Identification of genetic networks involved in the cell growth arrest and differentiation of a rat astrocyte cell line RCG‐12

Abstract

The purpose of the present study is to establish and characterize a conditionally immortalized astrocyte cell line and to clarify the genetic networks responsible for the cell growth arrest and differentiation. A conditionally immortalized astrocyte cell line, RCG-12, was established by infecting primary cultured rat cortical glia cells with a temperature-sensitive simian virus 40 large T-antigen. At a permissive temperature of 33 degrees C, the large T-antigen was expressed and cells grew continuously. On the other hand, the down-regulation of T-antigen at a non-permissive temperature of 39 degrees C led to growth arrest and differentiation. The cells expressed astrocyte-expressed genes such as glial fibrillary acidic protein. Interestingly, the differentiated condition induced by the non-permissive temperature significantly elevated the expression levels of several astrocyte-expressed genes. To identify the detailed mechanisms by which non-permissive temperature-induced cell growth arrest and differentiation, we performed high-density oligonucleotide microarray analysis and found that 556 out of 15,923 probe sets were differentially expressed 2.0-fold. A computational gene network analysis revealed that a genetic network containing up-regulated genes such as RB, NOTCH1, and CDKN1A was associated with the cellular growth and proliferation, and that a genetic network containing down-regulated genes such as MYC, CCNB1, and IGF1 was associated with the cell cycle. The established cell line RCG-12 retains some characteristics of astrocytes and should provide an excellent model for studies of astrocyte biology. The present results will also provide a basis for understanding the detailed molecular mechanisms of the growth arrest and differentiation of astrocytes.

MicroRNA-93 may control vascular endothelial growth factor A in circulating peripheral blood mononuclear cells in acute Kawasaki disease

“…Biotin-labeled cRNA was then synthesized from the cDNA using FT10AFYB Flash Taq Biotin RNA Labeling Kit (Affymetrix, Santa Clara, CA). After fragmentation, the biotinylated cRNA was hybridized to arrays (35). The arrays were washed, stained with streptavidin-phycoerythrin and scanned with a probe array scanner.…”

“…Hybridization intensity data were converted into a presence/absence call for each gene, and changes in gene expression between experiments were detected by comparison analysis. The data were further analyzed using GeneSpring GX 11.0 software (Silicon Genetics, Redwood City, CA) (35).…”

Hyperglycemia perturbs biochemical networks in human trophoblast BeWo cells

“…Gene expression was analyzed using a GeneChip system with a Human Genome U133 Plus 2.0 Array that was spotted with 54,000 probe sets (Affymetrix, Santa Clara, CA) as described [8]. Sample labeling, hybridization and staining were carried out according to the Eukaryotic Target Preparation protocol in the Affymetrix Technical Manual for GeneChip Expression Analysis.…”

Distinct Roles of Transforming Growth Factor-β-activated Kinase 1 (TAK1)-c-Rel and Interferon Regulatory Factor 4 (IRF4) Pathways in Human T Cell Lymphotropic Virus 1-transformed T helper 17 Cells Producing Interleukin-9

“…DNA Microarray-Total RNA was extracted from cells using RNAeasy Mini Kit (Qiagen, Valencia, CA). Gene expression was analyzed using a GeneChip system with Human Genome Array U133 plus 2.0 (Affymetrix, Santa Clara, CA) as described previously (22). In this study, six arrays were used: two for HuTsiLuc cells, two for HuT-siIRF4 cells, and two for HuT-siIRF3 cells (positive counter control).…”