Human coronaviruses (HCoVs) include seven species: HCoV-229E, HCoV-NL63, HCoV-OC43, HCoV-HKU1, MERS-CoV, SARS-CoV-1, and SARS-CoV-2. The last three, classified as Betacoronaviruses, are highly transmissible and have caused severe pandemics. HCoV infections primarily affect the respiratory system, leading to symptoms such as dry cough, fever, and breath shortness, which can progress to acute respiratory failure and death. Beyond respiratory effects, increasing evidence links HCoVs to neurological dysfunction. However, distinguishing direct neural complications from preexisting disorders, particularly in the elderly, remains challenging. This study examines the association between HCoVs and neurodegenerative diseases like Alzheimer disease, Parkinson disease, Lewy body dementia, amyotrophic lateral sclerosis, and Creutzfeldt-Jakob disease. It also presents the long-term neurological effects of HCoV infections and their differential impact across age groups and sexes. A key aspect of this study is the investigation of the sequence and structural similarities between amyloidogenic and HCoV spike proteins, which can provide insights into potential neuropathomechanisms.

The spike protein (SP) is an outward-projecting transmembrane glycoprotein on viral surfaces. SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2), responsible for COVID-19 (Coronavirus Disease 2019), uses SP to infect cells that express angiotensin converting enzyme 2 (ACE2) on their membrane. Remarkably, SP has the ability to cross the blood-brain barrier (BBB) into the brain and cause cerebral damage through various pathomechanisms. To combat the COVID-19 pandemic, novel gene-based products have been used worldwide to induce human body cells to produce SP to stimulate the immune system. This artificial SP also has a harmful effect on the human nervous system.

Narrative review.

This narrative review presents the crucial role of SP in neurological complaints after SARS-CoV-2 infection, but also of SP derived from novel gene-based anti-SARS-CoV-2 products (ASP).

Literature searches using broad terms such as "SARS-CoV-2", "spike protein", "COVID-19", "COVID-19 pandemic", "vaccines", "COVID-19 vaccines", "post-vaccination syndrome", "post-COVID-19 vaccination syndrome" and "proteinopathy" were performed using PubMed. Google Scholar was used to search for topic-specific full-text keywords.

The toxic properties of SP presented in this review provide a good explanation for many of the neurological symptoms following SARS-CoV-2 infection and after injection of SP-producing ASP. Both SP entities (from infection and injection) interfere, among others, with ACE2 and act on different cells, tissues and organs. Both SPs are able to cross the BBB and can trigger acute and chronic neurological complaints. Such SP-associated pathologies (spikeopathies) are further neurological proteinopathies with thrombogenic, neurotoxic, neuroinflammatory and neurodegenerative potential for the human nervous system, particularly the central nervous system. The potential neurotoxicity of SP from ASP needs to be critically examined, as ASPs have been administered to millions of people worldwide.

Long coronavirus disease 2019 (COVID-19)-a postacute consequence of severe acute respiratory syndrome coronavirus 2 infection-manifests with a broad spectrum of relapsing and remitting or persistent symptoms as well as varied levels of organ damage, which may be asymptomatic or present as acute events such as heart attacks or strokes and recurrent infections, hinting at complex underlying pathogenic mechanisms. Central to these symptoms is vascular dysfunction rooted in thrombotic endothelialitis. We review the scientific evidence that widespread endothelial dysfunction (ED) leads to chronic symptomatology. We briefly examine the molecular pathways contributing to endothelial pathology and provide a detailed analysis of how these cellular processes underpin the clinical picture. Noninvasive diagnostic techniques, such as flow-mediated dilation and peripheral arterial tonometry, are evaluated for their utility in identifying ED. We then explore mechanistic, cellular-targeted therapeutic interventions for their potential in treating ED. Overall, we emphasize the critical role of cellular health in managing Long COVID and highlight the need for early intervention to prevent long-term vascular and cellular dysfunction.

In the intricate realm of interactions between hosts and pathogens, Toll-like receptors (TLRs), which play a crucial role in the innate immune response, possess the ability to identify specific molecular signatures. This includes components originating from pathogens such as SARS-CoV-2, as well as the resulting damage-associated molecular patterns (DAMPs), the endogenous molecules released after cellular damage. A developing perspective suggests that TLRs play a central role in neuroinflammation, a fundamental factor in neurodegenerative conditions like Alzheimer's and Parkinson's disease (PD). This comprehensive review consolidates current research investigating the potential interplay between TLRs, their signaling mechanisms, and the processes of neurodegeneration following SARS-CoV-2 infection with an aim to elucidate the involvement of TLRs in the long-term neurological complications of COVID-19 and explore the potential of targeting TLRs as a means of implementing intervention strategies for the prevention or treatment of COVID-19-associated long-term brain outcomes.

Silver nanoparticles (AgNPs) have great potential in a broad range of applications because of their biochemical functionality, unique physical and optical properties. However, it is still a great challenge to synthesize small-size AgNPs with good stability and high performance for biosensors. In this work, triethanolamine and polyacrylic acid modified AgNPs (TEOA@AgNPs-PAA) with controllable size and high catalytic activity for sensitive detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are reported. Monodisperse TEOA@AgNPs-PAA are prepared by the one-pot synthesis using the TEOA and PAA as reducing agent and surfactant, respectively. The size of TEOA@AgNPs-PAA (the average size of 8.65 nm) is 6 times smaller than that of the TEOA@AgNPs (52 ± 1.5 nm) without PAA. The as-prepared TEOA@AgNPs-PAA possess catalytic activity and present mimicking property of horseradish peroxidase, which are employed to fabricate colorimetric biosensors by catalyzing the reaction between H2O2 and 3,3',5,5'-tetramethylbenzidine (TMB) to produce blue oxTMB for sensitive detection of SARS-CoV-2 spiking proteins. Significantly, the small-size TEOA@AgNPs-PAA can catalyze electroreduction process of K2S2O8 to enhance the cathodic ECL signal, and their surface cap abundant TEOA molecules, which can also act as a coreactant to enhance the anodic ECL of Ru(bpy)32+. Under optimal conditions, the fabricate immunosensors for anodic and cathodic ECL determination of SARS-CoV-2 present the detection limits of 9.2 fg/mL and 14.3 fg/mL (S/N = 3), respectively. This work exhibits a promising novel strategy for the development of multifunctional AgNPs as an efficient sensing platform for the clinical diagnosis and biosensing application.

