The effects of 1α,25-dihydroxyvitamin D3 on matrix metalloproteinase and prostaglandin E2 production by cells of the rheumatoid lesion

The biologically active metabolite of vitamin D₃, 1α,25-dihydroxyvitamin D₃ [1α,25(OH)₂D₃], acts through vitamin D receptors, which were found in rheumatoid tissues in the present study. IL-1β-activated rheumatoid synovial fibroblasts and human articular chondrocytes were shown to respond differently to exposure to 1α,25(OH)₂D₃, which has different effects on the regulatory pathways of specific matrix metalloproteinases and prostaglandin E₂.     

Transformation of vitamin D3 to 1α,25-dihydroxyvitamin D3 via 25-hydroxyvitamin D3 using Amycolata sp. strains

To enzymatically synthesize active metabolites of vitamin D₃, we screened about 500 bacterial strains and 450 fungal strains, of which 12 strains were able to convert vitamin D₃ to 1,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃] via 25-hyroxyvitamin D₃ [25(OH)D₃]. The conversion activity was only detected in strains belonging to the genus Amycolata among all the organisms tested. A preparative-scale conversion of vitamin D₃ to 25(OH)D₃ and 1,25(OH)₂D₃ in a 200-1 tank fermentor using A. autotrophica FERM BP-1573 was accomplished, yielding 8.3 mg 25(OH)D₃/l culture and 0.17 mg 1,25(OH)₂D₃/l culture. A related compound, vitamin D₂, could be also converted to 25-hydroxyvitamin D₂ and 1,25-dihydroxyvitamin D₂ using the same strain. The cytochrome P-450 of FERM BP-1573 was detected by reduced CO difference spectra in whole-cell suspensions. Vitamin D₃ in the culture induced cytochrome P-450 and the conversion activity simultaneously, suggesting that the hydroxylation at C-25 of vitamin D₃ and at C-1 of 25(OH)D₃ originates from cytochrome P-450.Correspondence to: J. Sasaki     

Transforming growth factor-β1 modulates the effect of 1α,25-dihydroxyvitamin D3 on leukemic cells

Summary The human leukemic cells HL-60, U937, KG-1 and THP-1 incubated with transforming growth factor-β1 (TGF-β1) were studied by examining cell surface antigens and macrophage-specific activities. The addition of 0.5 ng/ml (20 pM) of TGF-β1 with 1α,25-dihydroxyvitamin D₃ [1α, 25(OH)₂D₃] induced more Leu-M3 (CD14)-positive cells (approximately 80%) than 5×10⁻⁸ M 1α,25(OH)₂D₃ alone did (30 to 50%), although original HL-60 cells did not express any Leu-M3 antigen at all. Tumor necrosis factor-α (TNF-α) with TGF-β1 and 1α,25(OH)₂D₃ was found to potentiate the expression of these surface antigens. Furthermore, the phagocytic activity was also induced strongly. The expression of CR3 (CD11b) antigen was also increased, and all Leu-M3-positive cells were found CR3-positive when HL-60, U937, and THP-1 cells were treated with these stimulants. In contrast, CR3 but not Leu-M3 was induced in KG-1 cells after the same treatment. This may indicate that the responsiveness of leukemic cells to TGF-β1 and 1α,25(OH)₂D₃ might vary depending on a differentiation stage of the target cells. Furthermore, K562 cells originated from a more undifferentiated precursor, were not able to respond to these two inducers. These results suggested that some of TGF-β superfamily proteins might represent potent modulators in hematopoiesis, especially in the development of monocytes-macrophages or their precursors.     

