Fig. 5
Pancreatic cancer-derived PGE2 induces SC activation. a Representative IF images of DiI and PTGER1/2/4 in RSC96 cells. Scale bars: 20 μm (left), 5 μm (right). b Representative F-actin IF images of RSC96 and sNF96.2 cells after treatment with different doses of PGE2 (0 nM, 0.1 nM, 1 nM, or 10 nM). Scale bar: 50 μm. c, e–g Protein expression levels of p75NTR, c-Jun, and SOX2 in RSC96 cells treated with different doses of PGE2 (0 nM, 0.1 nM, 1 nM, or 10 nM). Relative expression levels were quantified as p75NTR/GAPDH and c-Jun/GAPDH; since the molecular weight of SOX2 (~35 kDa) is close to that of GAPDH (~36 kDa), the relative expression of SOX2/β-actin was quantified (n = 3). d, h–j Representative IF images of p75NTR, c-Jun, and SOX2 in RSC96 cells after PGE2 intervention. Scale bar: 100 μm. Statistics of the relative fluorescence intensity between the control and treatment groups are shown (n = 3). k, o-q Protein expression levels of p75NTR, c-Jun, and SOX2 in RSC96 cells (RSC96-PANC-1 coculture system) following CAY10526 treatment and PGE2 supplementation (n = 3). l, r–t Protein expression levels of p75NTR, c-Jun, and SOX2 in RSC96 cells (RSC96-BxPC-3 coculture system) following CAY10526 treatment and PGE2 supplementation (n = 3). m Effects of CAY10526 treatment and PGE2 supplementation on 3D cocultures of RSC96 cells with HPNE, PANC-1 or BxPC-3 cells (n = 3).Scale bar: 200 μm. +B/P/H: +BxPC-3/PANC-1/HPNE, + DMSO:+BxPC-3/PANC-1/HPNE + DMSO, + CAY:+BxPC-3/PANC-1/HPNE + DMSO + CAY10526, + CAY + PGE2:+BxPC-3/PANC-1/HPNE + DMSO + CAY10526 + PGE2. n, u, v effects of CAY10526 treatment and PGE2 supplementation on PANC-1/BxPC-3-DRG cocultures, along with quantification of the DRG growth index (n = 3). Scale bar: 500 μm. +B/P: +BxPC-3/PANC-1, +DMSO: +BxPC-3/PANC-1 + DMSO, +CAY: +BxPC-3/PANC-1 + DMSO + CAY10526, +CAY+. PGE2: +BxPC-3/PANC-1 + DMSO + CAY10526 + PGE2. All the results are presented as the means ± SDs. Each data point in the bar graphs represents an individual sample. Statistical significance was determined by one-way analysis of variance followed by Tukey’s HSD post hoc test (for e–g, o–t, and u, v) and a two-tailed unpaired t test (for Fig. 5h-j). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001
