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. 2017 Sep 21;8(52):89775–89792. doi: 10.18632/oncotarget.21128

Figure 6. SMA correlates to NCSC gene signature expression and aggressiveness in stage 4/M NBs.

Figure 6

(A-D) Association between SMA protein expression and histopathological features typical of stage 4/M NBs. (A) Representative images of anti-SMA immunohistochemistry in sections from Stage 1–2 vs. Stage 4 tumors. Scale bars: 100 μm. (B) Expression of SMA (% of stained area) was calculated as in Figure 4E. Aggressive neuroblastomas (Stage 4/M) showed higher expression levels than low-risk tumors (Stage 1 and 2). (C) Representative images of anti-SMA immunohistochemistry in stage 4/M tumor sections before and after treatment. Scale bars: 100 μm. (D) Quantification of SMA expression (% of stained area) in pre- and post-treatment tumor sections. Concomitantly to treatment-induced tumor differentiation (considered as an indicator of treatment response), we observed a significant decrease in SMA expression levels. (E) ACTA2 (gene encoding for SMA) expression correlates with mesenchymal genes (ENG, TGFB1, S100A10 and VIM). (F) Parametric analysis of gene set enrichment (PAGE) revealed that tumors with higher expression of ACTA2 were also enriched in expression of typical NCSC genes (gene set #1, described in Lee et al. [13]) and MSC genes (gene set #2, described in Boquest et al. [50]). Heat map (upper panel) shows the average z-scores for all genes in every single gene set, calculated for each individual tumor sample. (G) Quantification of the expression of NCSC or MSC gene signatures in ACTA2 high compared to ACTA2 low tumors. (H) Correlation between ACTA2 expression and NCSC gene signature average z-scores over the whole sample of tumors analyzed. (I) Tumors enriched in expression of NCSC genes (NCH) show higher expression levels of ACTA2. (J) Kaplan-Meier curves showing the association between high ACTA2 expression levels and lower event-free survival probability. (K) Kaplan-Meier curves showing the event-free survival probability of NCI tumors segregated into ACTA2 high (48%) and ACTA2 low (52%) tumors. (r-value = Pearson's correlation coefficient) (****p < 10–4, *p < 0.05; Mann-Whitney U-test). See also Supplementary Figure 6.