Controlling nanoparticle-induced endothelial leakiness with the protein corona.

Understanding nanoparticle-cell interaction is essential for advancing research in nanomedicine and nanotoxicology. Apart from the transcytotic pathway mediated by cellular recognition and energetics, nanoparticles (including nanomedicines) may harness the paracellular route for their transport by inducing endothelial leakiness at cadherin junctions. This phenomenon, termed as NanoEL, is correlated with the physicochemical properties of the nanoparticles in close association with cellular signalling, membrane mechanics, as well as cytoskeletal remodelling. However, nanoparticles in biological systems are transformed by the ubiquitous protein corona and yet the potential effect of the protein corona on NanoEL remains unclear. Using confocal fluorescence microscopy, biolayer interferometry, transwell, toxicity, and molecular inhibition assays, complemented by molecular docking, here we reveal the minimal to significant effects of the anionic human serum albumin and fibrinogen, the charge neutral immunoglobulin G as well as the cationic lysozyme on negating gold nanoparticle-induced endothelial leakiness in vitro and in vivo. This study suggests that nanoparticle-cadherin interaction and hence the extent of NanoEL may be partially controlled by pre-exposing the nanoparticles to plasma proteins of specific charge and topology to facilitate their biomedical applications.

Remediation of Metal Oxide Nanotoxicity with a Functional Amyloid.

Understanding the environmental health and safety of nanomaterials (NanoEHS) is essential for the sustained development of nanotechnology. Although extensive research over the past two decades has elucidated the phenomena, mechanisms, and implications of nanomaterials in cellular and organismal models, the active remediation of the adverse biological and environmental effects of nanomaterials remains largely unexplored. Inspired by recent developments in functional amyloids for biomedical and environmental engineering, this work shows their new utility as metallothionein mimics in the strategically important area of NanoEHS. Specifically, metal ions released from CuO and ZnO nanoparticles are sequestered through cysteine coordination and electrostatic interactions with beta-lactoglobulin (bLg) amyloid, as revealed by inductively coupled plasma mass spectrometry and molecular dynamics simulations. The toxicity of the metal oxide nanoparticles is subsequently mitigated by functional amyloids, as validated by cell viability and apoptosis assays in vitro and murine survival and biomarker assays in vivo. As bLg amyloid fibrils can be readily produced from whey in large quantities at a low cost, the study offers a crucial strategy for remediating the biological and environmental footprints of transition metal oxide nanomaterials.

Endothelial leakiness elicited by amyloid protein aggregation.

Alzheimer's disease (AD) is a major cause of dementia debilitating the global ageing population. Current understanding of the AD pathophysiology implicates the aggregation of amyloid beta (Aß) as causative to neurodegeneration, with tauopathies, apolipoprotein E and neuroinflammation considered as other major culprits. Curiously, vascular endothelial barrier dysfunction is strongly associated with Aß deposition and 80-90% AD subjects also experience cerebral amyloid angiopathy. Here we show amyloid protein-induced endothelial leakiness (APEL) in human microvascular endothelial monolayers as well as in mouse cerebral vasculature. Using signaling pathway assays and discrete molecular dynamics, we revealed that the angiopathy first arose from a disruption to vascular endothelial (VE)-cadherin junctions exposed to the nanoparticulates of Aß oligomers and seeds, preceding the earlier implicated proinflammatory and pro-oxidative stressors to endothelial leakiness. These findings were analogous to nanomaterials-induced endothelial leakiness (NanoEL), a major phenomenon in nanomedicine depicting the paracellular transport of anionic inorganic nanoparticles in the vasculature. As APEL also occurred in vitro with the oligomers and seeds of alpha synuclein, this study proposes a paradigm for elucidating the vascular permeation, systemic spread, and cross-seeding of amyloid proteins that underlie the pathogeneses of AD and Parkinson's disease.

Exploring Peptido-Nanocomposites in the Context of Amyloid Diseases.

