Islet Amyloid Polypeptide Amyloid Research Articles

Differential Effects of Aromatic Residues on Amyloid Formation and Cytotoxicity of Human IAPP.

Islet amyloid polypeptide (IAPP) is a 37-residue polypeptide hormone secreted by the pancreatic β-cells. IAPP plays a role in glycemic regulation, but in the pre-type-2 diabetic state, it aggregates to form an islet amyloid. The process of islet amyloid formation contributes to β-cell dysfunction and disease progression. The features of the IAPP sequence that modulate amyloid formation are still not understood. Human IAPP contains three aromatic residues, F15, F23, and Y37. F15 and Y37 are highly conserved, while F23 is more commonly a Leu or Ile in other species. The role of the aromatic residues in modulating the time course of amyloid formation and the cytotoxicity was examined using aromatic to Leu mutations. All three single and double mutants and the triple mutant were studied. F23 plays a dominant role in both amyloid formation and toxicity. An F15L mutant accelerated amyloid formation, a Y37L mutant had little effect, while an F23L replacement slowed amyloid formation by a factor of 2.6. Double mutants, which contained an F23L replacement, had a larger effect than those that did not, and there are non-additive effects between pairs of aromatic residues. F23 also plays a key role in toxicity. Single or multiple mutants that contain the F23L replacement were noticeably less toxic than the wild-type or mutants which did not include the F23L substitution. In contrast, the F15L mutant was more toxic than the wild-type one. The implications for IAPP amyloid formation and for the design of non-aggregating analogues of IAPP are discussed.

Substoichiometric Inhibition of Insulin against IAPP Aggregation Is Attenuated by the Incompletely Processed N-Terminus of proIAPP.

Substoichiometric aggregation inhibition of human islet amyloid polypeptide (IAPP), the hallmark of type 2 diabetes impacting millions of people, is crucial for developing clinic therapies, yet it remains challenging given that many candidate inhibitors require high doses. Intriguingly, insulin, the key regulatory polypeptide on blood glucose levels that are cosynthesized, costored, and cosecreted with IAPP by pancreatic β cells, has been identified as a potent inhibitor that can suppress IAPP amyloid aggregation at substoichiometric concentrations. Here, we computationally investigated the molecular mechanisms of the substoichiometric inhibition of insulin against the aggregation of IAPP and the incompletely processed IAPP (proIAPP) using discrete molecular dynamics simulations. Our results suggest that the amyloid aggregations of both IAPP and proIAPP might be disrupted by insulin through its binding with the shared amyloidogenic core sequences. However, the N-terminus of proIAPP competed with the amyloidogenic core sequences for the insulin interactions, resulting in attenuated inhibition by insulin. Moreover, insulin preferred to bind the elongation surfaces of IAPP seeds with fibril-like structure, with a stronger affinity than that of IAPP monomers. The capping of elongation surfaces by a small amount of insulin sterically prohibited the seed growth via monomer addition, achieving the substoichiometric inhibition. Together, our computational results provided molecular insights for the substoichiometric inhibition of insulin against IAPP aggregation, also the weakened effect on proIAPP. The uncovered substoichiometric inhibition by capping the elongation of amyloid seeds or fibrils may guide the rational designs of new potent inhibitors effective at low doses.

Silybins inhibit human IAPP amyloid growth and toxicity through stereospecific interactions

Type 2 Diabetes is a major public health threat, and its prevalence is increasing worldwide. The abnormal accumulation of islet amyloid polypeptide (IAPP) in pancreatic β-cells is associated with the onset of the disease. Therefore, the design of small molecules able to inhibit IAPP aggregation represents a promising strategy in the development of new therapies. Here we employ in vitro, biophysical, and computational methods to inspect the ability of Silybin A and Silybin B, two natural diastereoisomers extracted from milk thistle, to interfere with the toxic self-assembly of human IAPP (hIAPP). We show that Silybin B inhibits amyloid aggregation and protects INS-1 cells from hIAPP toxicity more than Silybin A. Molecular dynamics simulations revealed that the higher efficiency of Silybin B is ascribable to its interactions with precise hIAPP regions that are notoriously involved in hIAPP self-assembly i.e., the S20-S29 amyloidogenic core, H18, the N-terminal domain, and N35. These results highlight the importance of stereospecific ligand-peptide interactions in regulating amyloid aggregation and provide a blueprint for future studies aimed at designing Silybin derivatives with enhanced drug-like properties.

