25(OH)2D3 occupancy as the basis for the observed differences in VDR regulation. Finally, coidentity of the VDR-like sites in kidney versus testis was confirmed by competitive binding analysis comparing their relative affinities for 25(OH)D3 versus 1,25(OH)2D3 (30.5 +/- 6.4 vs. 35.6 +/- 3.6 in kidney and testis, respectively) and by immunoblot analysis using a highly specific monoclonal anti-rat VDR antibody. Thus, under a wide variety of experimental conditions, homologous up-regulation of the VDR occurs in the rat kidney in vivo, but not in several other target tissues which do not regulate plasma calcium homeostasis. Moreover, this differential VDR regulation did not result from secondary changes in plasma calcium, from differential 1,25(OH)2D3 responsiveness in the various tissues, nor from differences in endogenous 1,25(OH)2D3 occupancy of the VDR. These studies thus establish that, in contrast to observations in vitro, the widely described phenomenon of homologous VDR up-regulation in kidney and intestine is not a universal property of 1,25(OH)2D3 target tissues in vivo in the rat.     

Serum 1alpha,25-dihydroxyvitamin D3 accumulates into the fracture callus during rat femoral fracture healing

1,25-dihydroxyvitamin D3 (1,25(OH)2D3) is thought to be an important systemic factor in the fracture repair process, but the mechanism of action of 1,25(OH)2D3 has not been clearly defined. In this study, the role of 1,25(OH)2D3 in the fracture repair process was analyzed in a rat closed femoral fracture model. The plasma concentration of 1,25(OH)2D3 rapidly decreased on day 3 and continued to decrease to 10 days after fracture. We assessed whether this decrease was based on the accelerated degradation or retardation of the synthesis rate of 1,25(OH)2D3, from 25(OH)D3. After radiolabeled 3H-1,25(OH)2D3 or 3H-25(OH)D3 was injected i.v. into fractured or control (unfractured) rats, the concentrations of 25(OH)D3 and 1,25(OH)2D3 metabolites were measured by HPLC. The plasma concentrations of these radiolabeled metabolites in fractured group were similar to those in control rats early after operation. However, radioactivity in the femurs of fractured rats was higher than that of the control group. Furthermore, the radioactivity was concentrated in the callus of the fractured group analyzed by autoradiography. 1,25(OH)2D3 receptor gene expression was detected early after fracture and, additionally, both in the soft and hard callus on days 7 and 13 after fracture. These results showed that the rapid disappearance of 1,25(OH)2D3 in the early stages after fracture was not due to either increased degradation or decreased synthesis of 1,25(OH)2D3, but rather to increased consumption. Further, these results suggest the possibility that plasma 1,25(OH)2D3 becomes localized in the callus and may regulate cellular events in the process of fracture healing.     

Lack of effect of hypomagnesemia on elevated plasma 1,25-dihydroxycholecalciferol in acutely phosphate-deprived rats

The effect of hypomagnesemia occurring in phosphate deprivation on elevated levels of plasma 1,25-dihydroxycholecalciferol (1,25(OH)2D3) was evaluated in rats. Phosphate deprivation was associated with an elevated plasma 1,25(OH)2D3, hypophosphatemia, and hypomagnesemia. With oral magnesium supplement during phosphate deprivation, plasma magnesium did not fall, but plasma inorganic phosphate fell and 1,25(OH)2D3 rose to comparable levels seen in phosphate-deprived rats lacking magnesium supplement. These results indicate that a rise in plasma 1,25(OH)2D3 during phosphate deprivation is not due to concomitant hypomagnesemia and support the postulate that phosphate deprivation per se is responsible for the elevated plasma 1,25(OH)2D3.     