The rapid evolution and global impact of coronaviruses, notably SARS-CoV-1 and SARS-CoV-2, underscore the importance of understanding their molecular mechanisms in detail. This study focuses on the receptor-binding motif (RBM) within the spike protein of these viruses, a critical element for viral entry through interaction with the ACE2 receptor. We investigate the sequence variations in the RBM across SARS-CoV-1, SARS-CoV-2, and its early variants of concern (VOCs). Utilizing multicanonical simulations and microcanonical analysis, we examine how these variations influence the folding dynamics, thermostability, and solubility of the RBMs. Our methodology includes calculating the density of states (DoS) to identify structural phase transitions and assess thermodynamic properties. Furthermore, we solve the Poisson-Boltzmann equation to model the solubility of the RBMs in aqueous environments. This methodology is expected to elucidate structural and functional differences in viral evolution and pathogenicity, likely improving targeted treatments and vaccines.

The amyloid state of proteins occurs in many different contexts in Nature and in modern society, ranging from the pathological kind (neurodegenerative diseases and amyloidosis) via man-made forms (food processing and - to a much smaller extent - protein biologics) to functional versions (bacterial biofilm, peptide hormones and signal transmission). These classes all come together in the human body which endogenously produces amyloidogenic protein able to form pathological human amyloid (PaHA), hosts a microbiome which continuously makes functional bacterial amyloid (FuBA) and ingests food which can contain amyloid. This can have grave consequences, given that PaHA can spread throughout the body in a "hand-me-down" fashion from cell to cell through small amyloid fragments, which can kick-start growth of new amyloid wherever they encounter monomeric amyloid precursors. Amyloid proteins can also self- and cross-seed across dissimilar peptide sequences. While it is very unlikely that ingested amyloid plays a role in this crosstalk, FuBA-PaHA interactions are increasingly implicated in vivo amyloid propagation. We are now in a position to understand the structural and bioinformatic basis for this cross-talk, thanks to the very recently obtained atomic-level structures of the two major FuBAs CsgA (E. coli) and FapC (Pseudomonas). While there are many reports of homology-driven heterotypic interactions between different PaHA, the human proteome does not harbor significant homology to CsgA and FapC. Yet we and others have uncovered significant cross-stimulation (and in some cases inhibition) of FuBA and PaHA both in vitro and in vivo, which we here rationalize based on structure and sequence. These interactions have important consequences for the transmission and development of neurodegenerative diseases, not least because FuBA and PaHA can come into contact via the gut-brain interface, recurrent infections with microbes and potentially even through invasive biofilm in the brain. Whether FuBA and PaHA first interact in the gut or the brain, they can both stimulate and block each other's aggregation as well as trigger inflammatory responses. The microbiome may also affect amyloidogenesis in other ways, e.g. through their own chaperones which recognize and block growth of both PaHA and FuBA as we show both experimentally and computationally. Heterotypic interactions between and within PaHA and FuBA both in vitro and in vivo are a vital part of the amyloid phenomenon and constitute a vibrant and exciting frontier for future research.

The ongoing post-COVID-19 syndrome (PCS) epidemic, causing complications of diverse etiology, necessitates the search for new diagnostic markers and the development of widely accessible methods for their detection. This would enable the prognosis of PCS progression and faster implementation of targeted treatments. One potential marker is neutrophil elastase (NE), whose elevated levels in the blood during PCS may result from organ damage caused by increased secretion of severe inflammatory mediators or amyloidosis resulting from the interaction of NE with SARS-CoV-2. The aim of this publication is to present a step-by-step method for designing an enzymatic ELISA test, enabling the quantitative assessment of NE in the blood serum of patients.

NE was measured using the designed ELISA test.

The study outlines all the steps necessary for designing and optimizing the ELISA test, including the selection of standards, primary and secondary antibodies, and their dilutions. Using the test, elevated NE levels were demonstrated in patients with advanced-stage diabetic nephropathy after symptomatic COVID-19, compared to a relative group of patients sampled before COVID-19.

The undertaken efforts enabled the development of a test with high performance parameters (initially set sensitivity: ≥40 pg/μL; intra-assay precision: 7%; inter-assay precision <20%). No significant cross-reactivity with other tested proteins was observed. Serial dilution of plasma samples resulted in a proportional decrease in signal intensity.

In addition to the conventional symptoms reported for COVID-19, it is becoming increasingly clear that patients with long COVID are exhibiting new symptoms due to the emergence of autoantibodies against G-protein-coupled receptors, among which human muscarinic cholinergic receptors (CHRMs) have been prominently reported. With a chronic condition such as long COVID, additional symptoms caused by anti-CHRM autoantibodies (AAbs) have proven to be an added burden on these patients. The origins of these AAbs, their interactions with, and effects on the function of neural and non-neural cells within the nervous system have remained unknown. Furthermore, the specific symptom complex to which they contribute has not been clearly understood. In this context, we address these issues here and suggest methods to combat the autoantibodies that contribute to neurological symptoms in long COVID.

Since it first appeared, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has had a significant and lasting negative impact on the health and economies of millions of individuals all over the globe. At the level of individual health too, many patients are not recovering fully and experiencing a long-term condition now commonly termed 'long-COVID'. Long-COVID is a collection of symptoms which must last more than 12 weeks following initial COVID infection, and which cannot be adequately explained by alternate diagnoses. The neurological and psychosocial impact of long-COVID is itself now a global health crisis and therefore preventing, diagnosing, and managing these patients is of paramount importance. This review focuses primarily on: neurological functioning deficits; mental health impacts; long-term mood problems; and associated psychosocial issues, among patients suffering from long-COVID with an eye towards the neurological basis of these symptoms. A concise account of the clinical relevance of the neurological and psychosocial impacts of long-COVID, the effects on long-term morbidity, and varied approaches in managing patients with significant chronic neurological symptoms and conditions was extracted from the literature, analysed and reported. A comprehensive account of plausible pathophysiological mechanisms involved in the development of long-COVID, its management, and future research needs have been discussed.

