Bioactive peptides from barley protein hydrolysate stimulate cholecystokinin secretion <i>via</i> calcium-sensing receptor signaling in enteroendocrine cells

Anorexigenic hormone cholecystokinin (CCK), secreted by enteroendocrine cells (EECs), is known for its inhibitory role in appetite. The secretion of CCK is greatly affected by dietary ingredients. The search for functional components in food that stimulate CCK secretion is of great significance for weight control. Our previous study has demonstrated that barley protein hydrolysate (BPH) strongly stimulates CCK secretion from EECs <i>in vitro</i>, but the structural characteristics of the responsible components and their underlying mechanism of action have not yet been elucidated. Here, we monitor the plasma levels of CCK in mice after ingestion of BPH to demonstrate its <i>in vivo</i> activity in stimulating CCK secretion. Using size exclusion column chromatography and liquid chromatography-tandem mass spectrometry, we isolate and identify three novel CCK secretion-stimulating peptides, including VQVQIPF, VVTGVGGQ and PQQPQPFFQ. Among them, the peptide VVTGVGGQ exhibits the strongest activity in stimulating CCK secretion. Inhibition experiments show that the calcium-sensing receptor (CaSR) is required for VVTGVGGQ to increase CCK gene transcription and secretion. Blockage of CaSR by NPS-2143 completely abolishes the activity of VVTGVGGQ. Moreover, the downstream signaling molecules including CaSR coupled G-protein subtype G_q and intracellular Ca²⁺ are also involved in VVTGVGGQ-stimulated CCK secretion. Inhibition of G_q and intracellular Ca²⁺ completely and partially abolish this peptide-induced CCK secretion, respectively. Our findings highlight the potential of barley protein-derived hydrolysate and peptides as functional food ingredients for CCK secretion stimulation and weight control.