The influence of vitamin A on the utilization and amelioration of toxicity of cholecalciferol, 25-hydroxycholecalciferol, and 1,25 dihydroxycholecalciferol in young broiler chickens

Three experiments were conducted to determine the influence of vitamin A on the utilization and amelioration of toxicity of cholecalciferol (vitamin D3), 25-hydroxycholecalciferol [25-(OH)D3], and 1,25-dihydroxycholecalciferol [1,25-(OH)2D3] in young broiler chicks. Two levels of vitamin A (1,500 and 45,000 IU/kg or 450 and 13,500 microg) were fed in all experiments. In Experiment 1, chicks were fed six levels of vitamin D3 (0, 5, 10, 20, 40, and 80 microg/kg). High dietary vitamin A decreased bone ash (P < 0.001), and increased the incidence of rickets (P < or = 0.02). Linear and quadratic responses to vitamin D3 levels were significant (P < 0.01) for body weight, bone ash, incidence and severity of rickets, and plasma calcium. In Experiment 2, six levels of 25-(OH)D3 (0, 5, 10, 20, 40, and 80 microg/kg) were added to the basal diet. Adding 25-(OH)D3 increased (P < 0.001) body weight, bone ash, and plasma calcium, and decreased rickets and plasma vitamin A. Adding 25-(OH)D3 overcame the reduction in bone ash produced by high dietary vitamin A showing a significant (P < 0.02) interaction. In Experiment 3, six levels of 1,25-(OH)2D3 (0, 2, 4, 8, 16, and 32 microg/kg) were added to the basal diet. High dietary vitamin A increased (P < 0.01) the incidence and severity of rickets. Adding 1,25-(OH)2D3 increased (P < 0.01) body weight, bone ash, plasma calcium, and reduced rickets and plasma and liver vitamin A. Adding 1,25-(OH)2D3 overcame the reduction in bone ash, and the increase in rickets produced by high vitamin A was significant (P < or = 0.05). These results indicate that high dietary vitamin A (45,000 IU/kg) interferes with the utilization of vitamin D3, 25-(OH)D3 and 1,25-(OH)2D3, increasing the requirement for each of them. Moreover, 45,000 IU/kg of dietary vitamin A ameliorated the potential toxic effects of feeding high levels of vitamin D3, 25-(OH)D3 and 1,25-(OH)2D3 to young broiler chickens. Further work is necessary to find the minimum levels of these vitamins needed to cause these effects.     

Ethnic origin and serum levels of 1alpha,25-dihydroxyvitamin D3 are independent predictors of coronary calcium mass measured by electron-beam computed tomography

BACKGROUND: Blacks have been found to have lower amounts of coronary calcium as well as higher levels of the osteoregulatory steroid 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] than whites. We sought to determine if racial differences in coronary calcium mass could be explained by differences in serum levels of 1,25(OH)2D3. METHODS AND RESULTS: We evaluated standard coronary risk factors, quantified coronary calcium mass with electron-beam computed tomography (EBCT), and measured serum 1,25(OH)2D3 with radioimmunoassay in 283 high-risk subjects (51 [180%] black, 232 [82%] white). Black subjects had lower masses of coronary calcium than whites (14 versus 47 mg; P=.003). Serum 1,25(OH)2D3 levels were slightly higher in blacks (41 versus 38 pg/mL; P=.05). Log 1,25(OH)2D3 levels were inversely proportional to log-transformed calcium mass (r=-.19; P=.001) in both races. Multivariate linear regression demonstrated that both black race (P=.02) and 1,25(OH)2D3 levels (P=.007) contributed inversely and independently to coronary calcium mass. However, an interaction term of racex1,25(OH)2D3 did not significantly contribute to coronary calcium mass, indicating that other undetermined factors in addition to 1,25(OH)2D3 are responsible for ethnic differences in coronary calcium mass. CONCLUSIONS: Both black race and serum levels of 1,25(OH)2D3 are independent negative determinants of coronary calcium mass. Nevertheless, diminished amounts of coronary calcium in blacks are not accounted for by higher 1,25(OH)2D3 levels.     