Summer/winter differences in the serum 25-hydroxyvitamin D3 and parathyroid hormone levels of Japanese women

Serum 25-hydroxyvitamin D₃ [25(OH)D₃] is produced in the skin in response to exposure to ultraviolet radiation, and is a good indicator of vitamin D nutritional status. The aim of this study was to determine summer/winter differences in serum 25(OH)D₃ and parathyroid hormone (PTH) in Japanese women and how the summer and winter values are related. The subjects were 122 healthy Japanese women aged 45–81 years (average age: 65.7 years). They were medically examined twice, in September 1997 and February 1999. Serum 25(OH)D₃ and intact PTH were determined by high-performance liquid chromatography and a two-site immunoradiometric assay respectively. Lifestyle information was obtained through an interview. The seasonal differences (winter minus summer) in 25(OH)D₃ [Δ25(OH)D₃] and intact PTH concentrations were –18.8 nmol/l (SD 19.2, P<0.0001) and 0.98pmol/l (SD 1.02, P<0.0001) respectively. The correlation coefficient between summer (x) and winter (y) 25(OH)D₃ levels was 0.462 (P<0.0001), with a linearly fitted line of y=0.42x+26.4. This relationship was interpreted as subjects with higher summer 25(OH)D₃ values having greater reductions in winter 25(OH)D₃ concentrations. There were inter-individual differences in Δ25(OH)D₃, although the summer and winter 25(OH)D₃ concentrations were well-correlated. Since Δ25(OH)D₃ was not associated with any of the lifestyle factors, seasonal differences in the 25(OH)D₃ concentrations of an individual appeared to reflect her ability to produce 25(OH)D₃ photochemically in the skin. Sun bathing would be a less effective means of attaining adequate vitamin D nutritional status in a person with a small seasonal difference in 25(OH)D₃, i.e., one with a low 25(OH)D₃ level. Received: 17 December 1999 / Revised: 24 April 2000 / Accepted: 10 May 2000     

Comparative studies on the contents of vitamin D3, 25-hydroxy vitamin D3 and 7-dehydrocholesterol in fish liver

• 1.1. The contents of vitamin D₃, 25-hydroxyvitamin D₃ (25-OH-D₃) and 7-dehydrocholesterol (7-DHC) in 22 kinds of fish liver samples were determined by a high-performance liquid chromatographic (HPLC) method.
• 2.2. Vitamin D₃ was detected in all fish liver samples, but its contents varied from 84 to 264,000 ng/g wet tissue. The liver of fish belonging to Carangidae and Scombridae contained large amounts of the vitamin and therefore we deduced that vitamin D₃ levels in liver might have some relations with taxonomical positions of fishes.
• 3.3. 25-OH-D₃ was detected in 7 out of 22 kinds of fish liver samples, while 7-DHC was in 14 out of 22. The contents of the two sterols were generally much lower than those of vitamin D₃ and there was no special relationship between the contents of the sterols and the vitamin.

     

The vitamin D system in iguanian lizards

The apparent plasma concentration of vitamin D binding protein (DBP) in an iguanian lizard, Pogona barbata, and the affinity of this protein for 25-hydroxyvitamin D₃ (25(OH)D₃), 25-hydroxyvitamin D₂ (25(OH)D₂), and 1,25-dihydroxyvitamin D₃ (1,25(OH)D₃) was found to resemble more closely that of the domestic hen than that of the human. The characteristics of Pogona DBP, the pattern of vitamin D metabolites derived from injected radioactive vitamin D₃ and the plasma concentrations of endogenous 25-hydroxyvitamin D (25(OH)D) in a range of iguanian lizards have been examined. The findings suggest that 25-hydroxyvitamin D (25(OH)D) is the major metabolite of vitamin D, and that it may represent the storage form of vitamin D in these species in the same way as in mammals. High concentrations of vitamin D within iguanian embryos and egg yolks suggest a role for this compound in embryogenesis in these species, and perhaps indicates that there is a mechanism for vitamin D delivery to eggs comparable to that found in the domestic chicken.     

