Bone Marrow derived macrophages (BMDM) were grown using L929 conditioned medium. On day 7, BMDM were mock-treated (16x 15cm dishes) or treated (16x 15cm dishes) with 10U/ml interferon gamma at time zero. 4-thiouridine (4sU) was then used to label newly transcribed RNA every 30 minutes for 8h (0 to 30, 30 to 60 etc. to give 16 discrete successive labellings) in mock and interferon gamma-treated samples. 16x Mock and 16x treated total samples were then isolated (32 in total) and newly transcribed RNA isolated from each total RNA sample using a protocol described in Dolken et al (PMID: 18658122). For analysis, 32 (16x mock and 16x treated) newly-transcribed RNA samples (every 30 minutes for complete 8h time course) and 16 (8x mock and 8x treated) total RNA samples (every hour for 8h time course) were hybridized to Affymetrix GeneST v1.0 arrays.
Interferon gamma treatment of macrophages results in hundreds if not thousands of alterations in gene expression and an antiviral state being established in these cells. Little is known about relationship between transcript synthesis, abundance and decay in macrophages during the first hours after interferon gamma treatment and how these factors influence the antiviral cellular phenotype.
Microarrays were used to analyse alterations in transcript synthesis and abundance in BMDM treated with Interferon Gamma for 8h.
Robertson KA, Ghazal P (Division of Infection and Pathway Medicine, University of Edinburgh)