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Dynamic expression of DKK1 protein in the process whereby Epimedium-derived flavonoids up-regulate osteogenic and down-regulate adipogenic differentiation of bone marrow stromal cells in ovariectomized rats.

ABSTRACT: OBJECTIVE:To observe the dynamic expression of DKK1 protein in the process whereby Epimedium-derived flavonoids (EFs) regulate the balance between osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats, and to provide experimental evidence for the mechanism of EFs in the treatment of postmenopausal osteoporosis. METHODS:? Bone marrow stromal cells from ovariectomized rats were separated and cultivated in osteoinductive or liquid medium for 15 days in vitro. EFs (10?µg/mL) were applied to both cultures. Alkaline phosphatase (ALP) staining, ALP activity determination, Oil Red O staining and fluorescence quantitative polymerase chain reaction were used to determine the influence of EFs on osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats. Moreover, in order to explore the exact mechanism of EFs on osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats, enzyme linked immunosorbent assay was used to determine the dynamic expression of DKK1 protein in this process. RESULTS:EFs increased activity of ALP and mRNA expression of Runx2 (early osteoblast differentiation factor) and decreased mRNA expression of PPAR?-2 (key factor of fat generation). Importantly, EFs down-regulated expression of DKK1 protein in an osteogenic induction medium and inhibited up-regulation of DKK1 protein in an adipogenic induction medium. CONCLUSION:? EFs regulate the balance between osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats by down-regulating expression of DKK1 protein. This may be an important molecular mechanism of EFs in the context of treatment of postmenopausal osteoporosis.

SUBMITTER: Xu YX

PROVIDER: S-EPMC6583533 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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Dynamic expression of DKK1 protein in the process whereby Epimedium-derived flavonoids up-regulate osteogenic and down-regulate adipogenic differentiation of bone marrow stromal cells in ovariectomized rats.

Xu Ying-xing YX Xu Bing B Wu Cheng-liang CL Wu Yan Y Tong Pei-jian PJ Xiao Lu-wei LW

Orthopaedic surgery 20110501 2

<h4>Objective</h4>To observe the dynamic expression of DKK1 protein in the process whereby Epimedium-derived flavonoids (EFs) regulate the balance between osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats, and to provide experimental evidence for the mechanism of EFs in the treatment of postmenopausal osteoporosis.<h4>Methods</h4> Bone marrow stromal cells from ovariectomized rats were separated and cultivated in osteoinductive or liquid medium for 15 ...[more]

PMID: 22009597

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Project description:Bone marrow adipose tissue has brown fat characteristics. Several studies have demonstrated that total flavonoids of Epimedium (TFE) could prevent bone loss and reduce the white adiposity in bone marrow induced by ovariectomy (OVX) in rats. However, the effects of TFE on marrow brown fat in OVX rats remain unclear. In this word, we addressed this question expected to provide a new target for preventing and treating osteoporosis. Thirty-six 3-month-old female Sprague-Dawley rats were equally divided into Sham controls, OVX controls, and OVX treated with TFE. Chemical shift coding magnetic resonance was performed to detect marrow fat fraction at the left femur at baseline, 6 and 12 weeks post-OVX. Bone mineral density at the lumbar spine and femur was measured by dual-energy x-ray absorptiometry. Serum bone biomarkers by ELISA, trabecular bone microarchitecture at the proximal tibia by micro-CT, quantitative parameters of marrow adipocyte by hematoxylin, and eosin staining were evaluated. The marrow adipocyte gene and protein expressions profile were determined by real-time quantitative PCR and immunostaining in whole tibiae. We found that TFE treatment could decrease bone turnover rate and improved bone mineral density and trabecular microarchitecture in OVX rats. OVX resulted in marrow adipogenesis as evidenced by increased marrow fat fraction, larger marrow adipocyte size, increased adipocyte number and percentage of adipocyte area, marrow white adipocyte gene, and protein expression, including PPARγ2 and FABP4. These pathological changes induced by estrogen deficiency were restored by TFE treatment. TFE also increased brown adipocyte expressions of the transcription factor Ucp1 and Prdm16 in whole tibiae. There was no detectible protein expression of brown adipocyte markers in the proximal tibia. Taken together, TFE regulation of bone marrow adiposity in OVX rats is mediated, at least in part, via maintaining the reciprocity of white and brown adipose tissue.

Project description:Bone-marrow mesenchymal stem cells (BMSC) are considered the gold standard for use in tissue regeneration among mesenchymal stem cells (MSC). The abundance and ease of harvest make the adipose-derived stem cells (ASC) an attractive alternative to BMSC. The aim of the present study was to compare the transcriptome of ASC and BMSC, respectively isolated from subcutaneous adipose tissue and femur of 3 adult pigs, during in vitro osteogenic and adipogenic differentiation for up to four weeks. At 0, 2, 7, and 21 days of differentiation RNA was extracted for microarray analysis. A False Discovery Rate ?0.05 for overall interactions effect and P<0.001 between comparisons were used to determine differentially expressed genes (DEG). Ingenuity Pathway Analysis and DAVID performed the functional analysis of the DEG. Functional analysis of highest expressed genes in MSC and genes more expressed in MSC vs. fully differentiated tissues indicated low immunity and high angiogenic capacity. Only 64 genes were differentially expressed between ASC and BMSC before differentiation. The functional analysis uncovered a potential larger angiogenic, osteogenic, migration, and neurogenic capacity in BMSC and myogenic capacity in ASC. Less than 200 DEG were uncovered between ASC and BMSC during differentiation. Functional analysis also revealed an overall greater lipid metabolism in ASC, while BMSC had a greater cell growth and proliferation. The time course transcriptomic comparison between differentiation types uncovered <500 DEG necessary to determine cell fate. The functional analysis indicated that osteogenesis had a larger cell proliferation and cytoskeleton organization with a crucial role of G-proteins. Adipogenesis was driven by PPAR signaling and had greater angiogenesis, lipid metabolism, migration, and tumorigenesis capacity. Overall the data indicated that the transcriptome of the two MSC is relatively similar across the conditions studied. In addition, functional analysis data might indicate differences in therapeutic application.

Dataset Information

Dynamic expression of DKK1 protein in the process whereby Epimedium-derived flavonoids up-regulate osteogenic and down-regulate adipogenic differentiation of bone marrow stromal cells in ovariectomized rats.

Publications

Dynamic expression of DKK1 protein in the process whereby Epimedium-derived flavonoids up-regulate osteogenic and down-regulate adipogenic differentiation of bone marrow stromal cells in ovariectomized rats.

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