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Series GSE59775 Query DataSets for GSE59775
Status Public on Oct 31, 2014
Title Transcriptional analysis of follistatin influence on C2C12 mice myoblasts differentiation
Organism Mus musculus
Experiment type Expression profiling by array
Summary Background: Follistatin (FS) is an activin-binding glycoprotein, known for its antagonistic action on the TGF-β super-family of cytokines. It has ability to prevent muscle atrophy and to stimulate muscle growth through binding of myostatin (GDF8). FS does not cause these changes only by inhibition of GDF8 but this mechanism is still not well elucidated. In order to identify GDF8-dependent and GDF8-independent action of FS, we performed comprehensive transcriptomic analysis of differentiating C2C12 mouse myoblasts.
Results: To evaluate influence of FS-288 (200ng/ml) on differentiating C2C12 myoblasts was used immunofluorescence analysis of MyHC protein expression in the culture and counting of the number and type of myotubes. Differentiating myoblasts treated with FS showed increase in number and size of myotubes. Microarray studies performed by us identified 30 genes oppositely regulated by FS and GDF8. Analyses of the transcriptomic profiles showed that FS has highly significant influence, associated with GDF8 binding, on the regulation of cell cycle, transport processes, cell communication, cell adhesion, cell motion, developmental processes and binding functions. The signalling pathways that may play a primary role in the myogenesis stimulation by FS are associated with TGF-β, Delta-Notch and Wnt signalling pathways, integrin and actin cytoskeleton signalling and adipogenesis regulation. Changes in expression of Hgf, Sort1, Trdn, Stmn2, Hdac1 genes, significantly involved in the process of myoblasts differentiation were validated with Real-Time PCR method.
Conclusions: Our results implicate which aspects of transcriptional regulation of myogenesis process by FS could have significant and protective influence. We have shown that FS activity is associated not only with GDF8 binding but also with inhibited expression of genes involved in TGF-β signalling and by regulating of genes associated with vesicle transport apoptosis signalling pathway, inflammatory pathways and adipogenesis expression. Our analysis showed that follistatin GDF8-independent on differentiating myotubes involves regulation of localization processes, cell-matrix adhesion, mRNA processing, apoptosis, fatty acid metabolic processes, SNAP receptor activity functions and partial regulation of transcription processes. These results give a new and complete image of the follistatin role in the muscle cell differentiation and indicate target genes for FS which in the future may be a good therapeutic approach.
Overall design After scanning of hybridized microarrays, quantitation of slide images were performed using Feature Extraction Software (Agilent) using default parameters. Normalized raw data was exported to GeneSpring GX 11.0.5 (Agilent, Santa Clara, CA). For identification of genes significantly altered in cells treated with FSl compared with untreated cells. T total detected entities were filtered by flags (present, marginal) and error (coefficient of variation: CV < 50.0 percent) to remove very low signal entities and to select reproducible signal values of entities among the replicated experiments, respectively. In statistical analysis, separated for experiment with myoblasts treated with FS(FS vs. CTRL 1-4) was used t-test unpaired (p < 0.05) with multiple testing correction: Benjamini-Hochberg <0.05, all significant changes over fold change 1.6 were selected. Analysis of GO, GSEA and signaling pathway was carried out using GeneSpring GX 12 (Agilent) and the DAVID, KEEG and PANTHER Classification System (p<0.01). In the analysis of signaling pathways using GeneSpring GX 12 (Agilent) and Ariadne Pathway Studio.
Contributor(s) Wicik Z, Jank M, Motyl T
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Submission date Jul 25, 2014
Last update date Jan 12, 2017
Contact name Zofia Wicik
Organization name Mossakowski Medical Research Center
Department Department of Human Epigenetics
Street address Pawinskiego 5
City Warsaw
State/province mazovia
ZIP/Postal code 02-106
Country Poland
Platforms (1)
GPL7202 Agilent-014868 Whole Mouse Genome Microarray 4x44K G4122F (Probe Name version)
Samples (4)
GSM1446320 Myoblasts treated with follistatin-rep1
GSM1446321 Myoblasts treated with follistatin-rep2
GSM1446322 Myoblasts treated with follistatin-rep3
BioProject PRJNA256182

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Supplementary file Size Download File type/resource
GSE59775_RAW.tar 28.3 Mb (http)(custom) TAR (of TXT)
Processed data included within Sample table

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