In vivo regulation of chromogranin A messenger ribonucleic acid in the parathyroid by 1,25-dihydroxyvitamin D: studies in normal rats and in chronic renal insufficiency

Chromogranin-A (CgA) and PTH are the two major secretory products of the parathyroid gland. In vitro, 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] increases CgA, but decreases PTH messenger RNA (mRNA) levels. We investigated the physiological significance of the induced changes in CgA expression by examining the effects of 1,25-(OH)2D3 on parathyroid CgA mRNA levels in vivo. Normal rats were injected with 1,25-(OH)2D3 at 48 and 24 h before blood sampling and isolation of both parathyroid glands. Parathyroid total RNA was extracted and CgA and PTH mRNA quantified by Northern blot analysis. CgA mRNA levels increased 1.6-, 3.2- and 5.6-fold, whereas PTH mRNA levels decreased by 37, 63 and 97%, respectively, with 1,25-(OH)2D3 doses of 10, 50, and 250 pmol/100 g BW. Parathyroid gland CgA expression also was examined in rats with mild chronic renal insufficiency, induced by a 5/6 nephrectomy 5 weeks earlier. Chronic renal insufficiency rats, fed normal chow, had elevated serum urea, creatinine, and PTH levels and reduced 1,25-(OH)2D3 but normal serum levels of calcium and phosphate. PTH mRNA levels were elevated 4-fold and CgA mRNA levels were 50% lower in the uremic animals. This indicates that the regulation of CgA expression in normocalcemic rats occurs at physiological 1,25-(OH)2D3 concentrations. In summary, increases and decreases in serum 1,25-(OH)2D3 levels are associated with corresponding increases and decreases in CgA mRNA levels in the parathyroid glands of rats. Therefore, this study is the first to demonstrate the physiological relevance of the earlier in vitro observations.     

In vivo dihydrotachysterol2 metabolism in normal man: 1 alpha- and 1 beta-hydroxylation of 25-hydroxydihydrotachysterol2 and effects on plasma parathyroid hormone and 1 alpha,25-dihydroxyvitamin D3 concentrations

It has recently been shown that in the rat, dihydrotachysterol (DHT) is extensively metabolized in the side-chain in vivo along pathways similar to those of vitamin D. In addition 25-hydroxy-DHT2 [25OHDHT2] is hydroxylated at C1, producing both 1 alpha- and 1 beta- hydroxy compounds. An in vivo study in 1988 demonstrated that in normal adult subjects receiving oral DHT2, plasma 1 alpha,25-dihydroxyvitamin D [1,25-(OH)2D] concentrations fell, but with unchanged plasma PTH levels. Down-regulation of 1,25-(OH)2D3 production by 25-(OH)DHT2 or some other unknown metabolite was also suggested as an explanation for these observations. To investigate whether either of the newly characterized 1 alpha,25- or 1 beta,25-(OH)2DHT2 was formed in vivo in normal man, DHT2 (approximately 1 mg/day, orally) was administered to healthy volunteers (three males and one female). Plasma was analyzed by high performance liquid chromatography and gas chromatography-mass spectrometry, demonstrating the formation of both 1 alpha,25- and 1 beta,25-(OH)2DHT2 in vivo in normal human subjects. Plasma levels of 1,25-(OH)2D3, PTH, ionized and total calcium, inorganic phosphate, and alkaline phosphatase were monitored. The plasma concentrations of DHT2, 25OHDHT2, and 1 alpha,25- and 1 beta,25-(OH)2DHT2 were measured by gas chromatography-mass spectrometry. In all volunteers, plasma ionized calcium increased slightly during DHT2 administration; 1,25-(OH)2D3 and PTH concentrations fell. Plasma levels of DHT2 and its metabolites rose over the same period. The average fall in the level of plasma 1,25-(OH)2D (60-70 pmol/L) was mirrored by a rise in the concentration of 1 alpha,25-(OH)2DHT2 (550 pmol/L). This ratio is appropriate, because it has previously been shown that in a reconstituted COS cell, 1 alpha,25-(OH)2DHT3 has roughly one tenth the potency of 1,25-(OH)2D3. At maximum concentration, the ratios of DHT2/25OHDHT2/1 beta,25-(OH)2DHT2/1 alpha,25-(OH)2DHT2 were approximately 10:1:2:0.1. The concentration of 1 beta,25-(OH)2DHT2 was greater than that of 25OHDHT2, and the ratio of 1 alpha,25- to 1 beta,25-(OH)2DHT2 (1:20) was substantially lower than that in rat plasma (3:10). The data presented here suggest that the active DHT2 metabolite in man is 1 alpha,25-(OH)2DHT2 and that the fall in plasma 1,25-(OH)2D seen during DHT therapy may be partly the result of suppressed PTH secretion.     

