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11?-HSD1 plays a critical role in trabecular bone loss associated with systemic glucocorticoid therapy.

ABSTRACT: BACKGROUND:Despite their efficacy in the treatment of chronic inflammation, the prolonged application of therapeutic glucocorticoids (GCs) is limited by significant systemic side effects including glucocorticoid-induced osteoporosis (GIOP). 11?-Hydroxysteroid dehydrogenase type 1 (11?-HSD1) is a bi-directional enzyme that primarily activates GCs in vivo, regulating tissue-specific exposure to active GC. We aimed to determine the contribution of 11?-HSD1 to GIOP. METHODS:Wild type (WT) and 11?-HSD1 knockout (KO) mice were treated with corticosterone (100??g/ml, 0.66% ethanol) or vehicle (0.66% ethanol) in drinking water over 4 weeks (six animals per group). Bone parameters were assessed by micro-CT, sub-micron absorption tomography and serum markers of bone metabolism. Osteoblast and osteoclast gene expression was assessed by quantitative RT-PCR. RESULTS:Wild type mice receiving corticosterone developed marked trabecular bone loss with reduced bone volume to tissue volume (BV/TV), trabecular thickness (Tb.Th) and trabecular number (Tb.N). Histomorphometric analysis revealed a dramatic reduction in osteoblast numbers. This was matched by a significant reduction in the serum marker of osteoblast bone formation P1NP and gene expression of the osteoblast markers Alp and Bglap. In contrast, 11?-HSD1 KO mice receiving corticosterone demonstrated almost complete protection from trabecular bone loss, with partial protection from the decrease in osteoblast numbers and markers of bone formation relative to WT counterparts receiving corticosterone. CONCLUSIONS:This study demonstrates that 11?-HSD1 plays a critical role in GIOP, mediating GC suppression of anabolic bone formation and reduced bone volume secondary to a decrease in osteoblast numbers. This raises the intriguing possibility that therapeutic inhibitors of 11?-HSD1 may be effective in preventing GIOP in patients receiving therapeutic steroids.

SUBMITTER: Fenton CG

PROVIDER: S-EPMC6698000 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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11β-HSD1 plays a critical role in trabecular bone loss associated with systemic glucocorticoid therapy.

Fenton C G CG Doig C L CL Fareed S S Naylor A A Morrell A P AP Addison O O Wehmeyer C C Buckley C D CD Cooper M S MS Lavery G G GG Raza K K Hardy R S RS

Arthritis research & therapy 20190816 1

<h4>Background</h4>Despite their efficacy in the treatment of chronic inflammation, the prolonged application of therapeutic glucocorticoids (GCs) is limited by significant systemic side effects including glucocorticoid-induced osteoporosis (GIOP). 11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) is a bi-directional enzyme that primarily activates GCs in vivo, regulating tissue-specific exposure to active GC. We aimed to determine the contribution of 11β-HSD1 to GIOP.<h4>Methods</h4>Wild type ...[more]

