Genes regulated by TGF-beta in bovine articular chondrocytes
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ABSTRACT: Bovine articular chondrocytes were grown in micromass culture and were either untreated or treated with 5 ng TGF-b1/ml for 8 hours to identify genes regulated by TGF-b. Six total samples were analyzed. Three biological replicates of untreated bovine articular chondrocytes grown in micromass culture and three biological replicates of bovine articular chondrocytes grown in micromass culture and treated with 5ng TGF-b1/ ml for 8 hours.
Project description:Bovine articular chondrocytes were grown in micromass culture and were either untreated or treated with 5 ng TGF-b1/ml for 8 hours to identify genes regulated by TGF-b.
Project description:Very little is known about how intervertebral disc (IVD) is formed or maintained. Members of the TGF-beta superfamily are secreted signaling proteins that regulate many aspects of development including cellular differentiation. We recently showed that deletion of Tgfbr2 in Col2a expressing tissue results in alterations in development of IVD annulus fibrosus. The results suggested TGF-beta has an important role in regulating development of the axial skeleton, however, the mechanistic basis of TGF-beta action in these specialized joints is not known. To understand the mechanism of TGF-beta action in IVD development, we undertook a global analysis of gene expression comparing gene expression profiles in sclerotome cultures treated with TGF-beta or BMP4. As expected, treatment with BMP4 resulted in up-regulation of cartilage marker genes including Acan, Sox 5, Sox6, and Sox9. In contrast, treatment with TGF-beta1 did not regulate expression of cartilage markers but instead resulted in up-regulation of many IVD markers including Fmod and Adamtsl2. We propose TGF-beta has two functions in IVD development: 1) to prevent chondrocyte differentiation in the presumptive IVD and 2) to promote differentiation of annulus fibrosus from sclerotome. We have identified genes that are enriched in the IVD and regulated by TGF-beta that warrant further investigation as regulators of IVD development. Nine samples were analyzed. Three biological replicates of untreated sclerotome grown in micromass culture. Three biological replicates of cells treated with 50 ng/ml BMP4 for 8 hours and three biological replicates of cells treated with 5 ng/ ml TGF-beta1 for 8 hours.
Project description:Proliferative zone chondrocytes were microdissected from control and Ift88-deleted growth plates to determine gene expression profiles regulated by primary cilia. Four total samples were analyzed. Two biological replicates of proliferative zone chondrocytes microdissected from control mice and two biological replicates from Ift88 deleted (Col2aCre;Ift88fl/fl) mice. Control and experimental mice were in the Bl/6 background.
Project description:To identify DOT1L targets, associated signaling pathways and networks in chondrocytes, we used genome-wide gene expression microarray analysis in human articular chondrocytes of 5 different donors (without known or documented joint disease) treated with EPZ-5676 or vehicle for 4 days. It is known that DOT1L inhibitors require longer time of treatment in order to show effect and influence the expression of MLL target genes in leukemia cells, but we opted for this relatively short inhibition time to be able to identify early changes induced by DOT1L inhibition. Human articular chondrocytes were obtained from 5 non-OA hip fracture patients. The cells were treated with 3 μM EPZ-5676 or vehicle (DMSO) for 4 days.
Project description:Objective: To evalute the role of Ezh2 in chondrocyte differentiation. Method: Ezh2 was inactivated in chondrocytes using the Col2a1-Cre driver in vivo. Immature mouse chondrocytes (IMC) were isolated from wildtype and Ezh2 deficient mice, and plated in micromass culture. Chondrocytes were differentiated and RNA was isolated at days 3, 7, and 14 of culture. Isolated RNA was subjected to RNA sequencing analysis. Results: Ezh2 deficient chondrocytes exhibit enhanced expression of osteogenic genes compared to wildtype cells.
Project description:Primary micromass cultures derived from 11.5 day old mouse embryo limb buds were cultured for 15 days in differentiating conditions (beta-glycerophosphate and ascorbic acid). Total RNA from differentiating chondrocytes was isolated every three days i.e. days 3,6,9,12 and 15 and hybridized to MOE430A chips. Objective: Gain a view of the temporal gene expression changes occuring during chondrocyte differentiation. Keywords = micromass culture Keywords = temporal gene expression Keywords = mouse Keywords = chondrocyte differentiation Keywords: time-course
Project description:Very little is known about how intervertebral disc (IVD) is formed or maintained. Members of the TGF-ß superfamily are secreted signaling proteins that regulate many aspects of development including cellular differentiation. We recently showed that deletion of Tgfbr2 in Col2a expressing tissue results in alterations in development of IVD annulus fibrosus. The results suggested TGF-ß has an important role in regulating development of the axial skeleton, however, the mechanistic basis of TGF-ß action in these specialized joints is not known. One of the hurdles to understanding development of IVD is a lack of known markers. To identify genes that are enriched in the developing IVD and to begin to understand the mechanism of TGF-ß action in IVD development, we undertook a global analysis of gene expression comparing gene expression profiles in developing vertebrae and IVD. We also compared expression profiles in tissues from wild type and Tgfbr2 mutant mice. Lists of IVD and vertebrae enriched genes were generated. Expression patterns for several genes were verified either through in situ hybridization or literature/ database searches resulting in a list of genes that can be used as markers of IVD. Cluster analysis using genes listed under the Gene Ontology terms multicellular organism development and pattern specification indicated that mutant IVD more closely resembled vertebrae than wild type IVD. We propose TGF-ß has two functions in IVD development: 1) to prevent chondrocyte differentiation in the presumptive IVD and 2) to promote differentiation of annulus fibrosus from sclerotome. We have identified genes that are enriched in the IVD and regulated by TGF-ß that warrant further investigation as regulators of IVD development. Thirteen samples were analyzed. This includes three biological replicates of laser captured IVD from E13.5 day control mice, three biological replicates of laser captured vertebrae from the same E13.5 day control mice, three biological relicates of laser captured vertebrae from E13.5 day Col2aCre;Tgfbr2lox/lox mice, and four biological replicates of laser captured IVD from E13.5 day Col2aCre;Tgfbr2lox/lox mice.
Project description:The aim of the current study was to identify molecular markers for articular cartilage that can be used for the quality control of tissue engineered cartilage. Therefore a genom-wide expression analysis was performed using RNA isolated from articular and growth plate cartilage, both extracted from the knee joints of minipigs. Keywords: Native material or primary cells isolated from articular cartilage and growth plate cartilage Articular and growth plate cartilage were taken for RNA extraction and hybridization on Affymetrix microarrays. Furthermore chondrocytes from each type of cartilage were isolated and cell culture was started and terminated at day 10 or day 20. Total RNA from cultivated cells was extracted, and hybridization on Affymetrix microarrays was performed.
Project description:This SuperSeries is composed of the following subset Series: GSE18647: Gene expression in embryonic intervertebral disc and vertebrae. GSE18648: TGF-beta and BMP mediated gene expression in cultured sclerotome. Refer to individual Series