Therapeutic peptides are a major class of pharmaceutical drugs owing to their target-binding specificity as well as their versatility in inhibiting aberrant protein-protein interactions associated with human pathologies. Within the realm of amyloid diseases, the use of peptides and peptidomimetics tailor-designed to overcome amyloidogenesis has been an active research endeavor since the late 90s. In more recent years, incorporating nanoparticles for enhancing the biocirculation and delivery of peptide drugs has emerged as a frontier in nanomedicine, and nanoparticles have further demonstrated a potency against amyloid aggregation and cellular inflammation to rival strategies employing small molecules, peptides, and antibodies. Despite these efforts, however, a fundamental understanding of the chemistry, characteristics and function of peptido-nanocomposites is lacking, and a systematic analysis of such strategy for combating a range of amyloid pathogeneses is missing. Here we review the history, principles and evolving chemistry of constructing peptido-nanocomposites from bottom up and discuss their future application against amyloid diseases that debilitate a significant portion of the global population.

Nanoplastic Stimulates the Amyloidogenesis of Parkinson's Alpha-Synuclein NACore.

Environmental plastic wastes are potential health hazards due to their prevalence as well as their versatility in initiating physical, chemical, and biological interactions and transformations. Indeed, recent research has implicated the adverse effects of micro- and nano-plastics, including their neurotoxicity, yet how plastic particulates may impact the aggregation pathway and toxicity of amyloid proteins pertinent to the pathologies of neurological diseases remains unknown. Here, electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS) is employed to reveal the polymorphic oligomerization of NACore, a surrogate of alpha-synuclein that is associated with the pathogenesis of Parkinson's disease. These data indicate that the production rate and population of the NACore oligomers are modulated by their exposure to a polystyrene nanoplastic, and these cellular assays further reveal an elevated NACore toxicity in microglial cells elicited by the nanoplastic. These simulations confirm that the nanoplastic-NACore association is promoted by their hydrophobic interactions. These findings are corroborated by an impairment in zebrafish hatching, survival, and development in vivo upon their embryonic exposure to the nanoplastic. Together, this study has uncovered the dynamics and mechanism of amyloidogenesis elevated by a nanoplastic trigger, shedding a new light on the neurological burden of plastic pollution.

Spike Protein Fragments Promote Alzheimer's Amyloidogenesis.

Alzheimer's disease (AD) is a major cause of dementia inducing memory loss, cognitive decline, and mortality among the aging population. While the amyloid aggregation of peptide Aß has long been implicated in neurodegeneration in AD, primarily through the production of toxic polymorphic aggregates and reactive oxygen species, viral infection has a less explicit role in the etiology of the brain disease. On the other hand, while the COVID-19 pandemic is known to harm human organs and function, its adverse effects on AD pathobiology and other human conditions remain unclear. Here we first identified the amyloidogenic potential of 1058HGVVFLHVTYV1068, a short fragment of the spike protein of SARS-CoV-2 coronavirus. The peptide fragment was found to be toxic and displayed a high binding propensity for the amyloidogenic segments of Aß, thereby promoting the aggregation and toxicity of the peptide in vitro and in silico, while retarding the hatching and survival of zebrafish embryos upon exposure. Our study implicated SARS-CoV-2 viral infection as a potential contributor to AD pathogenesis, a little explored area in our quest for understanding and overcoming Long Covid.

Zinc-Epigallocatechin-3-gallate Network-Coated Nanocomposites against the Pathogenesis of Amyloid-Beta.

The aggregation of amyloid beta (Aß) is a hallmark of Alzheimer's disease (AD), a major cause of dementia and an unmet challenge in modern medicine. In this study, we constructed a biocompatible metal-phenolic network (MPN) comprising a polyphenol epigallocatechin gallate (EGCG) scaffold coordinated by physiological Zn(II). Upon adsorption onto gold nanoparticles, the MPN@AuNP nanoconstruct elicited a remarkable potency against the amyloid aggregation and toxicity of Aß in vitro. The superior performance of MPN@AuNP over EGCG@AuNP was attributed to the porosity and hence larger surface area of the MPN in comparison with that of EGCG alone. The atomic detail of Zn(II)-EGCG coordination was unraveled by density functional theory calculations and the structure and dynamics of Aß aggregation modulated by the MPN were further examined by discrete molecular dynamics simulations. As MPN@AuNP also displayed a robust capacity to cross a blood-brain barrier model through the paracellular pathway, and given the EGCG's function as an anti-amyloidosis and antioxidation agent, this MPN-based strategy may find application in regulating the broad AD pathology beyond protein aggregation inhibition.