Amyloidosis is characterized by the abnormal accumulation of misfolded proteins, called amyloid fibrils, leading to diverse clinical manifestations. Recent studies on the amyloidogenesis of SARS-CoV-2 protein segments have raised concerns on their potential link to post-infection neurodegeneration, however, the mechanisms remain unclear. Herein, we investigated the structure, stability, and amyloidogenic propensity of a nine-residue segment (SK9) of the SARS-CoV-2 envelope protein and their impact on neuronal protein α-synuclein (αSyn) aggregation. Specifically, the amino acid sequence of the SK9 wildtype has been modified from a basic and positively charged peptide (SFYVYSRVK), to a nearly neutral and more hydrophobic peptide (SAAVASAVK, labelled as SK9 var1), and to an acidic and negatively charged peptide (SDAVANAVK, labelled as SK9 var2). Our findings reveal that the SK9 wildtype exhibited a pronounced amyloidogenic propensity due to its disordered and unstable nature, while the SK9 variants possessed more ordered and stable structures preventing the amyloid formation. Significantly, the SK9 wildtype demonstrated distinct effect on αSyn aggregation kinetics and aggregate morphology to facilitate the formation of αSyn aggregates with enhanced resistance against enzymatic degradation. This study highlights the potential of modifying short peptide sequences to fine-tune their properties, providing insights into understanding and regulating viral-induced amyloid aggregations.

Under certain conditions, proteins may undergo misfolding and form long insoluble aggregates called amyloid fibrils. The presence of these aggregates is often associated with various diseases. The molecular mechanisms governing the aggregation process are yet to be fully understood. The self-assembly of amyloid protofilaments occurs over extended time frames, making the simulation of such events problematic. In this work, we describe a pipeline for multiscale modeling protofilament structures. In the first stage, the self-assembly of short fibrillar oligomers occurs during coarse-grained docking simulations of multiple copies of aggregating peptides. Subsequently, symmetry criteria are used to select the highest-ranked oligomer structures. Selected models are then reconstructed to an all-atom representation and used for the assembly of longer protofilaments. Models are optimized using molecular dynamics. Final structures are selected using various scoring protocols. We evaluated this modeling procedure through the test prediction of insulin amyloid protofilaments whose experimental structures have been published recently. The resulting insulin protofilament models closely resemble the experimental structures. This work provides a proof of concept for the proposed modeling procedure aiming to predict amyloid protofilament structures that exhibit in-register and parallel arrangement of β-sheets based solely on the amino acid sequence of aggregating peptides.

Human islet amyloid polypeptide (hIAPP), an intrinsically disordered protein (IDP), plays a significant role in the pathogenesis of type 2 diabetes through its aggregation. Recent studies have suggested that certain viral protein segments exhibit amyloidogenic potential and may influence its amyloid aggregations associated with pathogenesis. However, the potential link between recurrent SARS-CoV-2 infections and the exacerbation of type 2 diabetes remains poorly understood. In this study, we explore how the amyloidogenic segments of SARS-CoV-2, specifically SK9 and FI10, influence the aggregation of hIAPP and the toxicity of the resulting conformers in a membrane environment. To investigate this, we utilized a range of biophysical techniques, including circular dichroism, nuclear magnetic resonance, atomic force microscopy, dynamic light scattering, fluorescence assays, and cell cytotoxicity assays, complemented by molecular dynamics simulations. Our results indicate that SK9 and FI10 promote hIAPP aggregation in a membrane-mimicking environment, forming distinct aggregate structures. Specifically, SK9 accelerates rapid fibril formation due to inter-chain interactions, while FI10 stabilizes oligomeric aggregates primarily through intra-chain contacts. These results reveal the differential effects of viral protein segments on amyloid formation pathways and aggregate characteristics, providing new insights into the mechanisms of amyloid aggregation for developing better therapeutic strategies against amyloid-associated diseases, particularly diabetes.

The RBD of SARS-CoV-2 mediates viral entry into host cells by binding to the host receptor ACE2. SARS-CoV-2 infection is linked to various health issues resembling amyloid-related problems, persuading us to investigate the amyloidogenicity of the SARS-CoV-2 spike RBD.

The FoldAmyloid program was used to assess the amyloidogenic propensities in the RBD of Delta Plus and RBD of the Omicron variant, with and without the SUMO tag. After the expression of RBDs, purification, and dialysis steps were performed, subsequently the ThT assay, FTIR, and TEM were employed to check the RBD ability to form fibrils.

The ThT assay, TEM, and FTIR revealed the ability of RBD to self-assemble into β-sheet-rich aggregates (48.4% β-sheet content). Additionally, the presence of the SUMO tag reduced the formation of RBD amyloid-like fibrils. The amyloidogenic potential of Omicron RBD was higher than Delta Plus, according to both in silico and experimental analyses.

The SARS-CoV-2 RBD can assemble itself by forming aggregates containing amyloid-like fibrils and the presence of a SUMO tag can significantly decrease the formation of RBD amyloid-like fibrils. In silico analysis suggested that variation in the ThT fluorescence intensity of amyloid accumulations in the two SARS-CoV-2 strains arises from specific mutations in their RBD regions.