Synthesis and composition of vitamin D-3 metabolites in Solanum malacoxylon

The synthesis of vitamin D-3 hydroxylated metabolites in Solanum malacoxylon was investigated. When crude leaf homogenates and subcellular fractions were incubated with [³H]vitamin D-3 and [³H]25-hydroxy-vitamin D-3 under conditions described for animal vitamin D-3-25-hydroxylase and 25-hydroxy-vitamin D-3-1α-hydroxylase, respectively, labelled metabolites identified on the basis of their chromatographic properties as 25-hydroxy-vitamin D-3 and 1,25-dyhydroxy-vitamin D-3 were formed. Other unidentified product metabolites were also detected. Vitamin D-3-25-hydroxylase activity was localized in microsomes and 25-hydroxy-vitamin D-3-1α-hydroxylase in mitochondria and microsomes. Chromatography of sterols isolated from leaf extracts preincubated with β-glucosidase on Sephadex LH-20 columns permitted the isolation of three biologically active fractions with elution properties similar to vitamin D-3, 25-hydroxy-vitamin D-3 and 1,25-dihydroxy-vitamin D-3, respectively. Ultraviolet spectra characteristic of vitamin D-3 and its metabolites were obtained after purification of the fractions by TLC. Co-chromatography of individual fractions with authentic metabolites on TLC provided further evidence that the plant contains vitamin D-3, 25-hydroxy-vitamin D-3 and 1,25-dihydroxy-vitamin D-3 as glucoside derivatives. These results suggest that a similar pathway of vitamin D-3 hydroxylation as in animals may be operative in S. malacoxylon.     

1α,25-Dihydroxyvitamin D3 stimulates colony formation of chick embryo chondrocytes in soft agar

The effect of vitamin D metabolites on the growth of chick embryo chondrocytes in soft agar was examined. 1,25-Dihydroxyvitamin D₃ [1,25(OH)₂D₃]at 10⁻⁸-10⁻⁷ M induced colony formation by chick embryo chondrocytes in soft agar in the presence of 10% fetal bovine serum. Furthermore, 1,25(OH)₂D₃ increased the number of colonies in the presence of a maximal dose of basic fibroblast growth factor, a potent mitogen for chondrocytes in soft agar. However, 24R,25 (OH)₂D₃ and other metabolites had little effect on the soft agar growth of chondrocytes in the presence or absence of basic fibroblast growth factor. These results suggest that 1,25(OH)₂D₃ is an active metabolite which may be involved in supporting cartilage growth.     

Computer-aided de novo ligand design and docking/molecular dynamics study of Vitamin D receptor agonists

1α,25(OH)₂D₃, which is directly mediated by the vitamin D receptor (VDR), exerts a wide variety of biological actions. However, the treatment with 1α,25(OH)₂D₃ is limited because of its side effects. Many analogs and several nonsteroidal mimics with potent biological activity have been reported so far, and our rationale for designing the VDR agonists was on the basis of computer-aided drug design method by de novo design of A-ring and C/D-ring position of 1α,25(OH)₂D₃. Pyrimidine-2,4-diamine was selected as A-ring, and naphthalene and benzene were chosen as C/D-ring. By linking different components, a virtue compound library was obtained. To evaluate the contribution to activity of each component, we performed a series of automated molecular docking operations. Results revealed that the 19-dimethyl derivatives (the C-19 position correspond to C-20 in 1α,25(OH)₂D₃) show the favorable docking affinity to VDR. Moreover, the docking results are quite robust when further validated by molecular dynamics simulations. In addition, by free energy analysis using molecular mechanics-Poisson-Boltzmann surface area (MM-PBSA) method, the driving force of the binding between VDR and the ligands is proved to be hydrophobic interactions. Thus, a possible strategy to design new series of VDR agonists is proposed. The strategy can be successfully applied to explain the high potential activities of the 19-dimethyl derivatives. It is anticipated that the findings reported here may provide useful information for designing effective VDR agonists as well as the therapeutic treatment of VDR-related diseases.     

