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Time-resolved studies define the nature of toxic IAPP intermediates, providing insight for anti-amyloidosis therapeutics.

Abedini A, Plesner A, Cao P, Ridgway Z, Zhang J, Tu LH, Middleton CT, Chao B, Sartori DJ, Meng F, Wang H, Wong AG, Zanni MT, Verchere CB, Raleigh DP, Schmidt AM - Elife (2016)

Bottom Line: These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species.They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure.Aromatic interactions modulate, but are not required for toxicity.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Research Program, Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, New York University School of Medicine, New York, United States.

ABSTRACT
Islet amyloidosis by IAPP contributes to pancreatic β-cell death in diabetes, but the nature of toxic IAPP species remains elusive. Using concurrent time-resolved biophysical and biological measurements, we define the toxic species produced during IAPP amyloid formation and link their properties to induction of rat INS-1 β-cell and murine islet toxicity. These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species. They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure. Aromatic interactions modulate, but are not required for toxicity. Not all IAPP oligomers are toxic; toxicity depends on their partially structured conformational states. Some anti-amyloid agents paradoxically prolong cytotoxicity by prolonging the lifetime of the toxic species. The data highlight the distinguishing properties of toxic IAPP oligomers and the common features that they share with toxic species reported for other amyloidogenic polypeptides, providing information for rational drug design to treat IAPP induced β-cell death.

No MeSH data available.


Related in: MedlinePlus

Toxic h-IAPP lag phase intermediates induce β-cell apoptosis, but freshly dissolved h-IAPP (time-zero) and amyloid fibrils do not.Cleaved caspase-3 colorimetric assays of β-cells treated with exogenous h-IAPP lag phase intermediates (blue) show an increase in cleaved caspase-3, but time-zero h-IAPP species (black), h-IAPP amyloid fibrils (red), r-IAPP (green) and buffer (gold) do not. IAPP kinetic species were produced at 25°C in solutions containing 20 μM peptide. Final peptide concentration after dilution into β-cell assays was 14 μM. Data represent mean ± SEM of three replicates per condition.DOI:http://dx.doi.org/10.7554/eLife.12977.008
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fig3s2: Toxic h-IAPP lag phase intermediates induce β-cell apoptosis, but freshly dissolved h-IAPP (time-zero) and amyloid fibrils do not.Cleaved caspase-3 colorimetric assays of β-cells treated with exogenous h-IAPP lag phase intermediates (blue) show an increase in cleaved caspase-3, but time-zero h-IAPP species (black), h-IAPP amyloid fibrils (red), r-IAPP (green) and buffer (gold) do not. IAPP kinetic species were produced at 25°C in solutions containing 20 μM peptide. Final peptide concentration after dilution into β-cell assays was 14 μM. Data represent mean ± SEM of three replicates per condition.DOI:http://dx.doi.org/10.7554/eLife.12977.008

Mentions: Cellular stress and inflammation have been implicated in h-IAPP induced β-cell toxicity in vitro, in mouse models of metabolic disease and in human T2D (Westermark et al., 2011; Masters et al., 2010; Zraika et al., 2009; Janciauskiene and Ahrén, 2000; Konarkowska et al., 2005; Sakuraba et al., 2002). If the lag phase intermediates identified here are toxic species then they should upregulate pro-inflammatory mediators and the production of ROS. This is exactly what was observed. Along with a decrease in β-cell viability, h-IAPP lag phase intermediates also induce an increase in Ccl2 and Il1b mRNA expression, an increase in ROS production, upregulation of NADPH oxidase 1 (NOX1) protein expression, and an increase in cleaved caspase-3 production, consistent with h-IAPP induced β-cell stress, inflammation and apoptosis (Figure 3A–D and Figure 3—figure supplements 1 and 2). No significant upregulation of cytokines, ROS or cleaved caspase-3 production is induced by time-zero species or by h-IAPP amyloid fibrils, indicating that pro-inflammatory cellular responses are triggered specifically by pre-fibrillar lag phase intermediates. No toxicity or cytokine production is observed when non-amyloidogenic r-IAPP is added to cultured β-cells at any time point, consistent with previous reports (Westermark et al., 2011).10.7554/eLife.12977.006Figure 3.h-IAPP lag phase intermediates upregulate pro-inflammatory cytokines and oxidative stress.