The corona virus (SARS-CoV-2) pandemic and the resulting long-term neurological complications in patients, known as long COVID, have renewed interest in the correlation between viral infections and neurodegenerative brain disorders. While many viruses can reach the central nervous system (CNS) causing acute or chronic infections (such as herpes simplex virus 1, HSV-1), the lack of a clear mechanistic link between viruses and protein aggregation into amyloids, a characteristic of several neurodegenerative diseases, has rendered such a connection elusive. Recently, we showed that viruses can induce aggregation of purified amyloidogenic proteins via the direct physicochemical mechanism of heterogeneous nucleation (HEN). In the current study, we show that the incubation of HSV-1 and SARS-CoV-2 with human cerebrospinal fluid (CSF) leads to the amyloid aggregation of several proteins known to be involved in neurodegenerative diseases, such as APLP1 (amyloid β precursor like protein 1), ApoE, clusterin, α2-macroglobulin, PGK-1 (phosphoglycerate kinase 1), ceruloplasmin, nucleolin, 14-3-3, transthyretin, and vitronectin. Importantly, UV-inactivation of SARS-CoV-2 does not affect its ability to induce amyloid aggregation, as amyloid formation is dependent on viral surface catalysis via HEN and not its ability to replicate. Additionally, viral amyloid induction led to a dramatic drop in the soluble protein concentration in the CSF. Our results show that viruses can physically induce amyloid aggregation of proteins in human CSF and result in soluble protein depletion, thus providing a potential mechanism that may account for the association between persistent and latent/reactivating brain infections and neurodegenerative diseases.

Understanding the pathophysiology of neurological symptoms observed after severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection is essential to optimizing outcomes and therapeutics. To date, small sample sizes and narrow molecular profiling have limited the generalizability of findings. In this study, we profiled multiple cortical and subcortical regions in postmortem brains of patients with coronavirus disease 2019 (COVID-19) and controls with matched pulmonary pathology (total n = 42) using spatial transcriptomics, bulk gene expression and proteomics. We observed a multi-regional antiviral response without direct active SARS-CoV2 infection. We identified dysregulation of mitochondrial and synaptic pathways in deep-layer excitatory neurons and upregulation of neuroinflammation in glia, consistent across both mRNA and protein. Remarkably, these alterations overlapped substantially with changes in age-related neurodegenerative diseases, including Parkinson's disease and Alzheimer's disease. Our work, combining multiple experimental and analytical methods, demonstrates the brain-wide impact of severe acute/subacute COVID-19, involving both cortical and subcortical regions, shedding light on potential therapeutic targets within pathways typically associated with pathological aging and neurodegeneration.

The neurological symptoms of COVID-19, often referred to as neuro-COVID include neurological pain, memory loss, cognitive and sensory disruption. These neurological symptoms can persist for months and are known as Post-Acute Sequalae of COVID-19 (PASC). The molecular origins of neuro-COVID, and how it contributes to PASC are unknown, however a growing body of research highlights that the self-assembly of protein fragments from SARS-CoV-2 into amyloid nanofibrils may play a causative role. Previously, we identified two fragments from the SARS-CoV-2 proteins, Open Reading Frame (ORF) 6 and ORF10, that self-assemble into neurotoxic amyloid assemblies. Here we further our understanding of the self-assembly mechanisms and nano-architectures formed by these fragments and their biological responses. By solubilising the peptides in a fluorinated solvent, we eliminate insoluble aggregates in the starting materials (seeds) that change the polymorphic landscape of the assemblies. The resultant assemblies are dominated by structures with higher free energies (e.g. ribbons and amorphous aggregates) that are less toxic to cultured neurons but do affect their mitochondrial respiration. We also show the first direct evidence of cellular uptake of viral amyloids. This work highlights the importance of understanding the polymorphic behaviour of amyloids and the correlation to neurotoxicity, particularly in the context of neuro-COVID and PASC.

In classical amyloidoses, amyloid fibres form through the nucleation and accretion of protein monomers, with protofibrils and fibrils exhibiting a cross-β motif of parallel or antiparallel β-sheets oriented perpendicular to the fibre direction. These protofibrils and fibrils can intertwine to form mature amyloid fibres. Similar phenomena can occur in blood from individuals with circulating inflammatory molecules (and also some originating from viruses and bacteria). Such pathological clotting can result in an anomalous amyloid form termed fibrinaloid microclots. Previous proteomic analyses of these microclots have shown the presence of non-fibrin(ogen) proteins, suggesting a more complex mechanism than simple entrapment. We thus provide evidence against such a simple entrapment model, noting that clot pores are too large and centrifugation would have removed weakly bound proteins. Instead, we explore whether co-aggregation into amyloid fibres may involve axial (multiple proteins within the same fibril), lateral (single-protein fibrils contributing to a fibre), or both types of integration. Our analysis of proteomic data from fibrinaloid microclots in different diseases shows no significant quantitative overlap with the normal plasma proteome and no correlation between plasma protein abundance and their presence in fibrinaloid microclots. Notably, abundant plasma proteins like α-2-macroglobulin, fibronectin, and transthyretin are absent from microclots, while less abundant proteins such as adiponectin, periostin, and von Willebrand factor are well represented. Using bioinformatic tools, including AmyloGram and AnuPP, we found that proteins entrapped in fibrinaloid microclots exhibit high amyloidogenic tendencies, suggesting their integration as cross-β elements into amyloid structures. This integration likely contributes to the microclots' resistance to proteolysis. Our findings underscore the role of cross-seeding in fibrinaloid microclot formation and highlight the need for further investigation into their structural properties and implications in thrombotic and amyloid diseases. These insights provide a foundation for developing novel diagnostic and therapeutic strategies targeting amyloidogenic cross-seeding in blood clotting disorders.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces immune-mediated diseases. The pathophysiology of COVID-19 uses the following three mechanisms: (1) inflammasome activation mechanism; (2) cGAS-STING signaling mechanism; and (3) SAMHD1 tetramerization mechanism, which leads to IFN-I production. Interactions between the host and virus govern induction, resulting in multiorgan impacts. The NLRP3 with cGAS-STING constitutes the primary immune response. The expression of SARS-CoV-2 ORF3a, NSP6, NSP7, and NSP8 blocks innate immune activation and facilitates virus replication by targeting the RIG-I/MDA5, TRIF, and cGAS-STING signaling. SAMHD1 has a target motif for CDK1 to protect virion assembly, threonine 592 to modulate a catalytically active tetramer, and antiviral IFN responses to block retroviral infection. Plastic and allosteric nucleic acid binding of SAMHD1 modulates the antiretroviral activity of SAMHD1. Therefore, inflammasome activation, cGAS-STING signaling, and SAMHD1 tetramerization explain acute kidney injury, hepatic, cardiac, neurological, and gastrointestinal injury of COVID-19. It might be necessary to effectively block the pathological courses of diverse diseases.

