New insights into the roles of sulfated glycosaminoglycans in islet amyloid polypeptide amyloidogenesis and cytotoxicity

Abstract

Glycosaminoglycans (GAGs) are found in association with virtually all extracellular protein deposits related to amyloid diseases. Particularly, GAGs were shown to enhance fibrillogenesis of the islet amyloid polypeptide (IAPP), a peptide hormone whose aggregation is associated with Type-II diabetes pathogenesis. However, the exact molecular mechanism by which GAGs enhance IAPP amyloidogenesis remains unclear as well as the implications of cell surface GAGs in IAPP-mediated cytotoxicity. The aim of this study was to gain conformational and thermodynamics insights about GAGs-IAPP interactions as a function of IAPP protonation state and buffer ionic strength as well as to explore the roles of cell surface GAGs in IAPP cytotoxicity. Isothermal titration calorimetry revealed that protonation of residue His(18) increases the binding affinity of IAPP towards heparin and, in turn, strongly stimulates fibrillogenesis. Interaction of IAPP with heparin induces a random coil to helix conformational conversion and the helical intermediates could be on-pathway to amyloid fibrils formation. Using rat beta-cells INS-1 that were enzymatically treated with GAG lyases and a CHO cell line that is deficient in the biosynthesis of GAGs, we observed that the lack of GAGs at the plasma membrane does not prevent IAPP-induced toxicity, whereas the presence of soluble heparin in the cell media inhibits IAPP cytotoxicity. Overall, this study reinforces the postulate that sulfated GAGs are actively implicated in IAPP amyloidogenic process in vivo, where they could play a protective role by interacting with cytotoxic species and converting them into less culprit amyloid fibrils.

Amyloid Formation in Human Islets Is Enhanced by Heparin and Inhibited by Heparinase

“…The pro-amyloidogenic effect of heparin on IAPP fibril formation has been previously described (26,31). Here, we confirm this effect by thioflavin T assay and electron microscopy.…”

“…Prefibrillar species are thought to be the primary mediators of amyloid toxicity (10,34,35,42). Rapid amyloid formation in the presence of heparin may deplete these prefibrillar species, thus accounting for decreased toxicity in the presence of heparin after 6 h in our studies and an overall protective effect in another study (31). A similar effect was observed with amyloid-beta in the presence of methylene blue (43) and ellagic acid (44), compounds that promote fibrillization, protect against amyloid-associated toxicity, and inhibit oligomer accumulation.…”

“…Heparin is thought to act as a scaffold for amyloid formation (28) and inhibits amyloid fibril de-polymerization (29). Amyloid formation by IAPP in the presence of heparin results in increased fibril density and compaction (30) and thioflavin T fluorescence (31).…”

Heparan Sulfate Proteoglycans Are Important for Islet Amyloid Formation and Islet Amyloid Polypeptide-induced Apoptosis

“…In a recent paper, Carufel et al (56) demonstrated that the removal of cell surface GAGs had no effect on IAPP-induced cytotoxicity, measured 24 h after exposure to Ն25 M hIAPP. This is not unexpected because formation of fibrillar species and, therefore, cytotoxicity should occur at such supraphysiological concentrations of hIAPP well before 24 h, independent of the presence or absence of GAGs.…”

Modulation of amyloid assembly by glycosaminoglycans: from mechanism to biological significance

“…On its own, this peptide demonstrates no ability to form fibrils, yet upon addition of heparin to the solution, a prompt increase in thioflavin T (ThT) fluo-D r a f t 14 rescence was observed (Madine et al 2013). Similar effects were observed with polypeptides that are known for their propensity to aggregate and whose depositions are associated with pathological states, such as IAPP, TTR, β2-microglobulin and apomyoglobin (De Carufel et al 2013;Noborn et al 2011;Relini et al 2008;Vilasi et al 2011). Whether this occurs because GAGs directly influence the conformational transition between the native and amyloidogenic states and/or because GAGs increase the local concentration of pre-fibrillar species remains to be confirmed.…”