Figure 4. PRRX1 knock down decreased cell proliferation.
(A) Dot plots with median showing PRRX1a (black) and PRRX1b (white) mRNA expression relative to the siControl condition (red dashed line) in control (circle) and IPF (square) fibroblasts (n=8 per group) treated for 72 hr with PRRX1 siRNA (#1 or #2). (B) Immunoblot showing PRRX1 expression (n=4) in control and IPF fibroblasts treated 72 hr with PRRX1 siRNA (#1 or #2) or siControl. The quantification of PRRX1 expression relative to GAPDH (loading control) is displayed as dot plot with median. (C) Dot plots with median showing the relative growth ratio of control (n=6) and IPF (n=8) fibroblasts stimulated 72 hr with FCS 10% and treated with PRRX1 siRNA compared to siControl. (D) Dot plots with median showing the percent of cells in G1 (right) or S (left) phase in control and IPF fibroblasts (n=5) stimulated 72 hr with FCS and PRRX1 siRNA relative to siControl. (E) Dot plots with median showing mRNA expression of CCNA2 and CCNE2 relative to siControl in control and IPF fibroblasts stimulated 72 hr with FCS and treated with PRRX1 siRNA (n=10). (F) Dot plots with median showing the percent of cells positive for Ki67 marker in control and IPF fibroblasts stimulated 72 hr with FCS 10% and treated with PRRX1 siRNA relative to siControl (n=5). (G) ChIP analysis for PRRX1 recruitment at the promoter of GAPDH, CCNA2, CCNE2, and MKI67 in NHLF (n=5) relative to RNA POL-II occupancy, displayed as boxes with median and min to max. The diagrams of the different loci are showing the PRRX1 response element position relative to the TSS. The PCR amplified regions are underscored. Abbreviations: FCS (fetal calf serum); TSS (transcription starting site); IgG (Immunoglobulin); PRE (PRRX1 responses element); SRE (SRF response element), control siRNA sequence (siControl). Wilcoxon signed-rank test, $ p≤0.05, Wilcoxon matched-paired signed rank test ** p<0.01.
Figure 4—figure supplement 1. PRRX1 inhibition impacted fibroblast proliferation.
