Fig. 4.

INSM1 knockdown induces transcription of progenitor markers decreasing islet proliferation in wild-type p53 PanNET cells. (A) Graph showing cell viability of NT3 (p53 wild-type) and Bon1 (p53 mutated) cells. Values are mean of triplicates ± SD. (B, C) Western blotting analysis showing that INSM1 protein knockdown results in a decrease in ERK and AKT protein expression and phosphorylation in NT3 cells (B), but not in Bon1 cells (C). (D, E, F) NT3 (D) and Bon1 (E) cells were treated with INSM1 siRNA and analyzed by propidium iodide (PI) staining and flow cytometry to quantify cells in G1, S and G2 phase. Graphs shows cell cycle distribution in NT3 and Bon1 cells (F). Values represent mean of triplicates ± SD. ***P-value < 0.001 as determined by Fisher's exact test. (G, H) qRT-PCR analysis of cell cycle regulators (p38, MEN1, p57, p27, Cyclin D1) (G), and stem cell-related genes (CD133, FOXA2) (h) in NT3 and Bon1 cells treated with INSM1 or control siRNAs. Values were calculated using cDNA from untreated cells as the calibrator sample (logFC = 1). Values represent mean of triplicates ± SD. *P-value < 0.05, **P-value < 0.01, ***P-value < 0.001 as determined by t test.