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Cotton wool plaques (CWPs) have been described as features of the neuropathologic phenotype of dominantly inherited Alzheimer disease (DIAD) caused by some missense and deletion mutations in the presenilin 1 (PSEN1) gene. CWPs are round, eosinophilic amyloid-β (Aβ) plaques that lack an amyloid core and are recognizable, but not fluorescent, in Thioflavin S (ThS) preparations. Amino-terminally truncated and post-translationally modified Aβ peptide species are the main component of CWPs. Tau immunopositive neurites may be present in CWPs. In addition, neurofibrillary tangles coexist with CWPs. Herein, we report the structure of Aβ and tau filaments isolated from brain tissue of individuals affected by DIAD caused by the PSEN1 V261I and A431E mutations, with the CWP neuropathologic phenotype. CWPs are predominantly composed of type I Aβ filaments present in two novel arrangements, type Ic and type Id; additionally, CWPs contain type I and type Ib Aβ filaments. Tau filaments have the AD fold, which has been previously reported in sporadic AD and DIAD. The formation of type Ic and type Id Aβ filaments may be the basis for the phenotype of CWPs. Our data are relevant for the development of PET imaging methodologies to best detect CWPs in DIAD., (© 2024. The Author(s).)

Adult individuals with Down syndrome (DS) develop Alzheimer disease (AD). Whether there is a difference between AD in DS and AD regarding the structure of amyloid-β (Aβ) and tau filaments is unknown. Here we report the structure of Aβ and tau filaments from two DS brains. We found two Aβ 40 filaments (types IIIa and IIIb) that differ from those previously reported in sporadic AD and two types of Aβ 42 filaments (I and II) identical to those found in sporadic and familial AD. Tau filaments (paired helical filaments and straight filaments) were identical to those in AD, supporting the notion of a common mechanism through which amyloids trigger aggregation of tau. This knowledge is important for understanding AD in DS and assessing whether adults with DS could be included in AD clinical trials., (© 2024. The Author(s).)

Background: The Microtubule-Associated Protein Tau (MAPT) is one of the proteins that are central to neurodegenerative diseases. The nature of intracellular tau aggregates is determined by the cell types whether neuronal or glial, the participating tau isoforms, and the structure of the amyloid filament. The transmembrane protein 106B (TMEM106B) has recently emerged as another significant player in neurodegeneration and aging. In the central nervous system, the composition of the gray and white matter differs considerably. The gray matter consists of nerve cell bodies, dendrites, unmyelinated axons, synaptic terminals, astrocytes, oligodendrocytes (satellite cells) and microglia. The white matter differs from the gray for the presence of axonal tracts as the only neuronal component and for the absence of nerve cell bodies, dendrites and synaptic terminals. Cryogenic electron microscopy (cryo-EM) studies have unveiled the structure of tau and TMEM106B, from the cerebral cortex, in several neurodegenerative diseases; however, whether tau and TMEM106B filaments from the gray and white matter share a common fold requires additional investigation., Methods: We isolated tau and TMEM106B from the cerebral cortex and white matter of the frontal lobes of two individuals affected by multiple system tauopathy with presenile dementia (MSTD), a disease caused by the MAPT intron 10 mutation +3. We used immunostaining, biochemical, genetics and cryo-EM methods to characterize tau and TMEM106B., Results: We determined that tau filaments in the gray and the white matter of MSTD individuals can induce tau aggregation and have identical AGD type 2 folds. TMEM106B amyloid filaments were also found in the gray and white matter of MSTD; the filament folds were identical in the two anatomical regions., Conclusions: Our findings show for the first time that in MSTD two types of amyloid filaments extracted from the gray matter have identical folds to those extracted from the white matter. Whether in this genetic disorder there is a relationship in the pathogenesis of the tau and TMEM106B filaments, remains to be determined. Furthermore, additional studies are needed for other proteins and other neurodegenerative diseases to establish whether filaments extracted from the gray and white matter would have identical folds.

