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Comparison of Mineralizing Synchronous UMR106-01 cells with UMR106-01 cells treated with Mineralization Inhibitor AEBSF

ABSTRACT: UMR106-01 osteoblastic cells are a model for studying bone mineralization. We have shown that mineralization is temporally synchronized within cultures grown under defined conditions . Cells are plated at time zero and differentiate into osteoblastic phenotype by 64 h later. If an exogenous phosphate source is added to the cultures, the cells form and deposit hydroxyapatite mineral within distinct extracellular supramolecular lipid protein complexes termed biomineralization foci (BMF) starting 12 h later. Mineralization is largely complete by 24 h later (88 h after plating). We have also shown that AEBSF, covalent serine protease inhibitor, blocks mineralization within BMF and inhibits the fragmentation of several proteins related to biomineralization. The present experiment was designed to test whether AEBSF treatment for 12 h has an effect on transcription by UMR106-01 osteoblastic cells. AEBSF is known to inactivate several serine proteases including SKI-1 (site 1, subtilisin kexin protease-1).SKI-1 functions intracellularly to activate transmembrane bound transcription factor precursors releasing the transcriptionally active N-terminal portions to imported into the nucleus. Thus, if AEBSF blocks transcription of mineralization related genes, it would support a role for SKI-1 in gene regulation in mineralizing UMR106-01 osteoblastic cells.

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE21476 | GEO | 2010/11/13

SECONDARY ACCESSION(S): PRJNA126049

REPOSITORIES: GEO

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Comparison of Mineralizing Synchronous UMR106-01 cells with UMR106-01 cells treated with Mineralization Inhibitor AEBSF

Project description:UMR106-01 osteoblastic cells are a model for studying bone mineralization. We have shown that mineralization is temporally synchronized within cultures grown under defined conditions . Cells are plated at time zero and differentiate into osteoblastic phenotype by 64 h later. If an exogenous phosphate source is added to the cultures, the cells form and deposit hydroxyapatite mineral within distinct extracellular supramolecular lipid protein complexes termed biomineralization foci (BMF) starting 12 h later. Mineralization is largely complete by 24 h later (88 h after plating). We have also shown that AEBSF, covalent serine protease inhibitor, blocks mineralization within BMF and inhibits the fragmentation of several proteins related to biomineralization. The present experiment was designed to test whether AEBSF treatment for 12 h has an effect on transcription by UMR106-01 osteoblastic cells. AEBSF is known to inactivate several serine proteases including SKI-1 (site 1, subtilisin kexin protease-1).SKI-1 functions intracellularly to activate transmembrane bound transcription factor precursors releasing the transcriptionally active N-terminal portions to imported into the nucleus. Thus, if AEBSF blocks transcription of mineralization related genes, it would support a role for SKI-1 in gene regulation in mineralizing UMR106-01 osteoblastic cells. Comparison of triplicate cultures treated with mineralizing conditions with triplicate cultures treated with AEBSF protease inhibitor for 12 h.

2010-11-13 | E-GEOD-21476 | biostudies-arrayexpress

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Project description:Our previous study identified a small molecule, orally active ER-464195-01, inhibits the calreticulin binding to alpha integrins, and clearly suppresses the adherence ability of leukocytes. Our transcriptome analysis with the colon of dextran sodium sulfate (DSS)-treated mice reveals that the increased expression of pro-inflammatory genes was down-regulated by ER-464195-01. Male Balb/c mice (7 weeks) were allowed free access to distilled water (Otsuka Pharmaceutical). ER-464195-01 (10 mg/kg) or vehicle was orally administered once per day for 6 days.

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