693: Probing Amyloid-Like Protein Aggregates in Urine of Women with Preeclampsia (PE) Using Atomic Force Microscopy

Compelling evidence suggests amyloid-like misfolded protein aggregates exist in urine of women with PE and may play a pathogenic role analogous to soluble oligomers found in other protein aggregation disorders (e.g. Alzheimer’s disease). The physical characteristics of these protein aggregates may clarify the molecular mechanisms underlying the pathology of PE. Our objective was to apply atomic force microscopy (AFM) to provide a direct characterization of urine protein aggregates in PE. We investigated urine samples from women pregnant with singletons in the following groups: 1) PE severe features (sPE, n=5, GA 27.8 ± 4.3 wk); 2) HELLP patients (n=3, GA 25.2 ± 2.6 wk); 3) Healthy pregnant women served as control (CRL, n=3, GA 24.7 ± 3.3 wk). Urine was analyzed in the absence and presence of the amyloidophilic dye Congo red (CR), known to induce supramolecular self-assembly of PE aggregates. Spun urine along with its CR mixture were spotted onto freshly cleaved mica surfaces and incubated for protein adsorption. After washing, protein aggregates were visualized using Asylum Research AFM (MFP-3D Bio) on AC tapping mode in air with AC40-TS tips. Image data were processed with AR14 software using a particle threshold of 800 pm. Compared with CRL, sPE and HELLP were characterized by significant differences in AFM parameters such as average Z height, hull perimeter, particle volume, particle area, maximum distance and circle equivalent (CE) diameter (Table). In both sPE and HELLP, the particles were larger than in CRL. Compared to CRL, the AFM characteristics of the particles in sPE and HELLP appeared similar both in the presence and absence of CR. Interestingly, CR significantly decreased the size of the particles in sPE and HELLP urine (Fig). CR interacts with urine protein aggregates at molecular level likely affecting their folding and/or compaction characteristics. The high similarity between protein aggregates of women with sPE and HELLP syndrome supports a common pathogenic role at atomic force level.View Large Image Figure ViewerDownload Hi-res image Download (PPT)