Prion protein (PrP) aggregation and formation of PrP amyloid (APrP) are central events in the pathogenesis of prion diseases. In the dominantly inherited prion protein amyloidosis known as Gerstmann-Sträussler-Scheinker (GSS) disease, plaques made of PrP amyloid are present throughout the brain. The c.593t > c mutation in the prion protein gene (PRNP) results in a phenylalanine to serine amino acid substitution at PrP residue 198 (F198S) and causes the most severe amyloidosis among GSS variants. It has been shown that neurodegeneration in this disease is associated with the presence of extracellular APrP plaques and neuronal intracytoplasmic Tau inclusions, that have been shown to contain paired helical filaments identical to those found in Alzheimer disease. Using cryogenic electron microscopy (cryo-EM), we determined for the first time the structures of filaments of human APrP, isolated post-mortem from the brain of two symptomatic PRNP F198S mutation carriers. We report that in GSS (F198S) APrP filaments are composed of dimeric, trimeric and tetrameric left-handed protofilaments with their protomers sharing a common protein fold. The protomers in the cross-β spines consist of 62 amino acids and span from glycine 80 to phenylalanine 141, adopting a previously unseen spiral fold with a thicker outer layer and a thinner inner layer. Each protomer comprises nine short β-strands, with the β1 and β8 strands, as well as the β4 and β9 strands, forming a steric zipper. The data obtained by cryo-EM provide insights into the structural complexity of the PrP filament in a dominantly inherited human PrP amyloidosis. The novel findings highlight the urgency of extending our knowledge of the filaments' structures that may underlie distinct clinical and pathologic phenotypes of human neurodegenerative diseases., (© 2022. The Author(s).)

Mucosal-associated invariant T (MAIT) cells are unconventional T lymphocytes with a semi-conserved TCRα, activated by the presentation of vitamin B metabolites by the MHC-I related protein, MR1, and with diverse innate and adaptive effector functions. The role of MAIT cells in acute intestinal infections, especially at the mucosal level, is not well known. Here, we analyzed the presence and phenotype of MAIT cells in duodenal biopsies and paired peripheral blood samples, in patients during and after culture-confirmed Vibrio cholerae O1 infection. Immunohistochemical staining of duodenal biopsies from cholera patients (n = 5, median age 32 years, range 26-44, 1 female) identified MAIT cells in the lamina propria of the crypts, but not the villi. By flow cytometry (n = 10, median age 31 years, range 23-36, 1 female), we showed that duodenal MAIT cells are more activated than peripheral MAIT cells (p < 0.01 across time points), although there were no significant differences between duodenal MAIT cells at day 2 and day 30. We found fecal markers of intestinal permeability and inflammation to be correlated with the loss of duodenal (but not peripheral) MAIT cells, and single-cell sequencing revealed differing T cell receptor usage between the duodenal and peripheral blood MAIT cells. In this preliminary report limited by a small sample size, we show that MAIT cells are present in the lamina propria of the duodenum during V. cholerae infection, and more activated than those in the blood. Future work into the trafficking and tissue-resident function of MAIT cells is warranted., Competing Interests: The authors have declared that no competing interests exist.

Even with implementation of current influenza vaccines, influenza still claims up to 500,000 lives worldwide annually, indicating a need for a better vaccine strategy. We have developed a technology to generate unique S 60 -HA1 pseudovirus nanoparticles (PVNPs) that display the receptor-binding HA1 domains of influenza viruses. Each self-assembled S 60 -HA1 PVNP consists of a T = 1 icosahedral S 60 nanoparticle that resembles the inner shell of norovirus capsid and 60 surface-displayed HA1 antigens that are excellent vaccine targets. Soluble S 60 -HA1 PVNPs presenting HA1 antigens of H7N9 influenza virus subtypes have been produced efficiently in large amount. Their three-dimensional (3D) structures have been solved by cryogenic electron microscopy. The PVNP-displayed HA1 antigens react with HA-specific antibody, and retain authentic sialic acid binding specificity and hemagglutinate human erythrocytes. The PVNPs are highly immunogenic, eliciting high titers of HA1-specific antibodies in mice and the mouse sera strongly inhibited hemagglutinations of homologous and heterologous influenza virus HA proteins. Therefore, the S 60 -HA1 PVNPs may provide useful reagents to study influenza viruses and offer a potential new vaccine tactic to fight the deadly influenza disease., (© Tsinghua University Press and Springer-Verlag GmbH Germany, part of Springer Nature 2021.)