Post-COVID-19 condition (also known as long COVID) is generally defined as symptoms persisting for 3 months or more after acute COVID-19. Long COVID can affect multiple organ systems and lead to severe and protracted impairment of function as a result of organ damage. The burden of this disease, both on the individual and on health systems and national economies, is high. In this interdisciplinary Review, with a coauthor with lived experience of severe long COVID, we sought to bring together multiple streams of literature on the epidemiology, pathophysiology (including the hypothesised mechanisms of organ damage), lived experience and clinical manifestations, and clinical investigation and management of long COVID. Although current approaches to long COVID care are largely symptomatic and supportive, recent advances in clinical phenotyping, deep molecular profiling, and biomarker identification might herald a more mechanism-informed and personally tailored approach to clinical care. We also cover the organisation of services for long COVID, approaches to preventing long COVID, and suggestions for future research.

Long COVID, now unmistakably identified as a syndromic entity encompassing a complex spectrum of symptoms, demands immediate resolution of its elusive pathogenic underpinnings. The intricate interplay of diverse factors presents a complex puzzle, difficult to resolve, and thus poses a substantial challenge. As instances of long COVID manifest by repeated infections of SARS-CoV-2 and genetic predisposition, a detailed understanding in this regard is needed. This endeavor is a comprehensive exploration and analysis of the cascading pathogenetic events driven by viral persistence and replication. Beyond its morbidity, long COVID, more disabling than fatal, exacts one of the most substantial tolls on public health in contemporary times, with the potential to cripple national economies. The paper introduces a unified theory of long COVID, detailing a novel pathophysiological framework that interlinks persistent SARS-CoV-2 infection, autoimmunity, and systemic vascular pathology. We posit a model where viral reservoirs, immune dysregulation, and genetic predispositions converge to perpetuate disease. It challenges prevailing hypotheses with new evidence, suggesting innovative diagnostic and therapeutic approaches. The paper aims to shift the paradigm in long COVID research by providing an integrative perspective that encapsulates the multifaceted nature of the condition. We explain the immunological mechanisms, hypercoagulability states, and viral reservoirs in the skull that feed NeuroCOVID in patients with long COVID. Also, this study hints toward a patient approach and how to prioritize treatment sequences in long COVID patients in hospitals and clinics.

The COVID-19 pandemic, caused by infection with the SARS-CoV-2 virus, is associated with cognitive impairment and Alzheimer's disease (AD) progression. Once it enters the brain, the SARS-CoV-2 virus stimulates accumulation of amyloids in the brain that are highly toxic to neural cells. These amyloids may trigger neurological symptoms in COVID-19. The meningeal lymphatic vessels (MLVs) play an important role in removal of toxins and mediate viral drainage from the brain. MLVs are considered a promising target to prevent COVID-19-exacerbated dementia. However, there are limited methods for augmentation of MLV function. This review highlights new discoveries in the field of COVID-19-mediated amyloid accumulation in the brain associated with the neurological symptoms and the development of promising strategies to stimulate clearance of amyloids from the brain through lymphatic and other pathways. These strategies are based on innovative methods of treating brain dysfunction induced by COVID-19 infection, including the use of photobiomodulation, plasmalogens, and medicinal herbs, which offer hope for addressing the challenges posed by the SARS-CoV-2 virus.

Soon after the outbreak of coronavirus disease 2019 (COVID-19), unexplained sustained fatigue, cognitive disturbance, and muscle ache/weakness were reported in patients who had recovered from acute COVID-19 infection. This abnormal condition has been recognized as "long COVID (postacute sequelae of COVID-19 [PASC])" with a prevalence estimated to be from 10 to 20% of convalescent patients. Although the pathophysiology of PASC has been studied, the exact mechanism remains obscure. Microclots in circulation can represent one of the possible causes of PASC. Although hypercoagulability and thrombosis are critical mechanisms of acute COVID-19, recent studies have reported that thromboinflammation continues in some patients, even after the virus has cleared. Viral spike proteins and RNA can be detected months after patients have recovered, findings that may be responsible for persistent thromboinflammation and the development of microclots. Despite this theory, long-term results of anticoagulation, antiplatelet therapy, and vascular endothelial protection are inconsistent, and could not always show beneficial treatment effects. In summary, PASC reflects a heterogeneous condition, and microclots cannot explain all the presenting symptoms. After clarification of the pathomechanisms of each symptom, a symptom- or biomarker-based stratified approach should be considered for future studies.

The nuclear export protein of the influenza A virus (NEP) is involved in many important processes of the virus life cycle. This makes it an attractive target for the treatment of a disease caused by a virus. Previously it has been shown, that recombinant variants of NEP are highly prone to aggregation in solution under various conditions with the formation of amyloid-like aggregates. In the present work, the amyloid nature of NEP aggregates was evidenced by Congo red binding assays. Atomic force microscopy has shown that NEP can form two types of spherical nanoparticles, which provide an alternative pathway for the formation of amyloid-like fibrils. Type I of these "fibrillogenic" spheres, formed under physiological conditions, represents the micelle-like particles with height 10-60 nm, which can generate worm-like flexible fibrils with the diameter 2.5-4.0 nm, length 20-500 nm and the Young's modulus ~73 MPa. Type II spherical aggregates with size of about 400-1000 nm, formed at elevated temperatures, includes fractions of drop-like and vesicle-like particles, generating more rigid amyloid-like fibrils with height of ~8 nm, and length of up to 2 μm. The hypothetical mechanism of fibril formation via nanospherical structures was suggested. RESEARCH HIGHLIGHTS: AFM has revealed two types of the influenza A virus nuclear export protein spherical aggregates. They provide an alternative pathway for the formation of amyloid-like fibrils. The mechanism of fibril formation via spherical structures is suggested.

