Signaling Role of Acetoacetate in Accelerating Muscle Regeneration and Ameliorating Muscular Dystrophy
Ontology highlight
ABSTRACT: Acetoacetate (AA) is a ketone body and acts as a fuel to supply energy for cellular activity of various tissues. Here, we performed differential RNA-Seq analyses of proliferating and differentiating C2C12 myoblasts in the present or absent of AA. Differentially regulated genes and gene set enrichment analysis revealed that altered expression of the AA-mediated genes in both GM (growth medium) and DM (differentiation medium) were highly enriched in the signaling pathways associated with cell proliferation and differentiation, however, the genes encoding enzymes in biochemical pathways of ketolysis for energy production were not transcriptionally changed in the cells cultured in either GM or DM in response to AA. Notably the genes differentially expressed between GM and DM were also significantly regulated by AA. This indicates that AA-mediated myogenic cell function by transcriptional regulation of the genes required for cell proliferation and differentiation. Taken together, the data from our multi-parameter analyses consistently support the notion that AA plays a non-metabolic role in regulating muscle cell function. We sequenced mRNA from C2C12 cells in GM or DM treated with AA for 12 h or not to generate genes response to AA during C2C12 proliferation and differentiation. Three of duplication was performed. Examination of mRNA levels in proliferating or differentiating C2C12 cells between AA-treated groups and control groups.
Project description:Acetoacetate (AA) is a ketone body and acts as a fuel to supply energy for cellular activity of various tissues. Here, we performed differential RNA-Seq analyses of proliferating and differentiating C2C12 myoblasts in the present or absent of AA. Differentially regulated genes and gene set enrichment analysis revealed that altered expression of the AA-mediated genes in both GM (growth medium) and DM (differentiation medium) were highly enriched in the signaling pathways associated with cell proliferation and differentiation, however, the genes encoding enzymes in biochemical pathways of ketolysis for energy production were not transcriptionally changed in the cells cultured in either GM or DM in response to AA. Notably the genes differentially expressed between GM and DM were also significantly regulated by AA. This indicates that AA-mediated myogenic cell function by transcriptional regulation of the genes required for cell proliferation and differentiation. Taken together, the data from our multi-parameter analyses consistently support the notion that AA plays a non-metabolic role in regulating muscle cell function.
Project description:We want to know which genes are overexpressed or down regulated in Arabidopsis plants transformed with the sunflower hahb-4 homeodomain transcription factor with respect to non transformed ones in control and water stress conditions. This gene, hahb4, confers drought tolerance to transgenic arabidopsis plants. Aa water stress was applied to transgenic and non transformed three weeks old arabidopsis plants control and overexpressing line
Project description:This project investigates the function of p68/p72 in muscle gene expression and skeletal C2C12 cell differentiation. p68/p72 in C2C12 cells cultured in either growth medium (GM) or differentiation medium (DM) was knocked down using shRNA p68/p72 retrovirus, and globle gene expression was profiled using Affymetrix MOE 430A and 430B GeneChip. Keywords: Time-Series
Project description:This project investigates the function of p68/p72 in muscle gene expression and skeletal C2C12 cell differentiation. p68/p72 in C2C12 cells cultured in either growth medium (GM) or differentiation medium (DM) was knocked down using shRNA p68/p72 retrovirus, and globle gene expression was profiled using Affymetrix MOE 430A and 430B GeneChip. Experiment Overall Design: The specific aim is to study muscle gene expression changes and skeletal muscle differenciation using RNA interference of p68/p72 in C2C12 cells.
Project description:To characterize the differentiation by Sox2 KO, we performed microarray analyses of mouse ES cell line 2TS22C during the time-course being induced of Sox2 KO Inducible Sox2 KO cell line 2TS22C were assigned to eight experimental groups and cultured in GMEM medium (SIGMA) with LIF at 37°C in an atmosphere of 5% CO2 until the specified time point before the harvest: (1)for controls, cultured in the absence of Tc; (2) for 3h after the administration of Tc; (3) for 6h; (4) for 12h; (5) for 24h; (6) for 48h; (7) for 72h; (8) for 96h.
Project description:E12.5 mouse whole embryo and E12.5 placenta total RNA were pooled to create 25:75, 50:50, and 75:25 ratio mixtures, based on Bioanalyzer quantitation. These samples, along with the original unmixed RNAs, were used as templates for duplicate linear amplification labeling reactions. cRNA target mixtures were hybridized against a Universal Mouse Reference (Stratagene). Pairwise comparison using the NIA Microarray Analysis (ANOVA) software produced log ratios, which were compared to the expected log ratios for genes showing statistically significant (FDR<0.05) differential expression between unmixed embryo and placenta. Keywords: cell type comparison design,development or differentiation design,normalization testing design,reference design E12.5 mouse whole embryo and E12.5 placenta total RNA were pooled to create 25:75, 50:50, and 75:25 ratio mixtures, based on Bioanalyzer quantitation. These samples, along with the original unmixed RNAs, were used as templates for duplicate linear amplification labeling reactions. cRNA target mixtures were hybridized against a Universal Mouse Reference (Stratagene). Pairwise comparison using the NIA Microarray Analysis (ANOVA) software produced log ratios, which were compared to the expected log ratios for genes showing statistically significant (FDR<0.05) differential expression between unmixed embryo and placenta.
