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Effects of glucocorticoids on vitamin D3 24-hydroxylase (CYP24A1) in Saos-2 cells and primary human osteoblasts
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Vitamin D is essential for bone function and deficiency in active vitamin D hormone can lead to rickets, osteomalacia or osteoporosis. Long-term treatment with glucocorticoids is known to result in osteoporosis and a substantially increased risk of fractures. Although the actions of vitamin D and glucocorticoids play important roles for bone function and in the development of osteoporosis, much remains unclear regarding the effects of these compounds in cells of the bone. In the current study, the human osteosarcoma Saos-2 cell line and primary human osteoblast-like cells were found to express mRNA for the vitamin D receptor as well as both activating and deactivating enzymes in vitamin D3 metabolism. These bone cells exhibited the CYP24A1-mediated 24-hydroxylation, involved in deactivation of the active vitamin D3 form. However, bioactivating vitamin D3 hydroxylase activities could not be detected in either of these cells. Several commonly used therapeutic glucocorticoids, including prednisolone, down regulated mRNA expression for the CYP24A1 gene as well as the CYP24A1-mediated 24-hydroxylase activity in both Saos-2 and primary human osteoblast-like cells. Prednisolone had the strongest suppressive effect on CYP24A1 expression. Results from experiments with a human CYP24A1 promoter-luciferase reporter gene in Saos-2 cells, co-transfected with the glucocorticoid receptor, showed that treatment with prednisolone significantly suppresses the CYP24A1 promoter activity. Thus, the results of the present study showed suppression by glucocorticoids on expression of CYP24A1 mRNA, CYP24A1-mediated metabolism and CYP24A1 promoter activity in human osteoblast-like cells. As part of the present investigation we examined if glucocorticoids are formed locally in Saos-2 cells. The experiments indicate formation of 11-deoxycortisol, a steroid that has glucocorticoid activity and is able to bind to the glucocorticoid receptor.

Our data showing suppression by glucocorticoids on CYP24A1 expression in human osteoblast-like cells suggest a previously unknown mechanism for effects of glucocorticoids in human bone, where these compounds may act by increasing the normal levels of active vitamin D.

National Category
Chemical Sciences Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-333914OAI: oai:DiVA.org:uu-333914DiVA, id: diva2:1158234
Available from: 2017-11-18 Created: 2017-11-18 Last updated: 2017-11-19
In thesis
1. Expression and regulation of steroid metabolizing enzymes in cells of the nervous and skeletal systems: Special focus on vitamin D metabolism
Open this publication in new window or tab >>Expression and regulation of steroid metabolizing enzymes in cells of the nervous and skeletal systems: Special focus on vitamin D metabolism
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Little is known about the mechanisms of vitamin D actions in the brain and bone. In this study, the metabolism of vitamin D and its regulation in various cell cultures of the nervous and skeletal systems were examined.

Human osteosarcoma Saos-2 cells, human primary osteoblasts (hOB) and murine motor neuron-like NSC-34 cells were found to express mRNA for all enzymes required in vitamin D3 metabolism as well as the vitamin D receptor (VDR) that mediates vitamin D actions. Also, production of 24,25-dihydroxyvitamin D3 was found in these cells. Studies on vitamin D metabolism in NSC-34 cells and in primary neuron-enriched cells from rat cerebral cortex indicate formation of a previously unknown major metabolite formed from 25-hydroxyvitamin D3. Evaluation of the NSC-34 cells suggests that this cell line could be a novel model for studies of neuronal vitamin D metabolism and its regulation by endogenous and exogenous compounds.

Treatment with glucocorticoids down regulated mRNA expression for the CYP24A1 gene in Saos-2 and hOB cells. Additionally, the glucocorticoid prednisolone showed suppression of CYP24A1-mediated metabolism and CYP24A1 promoter activity in Saos-2 cells. In NSC-34 cells, CYP24A1 mRNA levels were up-regulated by prednisolone, 1α,25-dihydroxyvitamin D3 and its synthetic analogues, EB1089 and tacalcitol. Formation of an endogenous glucocorticoid, 11-deoxycortisol, was observed in Saos-2 cells. Effects of glucocorticoids on the vitamin D system in bone cells may contribute to the adverse side effects in long-term treatment with glucocorticoids. Also, there may be a correlation between the administration of corticosteroids and adverse effects in the CNS.

Expression and effects of vitamin D on steroidogenic enzymes were studied in primary neuron-enriched rat cortex cells, primary rat astrocytes and human neuroblastoma SH-SY5Y cells. These different cell cultures all expressed CYP17A1, whereas only astrocytes expressed 3β-hydroxysteroid dehydrogenase (3β-HSD). 1α,25-Dihydroxyvitamin D3 suppressed mRNA levels and enzyme activity of CYP17A1 in SH-SY5Y cells and astrocytes. 1α,25-Dihydroxyvitamin D3 suppressed enzyme activity and mRNA levels of 3β-HSD in astrocytes. The results suggest that vitamin D-mediated regulation of CYP17A1 and 3β-HSD may play a role in the nervous system.

The results presented here contribute to our understanding of vitamin D metabolism and effects of glucocorticoids in the brain and bone.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2018. p. 58
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 242
National Category
Chemical Sciences Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-333920 (URN)978-91-513-0154-9 (ISBN)
Public defence
2018-01-18, B7:101a, BMC, Husargatan 3, Uppsala, 13:15 (Swedish)
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Available from: 2017-12-21 Created: 2017-11-19 Last updated: 2018-03-08

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Almokhtar, MokhtarWikvall, KjellUbhayasekera, S. J. Kumari A.Bergqvist, JonasNorlin, Maria
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