Influence of exogenous porcine growth hormone on vitamin D metabolism and calcium and phosphorus absorption in intact pigs

The influence of growth hormone (GH) on vitamin D metabolism and calcium and phosphorus absorption in vivo is not clear. We, therefore, measured calcium and phosphorus balance, plasma 1,25-dihydroxyvitamin D (1,25(OH)2D), and intestinal vitamin D-dependent calcium-binding protein (CaBP 9k) in intact growing pigs given exogenous GH. Six 10-week-old pigs were given two daily subcutaneous injections of 50 micrograms porcine GH/kg body weight for 2 months; six control pigs were given vehicle. They were all fed a diet containing 1.1% Ca, 0.6% P, and 1000 IU vitamin D3/kg. Apparent Ca and P absorption and retention were measured in a 10-day balance trial at the end of the 2 months. The plasma levels of Ca, P, 1,25(OH)2D, IGF-I, and GH were determined, and the duodenal and jejunal mucosal CaBP 9k content was measured at slaughter. The plasma Ca and P of GH-treated pigs were unchanged, but all aspects of mineral metabolism, including the plasma 1,25(OH)2D concentration (40%), Ca absorption and retention (70%), P absorption (33%) and retention (45%), and jejunal CaBP 9k (40%), were stimulated, in addition to an increase in the circulating IGF-I concentration.     

Influence of metabolic acidosis on serum 1,25(OH)2D3 levels in chronic renal failure

Metabolic acidosis has been shown to alter vitamin D metabolism. There is also evidence that calcium may modulate 1,25(OH)2D3 by a parathyroid hormone (PTH)-independent mechanism. To investigate the effect of rapid correction of chronic metabolic acidosis on serum 1,25(OH)2D3 levels by free calcium clamp in chronic renal failure, 20 patients with mild to moderate metabolic acidosis (mean pH 7.31 +/- 0.04) and secondary hyperparathyroidism (mean intact PTH 156.47 +/- 84.20 ng/l) were enrolled in this study. None had yet received any dialysis therapy. Metabolic acidosis was corrected by continuous bicarbonate infusion for 3-4 h until plasma pH was around 7.4, while plasma ionized calcium was held at the preinfusion level by calcium solution infusion during the entire procedure. The plasma pH, bicarbonate, total CO2, sodium, and serum total calcium levels were significantly increased while serum concentrations of alkaline phosphatase and albumin were significantly decreased after bicarbonate infusion. The plasma ionized calcium, potassium, serum magnesium, inorganic phosphorus, and 25(OH)D levels showed no significant change before and after bicarbonate infusion. The serum 1,25(OH)2D3 levels were significantly increased (38.66 +/- 11.77 vs. 47.04 +/- 16.56 pmol/l, p < 0.05) after correction of metabolic acidosis. These results demonstrate that rapid correction of metabolic acidosis raises serum 1,25(OH)2D3 levels in vitamin D-deficient chronic renal failure patients, and may underline the importance of maintaining normal acid-base homeostasis in the presence of secondary hyperparathyroidism in chronic renal failure.     

