Fig 1: TGF-ß2 supports invasiveness and LD formation in acidosis-adapted cancer cells.a Heatmap representation of the top 20 genes upregulated in the indicated acidosis-adapted cancer cells. b mRNA expression for TGFB2 in native and acidosis-adapted tumor cells. c, d Levels of active form of TGF-ß2 in crude extracts (membrane-bound) from native and acidosis-adapted tumor cells (c) or in conditioned media (secreted) from native SiHa cells exposed to acidic pH 6.5 for 24 h (d). e Representative immunoblotting for phosphorylated and total forms of Smad2 (Ser465/467) and Smad3 (Ser423/425) in native and acidosis-adapted SiHa cells (with or without treatment with 4 ng/ml TGF-ß2 for 6 h). f mRNA expression for TGFB2 in native SiHa cells following treatment with 4 ng/ml TGF-ß2 for 24 h. g Representative immunoblotting for TSP-1 in native and acidosis-adapted tumor cells. h Levels of active form of TGF-ß2 in conditioned media from native and acidosis-adapted SiHa cells following transfection of TSP1-targeting (or control) siRNA for 72 h. i–k Representative immunoblotting for phosphorylated and total forms of Smad3 (Ser423/425) (i), invasion capacity in Matrigel-coated Boyden chambers for 24 h (j) and LD content (k) for native and acidosis-adapted SiHa cells following treatment with 10 µM TGFß2-specific antisense oligonucleotide Trabedersen for 7 days or 2 µM TGF-ßRI inhibitor SB431542 for 24 h. l LD content in acidosis-adapted SiHa following treatment with 10 µM TGFß2-specific antisense oligonucleotide Trabedersen for 7 days in absence or presence of 4 ng/ml TGF-ß2 for 24 h. Data are represented as mean ± SEM of three independent experiments (with =6 technical replicates). Significance was determined by Student’s t-test (d, f), one-way ANOVA (l) or two-way ANOVA (b–h, j, k) with Bonferroni multiple-comparison analysis. *p < 0.05; **p < 0.01; ***p < 0.001; ns, not significant. Source data are provided as a Source Data file.
Fig 2: TGF-ß2 promotes FA uptake and TG accumulation into LD.a–c Abundance of neutral lipids (NL), phospholipids (PL) and free fatty acids (FFA) (a), abundance of saturated and monounsaturated fatty acids (SFA and MUFA, respectively) in the neutral lipid fraction (b), and LD content (c) in native SiHa cells after treatment with 4 ng/ml TGF-ß2 for 6 h. d, e 14C-palmitate uptake for 10 min in SiHa cells after treatment with 4 ng/ml TGF-ß2 for 6 h (d) and in acidosis-adapted SiHa cells following treatment with 10 µM Trabedersen for 7 days in absence or presence of 4 ng/ml TGF-ß2 for 24 h (e). f–h Representative immunoblotting for cell surface-localized CD36 and total biotinylated proteins in native and acidosis-adapted SiHa cells following treatment with 4 ng/ml TGF-ß2 for 6 h and 24 h (f), following treatment with 10 µM Trabedersen for 7 days or 2 µM SB431542 for 24 h (g) or with 4 ng/ml TGF-ß2 and 10 µM PKC-? pseudo-substrate inhibitor for 24 h (h). i, j Quantification of surface-localized CD36 in native and acidosis-adapted SiHa cells treated as indicated in (h). k, l mRNA (k) and protein expression of DGAT1 (l) in native and acidosis-adapted tumor cells. m–o Representative immunoblotting for DGAT1 in native and acidosis-adapted SiHa cells following treatment with 10 µM Trabedersen for 7 days or 2 µM SB431542 for 24 h (m), with 10 µM PKC-? pseudo-substrate inhibitor for 24 h (n) or with 10 µM GW6471 for 48 h (o). p mRNA expression of PLIN2 in native and acidosis-adapted SiHa cells following treatment with 10 µM TGFß2-specific antisense oligonucleotide Trabedersen for 7 days. q Co-expression analysis of TGFB2, PLIN1, PLIN2, and PLIN3 genes in human healthy volunteers and colorectal cancer patient samples. Data are represented as mean ± SEM of three independent experiments (with =6 technical replicates). Significance was determined by Student’s t-test (c, d, j), by one-way ANOVA (e–i) or two-way ANOVA (a, k, p) with Bonferroni multiple-comparison analysis. *p < 0.05; **p < 0.01; ***p < 0.001; ns, not significant. Source data are provided as a Source Data file.
Supplier Page from Abcam for Recombinant human TGF beta 2 protein (Active)