Fig. 9: Effectiveness of Salinomycin on human Gastric Organoids (hGOs).

A Three different concentrations of Sal were used (6, 12, 18 µM) to evaluate its effectiveness on cancerous hGOs. After 48 h of treatment significant reduction of cell viability was recorded as compared with vehicle. Three independent experiments were performed. B Alteration of hGOs morphology was observed after 48 h of treatment (magnification 20×) with Sal 6 µM, concentration used for all further experiments. Three independent experiments were performed. C Activation of apoptotic RCD was assessed by measuring caspase 3/7 activity. The plot shows the ratios between luminescence recorded in Sal vs vehicle treatment. Only hGO-02 did not show caspase activation. Three independent experiments were performed. D hGOs were treated with Sal and vehicle for 48 h and then stained with JC-1 fluorescent probe. The alteration of mitochondria membrane potential was evaluated by measuring the red to green fluorescence intensities ratio. A marked shift of this ratio was recorded for hGO-02. E A strong reduction of ferroptosis markers (SLC7A11 and GPX4) expression was evident in hGO-02. F Autophagy markers upregulation was found in all hGOs lines. Vinculin was used as housekeeping gene for both western blots (E, F). One-sample t-test was employed to estimate significance (*p < 0.05, **p < 0.01, ***p < 0.001).