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Int J Clin Exp Pathol. 2015 May 1;8(5):4662-72. eCollection 2015.

Ghrelin inhibition of ethanol-induced gastric epithelial cell apoptosis is mediated by miR-21.

Author information

1
Department of Gastroenterology, Jinshan Hospital of Fudan University Shanghai, China.
2
Department of TCM, Jinshan Hospital of Fudan University Shanghai, China.

Abstract

AIM:

To investigate the underlying mechanism of ghrelin-induced gastro-protection in a cell culture model of ethanol-induced gastric epithelial cell injury.

METHODS:

The human gastric epithelial cell line GES-1 was incubated with ghrelin (0.01-1 µM), 1 µM ghrelin and 1 µM D-Lys3-growth hormone releasing peptide-6 (GHRP-6), or 1 µM ghrelin and 400 nM antagomiR-21 for 24 h, followed by treatment with 8% ethanol for 3 h to induce apoptosis. Cell viability was determined by MTT assays and flow cytometry was used for detection of apoptosis rates. miR-21 transcription was analyzed by qRT-PCR and Akt, Bcl-2, Bax and caspase 3 expressions were measured by Western blot.

RESULTS:

Flow cytometry and a quantitative RT-PCR analysis of the expression of miR-21 showed that ghrelin inhibited apoptosis in a dose dependent manner through a signaling pathway that was both growth hormone secretagogue receptor (GHS-R) and miR-21 dependent, as the antiapoptotic effect of ghrelin was blocked by both D-Lys3-GHRP-6 and antagomiR-21, respectively. Western blotting of Akt, Bcl-2, Bax, and caspase 3 showed that the levels of the antiapoptotic proteins, Akt and Bcl-2, in the cells pretreated with ghrelin alone were higher than those in the cells pretreated with D-Lys3-GHRP-6 or antagomiR-21. By contrast, the levels of the proapoptotic proteins, Bax and caspase 3, in the cells pretreated with ghrelin alone were lower than those in the cells pretreated with D-Lys3-GHRP-6 or antagomiR-21.

CONCLUSION:

Ghrelin inhibits GES-1 cell apoptosis through GHS-R-dependent signaling in which miR-21 activates the PI3K/Akt pathway, which upregulates Bcl-2 and downregulates Bax and caspase 3 expression.

KEYWORDS:

Ghrelin; PI3K/Akt; apoptosis; gastric epithelial cells; miR-21

PMID:
26191156
PMCID:
PMC4503028
[Indexed for MEDLINE]
Free PMC Article

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