Combination of human tau and islet amyloid polypeptide exacerbates metabolic dysfunction in transgenic mice.

Amyloid plaques and neurofibrillary tangles composed of hyperphosphorylated tau are important contributors to Alzheimer's disease (AD). Tau also impacts pancreatic beta cell function and glucose homeostasis. Amyloid deposits composed of islet amyloid polypeptide (IAPP) are a pathological feature of type 2 diabetes (T2D). The current study investigates the role of human tau (hTau) in combination with human IAPP (hIAPP) as a potential mechanism connecting AD and T2D. Transgenic mice expressing hTau and hIAPP in the absence of murine tau were generated to determine the impact of these pathological factors on glucose metabolism. Co-expression of hIAPP and hTau resulted in mice with increased hyperglycaemia, insulin resistance, and glucose intolerance. The hTau-hIAPP mice also exhibited reduced beta cell area, increased amyloid deposition, impaired insulin processing, and reduced insulin content in islets. Tau phosphorylation also increased after stimulation with high glucose. In addition, brain insulin content and signalling were reduced, and tau phosphorylation was increased in these animals. These data support a link between tau and IAPP amyloid, which seems to act co-ordinately to impair beta cell function and glucose homeostasis, and suggest that the combined pathological actions of these proteins may be a potential mechanism connecting AD and T2D. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Mechanisms of Transthyretin Inhibition of IAPP Amyloid Formation.

Amyloid-formation by the islet amyloid polypeptide (IAPP), produced by the β-cells in the human pancreas, has been associated with the development of type II diabetes mellitus (T2DM). The human plasma-protein transthyretin (TTR), a well-known amyloid-inhibiting protein, is interestingly also expressed within the IAPP producing β-cells. In the present study, we have characterized the ability of TTR to interfere with IAPP amyloid-formation, both in terms of its intrinsic stability as well as with regard to the effect of TTR-stabilizing drugs. The results show that TTR can prolong the lag-phase as well as impair elongation in the course of IAPP-amyloid formation. We also show that the interfering ability correlates inversely with the thermodynamic stability of TTR, while no such correlation was observed as a function of kinetic stability. Furthermore, we demonstrate that the ability of TTR to interfere is maintained also at the low pH environment within the IAPP-containing granules of the pancreatic β-cells. However, at both neutral and low pH, the addition of TTR-stabilizing drugs partly impaired its efficacy. Taken together, these results expose mechanisms of TTR-mediated inhibition of IAPP amyloid-formation and highlights a potential therapeutic target to prevent the onset of T2DM.

Role of lysine residue of islet amyloid polypeptide in fibril formation, membrane binding, and inhibitor binding

The aggregation of islet amyloid polypeptide (IAPP) is implicated in the pathogenesis of type 2 diabetes (T2D). In T2D, this peptide aggregates to form amyloid fibrils; the mechanism responsible for islet amyloid formation is unclear. However, it is known that the aggregation propensity of IAPP is highly related to its primary sequence. Several residues have been suggested to be critical in modulating IAPP amyloid formation, but role of the sole lysine residue at position 1 (Lys-1) in IAPP has not been discussed. In our previous study, we found that glycated IAPP can form amyloid faster than normal IAPP and induce normal IAPP to expedite the aggregation process. To gain more insight into the contribution of Lys-1 in the kinetics of fibril formation, we synthesized another two IAPP variants, K1E-IAPP and K1Nle-IAPP, in which the Lys residue was mutated to glutamate and norleucine, respectively. Interestingly, we observed that the negative or neutral charged side chain at this position was preferred for amyloid formation. The findings suggested this residue may take part in the inter- or intra-molecular interaction during IAPP aggregation, even though it was proposed not to be in part of fibril core structure. Our data also revealed that the inhibitory mechanism of some inhibitors for IAPP aggregation require reaction with Lys-1. Modifications of Lys-1, such as protein glycation, may affect the effectiveness of the inhibitory action of some potential drugs in the treatment of amyloidosis.

