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Aspirin, diabetes, and amyloid: re-examination of the inhibition of amyloid formation by aspirin and ketoprofen.

Tu LH, Noor H, Cao P, Raleigh DP - ACS Chem. Biol. (2014)

Bottom Line: The loss of β-cell function and β-cell death are key features of diabetes.There are no therapeutic strategies for the treatment or prevention of islet amyloidosis.Ketoprofen is similarly ineffective at inhibiting amylin amyloid formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Stony Brook University , Stony Brook, New York 11794-3400, United States.

ABSTRACT
The loss of β-cell function and β-cell death are key features of diabetes. A range of mechanisms are thought to contribute to β-cell loss, including islet amyloid formation by the neuropancreatic hormone amylin (islet amyloid polypeptide, IAPP). Islet amyloid deposition also contributes to the failure of islet transplants. There are no therapeutic strategies for the treatment or prevention of islet amyloidosis. Aspirin and the nonsteroid anti-inflammatory drug (NSAID) ketoprofen, at clinically relevant doses, have been proposed to inhibit amyloid formation by amylin and thus may hold promise for treatment of islet amyloidosis. These compounds are potentially attractive given the importance of inflammation in islet amyloidosis and given the fact that there are no anti-islet amyloid agents in the clinic. We show that aspirin, even in 20-fold excess, has no effect on the kinetics of amyloid formation by amylin as judged by thioflavin-T binding, right angle light scattering, and transmission electron microscopy, nor does it alter the morphology of resulting amyloid fibrils. Aspirin showed no ability to disaggregate preformed amylin amyloid fibrils under the conditions of these studies, 25 °C and pH 7.4. Ketoprofen is similarly ineffective at inhibiting amylin amyloid formation. The compounds do, however, interfere with circular dichroism- and Congo Red-based assays of amylin amyloid formation. This study highlights the importance of using multiple methods to follow amyloid formation when screening inhibitors.

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Aspirin does not inhibit amyloid formationby human amylin. (A)Right angle light scattering assays of the time course of amyloidformation in the absence (black) and in the presence (red) of a 20-foldexcess of aspirin. No thioflavin-T was added to either sample. (B)TEM image of the sample of amylin without aspirin recorded at theend of the kinetic experiment. (C) TEM image of the sample of amylinwith a 20-fold excess aspirin recorded at the end of the kinetic experiment.Scale bars represent 100 nm. Experiments were conducted at pH 7.4and 25 °C in 20 mM Tris buffer with 0.25% DMSO (v/v) in the absenceof any fluorinated alcohol cosolvent. The concentration of amylinwas 16 μM, and the concentration of aspirin was 320 μM.
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fig3: Aspirin does not inhibit amyloid formationby human amylin. (A)Right angle light scattering assays of the time course of amyloidformation in the absence (black) and in the presence (red) of a 20-foldexcess of aspirin. No thioflavin-T was added to either sample. (B)TEM image of the sample of amylin without aspirin recorded at theend of the kinetic experiment. (C) TEM image of the sample of amylinwith a 20-fold excess aspirin recorded at the end of the kinetic experiment.Scale bars represent 100 nm. Experiments were conducted at pH 7.4and 25 °C in 20 mM Tris buffer with 0.25% DMSO (v/v) in the absenceof any fluorinated alcohol cosolvent. The concentration of amylinwas 16 μM, and the concentration of aspirin was 320 μM.

Mentions: Initial reports of the abilityof aspirin to inhibit amylin amyloidformation did not use thioflavin-T assays or TEM, but rather usedCD and Congo red binding assays. It is possible, although unlikely,that thioflavin-T could displace aspirin from amylin and interferewith its effects. This does not seem plausible because thioflavin-Thas no such effect on a wide range of other inhibitors27,28 and does not bind to preamyloid intermediates. Nonetheless, we alsotested the effects of aspirin in the absence of thioflavin-T usingright angle light scattering (RALS) and TEM. Similar aggregation curvesare observed in the presence and absence of a 20-fold excess of aspirinusing RALS (Figure 3). TEM of the samples revealsthe presence of dense mats of fibrils in each sample. Note that thesesamples did not contain thioflavin-T.


