Figure 5.

Effects of increasing concentrations of exogenous PEA in in vitro duodenal biopsy specimens from controls. (A) Immunohistochemical images showing toluidine-positive cells (arrows) and (B) relative quantification of MCs in duodenal mucosa deriving from control cultured biopsy specimens at (1) pH = 3.0, in the presence of (2) exogenous PEA (0.1 μmol/L), co-incubated with either (3) PPARα antagonist MK866 (3 μmol/L) or (4) PPARγ antagonist (GW9662 9 nmol/L). Original magnification: 20×. Data show the number of MCs counted per square millimeter of tissue. Immunofluorescence staining of NeuN (green) and (C) tryptase-positive, (E) TRPV1-positive, and (G) TRPV4-positive cells (all red) and relative graph bars quantifying (D) tryptase-positive, (F) TRPV1-positive, (H) and TRPV4-positive cells deriving from controls biopsy specimens, cultured in the same experimental conditions. Original magnification: 20×. Data show the number of immune-reactive cells counted per square millimeter of tissue. (I) Immunoblot analysis and relative densitometric analysis (arbitrary units normalized on the expression of the housekeeping protein β-actin) quantifying (J) TRPV1 and (K) TRPV4 protein expression in tissue homogenates deriving from control cultured biopsy specimens at (1) pH = 3.0, in the presence of increasing concentrations of exogenous PEA (2) 0.001 μmol/L, (3) 0.01 μmol/L, (4) 0.1 μmol/L alone or co-incubated with either (5) PPARα antagonist MK866 (3 μmol/L) or (6) PPARγ antagonist (GW9662 9 nmol/L). (L–O) ELISA essays quantifying, respectively, the release of tryptase, NGF, histamine, and PGD2 in dyspeptic biopsy specimens, cultured in the same experimental conditions. ∗P < .05 for PEA 0.001 μmol/L, ∗∗P < .01 for PEA 0.01 μmol/L, and ∗∗∗P < .001 for PEA 0.1 μmol/L vs acid challenge and for co-incubation with PPARγ antagonist GW9662 vs acid challenge; °°°P < .001 for co-incubation with PPARα antagonist MK866 vs acid challenge. All results are means ± SD of n = 10 control subjects.