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Series GSE20625 Query DataSets for GSE20625
Status Public on Jul 06, 2010
Title Impact of MAA on mouse Leydig cell gene expression (Mus musculus)
Organism Mus musculus
Experiment type Expression profiling by array
Summary Methoxyacetic acid (MAA) is the active metabolite of the widely used industrial chemical ethylene glycol monomethyl ether, an established testicular toxicant. MAA induces the degradation of testicular germ cells in association with changes in gene expression in both germ cells and Sertoli cells of the testis. This study investigates the impact of MAA on gene expression in testicular Leydig cells, which play a critical role in germ cell survival and male reproductive function. Cultured mouse TM3 Leydig cells were treated with MAA for 3, 8, and 24 h and global gene expression was monitored by microarray analysis. A total of 3,912 MAA-responsive genes were identified. Ingenuity Pathway analysis identified reproductive system disease, inflammatory disease and connective tissue disorder as the top biological functions affected by MAA. The MAA-responsive genes were classified into 1,366 early responders, 1,387 mid-responders, and 1,138 late responders, based on the time required for MAA to elicit a response. Analysis of enriched functional clusters for each subgroup identified 106 MAA early response genes involved in transcription regulation, including 32 genes associated with developmental processes and 60 DNA-binding proteins that responded to MAA rapidly but transiently, and which may contribute to the downstream effects of MAA seen for large numbers of mid and late response genes. Genes within the phosphatidylinositol/phospholipase C/calcium signaling pathway, whose activity is required for potentiation of nuclear receptor signaling by MAA, were also enriched in the set of early MAA response genes. These findings on the progressive changes in gene expression induced by MAA in Leydig cells may help elucidate the signaling pathways perturbed by this testicular toxicant and explain its mechanism of toxicity at the gene level.
 
Overall design Mouse TM3 Leydig cells (American Type Culture Collection, Manassas, VA) were grown in DMEM-F12 medium containing 5% horse serum and 2.5% FBS. Cells were grown to ~60% confluence and treated with culture medium alone, or with culture medium containing 1 mM or 5 mM MAA for either 3, 8 or 24 h. Total RNA was then isolated using TRIzol reagent, followed by incubation with RQ1 RNAse-free DNAse for 1 h at 37°C and then heating at 75°C for 5 min using the manufacturer’s protocol. A total of 6 cultures of TM3 cells were independently treated with MAA under each of the 6 treatment conditions specified above (i.e., 1 mM or 5 mM MAA for either 3, 8 or 24 h), and the corresponding 6 sets of RNA samples were validated by RNA integrity analysis (Agilent Bioanalyzer). Each RNA sample was also validated by qPCR analysis using SYBR Green I-based chemistry and primers specific for 3 genes known to respond to MAA (Cyp17a1, Shbg, and Igfbp3) to verify consistency of the MAA responses. The 6 RNA samples were then used to prepare two independent pools (n=3 RNA samples each) for microarray analysis with dye swaps. Sample labeling, hybridization to microarrays, scanning and calculation of normalized expression ratios were carried out at the Wayne State University Institute of Environmental Health Sciences microarray facility using Alexa 555 and Alexa 647 aminoallyl-aRNA samples
 
Contributor(s) Bagchi G, Zhang Y, Waxman DJ
Citation(s) 20565877
Submission date Mar 04, 2010
Last update date May 10, 2018
Contact name David J. Waxman
E-mail(s) djw@bu.edu
Organization name Boston University
Department Department of Biology and Bioinformatics Program
Street address 5 Cummington Mall
City Boston
State/province MA
ZIP/Postal code 02215
Country USA
 
Platforms (1)
GPL4134 Agilent-014868 Whole Mouse Genome Microarray 4x44K G4122F (Feature Number version)
Samples (12)
GSM518026 3h-MAA 1mM/3h-Control mouse TM3 cells/Replicate #1
GSM518027 24h-MAA 5mM/24h-Control mouse TM3 cells/Replicate #1
GSM518028 24h-MAA 1mM/24h-Control mouse TM3 cells/Replicate #1
Relations
BioProject PRJNA124815

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE20625_RAW.tar 140.2 Mb (http)(custom) TAR (of TXT)
Processed data included within Sample table

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