Evidence for different in vitro oligomerization behaviors of synthetic hIAPP obtained from different sources

Abstract

Type 2 diabetes is characterized by the aggregation of human islet amyloid polypeptide (hIAPP), from monomer to amyloid deposits that are made of insoluble fibrils. Discrepancies concerning the nature of formed species or oligomerization kinetics among reported in vitro studies on hIAPP aggregation process have been highlighted. In this work, we investigated if the sample itself could be at the origin of those observed differences. To this aim, four hIAPP samples obtained from three different sources or suppliers have been analyzed and compared by ThT fluorescence spectroscopy and by two recently developed techniques, capillary electrophoresis (CE), and ESI-IMS-QToF-MS. Lots provided by the same supplier were shown to be very similar whatever the analytical technique used to characterize them. In contrast, several critical differences could be pointed out for hIAPP provided by different suppliers. We demonstrated that in several samples, some oligomerized peptides (e.g., dimer) were already present upon reception. Purity was also different, and the proneness of the peptide solution to form fibrils in vitro within 24 h could vary considerably from one sample source to another but not from lot to lot of the same source. All those results demonstrate that the initial state of conformation, oligomerization, and quality of the hIAPP can greatly impact the aggregation kinetics, and thus the information provided by these in vitro tests. Finally, a careful selection of the peptide batch and source is mandatory to perform relevant in vitro studies on hIAPP oligomerization and to screen new molecules modulating this pathological process.

Helical γ‐Peptide Foldamers as Dual Inhibitors of Amyloid‐β Peptide and Islet Amyloid Polypeptide Oligomerization and Fibrillization

“…Mass spectrometry (MS) and ion mobility spectrometry‐mass spectrometry (IMS‐MS) have been recently proposed for monitoring hIAPP species formed within the oligomerization process as well as for probing mechanisms of inhibition of hIAPP fibril formation [55–60] . Using the same buffer conditions as for CE experiments (50 μ m hIAPP, 50 m m ammonium acetate, pH 3.7 at room temperature) the MS spectrum showed the classically observed dynamic set of hIAPP species, mostly monomer to hexamer species in multiple charge states, (Figure 6 a,b).…”

“…We next evaluated the effect of foldamer 6 on the oligomerization process of Ab 1-42 and hIAPP peptides using capillary electrophoresis (CE). Some of us developed this technique for monitoring soluble species formed during the early Ab 1-42 [21][22][23]53] and hIAPP [54,55] aggregation processes. The typical electrophoretic profile over time for the Ab 1-42 control sample is shown in Figure 4a.A tt he beginning of the kinetics, the monomer ES was the major visible speciesa ssociated with a very small amount of small oligomers ES' (dimers to tetramers).…”

β-Hairpin Peptide Mimics Decrease Human Islet Amyloid Polypeptide (hIAPP) Aggregation

“…We then evaluated the effect of the three hairpins 3 , 8, and 14 on the oligomerization process of hIAPP peptide using our recently developed CE method able to monitor the soluble oligomeric species formed during the early hIAPP aggregation process ( Berardet et al, 2018 , 2020 ). The control electropherogram of hIAPP, shown in Figure 7A , displayed over time the typical profile, showing three groups of soluble species: peak 3 is mainly composed by monomers with possibly a small amount of dimers and trimers whereas peaks 1 and 2 are mainly assigned to oligomeric aggregates larger than 25-mers (peak 1 co-migrates with the electroosmotic flow EOF) ( Berardet et al, 2018 ).…”

“…A similar approach was used to characterize beta-amyloid oligomer-aggregates properties ( Iuraşcu et al, 2009 ). In addition, they also have been reported as powerful methods for studying the mechanism of hIAPP aggregation inhibitors ( Young et al, 2014 , 2015 , 2016 ; Li et al, 2015a ; Riba et al, 2015 ; Berardet et al, 2020 ). Our reported method, using the same buffer as for CE experiments ( Berardet et al, 2020 ; Kaffy et al, 2020 ), allowed us to study the effect of the hairpins 3 , 8, and 14 on the oligomerization process of hIAPP at three different hairpin/hIAPP ratios (10/1, 5/1, and 1/1).…”

“…The next step was to evaluate the ability of our compounds to decrease the presence of toxic oligomers and to maintain the presence of non-toxic monomers, as it is now well established that oligomers of hIAPP are toxic species ( Bram et al, 2014 ; Abedini et al, 2016 ; Akter et al, 2016 ; Kiriyama and Nochi, 2018 ; Rodriguez Camargo et al, 2018 ). The very early steps of the oligomerization process overtime and the analysis of the effect of our hairpins on these challenging first stages was thus performed by our reported CE and MS methods ( Berardet et al, 2018 , 2020 ). The most promising compounds 3 , 8, and 14 as inhibitors of hIAPP fibrillization had also a strong anti-oligomeric activity, and were able to stabilize the non-toxic monomeric species.…”