In human neurodegenerative diseases associated with the intracellular aggregation of Tau protein, the ordered cores of Tau filaments adopt distinct folds. Here, we analyze Tau filaments isolated from the brain of individuals affected by Prion-Protein cerebral amyloid angiopathy (PrP-CAA) with a nonsense mutation in the PRNP gene that leads to early termination of translation of PrP (Q160Ter or Q160X), and Gerstmann-Sträussler-Scheinker (GSS) disease, with a missense mutation in the PRNP gene that leads to an amino acid substitution at residue 198 (F198S) of PrP. The clinical and neuropathologic phenotypes associated with these two mutations in PRNP are different; however, the neuropathologic analyses of these two genetic variants have consistently shown the presence of numerous neurofibrillary tangles (NFTs) made of filamentous Tau aggregates in neurons. We report that Tau filaments in PrP-CAA (Q160X) and GSS (F198S) are composed of 3-repeat and 4-repeat Tau isoforms, having a striking similarity to NFTs in Alzheimer disease (AD). In PrP-CAA (Q160X), Tau filaments are made of both paired helical filaments (PHFs) and straight filaments (SFs), while in GSS (F198S), only PHFs were found. Mass spectrometry analyses of Tau filaments extracted from PrP-CAA (Q160X) and GSS (F198S) brains show the presence of post-translational modifications that are comparable to those seen in Tau aggregates from AD. Cryo-EM analysis reveals that the atomic models of the Tau filaments obtained from PrP-CAA (Q160X) and GSS (F198S) are identical to those of the Tau filaments from AD, and are therefore distinct from those of Pick disease, chronic traumatic encephalopathy, and corticobasal degeneration. Our data support the hypothesis that in the presence of extracellular amyloid deposits and regardless of the primary amino acid sequence of the amyloid protein, similar molecular mechanisms are at play in the formation of identical Tau filaments., (© 2021. The Author(s).)

Recent progress in the development of affinity grids for cryoelectron microscopy (cryo-EM) typically employs genetic engineering of the protein sample such as histidine or Spy tagging, immobilized antibody capture, or nonselective immobilization via electrostatic interactions or Schiff base formation. We report a powerful and flexible method for the affinity capture of target proteins for cryo-EM analysis that utilizes small-molecule ligands as bait for concentrating human target proteins directly onto the grid surface for single-particle reconstruction. This approach is demonstrated for human p97, captured using two different small-molecule high-affinity ligands of this AAA+ ATPase. Four electron density maps are revealed, each representing a p97 conformational state captured from solution, including a double-hexamer structure resolved to 3.6 Å. These results demonstrate that the noncovalent capture of protein targets on EM grids modified with high-affinity ligands can enable the structure elucidation of multiple configurational states of the target and potentially inform structure-based drug design campaigns.

The title hemihydrate, C 12 H 9 N 5 ·0.5H 2 O, was isolated from the condensation reaction of quinoline-2-carbaldehyde with 4-amino-4 H -1,2,4-triazole. The Schiff base mol-ecule adopts an E configuration about the C=N bond and is approximately planar, with a dihedral angle between the quinoline ring system and the 1,2,4-triazole ring of 12.2 (1)°. In the crystal, one water mol-ecule bridges two Schiff base mol-ecules via O-H⋯N hydrogen bonds. The Schiff base mol-ecules are inter-connected by π-π stacking inter-actions [centroid-centroid distances of 3.7486 (7) and 3.9003 (7) Å] into columns along [10]., (© M. Shahri et al. 2020.)

Vibrio cholerae O1 can cause life threatening diarrheal disease if left untreated. T cells can play critical roles in inducing B cell mediated immunity. As the mechanism of T cell dependent B cell maturation is not well established, we hypothesized that a specific population of T (follicular helper T, Tfh) cells, are involved in B cell maturation following cholera. We found flowcytometrically that V. cholerae infection induces significant increases in circulating Tfh cells expressing B cell maturation associated protein CD40L early in disease. The increased Tfh cells expressing CD40L recognize cholera toxin most prominently, with lessened responses to V. cholerae membrane preparation (MP) and V. cholerae cytolysin (VCC). We further showed that early induction of Tfh cells and CD40L was associated with later memory B cell responses to same antigens. Lastly, we demonstrated in vitro that Tfh cells isolated after cholera can stimulate class switching of co-cultured, isolated B cells from patients with cholera, leading to production of the more durable IgG antibody isotype colorimetrically. These studies were conducted on circulating Tfh cells; future studies will be directed at examining role of Tfh cells during cholera directly in gut mucosa of biopsied samples, at the single cell level if feasible., (Copyright © 2018 The Author(s). Published by Elsevier Masson SAS.. All rights reserved.)