Differential effects of serine side chain interactions in amyloid formation by islet amyloid polypeptide.

Islet amyloid polypeptide (IAPP), a 37 residue polypeptide, is the main protein component of islet amyloid deposits produced in the pancreas in Type 2 diabetes. Human IAPP contains five serine residues at positions 19, 20, 28, 29, and 34. Models of the IAPP amyloid fibril indicate a structure composed of two closely aligned columns of IAPP monomers with each monomer contributing to two intermolecular β-strands. Ser 19 and Ser 20 are in the partially ordered β-turn region, which links the two strands, whereas Ser 28, Ser 29, and Ser 34 are in the core region of the amyloid fibril. Ser 29 is involved in contacts between the two columns of monomers and is the part of the steric zipper interface. We undertook a study of individual serine substitutions with the hydrophobic isostere 2-aminobutyric acid (2-Abu) to examine the site-specific role of serine side chains in IAPP amyloid formation. All five variants formed amyloid. The Ser 19 to 2-Abu mutant accelerates amyloid formation by a factor of 3 to 4, while the Ser 29 to 2-Abu mutation modestly slows the rate of amyloid formation. 2-Abu replacements at the other sites had even smaller effects. The data demonstrate that the cross-column interactions made by residue 29 are not essential for amyloid formation and also show that cross-strand networks of hydrogen-bonded Ser side chains, so called Ser-ladders, are not required for IAPP amyloid formation. The effect of the Ser 19 to 2-Abu mutant suggests that residues in this region are important for amyloid formation by IAPP.

Analysis of Baboon IAPP Provides Insight into Amyloidogenicity and Cytotoxicity of Human IAPP

The polypeptide hormone islet amyloid polypeptide (IAPP) forms islet amyloid in type 2 diabetes, a process which contributes to pancreatic β-cell dysfunction and death. Not all species form islet amyloid, and the ability to do so correlates with the primary sequence. Humans form islet amyloid, but baboon IAPP has not been studied. The baboon peptide differs from human IAPP at three positions containing K1I, H18R, and A25T substitutions. The K1I substitution is a rare example of a replacement in the N-terminal region of amylin. The effect of this mutation on amyloid formation has not been studied, but it reduces the net charge, and amyloid prediction programs suggest that it should increase amyloidogenicity. The A25T replacement involves a nonconservative substitution in a region of IAPP that is believed to be important for aggregation, but the effects of this replacement have not been examined. The H18R point mutant has been previously shown to reduce aggregation in vitro. Baboon amylin forms amyloid on the same timescale as human amylin in vitro and exhibits similar toxicity toward cultured β-cells. The K1I replacement in human amylin slightly reduces toxicity, whereas the A25T substitution accelerates amyloid formation and enhances toxicity. Photochemical cross-linking reveals that the baboon amylin, like human amylin, forms low-order oligomers in the lag phase of amyloid formation. Ion-mobility mass spectrometry reveals broadly similar gas phase collisional cross sections for human and baboon amylin monomers and dimers, with some differences in the arrival time distributions. Preamyloid oligomers formed by baboon amylin, but not baboon amylin fibers, are toxic to cultured β-cells. The toxicity of baboon oligomers and lack of significantly detectable toxicity with exogenously added amyloid fibers is consistent with the hypothesis that preamyloid oligomers are the most toxic species produced during IAPP amyloid formation.

Identification of a hinge residue controlling islet amyloid polypeptide self-assembly and cytotoxicity