Time-resolved studies define the nature of toxic IAPP intermediates, providing insight for anti-amyloidosis therapeutics.

Abedini A, Plesner A, Cao P, Ridgway Z, Zhang J, Tu LH, Middleton CT, Chao B, Sartori DJ, Meng F, Wang H, Wong AG, Zanni MT, Verchere CB, Raleigh DP, Schmidt AM - Elife (2016)

Toxic h-IAPP lag phase intermediates induce β-cell apoptosis, but freshly dissolved h-IAPP (time-zero) and amyloid fibrils do not.Cleaved caspase-3 colorimetric assays of β-cells treated with exogenous h-IAPP lag phase intermediates (blue) show an increase in cleaved caspase-3, but time-zero h-IAPP species (black), h-IAPP amyloid fibrils (red), r-IAPP (green) and buffer (gold) do not. IAPP kinetic species were produced at 25°C in solutions containing 20 μM peptide. Final peptide concentration after dilution into β-cell assays was 14 μM. Data represent mean ± SEM of three replicates per condition.DOI:http://dx.doi.org/10.7554/eLife.12977.008
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940161&req=5

fig3s2: Toxic h-IAPP lag phase intermediates induce β-cell apoptosis, but freshly dissolved h-IAPP (time-zero) and amyloid fibrils do not.Cleaved caspase-3 colorimetric assays of β-cells treated with exogenous h-IAPP lag phase intermediates (blue) show an increase in cleaved caspase-3, but time-zero h-IAPP species (black), h-IAPP amyloid fibrils (red), r-IAPP (green) and buffer (gold) do not. IAPP kinetic species were produced at 25°C in solutions containing 20 μM peptide. Final peptide concentration after dilution into β-cell assays was 14 μM. Data represent mean ± SEM of three replicates per condition.DOI:http://dx.doi.org/10.7554/eLife.12977.008
Mentions: Cellular stress and inflammation have been implicated in h-IAPP induced β-cell toxicity in vitro, in mouse models of metabolic disease and in human T2D (Westermark et al., 2011; Masters et al., 2010; Zraika et al., 2009; Janciauskiene and Ahrén, 2000; Konarkowska et al., 2005; Sakuraba et al., 2002). If the lag phase intermediates identified here are toxic species then they should upregulate pro-inflammatory mediators and the production of ROS. This is exactly what was observed. Along with a decrease in β-cell viability, h-IAPP lag phase intermediates also induce an increase in Ccl2 and Il1b mRNA expression, an increase in ROS production, upregulation of NADPH oxidase 1 (NOX1) protein expression, and an increase in cleaved caspase-3 production, consistent with h-IAPP induced β-cell stress, inflammation and apoptosis (Figure 3A–D and Figure 3—figure supplements 1 and 2). No significant upregulation of cytokines, ROS or cleaved caspase-3 production is induced by time-zero species or by h-IAPP amyloid fibrils, indicating that pro-inflammatory cellular responses are triggered specifically by pre-fibrillar lag phase intermediates. No toxicity or cytokine production is observed when non-amyloidogenic r-IAPP is added to cultured β-cells at any time point, consistent with previous reports (Westermark et al., 2011).10.7554/eLife.12977.006Figure 3.h-IAPP lag phase intermediates upregulate pro-inflammatory cytokines and oxidative stress.

Bottom Line: These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species.They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure.Aromatic interactions modulate, but are not required for toxicity.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Research Program, Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, New York University School of Medicine, New York, United States.

ABSTRACT
Islet amyloidosis by IAPP contributes to pancreatic β-cell death in diabetes, but the nature of toxic IAPP species remains elusive. Using concurrent time-resolved biophysical and biological measurements, we define the toxic species produced during IAPP amyloid formation and link their properties to induction of rat INS-1 β-cell and murine islet toxicity. These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species. They do not bind 1-anilnonaphthalene-8-sulphonic acid and lack extensive β-sheet structure. Aromatic interactions modulate, but are not required for toxicity. Not all IAPP oligomers are toxic; toxicity depends on their partially structured conformational states. Some anti-amyloid agents paradoxically prolong cytotoxicity by prolonging the lifetime of the toxic species. The data highlight the distinguishing properties of toxic IAPP oligomers and the common features that they share with toxic species reported for other amyloidogenic polypeptides, providing information for rational drug design to treat IAPP induced β-cell death.

No MeSH data available.


Related in: MedlinePlus