Specificity of the polyclonal antibodies raised against a novel 25-hydroxyvitamin D3-bovine serum albumin conjugate linked through the C-11α position

To obtain a specific antibody for use in 25-hydroxyvitamin D₃ [25(OH)D₃] immunoassay, a novel hapten-carrier conjugate was prepared by coupling 11α-hemiglutaryloxy-25(OH)D₃ with bovine serum albumin (BSA). Three polyclonal antibodies (Ab₁₁) showing high titer and affinity for 25(OH)D₃ (K_a = 0.96−2.6 × 10⁹ M⁻¹) were elicited in rabbits by repeated immunization with the conjugate. Specificity of the Ab₁₁ was investigated by cross-reactivities with 11 related compounds in a radioimmunoassay using a tritium-labeled antigen and compared with that of conventional antibodies (Ab₃) raised against 25(OH)D₃ 3-hemiglutarate conjugated with BSA. The Ab₃ could not discriminate the A-ring modified metabolites [1,25(OH)₂D₃ (87–290%) and 25(OH)D₃ 3-sulfate (S) (130–180%)], although the cross-reactivities with the side chain modified metabolites were satisfactorily low [24,25(OH)₂D₃ (2.3–7.4%), 25(OH)D₂ (1.1%)]. On the contrary, the Ab₁₁ easily discriminated 1,25(OH)₂D₃ (0.10–2.4%) and 25(OH)D₃ 3S (<0.3%), whereas significant cross-reactivities were found with 24,25(OH)₂D₃ (110–120%) and 25,26(OH)₂D₃ (66–130%) having a dihydroxylated side chain. These results show that the Ab₁₁ are complementary to the A-ring portion of the 25(OH)D₃ molecule which is opposite from the side chain structure recognized by the Ab₃. Thus, the Ab₁₁ will compensate for insufficient specificity of the Ab₃ and are expected to be a useful tool for the pretreatment of biological samples in the development of various analyses of vitamin D metabolites including specific 25(OH)D₃ immunoassays using the Ab₃.     

Metabolism of 1α,25-dihydroxyvitamin D2 by human CYP24A1

The metabolism of 1α,25-dihydroxyvitamin D₂ (1α,25(OH)₂D₂) by human CYP24A1 was examined using the recombinant enzyme expressed in Escherichia coli cells. HPLC analysis revealed that human CYP24A1 produces at least 10 metabolites, while rat CYP24A1 produces only three metabolites, indicating a remarkable species-based difference in the CYP24A1-dependent metabolism of 1α,25(OH)₂D₂ between humans and rats. LC-MS analysis and periodate treatment of the metabolites strongly suggest that human CYP24A1 converts 1α,25(OH)₂D₂ to 1α,24,25,26(OH)₄D₂, 1α,24,25,28(OH)₄D₂, and 24-oxo-25,26,27-trinor-1α(OH)D₂ via 1α,24,25(OH)₃D₂. These results indicate that human CYP24A1 catalyzes the C₂₄-C₂₅ bond cleavage of 1α,24,25(OH)₂D₂, which is quite effective in the inactivation of the active form of vitamin D₂. The combination of hydroxylation at multiple sites and C-C bond cleavage could form a large number of metabolites. Our findings appear to be useful to predict the metabolism of vitamin D₂ and its analogs in the human body.     

Vitamin D Promotes Odontogenic Differentiation of Human Dental Pulp Cells via ERK Activation

