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

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• 作者：Christina M. Buchanan ; Anthony R.J. Phillips ; Garth J.S. Cooper
• 关键词：Insulin-like growth factor II ; Isoform ; Beta cell ; Secretory granule ; Cell line ; Cultured cells ; Metabolism
• 刊名：Growth Hormone IGF Research
• 出版年：2010
• 出版时间：October 2010
• 年：2010
• 卷：20
• 期：5
• 页码：360-366
• 全文大小：652 K

文摘

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 β-cells. Here we isolated secretory granules from a β-cell line, βTC6-F7, and characterized the nature of the IGF-II-LI located therein.

Design

Secretory granules were isolated from cultured mouse βTC6-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 NH₂-terminal sequencing and MALDI-TOF MS. Effects of glucose incorporation into muscle glycogen were determined by incubating with isolated rat soleus muscle strips.

Results

βTC6-F7 cells contained 60 ± 8 pmol of IGF-II-LI per 10⁶ 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 βTC6-F7 cells by intact-cell mass spectrometry. Mouse vesiculin evoked concentration-dependent stimulation of muscle glycogen synthesis ex vivo with an EC₅₀ 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 βTC6-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.