RNAi-mediated Ghrelin affects gastric H+–K+-ATPase activity and expression of GOAT–Ghrelin system in vitro

• shGhrelin inhibited ghrelin mRNA expression via an RNAi mechanism.
• shGhrelin reduced total and acylated ghrelin production and secretion (P < 0.05).
• GOAT mRNA expression was down-regulated (P < 0.05) in shGhrelin group.
• Ghrelin gene silencing inhibited the activity of H+–K+-ATPase and pepsin (P < 0.05).

Ghrelin has been implicated in the regulation of gastric functional development, and its physiological functions are mediated by Ghrelin-O-acyltransferase (GOAT) which is capable of generating the active form of this polypeptide hormone. However, whether and how ghrelin gene silencing may modify gastric acid secretion and GOAT–Ghrelin system is yet to be explored. The study was performed in gastric mucosal cells from weanling piglets in vitro. We evaluated the effect of ghrelin on gastric acid secretion, gene expression of GOAT and ghrelin as well as ghrelin levels by RNA interference assay. shGhrelin triggered the down-regulation of ghrelin mRNA expression (P < 0.05) via an RNAi mechanism, as observed by real-time RT-PCR. In addition, shGhrelin showed reduced total ghrelin production and secretion (P < 0.05) using ELISA in vitro. We also detected that GOAT mRNA expression was reduced in shGhrelin group (P < 0.05), compared with control groups. In accordance with the GOAT expression, acylated ghrelin production and secretion were reduced in gastric mucosal cells and culture medium (P < 0.05). Silencing of ghrelin gene achieved by RNAi-mediation inhibited the activity of H+–K+-ATPase and pepsin (P < 0.05) in gastric mucosal cells. These results indicated that RNAi of Ghrelin gene inhibited the gastric acid secretion with decreased GOAT mRNA and acylated Ghrelin in gastric mucosal cells.