Modulating Nanodroplet Formation En Route to Fibrillization of Amyloid Peptides with Designed Flanking Sequences.

Soluble oligomers populating early amyloid aggregation can be regarded as nanodroplets of liquid-liquid phase separation (LLPS). Amyloid peptides typically contain hydrophobic aggregation-prone regions connected by hydrophilic linkers and flanking sequences, and such a sequence hydropathy pattern drives the formation of supramolecular structures in the nanodroplets and modulates subsequent fibrillization. Here, we studied LLPS and fibrillization of coarse-grained amyloid peptides with increasing flanking sequences. Nanodroplets assumed lamellar, cylindrical micellar, and spherical micellar structures with increasing peptide hydrophilic/hydrophobic ratios, and such morphologies governed subsequent fibrillization processes. Adding glycine-serine repeats as flanking sequences to Aß16-22, the amyloidogenic core of amyloid-ß, our computational predictions of morphological transitions were corroborated experimentally. The uncovered inter-relationships between the peptide sequence pattern, oligomer/nanodroplet morphology, and fibrillization pathway, kinetics, and structure may contribute to our understanding of pathogenic amyloidosis in aging, facilitate future efforts ameliorating amyloidosis through peptide engineering, and aid in the design of novel amyloid-based functional nanobiomaterials and nanocomposites.

Rapid Oxygen-Tolerant Synthesis of Protein-Polymer Bioconjugates via Aqueous Copper-Mediated Polymerization.

The synthesis of protein-polymer conjugates usually requires extensive and costly deoxygenation procedures, thus limiting their availability and potential applications. In this work, we report the ultrafast synthesis of polymer-protein bioconjugates in the absence of any external deoxygenation via an aqueous copper-mediated methodology. Within 10 min and in the absence of any external stimulus such as light (which may limit the monomer scope and/or disrupt the secondary structure of the protein), a range of hydrophobic and hydrophilic monomers could be successfully grafted from a BSA macroinitiator, yielding well-defined polymer-protein bioconjugates at quantitative yields. Our approach is compatible with a wide range of monomer classes such as (meth) acrylates, styrene, and acrylamides as well as multiple macroinitiators including BSA, BSA nanoparticles, and beta-galactosidase from Aspergillus oryzae. Notably, the synthesis of challenging protein-polymer-polymer triblock copolymers was also demonstrated, thus significantly expanding the scope of our strategy. Importantly, both lower and higher scale polymerizations (from 0.2 to 35 mL) were possible without compromising the overall efficiency and the final yields. This simple methodology paves the way for a plethora of applications in aqueous solutions without the need of external stimuli or tedious deoxygenation.

Anionic nanoplastic exposure induces endothelial leakiness.

The global-scale production of plastics has been instrumental in advancing modern society, while the rising accumulation of plastics in landfills, oceans, and anything in between has become a major stressor on environmental sustainability, climate, and, potentially, human health. While mechanical and chemical forces of man and nature can eventually break down or recycle plastics, our understanding of the biological fingerprints of plastics, especially of nanoplastics, remains poor. Here we report on a phenomenon associated with the nanoplastic forms of anionic polystyrene and poly(methyl methacrylate), where their introduction disrupted the vascular endothelial cadherin junctions in a dose-dependent manner, as revealed by confocal fluorescence microscopy, signaling pathways, molecular dynamics simulations, as well as ex vivo and in vivo assays with animal model systems. Collectively, our results implicated nanoplastics-induced vasculature permeability as primarily biophysical-biochemical in nature, uncorrelated with cytotoxic events such as reactive oxygen species production, autophagy, and apoptosis. This uncovered route of paracellular transport has opened up vast avenues for investigating the behaviour and biological effects of nanoplastics, which may offer crucial insights for guiding innovations towards a sustainable plastics industry and environmental remediation.

Chemical and Biophysical Signatures of the Protein Corona in Nanomedicine.

