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Fig. 1 | Cell Communication and Signaling

Fig. 1

From: Cancer-associated fibroblasts promote prostate tumor growth and progression through upregulation of cholesterol and steroid biosynthesis

Fig. 1

Gene expression patterns of PCa spheroids with and without cancer-associated fibroblasts (CAFs). a Representative phase contrast images are shown for LNCaP and DuCaP PCa cells grown either in 75cm2 culture flasks (2D) or as 3D spheroids in 96 well hanging drop plates in the absence and (b) presence of GFP-expressing CAFs (magnification 10x, scale bar: 500 μm). Alterations in mRNA gene expression were determined via Illumina microarray analysis and the number of differentially expressed genes was represented in Venn diagrams. To determine gene expression changes that occur in PCa cells upon co-culture with CAFs, CAF-specific genes were subtracted. c Expression of HMGCS2 and AKR1C3 was validated by real-time RT-PCR in LNCaP and DuCaP monoculture spheroids and in LNCaP and DuCaP cells after spheroid co-culture with CAFs (LN/CAF, Du/CAF). Separation of cells was performed by flow cytometry-assisted cell sorting based on GFP expression in CAFs as described under methods. Values are expressed as relative expression of the gene of interest (GOI) normalized to hydroxymethylbilane synthase (HMBS). d Simplified overview of cholesterol and steroid biosynthesis metabolism. e Western blot analysis of HMGCS2 and AKR1C3 in mono vs co-culture spheroids. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as loading control. Quantification of bands was performed with Image Studio (Li-Cor) and the ratio between the protein of interest and GAPDH was blotted in a graph. Data are represented as mean ± SEM from at least three independent experiments. (* P < 0.05, ** P < 0.01)

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