Microscopy imaging has enabled us to establish the presence of fibrin(ogen) amyloid (fibrinaloid) microclots in a range of chronic, inflammatory diseases. Microclots may also be induced by a variety of purified substances, often at very low concentrations. These molecules include bacterial inflammagens, serum amyloid A, and the S1 spike protein of severe acute respiratory syndrome coronavirus 2. Here, we explore which of the properties of these microclots might be used to contribute to differential clinical diagnoses and prognoses of the various diseases with which they may be associated. Such properties include distributions in their size and number before and after the addition of exogenous thrombin, their spectral properties, the diameter of the fibers of which they are made, their resistance to proteolysis by various proteases, their cross-seeding ability, and the concentration dependence of their ability to bind small molecules including fluorogenic amyloid stains. Measuring these microclot parameters, together with microscopy imaging itself, along with methodologies like proteomics and imaging flow cytometry, as well as more conventional assays such as those for cytokines, might open up the possibility of a much finer use of these microclot properties in generative methods for a future where personalized medicine will be standard procedures in all clotting pathology disease diagnoses.

COVID-19 infection is associated with a variety of vascular occlusive morbidities. However, a comprehensive understanding of how this virus can induce vascular complications remains lacking. Here, we show that a peptide fragment of SARS-CoV-2 spike protein, S192 (sequence 192-211), is capable of forming amyloid-like aggregates that can induce agglutination of red blood cells, which was not observed with low- and non-aggregated S192 peptide. We subsequently screened eight amyloid-binding molecules and identified BAM1-EG6, a benzothiazole amphiphile, as a promising candidate capable of binding to aggregated S192 and partially inhibiting its agglutination activity. These results provide new insight into a potential molecular mechanism for the capability of spike protein metabolites to contribute to COVID-19-related blood complications and suggest a new therapeutic approach for combating microvascular morbidities in COVID-19 patients.

Apart from mediating viral entry, the function of the free HIV-1 envelope protein (gp120) has yet to be elucidated. Our group previously showed that EP2 derived from one β-strand in gp120 can form amyloid fibrils that increase HIV-1 infectivity. Importantly, gp120 contains ~30 β-strands. We examined whether gp120 might serve as a precursor protein for the proteolytic release of amyloidogenic fragments that form amyloid fibrils, thereby promoting viral infection. Peptide array scanning, enzyme degradation assays, and viral infection experiments in vitro confirmed that many β-stranded peptides derived from gp120 can indeed form amyloid fibrils that increase HIV-1 infectivity. These gp120-derived amyloidogenic peptides, or GAPs, which were confirmed to form amyloid fibrils, were termed gp120-derived enhancers of viral infection (GEVIs). GEVIs specifically capture HIV-1 virions and promote their attachment to target cells, thereby increasing HIV-1 infectivity. Different GAPs can cross-interact to form heterogeneous fibrils that retain the ability to increase HIV-1 infectivity. GEVIs even suppressed the antiviral activity of a panel of antiretroviral agents. Notably, endogenous GAPs and GEVIs were found in the lymphatic fluid, lymph nodes, and cerebrospinal fluid (CSF) of AIDS patients in vivo. Overall, gp120-derived amyloid fibrils might play a crucial role in the process of HIV-1 infectivity and thus represent novel targets for anti-HIV therapeutics.

Understanding the antibody response to SARS-CoV-2, the virus responsible for COVID-19, is crucial to comprehending disease progression and the significance of vaccine and therapeutic development. The emergence of highly contagious variants poses a significant challenge to humoral immunity, underscoring the necessity of grasping the intricacies of specific antibodies. This review emphasizes the pivotal role of antibodies in shaping immune responses and their implications for diagnosing, preventing, and treating SARS-CoV-2 infection. It delves into the kinetics and characteristics of the antibody response to SARS-CoV-2 and explores current antibody-based diagnostics, discussing their strengths, clinical utility, and limitations. Furthermore, we underscore the therapeutic potential of SARS-CoV-2-specific antibodies, discussing various antibody-based therapies such as monoclonal antibodies, polyclonal antibodies, anti-cytokines, convalescent plasma, and hyperimmunoglobulin-based therapies. Moreover, we offer insights into antibody responses to SARS-CoV-2 vaccines, emphasizing the significance of neutralizing antibodies in order to confer immunity to SARS-CoV-2, along with emerging variants of concern (VOCs) and circulating Omicron subvariants. We also highlight challenges in the field, such as the risks of antibody-dependent enhancement (ADE) for SARS-CoV-2 antibodies, and shed light on the challenges associated with the original antigenic sin (OAS) effect and long COVID. Overall, this review intends to provide valuable insights, which are crucial to advancing sensitive diagnostic tools, identifying efficient antibody-based therapeutics, and developing effective vaccines to combat the evolving threat of SARS-CoV-2 variants on a global scale.

The 2019 novel coronavirus disease (COVID-19) is an infectious disease that began to spread globally since 2019. Some COVID-19 patients have neurological complications, such as olfactory disorders and movement disorders, which coincide with the symptoms of Parkinson's disease (PD). Increasing imaging and autopsy evidence supports that the density of dopaminergic neurons in the nigrostriatal pathway is damaged in some COVID-19 patients. However, the underlying mechanism that causes PD-like symptoms remains unclear. PD is an age-related neurodegenerative disease with Lewy bodies (LBs) as its histopathologic feature. The main component of LBs is abnormally aggregated α-synuclein (α-syn). The prion-like propagation of α-syn aggregates plays a key role in the onset and progression of PD. The spike protein (S protein) of SARS-CoV-2 is a heparin-binding protein that mediates the entry of the virus into host cells. Here we found that the S1 domain interacts with α-syn and promotes α-syn aggregation. The S1 domain induces mitochondrial dysfunction, oxidative stress, and cytotoxicity. The S1-seeded α-syn fibrils show enhanced seeding activity and induce synaptic damage and cytotoxicity. Thus, the S1 domain of SARS-CoV-2 promotes the aggregation of α-syn in the cellular model of synucleinopathy and may contribute to the pathogenesis of PD.

