The Effects of Lipid Membranes, Crowding and Osmolytes on the Aggregation, and Fibrillation Propensity of Human IAPP

Figure 3

Structure, morphology, and cytotoxicity of hIAPP during its self-assembly. (a) CD spectrum of freshly prepared hIAPP (10 μM) in 10 mM phosphate buffer, pH 7.4 at 25°C, and secondary structure content obtained indicating a predominantly disordered structure of the monomeric peptide. (b) ThT assay of 100 μM hIAPP in 10 mM sodium acetate buffer containing 50 μM ThT, at pH 5.5, and 10°C showing a sigmoidal curve typical for a nucleation-dependent process. The fluorescence intensity was normalized to the intensity recorded at 400 h assuming the fibril formation to be completed. Time points at which hIAPP species were isolated are highlighted in gray. (c) Tapping mode AFM images of isolated hIAPP species at particular time points of the aggregation process showing the hIAPP morphology within the lag phase and the elongation phase. The scale bar included in the images represents 250 nm. (d) WST-1 cell proliferation assay of pancreatic INS-1E cells exposed to 10 μM isolated hIAPP and nonamyloidogenic ratIAPP species at different time points of the aggregation process, indicating the highest toxicity of hIAPP species within the lag phase. Adapted and modified from [17, 56] with permission from Wiley-VCH and Elsevier.

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