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Synthesis, bioactivity and specificity of glucagon-like peptide-1 (7-37)/polymer conjugate to isolated rat islets.

Abstract：

:In order to increase the functionality of islets encapsulated in a biohybrid artificial pancreas (BAP), it was proposed that co-encapsulation with insulinotropic agents would improve insulin secretion from islets. To prevent agents from leaking out, conjugation with high-molecular-weight polymers was inevitable. In this study, synthetic glucagon-like peptide-1 (GLP-1) (7-37) was conjugated to a water-soluble polymer, poly(N-vinyl-2-pyrroridone-co-acrylic acid) (5 mol% acrylic acid, M(w) 445 kDa), via poly(ethylene glycol, M(w) 3.4 kDa) spacer. The chemical conjugation was confirmed by reverse phase-HPLC and the GLP-1 content in the GLP-1/polymer conjugate (VAPG) was determined by UV spectrophotometry at 280 nm (ca. 29 wt/wt%). In a static insulin secretion test, the VAPG increased insulin secretion up to 200% over a control (no stimulation) at high glucose levels, although the insulinotropic activity of VAPG was slightly lower than that of native GLP-1. The bioactivity of VAPG was prolonged for at least 2 weeks, which was examined by co-encapsulation of the conjugate into islet microcapsules. Dose-response curve revealed that the half-maximal effective dose (ED(50)) of VAPG was about 55 nm (25 nm for native GLP-1). By N-terminal analysis using aminopeptidase and RP-HPLC, it was confirmed that the lowered bioactivity of VAPG stemmed from the polymer conjugation to N-terminal histidine moieties, which actively participate in binding to GLP-1 receptors, resulting in only 16% of N-terminal histidine remaining intact after the conjugation reaction. Finally, the specific interaction of the VAPG with isolated rat islets was investigated. Total cellular cyclic AMP levels were measured and confocal microscopy was conducted using GLP-1 and VAPG labeled with fluorescent probes. It was found that VAPG effectively increased the cAMP level in islet cells in a glucose concentration-dependent manner. Moreover, the confocal microscopy study showed that the binding of VAPG occurs at the same location where GLP-1 binds but with less affinity than that of native GLP-1. In summary, a GLP-1/polymer conjugate was synthesized for the first time, and its bioactivity was examined, which must result from its specific interaction with isolated islets.

journal_name

Biomaterials

journal_title

Biomaterials

authors

Kim S,Wan Kim S,Bae YH

doi

10.1016/j.biomaterials.2004.09.045

keywords:

subject

Has Abstract

pub_date

2005-06-01 00:00:00

pages

3597-606

issue

17

eissn

0142-9612

issn

1878-5905

pii

S0142-9612(04)00835-X

journal_volume

26

pub_type

杂志文章

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