A novel two-chain IGF-II-derived peptide from purified β-cell granules

Objective: Insulin-like growth factor II (IGF-II) is a potent mitogen that regulates prenatal growth and development in both humans and rodents. Its role in post-natal life is less clear although immunohistochemical studies have observed IGF-II-like immunoreactivity (IGF-II-LI) associated with insulin-producing pancreatic beta-cells. Here we isolated secretory granules from a line, beta TC6-F7, and characterized the nature of the ICE-II-LI located therein. Design: Secretory granules were isolated from cultured mouse beta TC6-F7 cells by ultracentrifugation. Granule protein content was separated by reversed-phase HPLC, and assayed for IGF-II (radioimmunoassay) prior to identification by gas-phase NH(2)-terminal sequencing and MALDI-TOF MS. Effects of glucose incorporation into muscle glycogen were determined by incubating with isolated rat soleus muscle strips. Results: beta TC6-F7 cells contained 60 +/- 8 pmol of IGF-II-LI per 10(6) cells compared to 340 +/- 44 pmol insulin-LI per 106 cells. IGF-II immunoreactive fractions were found to contain an IGF-II-like molecule with a molecular mass of 6847.6 Da. The protein was found to be a two-chain insulin-like product of Igf2 that corresponds to mouse des(37-40)IGF-II, which we termed 'vesiculin'. This molecule was also detectable in beta TC6-F7 cells by intact-cell mass spectrometry. Mouse vesiculin evoked concentration-dependent stimulation of muscle glycogen synthesis ex vivo with an EC(50) value of 131 nM +/- 1.35. Conclusions: Vesiculin, des(37-40)IGF-II, is a novel two-chain insulin-like hormone and the major "IGF-II-like" peptide found in purified mouse beta TC6-F7 secretory granules. It stimulated ex vivo muscle glycogen synthesis with an efficacy greater than or equal to the intrinsic potency of IGF-II when compared to insulin derived from the same species. (C) 2010 Growth Hormone Research Society. Published by Elsevier Ltd. All rights reserved.