An inconvenient hurdle in the practice of nanomedicine is the protein corona, a spontaneous collection of biomolecular species by nanoparticles in living systems. The protein corona is dynamic in composition and may entail improved water suspendability and compromised delivery and targeting to the nanoparticles. How much of this nonspecific protein ensemble is determined by the chemistry of the nanoparticle core and its surface functionalization, and how much of this entity is dictated by the biological environments that vary spatiotemporally in vivo? How do we "live with" and exploit the protein corona without significantly sacrificing the efficacy of nanomedicines in diagnosing and curing human diseases? This article discusses the chemical and biophysical signatures of the protein corona and ponders challenges ahead for the field of nanomedicine.

Inhibition of Amyloid Aggregation and Toxicity with Janus Iron Oxide Nanoparticles.

Amyloid aggregation is a ubiquitous form of protein misfolding underlying the pathologies of Alzheimer's disease (AD), Parkinson's disease (PD) and type 2 diabetes (T2D), three primary forms of human amyloid diseases. While much has been learned about the origin, diagnosis and management of these neurological and metabolic disorders, no cure is currently available due in part to the dynamic and heterogeneous nature of the toxic oligomers induced by amyloid aggregation. Here we synthesized beta casein-coated iron oxide nanoparticles (ßCas IONPs) via a BPA-P(OEGA-b-DBM) block copolymer linker. Using a thioflavin T kinetic assay, transmission electron microscopy, Fourier transform infrared spectroscopy, discrete molecular dynamics simulations and cell viability assays, we examined the Janus characteristics and the inhibition potential of ßCas IONPs against the aggregation of amyloid beta (Aß), alpha synuclein (αS) and human islet amyloid polypeptide (IAPP) which are implicated in the pathologies of AD, PD and T2D. Incubation of zebrafish embryos with the amyloid proteins largely inhibited hatching and elicited reactive oxygen species, which were effectively rescued by the inhibitor. Furthermore, Aß-induced damage to mouse brain was mitigated in vivo with the inhibitor. This study revealed the potential of Janus nanoparticles as a new nanomedicine against a diverse range of amyloid diseases.

Graphene quantum dots obstruct the membrane axis of Alzheimer's amyloid beta.

Alzheimer's disease (AD) is a primary form of dementia with debilitating consequences, but no effective cure is available. While the pathophysiology of AD remains multifactorial, the aggregation of amyloid beta (Aß) mediated by the cell membrane is known to be the cause for the neurodegeneration associated with AD. Here we examined the effects of graphene quantum dots (GQDs) on the obstruction of the membrane axis of Aß in its three representative forms of monomers (Aß-m), oligomers (Aß-o), and amyloid fibrils (Aß-f). Specifically, we determined the membrane fluidity of neuroblastoma SH-SY5Y cells perturbed by the Aß species, especially by the most toxic Aß-o, and demonstrated their recovery by GQDs using confocal fluorescence microscopy. Our computational data through discrete molecular dynamics simulations further revealed energetically favorable association of the Aß species with the GQDs in overcoming peptide-peptide aggregation. Overall, this study positively implicated GQDs as an effective agent in breaking down the membrane axis of Aß, thereby circumventing adverse downstream events and offering a potential therapeutic solution for AD.

Ultrasmall Molybdenum Disulfide Quantum Dots Cage Alzheimer's Amyloid Beta to Restore Membrane Fluidity.

Alzheimer's disease (AD) is a major cause of dementia characterized by the overexpression of transmembrane amyloid precursor protein and its neurotoxic byproduct amyloid beta (Aß). A small peptide of considerable hydrophobicity, Aß is aggregation prone catalyzed by the presence of cell membranes, among other environmental factors. Accordingly, current AD mitigation strategies often aim at breaking down the Aß-membrane communication, yet no data is available concerning the cohesive interplay of the three key entities of the cell membrane, Aß, and its inhibitor. Using a lipophilic Laurdan dye and confocal fluorescence microscopy, we observed cell membrane perturbation and actin reorganization induced by Aß oligomers but not by Aß monomers or amyloid fibrils. We further revealed recovery of membrane fluidity by ultrasmall MoS2 quantum dots, also shown in this study as a potent inhibitor of Aß amyloid aggregation. Using discrete molecular dynamics simulations, we uncovered the binding of MoS2 and Aß monomers as mediated by hydrophilic interactions between the quantum dots and the peptide N-terminus. In contrast, Aß oligomers and fibrils were surface-coated by the ultrasmall quantum dots in distinct testudo-like, reverse protein-corona formations to prevent their further association with the cell membrane and adverse effects downstream. This study offers a crucial new insight and a viable strategy for regulating the amyloid aggregation and membrane-axis of AD pathology with multifunctional nanomedicine.