PMID: 31420008

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Project description:Postmenopausal osteoporosis (PMOP) is a severe health issue faced by postmenopausal women. microRNA-128 (miR-128) is associated with aging, inflammatory signaling, and inflammatory diseases, such as PMOP. It has also been reported to modulate in vitro osteogenic/adipogenic differentiation. However, its function in osteoclast formation is unknown. Methods: First, the expression of miR-128 and nuclear factor of activated T cells 1 (Nfatc1, bone resorption master marker) was investigated in bone tissues derived from PMOP patients, while their correlation to each other was also investigated. The levels of miR-128 and Nfatc1 in bone specimens and bone marrow-derived macrophages (BMMs) from mice subjected to ovariectomy (OVX) were also assayed. Next, we employed mice BMMs modified for overexpression and inhibition of miR-128 levels to determine its effect on osteoclast differentiation. Moreover, we generated osteoclastic miR-128 conditional knockout (miR-128Oc-/- ) mice and isolated miR-128 deletion-BMMs to observe its biological function on bone phenotype and osteoclastogenesis in vivo, respectively. The miR-128Oc-/- BMMs were used to explore the downstream regulatory mechanisms using pull-down, luciferase reporter, and western-blotting assays. Finally, the impact of miR-128 deficiency on OVX-induced bone loss in mice was evaluated. Results: The miR-128 level was found to be positively correlated with the increase in Nfatc1 level in mouse/human bone specimens and mouse primary BMMs. In vitro experiments demonstrated miR-128 levels that were dependent on activity of osteoclast differentiation and miR-128 overexpression or inhibition in BMMs significantly increased or decreased osteoclastogenesis, respectively. In vivo, we revealed that osteoclastic miR-128 deletion remarkedly increased bone mass through the inhibition of osteoclastogenesis. Mechanistically, we identified sirtuin 1 (SIRT1) as the direct target of miR-128 at the post-transcriptional level during osteoclast differentiation. Increased levels of SIRT1 reduced nuclear factor ?B (NF-?B) activity by decreasing the level of acetylation of Lysine 310, as well as inhibiting tumor necrosis factor-? (Tnf-?) and interleukin 1 (IL-1) expressions. Lastly, osteoclastic deletion of miR-128 significantly suppressed OVX-triggered osteoclastogenesis and exerted a protective effect against bone loss in mice. Conclusions: Our findings reveal a critical mechanism for osteoclastogenesis that is mediated by the miR-128/SIRT1/NF-?B signaling axis, highlighting a possible avenue for the further exploration of diagnostic and therapeutic target molecules in PMOP.

Project description:BackgroundIntracellular metabolism of glucocorticoid hormones plays an important role in the pathogenesis of metabolic syndrome and regulates, among many physiological processes, collagen metabolism in skin. At the peripheral level the concentration of active glucocorticoids is mainly regulated by the 11?-hydroxysteroid dehydrogenase 1 (11?-HSD1) enzyme, involved in the conversion of cortisone into the biologically active hormone cortisol. Cortisol interacts with the glucocorticoid receptor and regulates the expression of different classes of genes within the nucleus. Due to its implication in glucocorticoid metabolism, the inhibition of 11?-HSD1 activity has become a dominant strategy for the treatment of metabolic syndrome. Moreover, inhibitors of this target enzyme can be used for development of formulations to counteract skin ageing. Here we present the construction of two yeast cell based assays that can be used for the screening of novel 11?-HSD1 inhibitors.ResultsThe yeast Saccharomyces cerevisiae is used as a host organism for the expression of human 11?-HSD1 as well as a genetically encoded assay system that allows intracellular screening of molecules with 11?-HSD1 inhibitory activity. As proof of concept the correlation between 11?-HSD1 inhibition and fluorescent output signals was successfully tested with increasing concentrations of carbenoxolone and tanshinone IIA, two known 11?-HSD1 inhibitors. The first assay detects a decrease in fluorescence upon 11?-HSD1 inhibition, whereas the second assay relies on stabilization of yEGFP upon inhibition of 11?-HSD1, resulting in a positive read-out and thus minimizing the rate of false positives sometimes associated with read-outs based on loss of signals. Specific inhibition of the ABC transporter Pdr5p improves the sensitivity of the assay strains to cortisone concentrations by up to 60 times.ConclusionsOur yeast assay strains provide a cost-efficient and easy to handle alternative to other currently available assays for the screening of 11?-HSD1 inhibitors. These assays are designed for an initial fast screening of large numbers of compounds and enable the selection of cell permeable molecules with target inhibitory activity, before proceeding to more advanced selection processes. Moreover, they can be employed in yeast synthetic biology platforms to reconstitute heterologous biosynthetic pathways of drug-relevant scaffolds for simultaneous synthesis and screening of 11?-HSD1 inhibitors at intracellular level.

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11?-HSD1 plays a critical role in trabecular bone loss associated with systemic glucocorticoid therapy.

Publications

11β-HSD1 plays a critical role in trabecular bone loss associated with systemic glucocorticoid therapy.

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