View source for MSC to adipocyte (mouse)

{{TimeCourse
|Time_Course=
|collaborators=Yasushi Okazaki
|description=mouse_ST2_adipocytes
|libraryids=CNhs13114,CNhs13115,CNhs13116,CNhs13117,CNhs13118,CNhs13120,CNhs13121,CNhs13122,CNhs13123,CNhs13125,CNhs13126,CNhs13127,CNhs13128,CNhs13129,CNhs13243,CNhs13244,CNhs13245,CNhs13247,CNhs13248,CNhs13249,CNhs13250,CNhs13251,CNhs13252,CNhs13253,CNhs13254,CNhs13255,CNhs13256,CNhs13257,CNhs13258,CNhs13292,CNhs13293,CNhs13295,CNhs13296,CNhs13297,CNhs13298,CNhs13299,CNhs13300,CNhs13301,CNhs13302,CNhs13303,CNhs13304,CNhs13305,CNhs13306,CNhs13307
|page_name=mouse_MSC_adipocytes
|series=IN_VITRO DIFFERENTIATION SERIES
|species=Mouse (Mus musculus)
|zenbu_config=http://fantom.gsc.riken.jp/zenbu/gLyphs/#config=eDHhE-APFBJsq9d4oP_U7
|TCOverview=Senile osteoporosis is the most common metabolic bone disease. This disease is often accompanied by increasing adipocytes in bone marrow tissues [1]. The ectopic adipocytes differentiation following bone loss seems to be caused by unbalanced differentiation of mesenchymal stem cells (MSCs) [2]. Although several differentiation regulators of MSCs have already been reported, little is known about the regulatory dynamics of bi-directional adipocytes/osteoblasts differentiation.<br>
To uncover the complex mechanism of osteoporosis and metabolic disease, we performed a variety of genome-wide analyses about gene expression and regulation that could influence adipocytes/osteoblasts differentiation from mouse ST2 cells (bone marrow-derived stromal cell line) [3,4]. In particular, we focused on the differentiation-specific non-cording RNAs and antisense transcripts as the novel regulator candidates of adipocytes/osteoblasts differentiation.<br>
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References:<br>
1. Burkhardt R, Kettner G, Bohm W, Schmidmeier M, Schlag R, et al. Changes in trabecular bone, hematopoiesis and bone marrow vessels in aplastic anemia, primary osteoporosis, and old age: a comparative histomorphometric study. Bone (1987) 8(3):157-164. <html><a href='http://www.ncbi.nlm.nih.gov/pubmed/3606907' target='_blank'>PMID:3606907</a></html><br>
2. Nuttall ME, Gimble JM. Controlling the balance between osteoblastogenesis and adipogenesis and the consequent therapeutic implications. Curr Opin Pharmacol (2004) 4(3):290–294. <a href='http://www.ncbi.nlm.nih.gov/pubmed/15140422' target='_blank'>PMID:15140422</a></html><br>
3. Tokuzawa Y, Yagi K, Yamashita Y, Nakachi Y, Nikaido I, et al. Id4, a new candidate gene for senile osteoporosis, acts as a molecular switch promoting osteoblast differentiation. PLoS Genet (2010) 6(7):e1001019. <html><a href='http://www.ncbi.nlm.nih.gov/pubmed/20628571' target='_blank'>PMID:20628571</a></html><br>
4. Mizuno Y, Yagi K, Tokuzawa Y, Kanesaki-Yatsuka Y, Suda T, et al. miR-125b inhibits osteoblastic differentiation by down-regulation of cell proliferation. Biochem Biophys Res Commun (2008) 368(2):267-272. <a href='http://www.ncbi.nlm.nih.gov/pubmed/18230348' target='_blank'>PMID:18230348</a></html>.<br>
|TCSample_description='''Cell line:'''<br>
ST2 cells were obtained from RIKEN BioResource Center (BRC, Tsukuba, Japan). These cell line is bone marrow-derived stromal cell line. ST2 differentiated most efficiently into both osteoblasts and adipocytes [3].<br>
'''Cell culture:'''<br>
ST2 cells were cultured according to the protocols supplied by BRC (RPMI1640 supplemented with 10% fetal bovine serum) [4].<br>
'''Differentiation induction:'''<br>
''Adipocyte differentation:''<br>
Adipogenic differentiation was induced by changing the medium to differentiation medium supplemented with 10% fetal bovine serum (FBS), 0.5 mM 3-isobutyl-1-methlxanthine, 0.25 mM dexamethasone, and insulin-transferrin-selenium-X supplement containing 5 mg/ml of insulin (Invitrogen, Carlsbad, CA) and 1 mM rosiglitazone. After 48 hr, the differentiation medium was replaced with conditional culture medium supplemented with 10% FBS [3]. Pre-conditioned medium which was originally same as culure medium, was carried by parallel culturing of ST2, to avoid the perturbation by "medium-change" shock as soon as possible.<br>
