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. 2017 Feb 9;8(10):16851–16874. doi: 10.18632/oncotarget.15170

Figure 4. Stromal and epithelial breast tumor cells under PRP supplementation (including cells attached to the plastic surface and cells attached to the network of fibrin bundles) were assessed by immunoblotting.

Figure 4

(A) Fibroadenoma stromal cells under PRP supplementation showed increased phospho-Src, phosphos-FAK(Y)397, and phospho-ERK1/2, and upregulated TGF-β1 expression. The graph represents the densitometry analyses from immunoblotting results. (B) Phyllodes stromal cells under PRP supplementation showed EMT reversion and upregulation of E-cadherin and downregulation of N-cadherin expression after treatment with P4 and the E2+P4 combination. Phospho-Src and phospho-FAK(Y)925, but not FAK(Y)397, showed increased phosphorylation. Phase contrast–confluent culture of phyllodes stromal cells showed morphology alteration in the presence of steroid hormones under PRP supplementation. (C–E) Luminal A and B and HER2+ epithelial cells under PRP supplementation showed EMT process with downregulation of E-cadherin and upregulation of N-cadherin expression. Phospho-Src and phospho-FAK(Y)925, but not FAK(Y)397, showed increased phosphorylation, and the α6β1 integrin subunits showed decreased expression. Conversely, the αvβ5 integrin subunits showed increased expression. The analysis of the TGF-β, Smad2, and Snail metastasis markers showed increased expression in the highest PRP concentrations. The results are represented as band intensities in arbitrary units relative to the respective total load of total proteins and control (β-actin). The statistical significance was evaluated using one-way ANOVA followed by the Tukey's test. These experiments were performed with cultured primary cells from specimens collected from patients, *p ≤ 0.05, **p ≤ 0.001. (F) Concentration of total TGF-β in CM from breast tumor cells. The conditioned medium was collected, centrifuged to remove platelets, and the presence of TGFβ1 in the supernatant measured by ELISA. Each bar represents the mean ± SEM of n = 2–6. ***p < 0.001 as determined by Student's t test.