Project description:Angiogenesis and lymphangiogenesis have important roles in cancer progression and chronic inflammatory diseases, but efficient therapies against these diseases have been hampered by the lack of identified vascular lineage-specific markers and growth factors. Using transcriptional profiling of matched pairs of human dermal blood vascular and lymphatic endothelial cells, we first identified 236 lymphatic and 342 blood vascular signature genes. In silico analyses of the biologic pathways associated with these genes revealed lineage-specific functions for each cell type. Using a selection of 85 identified vascular lineage-specific genes, we developed a TaqMan RT-PCR-based, microfluidic card-formatted low-density microvascular differentiation array (LD-MDA) that was used to reliably identify and quantify the degree of lineage-specific differentiation in different types of endothelial cells, and to detect admixture of lymphatic endothelial cells in commercial preparations of microvascular endothelial cells. Application of Prediction Relevance Ranking and analysis of variance of LD-MDA expression profiles of 43 lesional skin samples obtained from patients with the chronic inflammatory disease psoriasis led to identification of cytokines which are significantly associated with angiogenesis or lymphangiogenesis in vivo. In particular, interleukin-7 and fibroblast growth factor-12 were identified as novel (lymph)angiogenic factors. This technology provides a novel tool to quantify lineage-specific vascular differentiation and to characterize (lymph)angiogenesis in clinical samples obtained from angiogenic diseases. This SuperSeries is composed of the following subset Series: GSE11306: Quantification of vascular lineage-specific differentiation (cell type comparison) GSE11307: Quantification of vascular lineage-specific differentiation, psoriasis (chronic inflammation) study Keywords: SuperSeries Refer to individual Series
Project description:Kaposiâs sarcoma (KS) is the most frequently occurring malignant tumor in patients infected with the human immunodeficiency virus. Recent studies have revealed that infection of vascular endothelial cells with Kaposi's sarcoma-associated herpes virus in vitro results in a lymphatic re-programming of these cells, with potent induction of the lymphatic marker genes podoplanin and VEGFR-3 which is mediated by upregulation of the transcription factor Prox1. However, the potential effects of Prox1 expression on the biology of KS and, in particular, on the aggressive and invasive behavior of KS tumors in vivo have remained unknown. We stably expressed Prox1 cDNA in the two mouse hemangioendothelioma cell lines EOMA and Py-4-1, well-established murine models for kaposiform hemangioendothelioma. Surprisingly, we found that expression of Prox1 was sufficient to induce a more aggressive behavior of tumors growing in syngenic mice, leading to enhanced local invasion into the muscular layer and to cellular anaplasia. This enhanced malignant phenotype was associated with upregulation of several genes involved in proteolysis, cytoskeletal reorganisation and migration. Together, these results indicate that Prox1 plays an important, previously unanticipated role in mediating the aggressive behavior of vascular neoplasms such as Kaposi's sarcoma. Experiment Overall Design: Total cellular RNA was extracted from two stably Prox1-expressing Py-4-1 cell clones and from two control-transfected Py-4-1 cell clones.
Project description:E12.5 mouse whole embryo, E12.5 placenta, embryonic stem (ES) cells, and trophoblast stem (TS) cells were compared on a novel microarray design to validate the system. The in situ-synthesized 60-mer oligonucleotide microarray platform contains approximately 44,000 DNA features, and was designed to detect transcripts from all known genes, as well as about 5,000 potential genes, as identified by the NIA Mouse Gene Index 2.0. Seven external RNA controls were derived from intronic and intergenic yeast genome sequences, and 60-mer probes were spotted 10 time each on the slides. These spike-in RNA controls were diluted to cover 7 orders of magnitude in initial concentration, and used to define a function relating signal intensity to transcript abundance. Keywords: cell type comparison design,development or differentiation design,normalization testing design,reference design E12.5 mouse whole embryo, E12.5 placenta, embryonic stem (ES) cells, and trophoblast stem (TS) cells were compared on a novel microarray design to validate the system. The in situ-synthesized 60-mer oligonucleotide microarray platform contains approximately 44,000 DNA features, and was designed to detect transcripts from all known genes, as well as about 5,000 potential genes, as identified by the NIA Mouse Gene Index 2.0. Seven external RNA controls were derived from intronic and intergenic yeast genome sequences, and 60-mer probes were spotted 10 time each on the slides. These spike-in RNA controls were diluted to cover 7 orders of magnitude in initial concentration, and used to define a function relating signal intensity to transcript abundance.
Project description:To characterize Klf4 downstream genes, we performed microarray analyses of the ES cells knocked down by siRNA of Klf4 gene. Keywords: individual genetic characteristic design ES cells grow without feeders on gelatin coated in complete ES medium. Cells were incubated at 37 0C; 5% CO2. ES cells were transfected with siRNA and pCAGEGFP-IP plasmid. After transfection, cells were cultured for 24 hours, following to drag selection for more 24 hours in the presence of puromycin to ensure concentration of transfectants. Total RNA were extracted with Trizol reagent (Sigma) according to the manufacturerâ??s instructions.