Aspirin, diabetes, and amyloid: re-examination of the inhibition of amyloid formation by aspirin and ketoprofen.

Tu LH, Noor H, Cao P, Raleigh DP - ACS Chem. Biol. (2014)

Aspirin does not inhibit amyloid formationby human amylin. (A)Right angle light scattering assays of the time course of amyloidformation in the absence (black) and in the presence (red) of a 20-foldexcess of aspirin. No thioflavin-T was added to either sample. (B)TEM image of the sample of amylin without aspirin recorded at theend of the kinetic experiment. (C) TEM image of the sample of amylinwith a 20-fold excess aspirin recorded at the end of the kinetic experiment.Scale bars represent 100 nm. Experiments were conducted at pH 7.4and 25 °C in 20 mM Tris buffer with 0.25% DMSO (v/v) in the absenceof any fluorinated alcohol cosolvent. The concentration of amylinwas 16 μM, and the concentration of aspirin was 320 μM.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215902&req=5

fig3: Aspirin does not inhibit amyloid formationby human amylin. (A)Right angle light scattering assays of the time course of amyloidformation in the absence (black) and in the presence (red) of a 20-foldexcess of aspirin. No thioflavin-T was added to either sample. (B)TEM image of the sample of amylin without aspirin recorded at theend of the kinetic experiment. (C) TEM image of the sample of amylinwith a 20-fold excess aspirin recorded at the end of the kinetic experiment.Scale bars represent 100 nm. Experiments were conducted at pH 7.4and 25 °C in 20 mM Tris buffer with 0.25% DMSO (v/v) in the absenceof any fluorinated alcohol cosolvent. The concentration of amylinwas 16 μM, and the concentration of aspirin was 320 μM.
Mentions: Initial reports of the abilityof aspirin to inhibit amylin amyloidformation did not use thioflavin-T assays or TEM, but rather usedCD and Congo red binding assays. It is possible, although unlikely,that thioflavin-T could displace aspirin from amylin and interferewith its effects. This does not seem plausible because thioflavin-Thas no such effect on a wide range of other inhibitors27,28 and does not bind to preamyloid intermediates. Nonetheless, we alsotested the effects of aspirin in the absence of thioflavin-T usingright angle light scattering (RALS) and TEM. Similar aggregation curvesare observed in the presence and absence of a 20-fold excess of aspirinusing RALS (Figure 3). TEM of the samples revealsthe presence of dense mats of fibrils in each sample. Note that thesesamples did not contain thioflavin-T.

Bottom Line: The loss of β-cell function and β-cell death are key features of diabetes.There are no therapeutic strategies for the treatment or prevention of islet amyloidosis.Ketoprofen is similarly ineffective at inhibiting amylin amyloid formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Stony Brook University , Stony Brook, New York 11794-3400, United States.

ABSTRACT
The loss of β-cell function and β-cell death are key features of diabetes. A range of mechanisms are thought to contribute to β-cell loss, including islet amyloid formation by the neuropancreatic hormone amylin (islet amyloid polypeptide, IAPP). Islet amyloid deposition also contributes to the failure of islet transplants. There are no therapeutic strategies for the treatment or prevention of islet amyloidosis. Aspirin and the nonsteroid anti-inflammatory drug (NSAID) ketoprofen, at clinically relevant doses, have been proposed to inhibit amyloid formation by amylin and thus may hold promise for treatment of islet amyloidosis. These compounds are potentially attractive given the importance of inflammation in islet amyloidosis and given the fact that there are no anti-islet amyloid agents in the clinic. We show that aspirin, even in 20-fold excess, has no effect on the kinetics of amyloid formation by amylin as judged by thioflavin-T binding, right angle light scattering, and transmission electron microscopy, nor does it alter the morphology of resulting amyloid fibrils. Aspirin showed no ability to disaggregate preformed amylin amyloid fibrils under the conditions of these studies, 25 °C and pH 7.4. Ketoprofen is similarly ineffective at inhibiting amylin amyloid formation. The compounds do, however, interfere with circular dichroism- and Congo Red-based assays of amylin amyloid formation. This study highlights the importance of using multiple methods to follow amyloid formation when screening inhibitors.

Show MeSH
Related in: MedlinePlus