Yihua Loo

Natural tri- to hexapeptides self-assemble in water to amyloid β-type fiber aggregates by unexpected α-helical intermediate structures

Many fatal neurodegenerative diseases such as Alzheimer’s, Parkinson, the prion-related diseases, and non-neurodegenerative disorders such as type II diabetes are characterized by abnormal amyloid fiber aggregates, suggesting a common mechanism of pathogenesis. We have discovered that a class of systematically designed natural tri- to hexapeptides with a characteristic sequential motif can simulate the process of fiber assembly and further condensation to amyloid fibrils, probably via unexpected dimeric α-helical intermediate structures. The characteristic sequence motif of the novel peptide class consists of an aliphatic amino acid tail of decreasing hydrophobicity capped by a polar head. To our knowledge, the investigated aliphatic tripeptides are the shortest ever reported naturally occurring amino acid sequence that can adopt α-helical structure and promote amyloid formation. We propose the stepwise assembly process to be associated with characteristic conformational changes from random coil to α-helical intermediates terminating in cross-β peptide structures. Circular dichroism and X-ray fiber diffraction analyses confirmed the concentration-dependent conformational changes of the peptides in water. Molecular dynamics simulating peptide behavior in water revealed monomer antiparallel pairing to dimer structures by complementary structural alignment that further aggregated and stably condensed into coiled fibers. The ultrasmall size and the dynamic facile assembly process make this novel peptide class an excellent model system for studying the mechanism of amyloidogenesis, its evolution and pathogenicity. The ability to modify the properties of the assembled structures under defined conditions will shed light on strategies to manipulate the pathogenic amyloid aggregates in order to prevent or control aggregate formation.

From short peptides to nanofibers to macromolecular assemblies in biomedicine.

In the last few years, a variety of self-assembling short peptides that consist exclusively of simple amino acids have been designed and modified. These peptides exhibit self-assembling dynamic behaviors. At the molecular structural level, they form α-helical, β-sheet and β-hairpins structures in water. These structures further undergo spontaneous assembly to form nanofibers which aggregate into supramolecular scaffolds that entrap large volumes of water. Furthermore, nanostructures and supramolecular structures that self-organized from these short peptides also have a broad spectrum of biotechnological applications. They are useful as biological materials for 2D and 3D tissue cell cultures, regenerative and reparative medicine, tissue engineering as well as injectable drug delivery matrices that gel in situ. We have endeavored to do a comprehensive review of short peptides that form nanofibrous hydrogels. In particular, we have focused on recent advances in peptide assembly motifs and applications.

Peptide Bioink: Self-Assembling Nanofibrous Scaffolds for Three-Dimensional Organotypic Cultures

Printable scaffolds with adequate mechanical strength and stiffness are sought after to ensure viability of printed cells and tissues. We report the first peptide bioinks-lysine-containing hexapeptides that self-assemble into stable, nanofibrous three-dimensional hydrogels with unprecedented stiffness of up to 40 kPa. These biocompatible scaffolds support the three-dimensional culture of human stem cells and differentiation of primary cells into organotypic (gastrointestinal and skin) structures for high-throughput screening, diagnosis, and tissue engineering.

Bioprinting synthetic self-assembling peptide hydrogels for biomedical applications.

Three-dimensional (3D) bioprinting is a disruptive technology for creating organotypic constructs for high-throughput screening and regenerative medicine. One major challenge is the lack of suitable bioinks. Short synthetic self-assembling peptides are ideal candidates. Several classes of peptides self-assemble into nanofibrous hydrogels resembling the native extracellular matrix. This is a conducive microenvironment for maintaining cell survival and physiological function. Many peptides also demonstrate stimuli-responsive gelation and tuneable mechanical properties, which facilitates extrusion before dispensing and maintains the shape fidelity of the printed construct in aqueous media. The inherent biocompatibility and biodegradability bodes well for in vivo applications as implantable tissues and drug delivery matrices, while their short length and ease of functionalization facilitates synthesis and customization. By applying self-assembling peptide inks to bioprinting, the dynamic complexity of biological tissue can be recreated, thereby advancing current biomedical applications of peptide hydrogel scaffolds.

Creation of consistent burn wounds: a rat model.

Background Burn infliction techniques are poorly described in rat models. An accurate study can only be achieved with wounds that are uniform in size and depth. We describe a simple reproducible method for creating consistent burn wounds in rats. Methods Ten male Sprague-Dawley rats were anesthetized and dorsum shaved. A 100 g cylindrical stainless-steel rod (1 cm diameter) was heated to 100℃ in boiling water. Temperature was monitored using a thermocouple. We performed two consecutive toe-pinch tests on different limbs to assess the depth of sedation. Burn infliction was limited to the loin. The skin was pulled upwards, away from the underlying viscera, creating a flat surface. The rod rested on its own weight for 5, 10, and 20 seconds at three different sites on each rat. Wounds were evaluated for size, morphology and depth. Results Average wound size was 0.9957 cm2 (standard deviation [SD] 0.1845) (n=30). Wounds created with duration of 5 seconds were pale, with an indistinct margin of erythema. Wounds of 10 and 20 seconds were well-defined, uniformly brown with a rim of erythema. Average depths of tissue damage were 1.30 mm (SD 0.424), 2.35 mm (SD 0.071), and 2.60 mm (SD 0.283) for duration of 5, 10, 20 seconds respectively. Burn duration of 5 seconds resulted in full-thickness damage. Burn duration of 10 seconds and 20 seconds resulted in full-thickness damage, involving subjacent skeletal muscle. Conclusions This is a simple reproducible method for creating burn wounds consistent in size and depth in a rat burn model.