Microarray analysis of defective cartilage in Hoxc8- and Hoxd4-transgenic mice
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Claudia Kappen
PROVIDER: E-GEOD-19002 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Homeobox genes of the Hox class are required for proper patterning of skeletal elements and play a role in cartilage differentiation. In transgenic mice with overexpression of Hoxc8 during cartilage development, we observed severe defects, namely physical instability of cartilage, accumulation of immature chondrocytes, and decreased maturation to hypertrophy. To define the molecular basis underlying these defects, we performed gene expression profiling using the Affymetrix microarray platform. Embryos were dissected from four different mouse litters at E18.5. Embryos from each litter were grouped according their genotype: TA=control; TR=transgenic.
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Project description:BBF2H7 (BBF2 human homolog on chromosome 7), an ER-resident basic leucine zipper transcription factor, is activated in response to ER stress and abundantly expresses in chondrocytes. While BBF2H7 is widely expressed in many tissues and organs, the most intense signals were detected in the proliferating zone of the cartilage. We compared gene expressions in primary cultured chondrocytes prepared from rib cartilage between WT and BBF2H7-/- mice at E18.5. Primary cultured chondrocytes were prepared from E18.5 rib cartilage of WT and BBF2H7-/- mice. Chondrocytes were isolated using 0.2% collagenase D (Roche) after adherent connective tissue was removed by 0.2% trypsin (Sigma) and collagenase pretreatment. Isolated chondrocytes were maintained in α-MEM (Gibco) supplemented with 10% FCS and 50 µg/mL ascorbic acid. Adenovirus vectors expressing the mouse p60 BBF2H7 (1-377 aa, BBF-N) were constructed with the AdenoX Expression system (Clontech), according to the manufacturerâs protocol. The cells were infected with adenoviruses 30 h before analysis. We compared gene expressions in primary cultured chondrocytes prepared from rib cartilage between WT and BBF2H7-/- mice at E18.5 using a microarray and various genes associated with protein secretory pathway and ER biogenesis were significantly down-regulated in BBF2H7-/- chondrocytes. We infected primary cultured chondrocytes prepared from BBF2H7-/- mice with adenovirus expressing p60 BBF2H7. Several genes were up-regulated and we picked up them as the direct target of BBF2H7.
2010-05-05 | E-GEOD-18052 | biostudies-arrayexpress
Project description:Overexpression of Hoxc8 transcription factor alters transcriptional profiles in mouse chondrocytes at E18.5
| PRJNA123735 | ENA
Project description:A comprehensive analysis of Sox9 binding profiles in developing chondrocytes identified marked enrichment of an AP-1-like motif (Ohba et al. 2015). Here, we have explored the functional interplay between Sox9 and AP-1 in mammalian chondrocyte development. Among AP-1 family members, Jun and Fosl2 were highly expressed within prehypertrophic and early hypertrophic chondrocytes. Chromatin immunoprecipitation followed by DNA sequencing (ChIP-seq) showed a striking overlap in Jun- and Sox9-bound regions throughout the chondrocyte genome, a reflection of direct binding of each factor to target motifs in shared enhancers, and physical interactions of AP-1 with Sox9. In vitro expression analysis indicates that direct co-binding of Sox9 and AP-1 at target motifs enhanced target gene expression, while protein-protein interactions suppressed AP-1- and Sox9-driven transcription. Analysis of prehypertrophic chondrocyte removal of Sox9 demonstrated Sox9 was essential for hypertrophic chondrocyte development, while in vitro and ex vivo analyses showed AP-1 promotes chondrocyte hypertrophy. Sox9 and Jun co-bound and co-activated a Col10a1 enhancer in Sox9 and AP-1 motif-dependent manners consistent with their combined action promoting hypertrophic gene expression. Together, the data support a model where AP-1-family members promote Sox9-action in the transition of chondrocytes to a terminal hypertrophic program. Intersection of ChIP-seq data from Sox9 and AP-1 factor Jun, RNA-seq data from developing rib chondrocytes and Col10a1mCherry positive hypertrophic chondrocytes in neonatal mice to uncover regulation of Sox9 by AP-1 factors during chondrocyte hypertrophy.
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2008-06-15 | E-GEOD-7683 | biostudies-arrayexpress
Project description:We have studied the expression profile of 3D cultured human chondrocytes that were stimulated with supernatant of synovial fibroblasts derived from a RA patient (RASF=HSE cell line) and from a normal donor (NDSF=K4IM cell line), respectively. For this purpose, passage 2 human chondrocytes were cultured for 14 days in alginate beads and subsequently stimulated for 48 hours with supernatant of RASF and NDSF. Baseline expression was determined of unstimulated chondrocytes. Differential genome-wide microarray analysis of RASF and NDSF stimulated chondrocytes disclosed a distinct expression profile related to cartilage destruction involving marker genes of inflammation (COX-2), NF-kappa B signaling pathway (TLR2), cytokines/chemokines and receptors (CXCL1-3, CCL20, CXCL8, CXCR4, IL-6, IL-1beta), matrix degradation (MMP-10, MMP-12) and suppressed matrix synthesis (COMP). Thus, transcriptome profiling of RASF and NDSF stimulated chondrocytes revealed a disturbed catabolic-anabolic homeostasis of chondrocyte function. This study provides a comprehensive insight into the molecular regulatory processes induced in human chondrocytes during RA-related cartilage destruction. Experiment Overall Design: To reveal the RA-related chondrocyte gene expression profile, genome-wide microarray analysis of RASF stimulated human chondrocytes compared to NDSF stimulation was performed. Unstimulated chondrocytes were analyzed for baseline gene expression. For microarray analysis of RASF and NDSF stimulated and unstimulated chondrocytes, respectively, two RNA pools were analyzed, each pool consisting of equal amounts of RNA from three different donors.
2008-06-11 | E-GEOD-10024 | biostudies-arrayexpress
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Project description:Overexpression of Hoxd4 transcription factor alters transcriptional profiles in mouse chondrocytes at E18.5
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