Fig. 4

Pancreatic organoid-derived epithelial monolayers recapitulate ductal anion secretory pathways. A The effect of forskolin (F), secretin (S) and VIP (V) on PKA-mediated phosphorylation of VASP (top panel) and total VASP (bottom panel) levels was assessed by Western blot analysis. Detection of β-actin served as a loading control. Note that phosphorylation of Ser-157 decreases the electrophoretic mobility of VASP. Outer left lane shows the position of molecular weight markers, with an approximate molecular mass as indicated. B Representative experiments showing the effect of secretin, VIP, forskolin and PPQ-102 on the anion secretory current. C The cAMP-dependent Isc responses in nine organoid lines obtained from different animals. Each data point represents one technical replicate. D Forskolin-dependent Isc responses in the presence or absence of bumetanide (Bu). Each data point represents one technical replicate. Data were obtained from two organoid lines. **P= 0.006. Statistical analysis: paired t-test. E Forskolin-dependent Isc responses of monolayers bathed in Meyler solution (M), or in bathing solution containing only bicarbonate as the CFTR-permeating anion. Bicarbonate secretion was assayed at luminal bicarbonate concentrations of 20 or 100 mmol/L, as indicated. Each data point represents one organoid line. F Representative experiment demonstrating the forskolin-dependent Isc response at luminal bicarbonate concentrations of 20 or 100 mmol/L. G Effect of UTP on the anion secretory current. The inset shows the peak UTP-dependent Isc response, assessed in the presence of PPQ-102, in nine organoid lines