Spontaneous Formation of ß-sheet Nano-barrels during the Early Aggregation of Alzheimer's Amyloid Beta.

Soluble low-molecular-weight oligomers formed during the early aggregation of amyloid peptides have been hypothesized as a major toxic species of amyloidogenesis. Herein, we performed the first synergic in silico, in vitro and in vivo validations of the structure, dynamics and toxicity of Aß42 oligomers. Aß peptides readily assembled into ß-rich oligomers comprised of extended ß-hairpins and ß-strands. Nanosized ß-barrels were observed with certainty with simulations, transmission electron microscopy and Fourier transform infrared spectroscopy, corroborated by immunohistochemistry, cell viability, apoptosis, inflammation, autophagy and animal behavior assays. Secondary and tertiary structural proprieties of these oligomers, such as the sequence regions with high ß-sheet propensities and inter-residue contact frequency patterns, were similar to the properties known for Aß fibrils. The unambiguous spontaneous formation of ß-barrels in the early aggregation of Aß42 supports their roles as the common toxic intermediates in Alzheimer's pathobiology and a target for Alzheimer's therapeutics.

The membrane axis of Alzheimer's nanomedicine.

Alzheimer's disease (AD) is a major neurological disorder impairing its carrier's cognitive function, memory and lifespan. While the development of AD nanomedicine is still nascent, the field is evolving into a new scientific frontier driven by the diverse physicochemical properties and theranostic potential of nanomaterials and nanocomposites. Characteristic to the AD pathology is the deposition of amyloid plaques and tangles of amyloid beta (Aß) and tau, whose aggregation kinetics may be curbed by nanoparticle inhibitors via sequence-specific targeting or nonspecific interactions with the amyloidogenic proteins. As literature implicates cell membrane as a culprit in AD pathogenesis, here we summarize the membrane axis of AD nanomedicine and present a new rationale that the field development may greatly benefit from harnessing our existing knowledge of Aß-membrane interaction, nanoparticle-membrane interaction and Aß-nanoparticle interaction.

Accelerated Amyloid Beta Pathogenesis by Bacterial Amyloid FapC.

The gut-brain axis has attracted increasing attention in recent years, fueled by accumulating symptomatic, physiological, and pathological findings. In this study, the aggregation and toxicity of amyloid beta (Aß), the pathogenic peptide associated with Alzheimer's disease (AD), seeded by FapC amyloid fragments (FapCS) of Pseudomonas aeruginosa that colonizes the gut microbiome through infections are examined. FapCS display favorable binding with Aß and a catalytic capacity in seeding the peptide amyloidosis. Upon seeding, twisted Aß fibrils assume a much-shortened periodicity approximating that of FapC fibrils, accompanied by a 37% sharp rise in the fibrillar diameter, compared with the control. The robust seeding capacity for Aß by FapCS and the biofilm fragments derived from P. aeruginosa entail abnormal behavior pathology and immunohistology, as well as impaired cognitive function of zebrafish. Together, the data offer the first concrete evidence of structural integration and inheritance in peptide cross-seeding, a crucial knowledge gap in understanding the pathological correlations between different amyloid diseases. The catalytic role of infectious bacteria in promoting Aß amyloidosis may be exploited as a potential therapeutic target, while the altered mesoscopic signatures of Aß fibrils may serve as a prototype for molecular assembly and a biomarker for screening bacterial infections in AD.

Nanomaterial synthesis, an enabler of amyloidosis inhibition against human diseases.

Amyloid diseases are global epidemics with no cure currently available. In the past decade, the use of engineered nanomaterials as inhibitors or probes against the pathogenic aggregation of amyloid peptides and proteins has emerged as a new frontier in nanomedicine. In this Minireview, we summarize for the first time the pivotal role of chemical synthesis in enabling the development of this multidisciplinary field.