Figure 2

TNFα treatment increases expression of IL-1β, IL-6 and COX-2 expression in differentiated adipocytes. 3T3-L1 preadipocytes were differentiated into adipocytes as in Fig. 1. Following differentiation, cells were incubated with TNFα for the indicated times. Total RNA was extracted and gene expression levels were measured by qRT-PCR using the Comparative C_T method. Amplification of β-actin sequence was performed in parallel and used to normalize values obtained for target genes. Expression levels for IL-6 and COX-2 were determined by comparing values obtained at each time point following TNFα stimulation to values obtained from untreated cells. Expression of these genes in untreated adipocytes was assigned a value of 1 to determine fold-changes in expression due to TNFα stimulation. Expression of IL-1β resulting from TNFα stimulation was assessed by comparing later time points (20 and 62 h) to 10 h TNFα treatment time since no IL-1β expression could be measured in untreated adipocytes. For this analysis, IL-1β expression measured at 10 h post-TNFα treatment was assigned a value of 1 to determine fold-changes in expression at later time points.