The islet amyloid polypeptide (IAPP) is a 37-residue peptide hormone whose deposition as amyloid fibrils in the pancreatic islets is associated with type 2 diabetes. Previous studies have suggested that residue Asn-21 plays a critical role in the in vitro self-assembly of IAPP. Herein, we studied structure-self-assembly relationships focusing on position 21 to gain detailed insights into the molecular mechanisms of IAPP self-assembly and to probe the conformational nature of the toxic assemblies associated with β-cell death. Thioflavin T (ThT) fluorescence, CD spectroscopy, and transmission EM analysis revealed that the Asn-21 amide side chain is not required for IAPP nucleation and amyloid elongation, as N21A and N21G variants assembled into prototypical fibrils. In contrast, Asn-21 substitution with the conformationally constrained and turn-inducing residue Pro accelerated IAPP self-assembly. Successive substitutions with hydrophobic residues led to the formation of ThT-negative β-sheet-rich aggregates having high surface hydrophobicity. Cell-based assays revealed no direct correlation between the in vitro amyloidogenicity of these variants and their toxicity. In contrast, leakage of anionic lipid vesicles disclosed that membrane disruption is closely associated with cytotoxicity. We observed that the N21F variant self-assembles into worm-like aggregates, causing loss of lipid membrane structural integrity and inducing β-cell apoptosis. These results indicate that specific intra- and intermolecular interactions involving Asn-21 promote IAPP primary nucleation events by modulating the conformational conversion of the oligomeric intermediates into amyloid fibrils. Our study identifies position 21 as a hinge residue that modulates IAPP amyloidogenicity and cytotoxicity.

Ultraeffective Inhibition of Amyloid Fibril Assembly by Nanobody-Gold Nanoparticle Conjugates.

Amyloid fibril assembly is associated with many human disorders, and to approach an inhibitor of amyloid formation that is effective at ultralow stoichiometric concentrations remains a big challenge. Taking fibril assembly of human islet amyloid polypeptide (IAPP) as a model system, we demonstrate here that conjugating a rationally designed sequence-specific nanobody inhibitor M1 with gold nanoparticles (AuNPs) can significantly enhance the inhibition potency of M1, leading to complete inhibition of IAPP amyloid fibrillation at very low M1:IAPP molar ratios. Thioflavin T kinetics fluorescence assays, dynamic light scattering measurements, far-UV circular dichroism, and transmission electron microscopy all indicate that M1-AuNP conjugates prevent IAPP fibrillation at M1:IAPP molar ratios of as low as 1:50, while free M1 is unable to prevent fibrillation at the same substoichiometric concentrations. This strategy represents a prototype of the facile development of a variety of highly potent amyloid inhibitors with enhanced therapeutic effects.

Oligotyrosines Inhibit Amyloid Formation of Human Islet Amyloid Polypeptide in a Tyrosine-Number-Dependent Manner.

Misfolding and amyloid formation of human islet amyloid polypeptide (IAPP) are believed to be critical in the pathogenesis of type 2 diabetes. Inhibitors that can effectively prevent protein aggregation and fibrillation are considered as potential therapeutics for the prevention and treatment of type 2 diabetes. Here, we report that oligotyrosines manipulate IAPP amyloid formation in vitro and modulate IAPP-induced cytotoxicity in a manner that is related to the number of tyrosine units. Tyr2 and Tyr3 can effectively inhibit the aggregation of IAPP, either in bulk solution or in the presence of lipid membranes, and alleviate IAPP-mediated cytotoxicity. On the contrary, Tyr, Tyr4, and Tyr6 do not show significant inhibitory effects on the IAPP aggregation at the same conditions. To the best of our knowledge, this is the first report of a residue-number-dependent inhibition of IAPP aggregation by oligotyrosines, and Tyr2 and Tyr3 are proved to be potent inhibitors of IAPP amyloid formation. The interactions between oligotyrosines and IAPP were simulated through molecular docking, which provides us a new insight about the inhibition mechanism of IAPP amyloid formation that will be helpful for developing antidiabetic drug candidates.

Nanoscale inhibition of polymorphic and ambidextrous IAPP amyloid aggregation with small molecules.

Understanding how small molecules interface amyloid fibrils on the nanoscale is of importance for developing therapeutic treatment against amyloid-based diseases. Here we show, for the first time, that human islet amyloid polypeptide (IAPP) in the fibrillar form is polymorphic and ambidextrous possessing multiple periodicities. Upon interfacing with small molecule epigallocatechin gallate (EGCG), IAPP aggregation was rendered off pathway assuming the form of soft and disordered clusters, while mature IAPP fibrils displayed kinks and branching but conserved the twisted fibril morphology. These nanoscale phenomena resulted from competitive interactions between EGCG and the IAPP amyloidogenic region, as well as end capping of the fibrils by the small molecule. This information is crucial to delineating IAPP toxicity implicated in type 2 diabetes and developing new inhibitors against amyloidogenesis.