Antibody secreting cells (ASCs) generate antibodies in an antigen-specific manner as part of the adaptive immune response to infections, and these cells increase their surface expression of HLA-DR. We have studied this parameter (HLA-DR+ ASC) in patients with recent diarrheal infection using immuno-magnetic cell sorting and an enzyme linked immunospot (ELISPOT) technique that requires only one milliliter of blood. We validated this approach in adult patients with cholera (n = 15) or ETEC diarrhea (n = 30) on days 2, 7 and 30 after showing clinical symptom at the International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b) hospital in Dhaka, and we compared responses to age-matched healthy controls (n = 7). We found that HLA-DR+ ASC (DR+ASC) responses specific both for T cell-dependent (cholera toxin B subunit), and T cell-independent (lipopolysaccharide) antigens were elevated at day 7 after showing clinical cholera symptom. Similarly, DR+ASCs were elevated against both heat-labile toxin and colonization factors following ETEC infection. We observed significant correlations between antigen-specific DR+ASC responses and antigen-specific, gut homing ASC and plasma antibody responses. This study demonstrates that a simple ELISPOT procedure allows determination of antigen-specific ASC responses using a small volume of whole blood following diarrhea. This technique may be particularly useful in studying DR+ASC responses in young children and infants, either following infection or vaccination., (Copyright © 2017 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

We characterized the acute B cell response in adults with cholera by analyzing the repertoire, specificity, and functional characteristics of 138 monoclonal antibodies (MAbs) generated from single-cell-sorted plasmablasts. We found that the cholera-induced responses were characterized by high levels of somatic hypermutation and large clonal expansions. A majority of the expansions targeted cholera toxin (CT) or lipopolysaccharide (LPS). Using a novel proteomics approach, we were able to identify sialidase as another major antigen targeted by the antibody response to Vibrio cholerae infection. Antitoxin MAbs targeted both the A and B subunits, and most were also potent neutralizers of enterotoxigenic Escherichia coli heat-labile toxin. LPS-specific MAbs uniformly targeted the O-specific polysaccharide, with no detectable responses to either the core or the lipid moiety of LPS. Interestingly, the LPS-specific antibodies varied widely in serotype specificity and functional characteristics. One participant infected with the Ogawa serotype produced highly mutated LPS-specific antibodies that preferentially bound the previously circulating Inaba serotype. This demonstrates durable memory against a polysaccharide antigen presented at the mucosal surface and provides a mechanism for the long-term, partial heterotypic immunity seen following cholera., Importance: Cholera is a diarrheal disease that results in significant mortality. While oral cholera vaccines are beneficial, they do not achieve equivalent protection compared to infection with Vibrio cholerae Although antibodies likely mediate protection, the mechanisms of immunity following cholera are poorly understood, and a detailed understanding of antibody responses to cholera is of significance for human health. In this study, we characterized the human response to cholera at the single-plasmablast, monoclonal antibody level. Although this approach has not been widely applied to the study of human bacterial infection, we were able to uncover the basis of cross-reactivity between different V. cholerae serotypes and the likely impact of prior enterotoxigenic Escherichia coli exposure on the response to cholera, as well as identify novel antigenic targets. In addition to improving our understanding of the repertoire and function of the antibody response to cholera in humans, this study has implications for future cholera vaccination efforts., (Copyright © 2016 Kauffman et al.)