Severe deficiency of 1,25-dihydroxyvitamin D3 in human immunodeficiency virus infection: association with immunological hyperactivity and only minor changes in calcium homeostasis

The serum level of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D], the biologically most potent metabolite of vitamin D, is tightly regulated within narrow limits in human healthy adults. 1,25-(OH)2D deficiency is rare and is associated with disturbances in calcium and bone metabolism. We have previously reported a marked decrease in serum levels of 1,25-(OH)2D in human immunodeficiency virus (HIV)-infected patients. The present study was designed to further examine the causes and consequences of severe 1,25-(OH)2D deficiency in these patients. The design was a prospective cohort study. Fifty-four HIV-infected patients clinically classified according to the revised criteria from Centers for Disease Control and Prevention and healthy controls were studied. Parameters related to vitamin D and calcium metabolism as well as immunological and nutritional status were determined. Twenty-nine of the patients (54%) had serum levels of 1,25-(OH)2D below the lower reference limit, and 18 of these had undetectable levels. In contrast, HIV-infected patients had normal serum levels of 25-hydroxyvitamin D and vitamin D-binding protein. HIV-infected patients as a group had modestly depressed serum calcium and PTH levels. There were, however, no correlations between these parameters and serum levels of 1,25-(OH)2D. There were no differences in serum calcium or PTH levels or nutritional status when patients with severe 1,25-(OH)2D deficiency were compared to other patients, but patients with undetectable 1,25-(OH)2D had significantly elevated serum phosphate levels. Furthermore, patients with undetectable 1,25-(OH)2D levels were characterized by advanced clinical HIV infection, low CD4+ lymphocyte counts, and high serum levels of tumor necrosis factor-alpha (TNFalpha). We conclude that inadequate 1alpha-hydroxylation of 25-hydroxyvitamin D seems to be the most likely cause of 1,25-(OH)2D deficiency in HIV-infected patients, possibly induced by an inhibitory effect of TNFalpha. The low 1,25-(OH)2D and high TNFalpha levels observed may impair the immune response in HIV-infected patients both independently and in combination and may represent an important feature of the pathogenesis of HIV-related immunodeficiency. Markedly depressed 1,25-(OH)2D serum levels are also present in certain other disorders characterized by immunological hyperactivity. Thus, the findings in the present study may not only represent a previously unrecognized immune-mediated mechanism for induction of 1,25-(OH)2D deficiency in human disease, but may also reflect the importance of adequate serum levels of 1,25-(OH)2D for satisfactory performance of the immune system in man.     

Impaired vitamin D metabolism and response in spontaneously diabetic GK rats

Several studies of diabetes mellitus patients have demonstrated abnormalities in calcium, phosphate and vitamin D metabolism. In an earlier study, the authors reported impaired renal processing of phosphate in spontaneously diabetic GK rats, an animal model of type II diabetes mellitus. In the present study, which represents an extension of the earlier study, vitamin D metabolism and response are examined in 20-week-old GK rats. Serum 1,25-dihydroxyvitamin D [1,25-(OH)2D] was found to be lower in GK rats than in Wistar rats. After intraperitoneal administration of 0.5 micrograms/kg 1,25-(OH)2D, serum calcium increased in GK rats, but not in Wistar rats, while serum phosphate remained unchanged in GK rats, but increased in Wistar rats. Although serum 1,25-(OH)2D rose abruptly in 3 h and decreased thereafter in both GK and Wistar rats, the decrease in serum 1,25-(OH)2D at 6 h was more marked in GK rats than in Wistar rats. Serum 24,25-dihydroxyvitamin D was consistently higher in GK rats than in Wistar rats. Northern blotting and dot blotting with use of a cDNA probe for the 24-hydroxylase gene showed an increased expression of the gene in the kidney of GK rats. These results demonstrate impaired vitamin D metabolism in GK rats. Increased activity of 24-hydroxylase, in addition to impaired phosphate metabolism, may play a role in impaired vitamin D metabolism in GK rats.     

Vitamin D metabolism: update for the clinician

Vitamin D3 must undergo two hydroxylation steps before it becomes fully active: 25-hydroxylation in the liver and 1- or 24-hydroxylation in the kidney. Parathyroid hormone, serum phosphate, and serum calcium are important in regulation of renal production of 1,25-dihydroxy vitamin D3 (1,25-[OH]2D3) and 24,25-dihydroxy vitamin D3. An enzyme involved in renal hydroxylation is deficient or defective in patients with chronic renal failure, the Fanconi syndrome, vitamin D-dependent rickets, hypoparathyroidism, and pseudohypoparathyroidism. Altered vitamin D metabolism also occurs in various hepatic diseases, postmenopausal osteoporosis, and anticonvulsant osteomalacia. Recently, 1,25-(OH)2D3 was approved for treatment of renal osteodystrophy. In physiologic doses, it predictably corrects many of the clinical and biochemical abnormalities associated with this disorder.     