Sterol Structure Strongly Modulates Membrane-Islet Amyloid Polypeptide Interactions.

Amyloid formation has been implicated in a wide range of human diseases, and the interaction of amyloidogenic proteins with membranes are believed to be important for many of them. In type-2 diabetes, human islet amyloid polypeptide (IAPP) forms amyloids, which contribute to β-cell death and dysfunction in the disease. IAPP-membrane interactions are potential mechanisms of cytotoxicity. In vitro studies have shown that cholesterol significantly modulates the ability of model membranes to induce IAPP amyloid formation and IAPP-mediated membrane damage. It is not known if this is due to the general effects of cholesterol on membranes or because of specific sterol-IAPP interactions. The effects of replacing cholesterol with eight other sterols/steroids on IAPP binding to model membranes, membrane disruption, and membrane-mediated amyloid formation were examined. The primary effect of the sterols/steroids on the IAPP-membrane interactions was found to reflect their effect upon membrane order as judged by fluorescence anisotropy measurements. Specific IAPP-sterol/steroid interactions have smaller effects. The fraction of vesicles that bind IAPP was inversely correlated with the sterols/steroids' effect on membrane order, as was the extent of IAPP-induced membrane leakage and the time to form amyloids. The correlation between the fraction of vesicles binding IAPP and membrane leakage was particularly tight, suggesting the restriction of IAPP to a subset of vesicles is responsible for incomplete leakage.

Extracellular vesicles from human pancreatic islets suppress human islet amyloid polypeptide amyloid formation

Extracellular vesicles (EVs) are small vesicles released by cells to aid cell-cell communication and tissue homeostasis. Human islet amyloid polypeptide (IAPP) is the major component of amyloid deposits found in pancreatic islets of patients with type 2 diabetes (T2D). IAPP is secreted in conjunction with insulin from pancreatic β cells to regulate glucose metabolism. Here, using a combination of analytical and biophysical methods in vitro, we tested whether EVs isolated from pancreatic islets of healthy patients and patients with T2D modulate IAPP amyloid formation. We discovered that pancreatic EVs from healthy patients reduce IAPP amyloid formation by peptide scavenging, but T2D pancreatic and human serum EVs have no effect. In accordance with these differential effects, the insulin:C-peptide ratio and lipid composition differ between EVs from healthy pancreas and EVs from T2D pancreas and serum. It appears that healthy pancreatic EVs limit IAPP amyloid formation via direct binding as a tissue-specific control mechanism.

Induction of IAPP amyloid deposition and associated diabetic abnormalities by a prion-like mechanism.

Although a large proportion of patients with type 2 diabetes (T2D) accumulate misfolded aggregates composed of the islet amyloid polypeptide (IAPP), its role in the disease is unknown. Here, we show that pancreatic IAPP aggregates can promote the misfolding and aggregation of endogenous IAPP in islet cultures obtained from transgenic mouse or healthy human pancreas. Islet homogenates immunodepleted with anti-IAPP-specific antibodies were not able to induce IAPP aggregation. Importantly, intraperitoneal inoculation of pancreatic homogenates containing IAPP aggregates into transgenic mice expressing human IAPP dramatically accelerates IAPP amyloid deposition, which was accompanied by clinical abnormalities typical of T2D, including hyperglycemia, impaired glucose tolerance, and a substantial reduction on β cell number and mass. Finally, induction of IAPP deposition and diabetic abnormalities were also induced in vivo by administration of IAPP aggregates prepared in vitro using pure, synthetic IAPP. Our findings suggest that some of the pathologic and clinical alterations of T2D might be transmissible through a similar mechanism by which prions propagate in prion diseases.

Role of Site-Specific Asparagine Deamidation in Islet Amyloid Polypeptide Amyloidogenesis: Key Contributions of Residues 14 and 21.

