N-glycosylation of somatostatin receptor type 2 protects rats from acute pancreatitis

BACKGROUND: The growth hormone inhibitor somatostatin and its analogs are promising therapeutic agents for acute pancreatitis. However, the therapeutic effects remain controversial. Somatostatin analogs preferentially bind to somatostatin receptor 2 (SSTR2), and this study aimed to investigate whether N-glycosylation affects SSTR2 stability, membrane trafficking, and signal transduction. METHODS: Western blot analysis following PNGase F digestion was performed to confirm N-glycosylation of SSTR2 in rat pancreatic acinar AR42J cells. Rats were subjected to 4 hourly intraperitoneal injections of cerulein (50 µg/kg) plus LPS (5 µg/mL) to induce acute pancreatitis. Mass spectrometry was conducted to identify the glycosylation sites of SSTR2, and immunofluorescent staining was carried out to examine the localization of wild-type and asparagine 9 (N9)Q-mutant SSTR2. Proteasome inhibitor MG132 was employed to assess the stability of SSTR2, and overexpression of wild-type or N9Q-mutant SSTR2 was used to examine the role of N-glycosylation in SSTR2 signal transduction in vitro and its therapeutic effect on pancreatitis in rats. RESULTS: SSTR2 protein in AR42J cells was N-glycosylated at the N9 residue. Wild-type SSTR2 localized in the plasma membrane whereas N9Q-mutant SSTR2 was retained in the endoplasmic reticulum (ER). MG132 rapidly restored the ectopic expression of mutant but not wild-type SSTR2 protein in AR42J cells. Overexpression of wild-type but not N9Q-mutant SSTR2 activated NF-κB signaling and facilitated nuclear transportation of p65 in AR42J cells upon cerulein plus LPS stimulation. Furthermore, pretreatment with wild-type but not N9Q-mutant SSTR2-overexpressing vectors significantly ameliorated biochemical parameters and pancreatic histological impairments in rats with pancreatitis. CONCLUSIONS: N-glycosylation of SSTR2 is essential for membrane localization, stability, and signal transduction of SSTR2 in pancreatic cells, playing a protective role in experimental acute pancreatitis.