Partial prevention of active Heymann nephritis by 1 alpha, 25 dihydroxyvitamin D3

The hormone 1 alpha, 25 dihydroxyvitamin D3 (1,25(OH)2D3) has potent immunosuppressive effects in vitro. Recent publications also described a protective effect of the hormone in various animal models of immune-mediated diseases. To test its in vivo activity we induced active Heymann nephritis in Lewis rats that were either untreated or treated with 1,25(OH)2D3 or its synthetic 20-epi analogue, KH1060. Treatment with cyclosporine A (CsA) was used as an immunosuppressive control. In this nephrotic model the administration of 1,25(OH)2D3 (0.5 microgram/kg body weight) given on alternate days during the first 13 days after active immunization significantly reduced the proteinuria as measured by weeks 7-9. This reduction was comparable to the reduction observed in rats treated with CsA (20 mg/kg) on alternate days. A second series of experiments with 1,25(OH)2D3 confirmed these findings. The level of autoantibodies was found to be significantly suppressed during the treatment time in the CsA (20 mg/kg) group, whereas the limit of significance (P = 0.06) was reached in the 1,25(OH)2D3 (0.5 microgram/kg) group. The size of the immune deposits also was found to be substantially smaller in the groups that developed less proteinuria. The administration of 1,25(OH)2D3 transiently increased the mean serum calcium concentration with 2.5 mg/dl above the pretreatment values, and the urinary calcium excretion by a factor of 3-5 during the short treatment time. Treatment with the analogue KH1060 did not reduce the proteinuria significantly. Our experiments add evidence to the hypothesis that 1,25(OH)2D3 in pharmacological doses has immunosuppressive potency.     

EB 1089, a novel vitamin D analogue, has strong antiproliferative and differentiation inducing effects on cancer cells

EB 1089 is a novel vitamin D analogue which in vitro strongly inhibits the proliferation of U937 histiocytic lymphoma cells and MCF-7 breast cancer cells, with a potency of 50 to 100 times that of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]. Studies of c-myc and c-fos expression in MCF-7 cells and of differentiation markers in U937 cells show that growth inhibition by EB 1089 is accompanied by induction of differentiation. The ability of EB 1089 to affect calcium metabolism in vivo in rats is decreased, compared to 1,25(OH)2D3. This low calcemic effect combined with the strong biological effect on cancer cells in vitro, makes EB 1089 an interesting candidate for treatment of cancer.     

Changes in 1,25-(OH)2D3 synthesis and its receptor expression in spleen cell subpopulations of mice infected with LPBM5 retrovirus

This study examines the influence of chronic retroviral infection of mice with a LPBM5 virus mixture on the paracrine system involving immune cells and 1,25-(OH)2D3 in the spleen. Plasma ionized calcium, 25-(OH)D and 1,25-(OH)2D of infected mice were unchanged. In contrast, the specific binding of 1,25-(OH)2D3 to spleen cytosol and the number of monocyte/macrophages expressing 1,25-(OH)2D3 receptors (VDR) were markedly increased. The retroviral infection also influenced the local production of 1,25-(OH)2D3 in the spleen. It did not alter this production in monocyte/macrophages but increased that in isolated T cells. Isolated B cells in control mice did not produce 1,25-(OH)2D3, but they increased the ability of isolated T cells to produce this metabolite during coculture incubations. Infection altered this cell interaction as 1,25-(OH)2D3 production in infected T cells decreased when these cells were cocultured with infected B cells. Thus, chronic retroviral infection alters both the local vitamin D metabolism and VDR expression by immune cells in mice. These findings suggest close local interactions between 1,25-(OH)2D3 and immune system activation during retroviral infection.