The active metabolite of vitamin D such as 1α,25-dihydroxyvitamin D₃ (1α,25(OH)₂D₃) is a well-known key regulatory factor in bone metabolism. However, little is known about the potential of vitamin D as an odontogenic inducer in human dental pulp cells (HDPCs) in vitro. The purpose of this study was to evaluate the effect of vitamin D₃ metabolite, 1α,25(OH)₂D₃, on odontoblastic differentiation in HDPCs. HDPCs extracted from maxillary supernumerary incisors and third molars were directly cultured with 1α,25(OH)₂D₃ in the absence of differentiation-inducing factors. Treatment of HDPCs with 1α,25(OH)₂D₃ at a concentration of 10 nM or 100 nM significantly upregulated the expression of dentin sialophosphoprotein (DSPP) and dentin matrix protein1 (DMP1), the odontogenesis-related genes. Also, 1α,25(OH)₂D₃ enhanced the alkaline phosphatase (ALP) activity and mineralization in HDPCs. In addition, 1α,25(OH)₂D₃ induced activation of extracellular signal-regulated kinases (ERKs), whereas the ERK inhibitor U0126 ameliorated the upregulation of DSPP and DMP1 and reduced the mineralization enhanced by 1α,25(OH)₂D₃. These results demonstrated that 1α,25(OH)₂D₃ promoted odontoblastic differentiation of HDPCs via modulating ERK activation.     

Culture requirements for optimal expression of 1α,25-dihydroxyvitamin D3-enhanced thyrotropin secretion

Summary An effect of the hormone, 1α,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃] on hormone secretion by normal rat pituitary cells was investigated in vitro. Based on previous findings using GH₄C₁ cells, dispersed anterior pituitary cell cultures were prepared and maintained in serum-free conditions for up to 6 d. Under these circumstances, there was no effect of 1,25(OH)₂D₃ to alter medium or cell-associated levels of thyrotropin (TSH), prolactin (PRL), or growth hormone (GH). Cultures maintained under these conditions had lower medium and cell-associated hormone levels and lesser responses to agonists than cultures maintained in serum-supplemented medium. In the presence of 10% charcoal-treated fetal bovine serum, treatment with 10⁻⁸ M 1,25(OH)₂D₃ for 24 h selectively increased TRH (10⁻¹⁰ to 10⁻⁷ M)-induced TSH secretion (P<0.001), with maximal enhancement observed at 10⁻⁹ M TSH-releasing hormone (TRH). Enhancement of TSH secretion by 1,25(OH)₂D₃ was detected after 15 min exposure to TRH. There was no effect on agonist-induced PRL or GH secretion or on cell-associated hormone levels. The effect was evident after 24 h treatment with 1,25(OH)₂D₃, and decreased thereafter. Several other steroid hormones had no effect on 10⁻⁹ M TRH-induced TSH secretion. These data contrast with the effect of 1,25(OH)₂D₃ in GH cells. They suggest that 1,25(OH)₂D₃ may act selectively in the normal pituitary to modulate TSH secretion.     

Simultaneous measurement of plasma vitamin D(3) metabolites, including 4β,25-dihydroxyvitamin D(3), using liquid chromatography-tandem mass spectrometry

Simultaneous and accurate measurement of circulating vitamin D metabolites is critical to studies of the metabolic regulation of vitamin D and its impact on health and disease. To that end, we have developed a specific liquid chromatography–tandem mass spectrometry (LC–MS/MS) method that permits the quantification of major circulating vitamin D₃ metabolites in human plasma. Plasma samples were subjected to a protein precipitation, liquid–liquid extraction, and Diels–Alder derivatization procedure prior to LC–MS/MS analysis. Importantly, in all human plasma samples tested, we identified a significant dihydroxyvitamin D₃ peak that could potentially interfere with the determination of 1α,25-dihydroxyvitamin D₃ [1α,25(OH)₂D₃] concentrations. This interfering metabolite has been identified as 4β,25-dihydroxyvitamin D₃ [4β,25(OH)₂D₃] and was found at concentrations comparable to 1α,25(OH)₂D₃. Quantification of 1α,25(OH)₂D₃ in plasma required complete chromatographic separation of 1α,25(OH)₂D₃ from 4β,25(OH)₂D₃. An assay incorporating this feature was used to simultaneously determine the plasma concentrations of 25OHD₃, 24R,25(OH)₂D₃, 1α,25(OH)₂D₃, and 4β,25(OH)₂D₃ in healthy individuals. The LC–MS/MS method developed and described here could result in considerable improvement in quantifying 1α,25(OH)₂D₃ as well as monitoring the newly identified circulating metabolite, 4β,25(OH)₂D₃.     

