Figure S7 from Elevated Heme Synthesis and Uptake Underpin Intensified Oxidative Metabolism and Tumorigenic Functions in Non–Small Cell Lung Cancer Cells

Abstract

<p>Fig. S7 HSPs inhibit tumorigenic functions in A549 NSCLC cells. (A) HSPs inhibit migration in A549 NSCLC cells. (B) HSPs inhibit invasion in A549 NSCLC cells. The potent inhibitor of heme synthesis, succinyl acetone (SA), is shown for comparison. Addition of 10 μM heme largely reversed the effects of SA and HSPs. The images shown are cells that had migrated across Transwell inserts (in A) or had crossed invasion chambers coated with Corning Matrigel matrix and also passed Transwell inserts (in B). Data are plotted as mean {plus minus} SD. For statistical analysis, the levels in treated cells were compared to the levels in untreated cells with a Welch 2-sample t-test.*, p-value, 0.05, **, p-value < 0.005. For heme add-back experiments, the levels in cells treated with heme and SA or HSPs were compared to the levels in cells treated with only SA or HSPs. [**], p-value < 0.005. The effects SA, HSPs, heme oxygenase inhibitors, and iron chelator DFX on NSCLC cell colony formation are shown in C-G. (C) The effect of SA on colony formation in A549 cells. (D) The effect of HSP1 on colony formation in A549 cells. (E) The effect of HSP2 on colony formation in A549 cells. (F) The effects of heme oxygenase inhibitor SnPP and ZnPP on colony formation in A549 cells. (G) The effect of DFX on colony formation in A549 cells. (H) Comparison of the effects of HSP2 and DFX treatments on the levels of transferrin receptor TFRC. (I) Comparison of the effects of HSP2 and DFX treatments on the levels of ferroportin SLC40A1. In H & I, data from Western blotting analysis of proteins prepared from NSCLC cells treated with HSP2 or DFX for 6 days were shown. For statistical analysis, the levels in treated cells were compared to the levels in untreated cells (Control) with a Welch 2-sample t-test. **, p-value < 0.005.</p>