Deamidation of an asparagine residue is a spontaneous non-enzymatic post-translational modification that results in the conversion of asparagine into a mixture of aspartic acid and isoaspartic acid. This chemical conversion modulates protein conformation and physicochemical properties, which could lead to protein misfolding and aggregation. In this study, we investigated the effects of site-specific Asn deamidation on the amyloidogenicity of the aggregation-prone peptide islet amyloid polypeptide (IAPP). IAPP is a 37-residue peptidic hormone whose deposition as insoluble amyloid fibrils is closely associated with type 2 diabetes. Asn residues were successively substituted with an Asp or isoAsp, and amyloid formation was evaluated by a thioflavin T fluorescence assay, circular dichroism spectroscopy, atomic force microscopy, and transmission electron microscopy. Whereas deamidation at position 21 inhibited IAPP conformational conversion and amyloid formation, the N14D mutation accelerated self-assembly and led to the formation of long and thick amyloid fibrils. In contrast, IAPP was somewhat tolerant to the successive deamidation of Asn residues 22, 31, and 35. Interestingly, a small molar ratio of IAPP deamidated at position 14 promoted the formation of nucleating species and the elongation from unmodified IAPP. Besides, using the rat pancreatic β cell line INS-1E, we observed that site-specific deamidation did not significantly alter IAPP-induced toxicity. These data indicate that Asn deamidation can modulate IAPP amyloid formation and fibril morphology and that the site of modification plays a critical role. Above all, this study reinforces the notion that IAPP amyloidogenesis is governed by precise intermolecular interactions involving specific Asn side chains.

Effects of Protein Corona on IAPP Amyloid Aggregation, Fibril Remodelling, and Cytotoxicity

Aggregation of islet amyloid polypeptide (IAPP), a peptide hormone co-synthesized and co-stored with insulin in pancreatic cells and also co-secreted to the circulation, is associated with beta-cell death in type-2 diabetes (T2D). In T2D patients IAPP is found aggregating in the extracellular space of the islets of Langerhans. Although the physiological environments of these intra- and extra-cellular compartments and vascular systems significantly differ, the presence of proteins is ubiquitous but the effects of protein binding on IAPP aggregation are largely unknown. Here we examined the binding of freshly-dissolved IAPP as well as pre-formed fibrils with two homologous proteins, namely cationic lysozyme (Lys) and anionic alpha-lactalbumin (aLac), both of which can be found in the circulation. Biophysical characterizations and a cell viability assay revealed distinct effects of Lys and aLac on IAPP amyloid aggregation, fibril remodelling and cytotoxicity, pointing to the role of protein “corona” in conferring the biological impact of amyloidogenic peptides. Systematic molecular dynamics simulations further provided molecular and structural details for the observed differential effects of proteins on IAPP amyloidosis. This study facilitates our understanding of the fate and transformation of IAPP in vivo, which are expected to have consequential bearings on IAPP glycemic control and T2D pathology.

The human serum protein C4b-binding protein inhibits pancreatic IAPP-induced inflammasome activation

Aims/hypothesisInflammasome activation and subsequent IL-1β production is a driver of islet pathology in type 2 diabetes. Oligomers, but not mature amyloid fibrils, of human islet amyloid polypeptide (IAPP), which is co-secreted with insulin, trigger NOD-like receptor pyrin domain containing-3 (NLRP3) inflammasome activation. C4b-binding protein (C4BP), present in serum, binds to IAPP and affects transition of IAPP monomers and oligomers to amyloid fibrils. We therefore hypothesised that C4BP inhibits IAPP-mediated inflammasome activation and IL-1β production.MethodsMacrophages were exposed to IAPP in the presence or absence of plasma-purified human C4BP, and inflammasome activation was assessed by IL-1β secretion as detected by ELISA and reporter cell lines. IAPP fibrillation was assessed by thioflavin T assay. Uptake of IAPP–C4BP complexes and their effects on phagolysosomal stability were assessed by flow cytometry and confocal microscopy. The effect of C4BP regulation of IAPP-mediated inflammasome activation on beta cell function was assessed using a clonal rat beta cell line. Immunohistochemistry was used to examine the association of IAPP amyloid deposits and macrophage infiltration in isolated human and mouse pancreatic islets, and expression of C4BP from isolated human pancreatic islets was assessed by quantitative PCR, immunohistochemistry and western blot.ResultsC4BP significantly inhibited IAPP-mediated IL-1β secretion from primed macrophages at physiological concentrations in a dose-dependent manner. C4BP bound to and was internalised together with IAPP. C4BP did not affect IAPP uptake into phagolysosomal compartments, although it did inhibit its formation into amyloid fibrils. The loss of macrophage phagolysosomal integrity induced by IAPP incubation was inhibited by co-incubation with C4BP. Supernatant fractions from macrophages activated with IAPP inhibited both insulin secretion and viability of clonal beta cells in an IL-1β-dependent manner but the presence of C4BP during macrophage IAPP incubation rescued beta cell function and viability. In human and mouse islets, the presence of amyloid deposits correlated with higher numbers of infiltrating macrophages. Isolated human islets expressed and secreted C4BP, which increased with addition of IL-1β.Conclusions/interpretationIAPP deposition is associated with inflammatory cell infiltrates in pancreatic islets. C4BP blocks IAPP-induced inflammasome activation by preventing the loss of macrophage phagolysosomal integrity required for NLRP3 activation. The consequence of this is the preservation of beta cell function and viability. C4BP is secreted directly from human pancreatic islets and this increases in response to inflammatory cytokines. We therefore propose that C4BP acts as an extracellular chaperone protein that limits the proinflammatory effects of IAPP.