During the COVID-19 pandemic, excess deaths including cancer have become a concern in Japan, which has a rapidly aging population. Thus, this study aimed to evaluate how age-adjusted mortality rates (AMRs) for different types of cancer in Japan changed during the COVID-19 pandemic (2020-2022). Official statistics from Japan were used to compare observed annual and monthly AMRs with predicted rates based on pre-pandemic (2010-2019) figures using logistic regression analysis. No significant excess mortality was observed during the first year of the pandemic (2020). However, some excess cancer mortalities were observed in 2021 after mass vaccination with the first and second vaccine doses, and significant excess mortalities were observed for all cancers and some specific types of cancer (including ovarian cancer, leukemia, prostate cancer, lip/oral/pharyngeal cancer, pancreatic cancer, and breast cancer) after mass vaccination with the third dose in 2022. AMRs for the four cancers with the most deaths (lung, colorectal, stomach, and liver) showed a decreasing trend until the first year of the pandemic in 2020, but the rate of decrease slowed in 2021 and 2022. This study discusses possible explanations for these increases in age-adjusted cancer mortality rates.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection initially results in respiratory distress symptoms but can also lead to central nervous system (CNS) and neurological manifestations, significantly impacting coronavirus disease 2019 (COVID-19) patients with neurodegenerative diseases. Additionally, strict lockdown measures introduced to curtail the spread of COVID-19 have raised concerns over the wellbeing of patients with dementia and/or Alzheimer's disease. The aim of this review was to discuss the overlapping molecular pathologies and the potential bidirectional relationship between COVID-19 and Alzheimer's dementia, as well as the impact of lockdown/restriction measures on the neuropsychiatric symptoms (NPS) of patients with Alzheimer's dementia. Furthermore, we aimed to assess the impact of lockdown measures on the NPS of caregivers, exploring its potential effects on the quality and extent of care they provide to dementia patients.We utilized the PubMed and Google Scholar databases to search for articles on COVID-19, dementia, Alzheimer's disease, lockdown, and caregivers. Our review highlights that patients with Alzheimer's disease face an increased risk of COVID-19 infection and complications. Additionally, these patients are likely to experience greater cognitive decline. It appears that these issues are primarily caused by the SARS-CoV-2 infection and appear to be further exacerbated by restrictive/lockdown measures. Moreover, lockdown measures introduced during the pandemic have negatively impacted both the NPSs of caregivers and their perception of the wellbeing of their Alzheimer's patients. Thus, additional safeguard measures, along with pharmacological and non-pharmacological approaches, are needed to protect the wellbeing of dementia patients and their caregivers in light of this and possible future pandemics.

Parkinson's disease (PD) is a neurodegenerative disease characterized by the degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and the formation of Lewy bodies (LBs). The main proteinaceous component of LBs is aggregated α-synuclein (α-syn). However, the mechanisms underlying α-syn aggregation are not yet fully understood. Converging lines of evidence indicate that, under certain pathological conditions, various proteins can interact with α-syn and regulate its aggregation. Understanding these protein-protein interactions is crucial for unraveling the molecular mechanisms contributing to PD pathogenesis. In this review we provide an overview of the current knowledge on protein-protein interactions that regulate α-syn aggregation. Additionally, we briefly summarize the methods used to investigate the influence of protein-protein interactions on α-syn aggregation and propagation.

It is unclear how severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection leads to the strong but ineffective inflammatory response that characterizes severe Coronavirus disease 2019 (COVID-19), with amplified immune activation in diverse cell types, including cells without angiotensin-converting enzyme 2 receptors necessary for infection. Proteolytic degradation of SARS-CoV-2 virions is a milestone in host viral clearance, but the impact of remnant viral peptide fragments from high viral loads is not known. Here, we examine the inflammatory capacity of fragmented viral components from the perspective of supramolecular self-organization in the infected host environment. Interestingly, a machine learning analysis to SARS-CoV-2 proteome reveals sequence motifs that mimic host antimicrobial peptides (xenoAMPs), especially highly cationic human cathelicidin LL-37 capable of augmenting inflammation. Such xenoAMPs are strongly enriched in SARS-CoV-2 relative to low-pathogenicity coronaviruses. Moreover, xenoAMPs from SARS-CoV-2 but not low-pathogenicity homologs assemble double-stranded RNA (dsRNA) into nanocrystalline complexes with lattice constants commensurate with the steric size of Toll-like receptor (TLR)-3 and therefore capable of multivalent binding. Such complexes amplify cytokine secretion in diverse uninfected cell types in culture (epithelial cells, endothelial cells, keratinocytes, monocytes, and macrophages), similar to cathelicidin's role in rheumatoid arthritis and lupus. The induced transcriptome matches well with the global gene expression pattern in COVID-19, despite using <0.3% of the viral proteome. Delivery of these complexes to uninfected mice boosts plasma interleukin-6 and CXCL1 levels as observed in COVID-19 patients.

Virus-like particles (VLPs) are nanometric structures composed of structural components of virions, keeping most of the cellular recognition and internalization properties, but are non-infective as they are deprived of their genetic material. VLPs have been a versatile platform for developing vaccines by carrying their own or heterologous antigenic epitopes. Moreover, VLPs can also be used as nanovessels for encapsulating molecules with therapeutic applications, like enzymes, nucleic acids, and drugs. Parvovirus B19 (B19V) VLPs can be self-assembled in vitro from the denatured major viral particle protein VP2 by equilibrium dialysis. Despite its fair productivity, this process is currently a time-consuming task. Affinity chromatography is used as an efficient step for concentration and purification, but it is only sometimes seen as a method that facilitates the oligomerization of proteins. In this research, we report a novel approach for the in vitro assembly of B19V VLPs through the immobilization of the denatured VP2 into an immobilized metal affinity chromatography (IMAC) column, followed by the on-column folding and the final VLP assembly upon protein elution. This method is suitable for the fast production of B19V VLPs. KEY POINTS: • Biotechnological applications for inclusion bodies • Efficient single-step purification and immobilization strategies • Rapid VLP assembly strategy.

