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ABSTRACT:
PROVIDER: PRJNA817675 | ENA |
REPOSITORIES: ENA
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Project description:To investigate the function of lymphotoxin alpha in the regulation osteoblasts, murine pre-osteoblast cell line MC3T3-E1 clone #4 was treated with recombinant human lymphotoxin alpha1/beta2 for different times. Subsequently, we extracted total RNA from the treated cells, performed reverse transcription and cDNA library for RNA-Seq analysis.
2024-07-01 | GSE241494 | GEO
Project description:Osteoblasts are responsive to shear stress. We investigated the effect of of laminar fluid flow (LFF) on osteoblast-like MC3T3-E1 cells at two timepoints. We used microarray analysis to detail the global gene expression of MC3T3-E1 cells in response to 1 hour of laminar fluid flow directly and 4 hours after treatment.
2021-02-15 | GSE138160 | GEO
Project description:The biological effects of 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) on osteoblast differentiation and function differ significantly depending upon the cellular state of maturation. To explore this phenomenon mechanistically, we examined the impact of 1,25(OH)2D3 on the transcriptomes of both pre-osteoblastic (POBs) and differentiated osteoblastic (OBs) MC3T3-E1 cells, and assessed localization of the vitamin D receptor (VDR) at sites of action on a genome-scale using ChIP-seq analysis. We observed that the 1,25(OH)2D3-induced transcriptomes of POBs and OBs were quantitatively and qualitatively different, supporting not only the altered biology observed but the potential for a change in VDR interaction at the genome as well. This idea was confirmed through discovery that VDR cistromes in POBs and OBs were also strikingly different. Depletion of VDR binding sites in OBs, due in part to reduced VDR expression, was the likely cause of the loss of VDR-target gene interaction. Continued novel regulation by 1,25(OH)2D3, however, suggested that factors in addition to the VDR might also be involved. Accordingly, we show that transcriptomic modifications are also accompanied by changes in genome binding of the master osteoblast regulator RUNX2 and the chromatin remodeler C/EBPβ. Importantly, genome occupancy was also highlighted by the presence of epigenetic enhancer signatures which were selectively changed in response to both differentiation and 1,25(OH)2D3. The impact of VDR, RUNX2, and C/EBPβ on osteoblast differentiation is exemplified by their actions at the Runx2 and Sp7 gene loci. We conclude that each of these mechanisms may contribute to the diverse actions of 1,25(OH)2D3 on differentiating osteoblasts. RNA was isolated and applied to gene expression microarrays in undifferentiated MC3T3-E1 cells as well as post 15 day osteogenic differentiation MC3T3-E1 cells, which were treated for 24 hours with 10-7M 1,25(OH)2D3. For the vehicle matched samples, please refer to study GSE41955. The samples were completed in biological triplicate.
2014-06-02 | E-GEOD-51514 | biostudies-arrayexpress
Project description:The biological effects of 1?,25-dihydroxyvitamin D3 (1,25(OH)2D3) on osteoblast differentiation and function differ significantly depending upon the cellular state of maturation. To explore this phenomenon mechanistically, we examined the impact of 1,25(OH)2D3 on the transcriptomes of both pre-osteoblastic (POBs) and differentiated osteoblastic (OBs) MC3T3-E1 cells, and assessed localization of the vitamin D receptor (VDR) at sites of action on a genome-scale using ChIP-seq analysis. We observed that the 1,25(OH)2D3-induced transcriptomes of POBs and OBs were quantitatively and qualitatively different, supporting not only the altered biology observed but the potential for a change in VDR interaction at the genome as well. This idea was confirmed through discovery that VDR cistromes in POBs and OBs were also strikingly different. Depletion of VDR binding sites in OBs, due in part to reduced VDR expression, was the likely cause of the loss of VDR-target gene interaction. Continued novel regulation by 1,25(OH)2D3, however, suggested that factors in addition to the VDR might also be involved. Accordingly, we show that transcriptomic modifications are also accompanied by changes in genome binding of the master osteoblast regulator RUNX2 and the chromatin remodeler C/EBP?. Importantly, genome occupancy was also highlighted by the presence of epigenetic enhancer signatures which were selectively changed in response to both differentiation and 1,25(OH)2D3. The impact of VDR, RUNX2, and C/EBP? on osteoblast differentiation is exemplified by their actions at the Runx2 and Sp7 gene loci. We conclude that each of these mechanisms may contribute to the diverse actions of 1,25(OH)2D3 on differentiating osteoblasts. 4 transcription factors and 5 histone modifications were examined in undifferentiated MC3T3-E1 cells as well as post 15 day osteogenic differentiation MC3T3-E1 cells, which were treated for 3 hours prior to ChIP assay with ethanol vehicle or with 10-7M 1,25(OH)2D3. For the vehicle matched samples for RUNX2, CEBP beta and histones, please refer to study GSE41955. The samples were completed in biological replicate and examined separately.
