Fig. 4

Panel a: CD248 knockdown with specific siRNA significantly reduced BrdU uptake in NHLFs (n = 3 different NHLF batches, total replicates 18, p < 0.001 with Student’s non-equal varienace t-test). Cells were incubated with 30 pmol of control or CD248-specific siRNA for 48 h. BrdU was added according to the manufacturer’s recommendation (Calbiochem). BrdU incorporation was detected using an indirect ELISA method. Panel b. CD248 expression in NHLF cells measured with flow cytometry (cells from n = 5 different donors). CD248-specific siRNA caused significant (*p < 0.05, non-equal variance t-test) drop in the protein expression levels when compared to treatment with scrambled control siRNA. Median Fluorescent intensity and Standard error of the mean (SEM) is plotted on the graph. Panel c. Intracellular Collagen 1a1 expression levels in NHLF cells measured by flow cytometry (cells from n = 5 different donors). TGF-β1 treatment caused a significant elevation in collagen 1 synthesis (*p < 0.01, non-equal variance t-test) but we measured no significant difference when cells were treated with CD248-specific siRNA. Median Fluorescent intensity and Standard error of the mean (SEM) is plotted on the graph. Panel d. Intracellular alpha smooth muscle (aSMA) expression levels in NHLF cells measured by flow cytometry (cells from n = 5 different donors). TGF-β1 treatment caused significant increase in aSMA expression in NHLF cells regardless of siRNA treatment siRNA (p < 0.05, non-equal variance t-test). Median Fluorescent intensity and Standard error of the mean (SEM) is plotted on the graph