Environmental enteropathy (EE) is a poorly understood condition that refers to chronic alterations in intestinal permeability, absorption, and inflammation, which mainly affects young children in resource-limited settings. Recently, EE has been linked to suboptimal oral vaccine responses in children, although immunological mechanisms are poorly defined. The objective of this study was to determine host factors associated with immune responses to an oral cholera vaccine (OCV). We measured antibody and memory T cell immune responses to cholera antigens, micronutrient markers in blood, and EE markers in blood and stool from 40 Bangladeshi children aged 3-14 years who received two doses of OCV given 14 days apart. EE markers included stool myeloperoxidase (MPO) and alpha anti-trypsin (AAT), and plasma endotoxin core antibody (EndoCab), intestinal fatty acid binding protein (i-FABP), and soluble CD14 (sCD14). We used multiple linear regression analysis with LASSO regularization to identify host factors, including EE markers, micronutrient (nutritional) status, age, and HAZ score, predictive for each response of interest. We found stool MPO to be positively associated with IgG antibody responses to the B subunit of cholera toxin (P = 0.03) and IgA responses to LPS (P = 0.02); plasma sCD14 to be positively associated with LPS IgG responses (P = 0.07); plasma i-FABP to be positively associated with LPS IgG responses (P = 0.01) and with memory T cell responses specific to cholera toxin (P = 0.01); stool AAT to be negatively associated with IL-10 (regulatory) T cell responses specific to cholera toxin (P = 0.02), and plasma EndoCab to be negatively associated with cholera toxin-specific memory T cell responses (P = 0.02). In summary, in a cohort of children 3-14 years old, we demonstrated that the majority of biomarkers of environmental enteropathy were positively associated with immune responses after vaccination with an OCV., Competing Interests: The authors have declared that no competing interests exist.

Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC) are noninvasive mucosal pathogens that cause acute watery diarrhea in people in developing countries. Direct assessment of the mucosal immune responses to these pathogens is problematic. Surrogate markers of local mucosal responses in blood are increasingly being studied to determine the mucosal immune responses after infection. However, the volume of blood available in children and infants has limited this approach. We assessed whether an approach that first isolates β7-positive cells from a small volume of blood would allow measurement of the antigen-specific immune responses in patients with cholera and ETEC infection. β7 is a cell surface marker associated with mucosal homing. We isolated β7-expressing cells from blood on days 2, 7, and 30 and used an enzyme-linked immunosorbent spot (ELISPOT) assay to assess the gut-homing antibody-secreting cells (ASCs) specific to pathogen antigens. Patients with ETEC diarrhea showed a significant increase in toxin-specific gut-homing ASCs at day 7 compared to the levels at days 2 and 30 after onset of illness and to the levels in healthy controls. Similar elevations of responses to the ETEC colonization factors (CFs) CS6 and CFA/I were observed in patients infected with CS6- and CFA/I-positive ETEC strains. Antigen-specific gut-homing ASCs to the B subunit of cholera toxin and cholera-specific lipopolysaccharides (LPS) were also observed on day 7 after the onset of cholera using this approach. This study demonstrates that a simple ELISPOT assay can be used to study the mucosal immunity to specific antigens using a cell-sorting protocol to isolate mucosal homing cells, facilitating measurement of mucosal responses in children following infection or vaccination., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Background: Mucosal Associated Invariant T (MAIT) cells are innate-like T cells found in abundance in the intestinal mucosa, and are thought to play a role in bridging the innate-adaptive interface., Methods: We measured MAIT cell frequencies and antibody responses in blood from patients presenting with culture-confirmed severe cholera to a hospital in Dhaka, Bangladesh at days 2, 7, 30, and 90 of illness., Results: We found that MAIT (CD3+CD4-CD161hiVα7.2+) cells were maximally activated at day 7 after onset of cholera. In adult patients, MAIT frequencies did not change over time, whereas in child patients, MAITs were significantly decreased at day 7, and this decrease persisted to day 90. Fold changes in MAIT frequency correlated with increases in LPS IgA and IgG, but not LPS IgM nor antibody responses to cholera toxin B subunit., Conclusions: In the acute phase of cholera, MAIT cells are activated, depleted from the periphery, and as part of the innate response against V. cholerae infection, are possibly involved in mechanisms underlying class switching of antibody responses to T cell-independent antigens.

This cross sectional study was conducted in Sir Salimullah Medical College Hospital, Mitford and Dhaka Medical College Hospital, Dhaka, Bangladesh from January 2009 to December 2009 by purposive sampling method. Among the 50 cases, involvement of supraglottic (72%) region is significantly higher than glottic (28%) region. Affected peoples are mainly male and mean age was 58.1±10.4 years. Frequency of regional metastasis of supraglottic carcinoma (47.22%) was higher than glottic carcinoma (7.14%). Most of the lymphnode involvement was at level-II to IV. Most of the patients presented at stage-I & stage-II.