2014-06-02 | E-GEOD-51513 | biostudies-arrayexpress
Project description:Developing osteoblasts undergo a sequence of three consecutive phases: cell proliferation, extracellular matrix maturation, and mineralization. We investigated pH effects on these phases using the osteoblast-like cell line MC3T3-E1.
2016-12-28 | GSE84907 | GEO
Project description:RUNX2 is a transcription factor that is first expressed in early osteoblast-lineage cells and represents a primary determinant of osteoblastogenesis. While numerous target genes are regulated by RUNX2, little is known of sites on the genome occupied by RUNX2 or of the gene networks that are controlled by these sites. To explore this, we conducted a genome-wide analysis of the RUNX2 cistrome in both pre-osteoblastic MC3T3-E1 cells (POB) and their mature osteoblast progeny (OB), characterized the two cistromes and assessed their relationship to changes in gene expression. We found that although RUNX2 was widely bound to the genome in POB cells, this binding profile was reduced upon differentiation to OBs. Numerous sites were lost upon differentiation, new sites were also gained; many sites remained common to both cell states. Additional features were identified as well including location relative to potential target genes, abundance with respect to single genes, the frequent presence of a consensus TGTGGT RUNX2 binding motif, co-occupancy by C/EBPβ and the presence of a typical epigenetic histone enhancer signature. This signature was changed quantitatively following differentiation. While RUNX2 binding sites were associated extensively with adjacent genes, the distal nature of the majority of these sites prevented assessment of whether they represented direct targets of RUNX2 action. Changes in gene expression, however, revealed an abundance of genes that contained RUNX2 binding sites and were regulated in concert. These studies establish a basis for further analysis of the role of RUNX2 activity and its function during osteoblast lineage maturation. 2 transcription factors and 5 histone modifications were examined in undifferentiated MC3T3-E1 cells as well as post 15 day osteogenic differentiation MC3T3-E1 cells. The samples were completed in biological replicate and examined separately.
2014-04-24 | E-GEOD-41920 | biostudies-arrayexpress
Project description:RUNX2 is a transcription factor that is first expressed in early osteoblast-lineage cells and represents a primary determinant of osteoblastogenesis. While numerous target genes are regulated by RUNX2, little is known of sites on the genome occupied by RUNX2 or of the gene networks that are controlled by these sites. To explore this, we conducted a genome-wide analysis of the RUNX2 cistrome in both pre-osteoblastic MC3T3-E1 cells (POB) and their mature osteoblast progeny (OB), characterized the two cistromes and assessed their relationship to changes in gene expression. We found that although RUNX2 was widely bound to the genome in POB cells, this binding profile was reduced upon differentiation to OBs. Numerous sites were lost upon differentiation, new sites were also gained; many sites remained common to both cell states. Additional features were identified as well including location relative to potential target genes, abundance with respect to single genes, the frequent presence of a consensus TGTGGT RUNX2 binding motif, co-occupancy by C/EBPβ and the presence of a typical epigenetic histone enhancer signature. This signature was changed quantitatively following differentiation. While RUNX2 binding sites were associated extensively with adjacent genes, the distal nature of the majority of these sites prevented assessment of whether they represented direct targets of RUNX2 action. Changes in gene expression, however, revealed an abundance of genes that contained RUNX2 binding sites and were regulated in concert. These studies establish a basis for further analysis of the role of RUNX2 activity and its function during osteoblast lineage maturation. RNA was isolated and applied to gene expression microarrays in undifferentiated MC3T3-E1 cells as well as post 15 day osteogenic differentiation MC3T3-E1 cells. The samples were completed in biological triplicate.