Since the SARS-CoV-2 virus started spreading worldwide, evidence pointed toward an impact of the infection on the nervous system. COVID-19 patients present neurological manifestations and have an increased risk of developing brain-related symptoms in the long term. In fact, evidence in support of the neuroinvasive potential of SARS-CoV-2 has emerged. Considering that viral parkisonism was observed as a consequence of encephalopathies caused by viral infections, it has been already suggested that COVID-19 could affect the dopaminergic neurons and contribute to neurodegeneration in Parkinson's disease (PD), by promoting the formation of amyloid fibrils constituted by the PD-related protein α-synuclein. Here, we observe not only that SARS-CoV-2 viral spike protein and nucleocapsid protein can alone promote α-synuclein aggregation but also that the spike protein organization in a corona shape on the viral envelope may be crucial in triggering fast amyloid fibrils formation, thus possibly contributing to PD pathogenesis.

Our understanding of COVID-19 vaccinations and their impact on health and mortality has evolved substantially since the first vaccine rollouts. Published reports from the original randomized phase 3 trials concluded that the COVID-19 mRNA vaccines could greatly reduce COVID-19 symptoms. In the interim, problems with the methods, execution, and reporting of these pivotal trials have emerged. Re-analysis of the Pfizer trial data identified statistically significant increases in serious adverse events (SAEs) in the vaccine group. Numerous SAEs were identified following the Emergency Use Authorization (EUA), including death, cancer, cardiac events, and various autoimmune, hematological, reproductive, and neurological disorders. Furthermore, these products never underwent adequate safety and toxicological testing in accordance with previously established scientific standards. Among the other major topics addressed in this narrative review are the published analyses of serious harms to humans, quality control issues and process-related impurities, mechanisms underlying adverse events (AEs), the immunologic basis for vaccine inefficacy, and concerning mortality trends based on the registrational trial data. The risk-benefit imbalance substantiated by the evidence to date contraindicates further booster injections and suggests that, at a minimum, the mRNA injections should be removed from the childhood immunization program until proper safety and toxicological studies are conducted. Federal agency approval of the COVID-19 mRNA vaccines on a blanket-coverage population-wide basis had no support from an honest assessment of all relevant registrational data and commensurate consideration of risks versus benefits. Given the extensive, well-documented SAEs and unacceptably high harm-to-reward ratio, we urge governments to endorse a global moratorium on the modified mRNA products until all relevant questions pertaining to causality, residual DNA, and aberrant protein production are answered.

The effects of SARS-CoV-2 in COVID-19 on the nervous system are incompletely understood. SARS-CoV-2 can infect endothelial cells, neurons, astrocytes, and oligodendrocytes with consequences for the host. There are indications that infection of these CNS-resident cells may result in long-term effects, including emergence of neurodegenerative diseases. Indirect effects of infection with SARS-CoV-2 relate to the induction of autoimmune disease involving molecular mimicry or/and bystander activation of T- and B cells and emergence of autoantibodies against various self-antigens. Data obtained in preclinical models of coronavirus-induced disease gives important clues for the understanding of nervous system-related assault of SARS-CoV-2. The pathophysiology of long-COVID syndrome and post-COVID syndrome in which autoimmunity and immune dysregulation might be the driving forces are still incompletely understood. A better understanding of nervous-system-related immunity in COVID-19 might support the development of therapeutic approaches. In this review, the current understanding of SARS-CoV-2 tropism for the nervous system, the associated immune responses, and diseases are summarized. The data indicates that there is viral tropism of SARS-CoV-2 in the nervous system resulting in various disease conditions. Prevention of SARS-CoV-2 infection by means of vaccination is currently the best strategy for the prevention of subsequent tissue damage involving the nervous system.

Increasing evidence suggests a potential role for infectious pathogens in the etiology of synucleinopathies, a group of age-related neurodegenerative disorders including Parkinson's disease (PD), multiple system atrophy and dementia with Lewy bodies. In this review, we discuss the link between infections and synucleinopathies from a historical perspective, present emerging evidence that supports this link, and address current research challenges with a focus on neuroinflammation. Infectious pathogens can elicit a neuroinflammatory response and modulate genetic risk in PD and related synucleinopathies. The mechanisms of how infections might be linked with synucleinopathies as well as the overlap between the immune cellular pathways affected by virulent pathogens and disease-related genetic risk factors are discussed. Here, an important role for α-synuclein in the immune response against infections is emerging. Critical methodological and knowledge gaps are addressed, and we provide new future perspectives on how to address these gaps. Understanding how infections and neuroinflammation influence synucleinopathies will be essential for the development of early diagnostic tools and novel therapies.

The long-term impact of the COVID-19 pandemic concerns risk to human health, particularly its potential association with protein misfolding and amyloidosis. This review article explores the causality relationship between SARS-CoV-2 infection, and protein misfolding, leading to amyloid-related conditions. It delves into the mechanisms by which viral proteins may accelerate amyloid formation, exacerbating post-infection complications, including neurological sequelae. Drawing from interdisciplinary research and clinical observations, the potential links between COVID-19, vaccination, and amyloidosis, emphasize the importance of understanding the longterm effect of post-COVID symptoms. This review examines the potential role of COVID-19-related proteins in the formation of amyloid in other related proteins of amyloidosis.

Despite advances in machine learning-based protein structure prediction, we are still far from fully understanding how proteins fold into their native conformation. The conventional notion that polypeptides fold spontaneously to their biologically active states has gradually been replaced by our understanding that cellular protein folding often requires context-dependent guidance from molecular chaperones in order to avoid misfolding. Misfolded proteins can aggregate into larger structures, such as amyloid fibrils, which perpetuate the misfolding process, creating a self-reinforcing cascade. A surge in amyloid fibril structures has deepened our comprehension of how a single polypeptide sequence can exhibit multiple amyloid conformations, known as polymorphism. The assembly of these polymorphs is not a random process but is influenced by the specific conditions and tissues in which they originate. This observation suggests that, similar to the folding of native proteins, the kinetics of pathological amyloid assembly are modulated by interactions specific to cells and tissues. Here, we review the current understanding of how intrinsic protein conformational propensities are modulated by physiological and pathological interactions in the cell to shape protein misfolding and aggregation pathology.