Role of ghrelin in the pancreatic exocrine secretion via mitogen-activated protein kinase signaling in rats

Abstract

Background: This study was performed to investigate the impact of exogenous ghrelin on the pancreatic α-amylase outputs and responses of pancreatic proteins to ghrelin that may relate to pancreatic exocrine.

Methods: Sprague-Dawley male rats (9 weeks old, 300 ± 10 g) were injected with ghrelin via intraperitoneal (i.p.) infusion at dosage of 0, 0.1, 1.0 and 10.0 μg/kg body weight (BW), respectively. The plasma ghrelin and cholecystokinin (CCK) level were determined using enzyme immunoassay kit; the mRNA expression of ghrelin receptor (GHSR-1α) and growth hormone (GH) receptor were assessed by reverse transcription PCR; the expressions of pancreatic α-amylase activity, extracellular-signal-regulated kinases (ERK), phosphorylated extracellular-signal-regulated kinases (pERK) and c-Jun N-terminal kinase (JNK) were evaluated by western blotting; moreover the responses of pancreatic proteins to ghrelin were analyzed using the two-dimensional gel electrophoresis system.

Results: The exogenous ghrelin (1.0 and 10.0 μg/kg BW) elevated the level of plasma ghrelin (p < 0.05), and suppressed the expression of pancreatic α-amylase at a dose of 10.0 μg/kg BW (p < 0.05). No difference in the level of plasma CCK was observed, even though rats were exposed to any dose of exogenous ghrelin. In addition, a combination of western blot and proteomic analysis revealed exogenous ghrelin (10.0 μg/kg BW) induced increasing the JNK and ERK expressions (p < 0.05) and four proteins such as Destrin, Anionic trypsin-1, Trypsinogen, and especially eukaryotic translation initiation factor 3 in rat pancreas.

Conclusions: Taken together, exogenous ghrelin by i.p. infusion plays a role in the pancreatic exocrine secretion via mitogen-activated protein kinase signaling pathway.

Keywords: Cholecystokinin; Ghrelin; Pancreatic exocrine; Sprague-Dawley rats; α-Amylase activity.

Fig. 1

Concentration of plasma ghrelin (a) and cholecystokinin (CCK) (b) in rat exposure to exogenous ghrelin (0, 0.1, 1.0 and 10.0 μg/kg BW). The values are expressed as the average with SEM (n = 4). Asterisk shows significant difference with the ghrelin-untreated group (*p < 0.05, **p < 0.01)

Fig. 2

Western blot analysis of α-amylase expressions in tissues of pancreas of rat exposure to exogenous ghrelin (0, 0.1, 1.0 and 10.0 μg/kg BW). The values are expressed as the average with SEM (n = 4). Asterisk shows significant difference with the ghrelin-untreated group (*p < 0.05)

Fig. 3

mRNA expressions of pancreatic ghrelin receptor (GHSR-1α) and growth hormone (GH) receptor in tissues of pancreas, liver, and pituitary of rat exposure to exogenous ghrelin (0 and 10.0 μg/kg BW) using RT-PCR

Fig. 4

Level of pancreatic extracellular-signal-regulated kinases (ERK), phosphorylated extracellular-signal-regulated kinases (pERK) and c-Jun N-terminal kinase (JNK) expression by using western blotting. Protein sample from pancreatic tissue of rats treated with ghrelin (10.0 μg/kg BW) was analyzed by western blot (a) and their bands were determined by pixel intensity analysis using NIH Image J (b). The values are expressed as the average with SEM (n = 4). Asterisk shows significant difference with the ghrelin-untreated group (*p < 0.05, **p < 0.01)

Fig. 5

Identification of differentially altered protein spots in rat pancreas by ESI/Q-TOF MS. Representative silver-stained 2-DE images of the normal rat pancreas (left) and 10.0 μg/kg BW ghrelin-infused rat pancreas (right) (a). The spots that differentially expressed proteins in rat pancreas were identified by ESI/Q-TOF MS (b). ¹⁾ The values of the protein expression were presented as the ghrelin-treated group against ghrelin-untreated group