2014-04-24 | E-GEOD-41954 | biostudies-arrayexpress
Project description:Purpose: To identify gene expression changes (using RNAsequencing) in MC3T3-E1 cells where Rorβ has been deleted using CRISPR/Cas9 (and a control cell line) Method: MC3T3-Control and MC3T3-Rorβ-CRISPR(KO) cells were osteoblast differentiated for 2 days, and RNA was harvested (n=3 for each cell line) Results: We identified genes and pathways (using IPA) that were significantly altered in the Rorβ-CRISPR(KO) cell line, compared with the Control cell line Conclusions: Many pathways were changed, namely the Wnt pathway which is important to the biology of Rorβ and to osteoblast differentiation in general
2017-12-09 | GSE107849 | GEO
Project description:The miRNAs play important roles in regulating gene expression at post-transcriptional level through fine-tuning of protein-encoding gene expression, and are involved in regulating important biological processes in different cells and tissue types, including osteoblast differentiation. Canonical Wnt signaling is also known to play pleiotropic roles in regulating cell differentiation during mammalian osteogenesis, although the role of miRNAs is not established for Wnt signaling in osteoblast differentiation. Here we examined the role of candidate miRNAs expressed differentially after Wnt3a expression during osteoblast differentiation. Overexpression of the Wnt3a gene resulted in increased transcription of the ALP gene, but in decreased transcription of the Runx2, Col1a and OCN genes in osteoblastic MC3T3-E1 cells. Analysis of miRNA expression profile showed that 14 miRNAs were up-regulated and 21 miRNAs were down-regulated after Wnt3a overexpression in MC3T3-E1 cells. TGF-beta3 is presumed to be a candidate target gene of one of the down-regulated miRNAs with database search. Transfection of the miRNA mimic into MC3T3-E1 cells significantly inhibited the TGF-beta3 mRNA level and resultant protein expression, although overexpression of the Wnt3a gene could reverse the effect of the miRNA, resulting in the increased TGF-beta3 mRNA and protein levels. These results suggest that the miRNA is involved in osteoblast differentiation as a critical regulatory factor, mediating crosstalk between Wnt3a signaling pathway and TGF-beta3 signaling pathway.
2018-03-20 | GSE96757 | GEO
Project description:Background:; Osteoblast differentiation requires the coordinated stepwise expression of multiple genes. Histone deacetylase inhibitors (HDIs) accelerate the osteoblast differentiation process by blocking the activity of histone deacetylases (HDACs), which alter gene expression by modifying chromatin structure. We previously demonstrated that HDIs and HDAC3 shRNAs accelerate matrix mineralization and the expression of osteoblast maturation genes (e.g. alkaline phosphatase, osteocalcin). Identifying other genes that are differentially regulated by HDIs might identify new pathways that contribute to osteoblast differentiation. Results:; To identify other osteoblast genes that are altered early by HDIs, we incubated MC3T3-E1 preosteoblasts with HDIs (trichostatin A, MS-275, or valproic acid) for 18 hours in osteogenic conditions. The promotion of osteoblast differentiation by HDIs in this experiment was confirmed by osteogenic assays. Gene expression profiles relative to vehicle-treated cells were assessed by microarray analysis with Affymetrix GeneChip 430 2.0 arrays. The regulation of several genes by HDIs in MC3T3-E1 cells and primary osteoblasts was verified by quantitative real-time PCR. Nine genes were differentially regulated by at least two-fold after exposure to each of the three HDIs and six were verified by PCR in osteoblasts. Four of the verified genes (solute carrier family 9 isoform 3 regulator 1 (Slc9a3r1), sorbitol dehydrogenase 1, a kinase anchor protein, and glutathione S-transferase alpha 4) were induced. Two genes (proteasome subunit, beta type 10 and adaptor-related protein complex AP-4 sigma 1) were suppressed. We also identified eight growth factors and growth factor receptor genes that are significantly altered by each of the HDIs, including Frizzled related proteins 1 and 4, which modulate the Wnt signaling pathway. Conclusions: This study identifies osteoblast genes that are regulated early by HDIs and indicates pathways that might promote osteoblast maturation following HDI exposure. One gene whose upregulation following HDI treatment is consistent with this notion is Slc9a3r1. Also known as NHERF1, Slc9a3r1 is required for optimal bone density. Similarly, the regulation of Wnt receptor genes indicates that this crucial pathway in osteoblast development is also affected by HDIs. These data support the hypothesis that HDIs regulate the expression of genes that promote osteoblast differentiation and maturation. Experiment Overall Design: To identify other osteoblast genes that are altered early by HDIs, we incubated MC3T3-E1 preosteoblasts with HDIs (trichostatin A, MS-275, or valproic acid) or the vehicle control (DMSO) for 18 hours in osteogenic conditions. Gene expression profiles relative to vehicle-treated cells were assessed in triplicate (in some cases quadruplicate) samples by microarray analysis with Affymetrix GeneChip 430 2.0 arrays.
2008-06-16 | E-GEOD-9247 | biostudies-arrayexpress