1α ,25-Dihydroxyvitamin D3 activates T cells of tumor bearers through protein phosphatase 2A

 The sterol 1α,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃] can inhibit T cell activation as well as restore the functional competence of suppressed T cells. The present studies determined whether 1,25(OH)₂D₃ had a differential effect on the activation of normal T cells or of suppressed T cells from mice bearing Lewis lung carcinoma tumors. Normal spleen cell proliferation in response to immobilized anti-CD3 was unaffected by the lower doses of 0.1 – 10 nM 1,25(OH)₂D₃, and was inhibited by the higher dose of 100 nM 1,25(OH)₂D₃. In contrast, 1,25(OH)₂D₃ increased proliferation and interferon γ secretion by T cells of tumor bearers in response to stimulation through T cell receptor/CD3. Assessment of mechanisms associated with the 1,25(OH)₂D₃ stimulation of tumor-bearer T cells implicated protein phosphatase 2A (PP-2A). First, PP-2A activity of spleen cells from tumor bearers was reduced compared to that of normal spleen cells but was increased by 1,25(OH)₂D₃. Second, 1,25(OH)₂D₃ stimulation of tumor-bearer T cell proliferation was dependent on this PP-2A activity as it was blocked by doses of okadaic acid that selectively inhibit PP-2A. These results suggest that 1,25(OH)₂D₃ preferentially enhances the responsiveness of immunosuppressed T cells from tumor bearers to TCR/CD3 stimulation by restoring PP-2A activity. Received: 7 November 1996 / Accepted: 2 January 1997     

Evaluation of vitamin D3 metabolites in Callithrix jacchus (common marmoset)

Vitamin D₃ (cholecalciferol) is endogenously produced in the skin of primates when exposed to the appropriate wavelengths of ultraviolet light (UV-B). Common marmosets (Callithrix jacchus) maintained indoors require dietary provision of vitamin D₃ due to lack of sunlight exposure. The minimum dietary vitamin D₃ requirement and the maximum amount of vitamin D₃ that can be metabolized by marmosets is unknown. Observations of metabolic bone disease and gastrointestinal malabsorption have led to wide variation in dietary vitamin D₃ provision amongst research institutions, with resulting variation in circulating 25-hydroxyvitamin D₃ (25(OH)D₃), the accepted marker for vitamin D sufficiency/deficiency. Multiple studies have reported serum 25(OH)D₃ in captive marmosets, but 25(OH)D₃ is not the final product of vitamin D₃ metabolism. In addition to serum 25(OH)D_3, we measured the most physiologically active metabolite, 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃), and the less well understood metabolite, 24,25-dihydroxyvitamin D₃ (24,25(OH)₂D₃) to characterize the marmoset's ability to metabolize dietary vitamin D₃. We present vitamin D₃ metabolite and related serum chemistry value colony reference ranges in marmosets provided diets with 26,367 (Colony A, N = 113) or 8,888 (Colony B, N = 52) international units (IU) of dietary vitamin D₃ per kilogram of dry matter. Colony A marmosets had higher serum 25(OH)D₃ (426 ng/ml [SD 200] vs. 215 ng/ml [SD 113]) and 24,25(OH)₂D₃ (53 ng/ml [SD 35] vs. 7 ng/ml [SD 5]). There was no difference in serum 1,25(OH)₂D₃ between the colonies. Serum 1,25(OH)₂D₃ increased and 25(OH)D₃ decreased with age, but the effect was weak. Marmosets tightly regulate metabolism of dietary vitamin D₃ into the active metabolite 1,25(OH)₂D₃; excess 25(OH)D₃ is metabolized into 24,25(OH)₂D₃. This ability explains the tolerance of high levels of dietary vitamin D₃ by marmosets, however, our data suggest that these high dietary levels are not required.     

