Tigerinin-1R: a potent, non-toxic insulin-releasing peptide isolated from the skin of the Asian frog, Hoplobatrachus rugulosus

Abstract

Characterization of peptides in the skin of the Vietnamese common lowland frog Hoplobatrachus rugulosus with the ability to stimulate insulin release in vitro and improve glucose tolerance in vivo. Peptides in an extract of skin were purified by reversed-phase HPLC, and their abilities to stimulate the release of insulin and the cytosolic enzyme lactate dehydrogenase were determined using BRIN-BD11 clonal β cells. Insulin-releasing potencies of synthetic peptides and their effects on membrane potential and intracellular Ca²⁺ concentration were also measured using BRIN-BD11 cells. Effects on glucose tolerance and insulin release in vivo were determined in mice fed a high-fat diet to induce obesity and insulin resistance. A cyclic dodecapeptide (RVCSAIPLPICH.NH₂), termed tigerinin-1R, was isolated from the skin extract that lacked short-term cytotoxic and haemolytic activity but significantly (p < 0.01) stimulated the rate of release of insulin from BRIN-BD11 cells at concentrations ≥ 0.1 nM. The maximum response was 405% of the basal rate at 5.6 mM ambient glucose concentration and 290% of basal rate at 16.7 mM glucose. C-terminal α-amidation was necessary for high potency and a possible mechanism of action of the peptide-involved membrane depolarization and an increase in intracellular Ca²⁺ concentration. Administration of tigerinin-1R (75 nmol/kg body weight) to high fat-fed mice significantly (p < 0.05) enhanced insulin release and improved glucose tolerance during the 60-min period following an intraperitoneal glucose load. Tigerinin-1R is a potent, non-toxic insulin-releasing peptide that shows potential for development into an agent for the treatment of type 2 diabetes.