Islet Amyloid Polypeptide Membrane Interactions: Effects of Membrane Composition.

Amyloid formation by islet amyloid polypeptide (IAPP) contributes to β-cell dysfunction in type 2 diabetes. Perturbation of the β-cell membrane may contribute to IAPP-induced toxicity. We examine the effects of lipid composition, salt, and buffer on IAPP amyloid formation and on the ability of IAPP to induce leakage of model membranes. Even low levels of anionic lipids promote amyloid formation and membrane permeabilization. Increasing the percentage of the anionic lipids, 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine (POPS) or 1,2-dioleoyl-sn-glycero-3-phospho(1'-rac-glycerol), enhances the rate of amyloid formation and increases the level of membrane permeabilization. The choice of zwitterionic lipid has no noticeable effect on membrane-catalyzed amyloid formation but in most cases affects leakage, which tends to decrease in the following order: 1,2-dioleoyl-sn-glycero-3-phosphocholine > 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine > sphingomyelin. Uncharged lipids that increase the level of membrane order weaken the ability of IAPP to induce leakage. Leakage is due predominately to pore formation rather than complete disruption of the vesicles under the conditions used in these studies. Cholesterol at or below physiological levels significantly reduces the rate of vesicle-catalyzed IAPP amyloid formation and decreases the susceptibility to IAPP-induced leakage. The effects of cholesterol on amyloid formation are masked by 25 mol % POPS. Overall, there is a strong inverse correlation between the time to form amyloid and the extent of vesicle leakage. NaCl reduces the rate of membrane-catalyzed amyloid formation by anionic vesicles, but accelerates amyloid formation in solution. The implications for IAPP membrane interactions are discussed, as is the possibility that the loss of phosphatidylserine asymmetry enhances IAPP amyloid formation and membrane damage in vivo via a positive feedback loop.

Cross-talk between amyloidogenic proteins in type-2 diabetes and Parkinson’s disease

In type-2 diabetes (T2D) and Parkinson's disease (PD), polypeptide assembly into amyloid fibers plays central roles: in PD, α-synuclein (aS) forms amyloids and in T2D, amylin [islet amyloid polypeptide (IAPP)] forms amyloids. Using a combination of biophysical methods in vitro we have investigated whether aS, IAPP, and unprocessed IAPP, pro-IAPP, polypeptides can cross-react. Whereas IAPP forms amyloids within minutes, aS takes many hours to assemble into amyloids and pro-IAPP aggregates even slower under the same conditions. We discovered that preformed amyloids of pro-IAPP inhibit, whereas IAPP amyloids promote, aS amyloid formation. Amyloids of aS promote pro-IAPP amyloid formation, whereas they inhibit IAPP amyloid formation. In contrast, mixing of IAPP and aS monomers results in coaggregation that is faster than either protein alone; moreover, pro-IAPP can incorporate aS monomers into its amyloid fibers. From this intricate network of cross-reactivity, it is clear that the presence of IAPP can accelerate aS amyloid formation. This observation may explain why T2D patients are susceptible to developing PD.