Effects of 1,25(OH)2D3 and 25(OH)D3 on C2C12 Myoblast Proliferation,Differentiation, and Myotube Hypertrophy

An adequate vitamin D status is essential to optimize muscle strength. However, whether vitamin D directly reduces muscle fiber atrophy or stimulates muscle fiber hypertrophy remains subject of debate. A mechanism that may affect the role of vitamin D in the regulation of muscle fiber size is the local conversion of 25(OH)D to 1,25(OH)₂D by 1α‐hydroxylase. Therefore, we investigated in a murine C2C12 myoblast culture whether both 1,25(OH)₂D₃ and 25(OH)D₃ affect myoblast proliferation, differentiation, and myotube size and whether these cells are able to metabolize 25(OH)D₃ and 1,25(OH)₂D₃. We show that myoblasts not only responded to 1,25(OH)₂D₃, but also to the precursor 25(OH)D₃ by increasing their VDR mRNA expression and reducing their proliferation. In differentiating myoblasts and myotubes 1,25(OH)₂D₃ as well as 25(OH)D₃ stimulated VDR mRNA expression and in myotubes 1,25(OH)₂D₃ also stimulated MHC mRNA expression. However, this occurred without notable effects on myotube size. Moreover, no effects on the Akt/mTOR signaling pathway as well as MyoD and myogenin mRNA levels were observed. Interestingly, both myoblasts and myotubes expressed CYP27B1 and CYP24 mRNA which are required for vitamin D₃ metabolism. Although 1α‐hydroxylase activity could not be shown in myotubes, after treatment with 1,25(OH)₂D₃ or 25(OH)D₃ myotubes showed strongly elevated CYP24 mRNA levels compared to untreated cells. Moreover, myotubes were able to convert 25(OH)D₃ to 24R,25(OH)₂D₃ which may play a role in myoblast proliferation and differentiation. These data suggest that skeletal muscle is not only a direct target for vitamin D₃ metabolites, but is also able to metabolize 25(OH)D₃ and 1,25(OH)₂D₃. J. Cell. Physiol. 231: 2517–2528, 2016. © 2016 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.     

Biphasic action of prostaglandin E2 on conversion of 25-hydroxyvitamin D3 to 1,25-dihydroxyvitamin D3 in chick renal tubules

The effect of PGE₂ on the conversion of 25-hydroxyvitamin D₃ (25 OH D₃) to 1,25-dihydroxyvitamin D₃ (1,25- (OH) ₂D₃) by isolated renal tubules from vitamin D deficient chicks was studied under a variety of experimental conditions. In the absence of added vitamin D metabolites, PGE₂ (2 × 10⁻⁶M) caused an immediate inhibition of formation of 1,25-(OH) ₂D₃, followed by a delayed stimulation, apparent after 15 h exposure to PGE₂. Pretreatment of the tubules with 1,25-(OH) ₂D₃ prevented the immediate inhibitory action of PGE₂, and allowed the stimulation to be apparent after 4 h exposure to PGE₂. The cyclic nucleotide phosphodiesterase inhibitor 3-isobutyl-1-methyl xanthine (IBMX) significantly stimulated the formation of 1,25-(OH) ₂D₃. PGE₂ significantly inhibited 1,25-(OH) ₂D₃ formation in tubules which had been stimulated by IBMX. PGE₂ stimulated the adenylate cyclase activity in a crude particulate fraction from the chick kidney, and raised cyclic adenosine 3′, 5′-monophosphate (cyclic AMP) levels in the renal tubules.It is concluded that PGE₂ can either stimulate or inhibit 1,25-(OH) ₂D₃ formation in chick renal tubules. The stimulatory effect may be partly due to elevation of cyclic AMP. The mechanism of the inhibitory effect requires further investigation.