Fig. 3: Effects of PRRX1 knockdown on the proliferation or tumorigenesis of human MPNST cells.

a Comparison of PRRX1 mRNA among human MPNST cell lines by qPCR analysis. Total RNA was extracted from human foetal lung fibroblasts (MRC-5) or the human MPNST cell lines FMS-1, HS-Sch-2 and HS-PSS to compare the expression level of PRRX1 mRNA. All values were normalised to ACTB mRNA levels (n = 3). b Western blot analysis after PRRX1 knockdown. HS-PSS cells were infected with lentivirus encoding each shPRRX1 clone and total cell lysates were extracted to compare the expression level of PRRX1. c Comparison of proliferative capacity after PRRX1 knockdown by WST-8 assay. (n = 7, three independent experiments). d Comparison of the RNA transcriptome after PRRX1 knockdown. RNA-seq analysis of HS-PSS/Control and HS-PSS/shPRRX1#1 and #2 was performed and data were compared by gene set enrichment analysis (GSEA). Genes upregulated in HS-PSS/Control are clustered on the left (n = 2, two independent experiments). e, f, g Comparison of tumour volume and weight after PRRX1 knockdown. Tumour volume was measured at each indicated time point after subcutaneous transplantation into NOD-SCID mice. At 56 days after transplantation, mice were sacrificed and the weights of the developed tumours were measured (n = 6, three independent experiments). h Immunohistological analysis of developed xenograft tumours. Tumour sections were stained with hematoxylin and eosin (H&E), immunostained for PRRX1 or Ki-67, or stained by TUNEL. Representative photomicrographs are shown. Data are presented as the means ± SEMs. * p < 0.05; ** p < 0.01; *** p < 0.001.