Possible involvement of pregnane X receptor–enhanced CYP24 expression in drug-induced osteomalacia
Jean Marc Pascussi, Agnes Robert, Minh Nguyen, Odile Walrant-Debray, Michèle Garabedian, Pascal Martin, Thierry Pineau, Jean Saric, Fréderic Navarro, Patrick Maurel, Marie Josè Vilarem
Jean Marc Pascussi, Agnes Robert, Minh Nguyen, Odile Walrant-Debray, Michèle Garabedian, Pascal Martin, Thierry Pineau, Jean Saric, Fréderic Navarro, Patrick Maurel, Marie Josè Vilarem
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Article Endocrinology

Possible involvement of pregnane X receptor–enhanced CYP24 expression in drug-induced osteomalacia

Abstract

Vitamin D controls calcium homeostasis and the development and maintenance of bones through vitamin D receptor activation. Prolonged therapy with rifampicin or phenobarbital has been shown to cause vitamin D deficiency or osteomalacia, particularly in patients with marginal vitamin D stores. However, the molecular mechanism of this process is unknown. Here we show that these drugs lead to the upregulation of 25-hydroxyvitamin D3-24-hydroxylase (CYP24) gene expression through the activation of the nuclear receptor pregnane X receptor (PXR; NR1I2). CYP24 is a mitochondrial enzyme responsible for inactivating vitamin D metabolites. CYP24 mRNA is upregulated in vivo in mice by pregnenolone 16α-carbonitrile and dexamethasone, 2 murine PXR agonists, and in vitro in human hepatocytes by rifampicin and hyperforin, 2 human PXR agonists. Moreover, rifampicin increased 24-hydroxylase activity in these cells, while, in vivo in mice, pregnenolone 16α-carbonitrile increased the plasma concentration of 24,25-dihydroxyvitamin D3. Transfection of PXR in human embryonic kidney cells resulted in rifampicin-mediated induction of CYP24 mRNA. Analysis of the human CYP24 promoter showed that PXR transactivates the sequence between –326 and –142. We demonstrated that PXR binds to and transactivates the 2 proximal vitamin D–responsive elements of the human CYP24 promoter. These data suggest that xenobiotics and drugs can modulate CYP24 gene expression and alter vitamin D3 hormonal activity and calcium homeostasis through the activation of PXR.

Authors

Jean Marc Pascussi, Agnes Robert, Minh Nguyen, Odile Walrant-Debray, Michèle Garabedian, Pascal Martin, Thierry Pineau, Jean Saric, Fréderic Navarro, Patrick Maurel, Marie Josè Vilarem

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In vivo modulation of CYP24 by PXR agonists. (A and B) Effect of pregnen...
In vivo modulation of CYP24 by PXR agonists. (A and B) Effect of pregnenolone 16α-carbonitrile and dexamethasone on cyp24 mRNA abundance. Mice (n = 5) were injected i.p. for 6 consecutive days with dexamethasone (DEX; 10 mg/kg/d), pregnenolone 16α-carbonitrile (PCN; 100 mg/kg), or corn oil (UT). Total RNA from liver or kidney was prepared, and 1 μg was reverse-transcribed. The relative levels of cyp3a11 and cyp24 mRNAs were determined in duplicate for each mouse by real-time PCR using cyp3a11-specific (A) and cyp24-specific (B) primers. Cyclophilin mRNA levels were used as a reference standard. Data are means ± SE of the ratio of mRNA levels in treated mice to corresponding levels in untreated mice, normalized with respect to cyclophilin mRNA levels. (C and D) Effect of pregnenolone 16α-carbonitrile on vitamin D3 metabolites in mouse plasma. Mice (n = 5) were injected i.p. for 6 consecutive days with pregnenolone 16α-carbonitrile (100 mg/kg) or corn oil and plasma samples. Pooled mouse plasma (1–2 mice, 300 μl, n = 3) or 50-μl plasma samples (n = 5) were analyzed for 24,25(OH)2D3 (C) or 25(OH)D3 metabolites (D), respectively, as described in Methods. Statistically significant expressions compared with untreated mice are marked with asterisks: *P < 0.05, **P < 0.01, and ***P < 0.005. Fold change relative to control mice is indicated.