<html><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/NileRed4d.jpg'></html><br>
Figure 1. Histological staining of ST2 cells using Nile Red staining during adipocyte differentiation.<br>
The number of Nile Red stained lipid droplets increased in ST2 cell (4 days after adipocyte induction). Bar: 100 μm.<br>
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''Osteoblast differentiation:''<br>
Osteogenic differentiation was induced by changing the medium every three days to culture medium supplemented with 100 ng/ml of bone morphogenetic protein 4 (BMP4, R&D Systems, Mineapolis, MN) [3]. Pre-conditioned medium which was originally same as culure medium, was carried by parallel culturing of ST2, to avoid the perturbation by "medium-change" shock as soon as possible.<br>
<html><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/ALP6d.png'></html><br>
Figure 2. ALP staining of ST2 cells for 0, 6 and 20 days after osteoblast induction.<br>
The ALP activity of ST2 cells (right, 20 days) were more prominently increased in the presence of BMP4 than 0 day (left).<br>
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'''Sampling:'''<br>
ST2 cells are sampled during adipocyte or osteoblast differentiation (15min, 30min, 1-3hr, 6,12,18,24,36,48hr, 3-6day) and non-treatment control (2 points; 0,6day).<br>
<html><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/500px-Figure1.png'></html><br>
Figure 3. Sampling points for ST2 time-course CAGE data<br>
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References:<br>
3. Tokuzawa Y, Yagi K, Yamashita Y, Nakachi Y, Nikaido I, et al. Id4, a new candidate gene for senile osteoporosis, acts as a molecular switch promoting osteoblast differentiation. PLoS Genet (2010) 6(7):e1001019. PMID:20628571<br>
4. Mizuno Y, Yagi K, Tokuzawa Y, Kanesaki-Yatsuka Y, Suda T, et al. miR-125b inhibits osteoblastic differentiation by down-regulation of cell proliferation. Biochem Biophys Res Commun (2008) 368(2):267-272. PMID:18230348.<br>
|TCQuality_control='''Marker gene expression:'''<br>
Adipocyte differentation, hCAGE:<br>
<html><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo1.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo2.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'></html><br>
<html><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/500px-Figure1.png'></html><br>
Figure 3. Sampling points for ST2 time-course CAGE data<br>
|TCQuality_control='''Marker gene expression:'''<br>
Adipocyte differentation, hCAGE:<br>
<html><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo1.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo2.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'><br>
<img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo3.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo4.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'><br>
<img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo5.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Adipo6.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'><br>
<img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Osteo1.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Osteo2.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'><br>
<img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Osteo3.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Osteo4.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'><br>
<img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Osteo5.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Osteo6.jpg'><img src='https://fantom5-collaboration.gsc.riken.jp/resource_browser/images/TC_qc/Legend.jpg'><br>
</html><br>
Figurea 4. Gene expression (TPM) of key regulators (Hes1, Id1, Id2,Sp7) and differentiation markers (Alpl, Bglap).<br>
}}