Functional and Structural Biological Methods for Palytoxin Detection

Palytoxin (PLTX) and its analogues are marine polyethers identified in Palythoa and Zoanthus corals, Ostreopsis dinoflagellates, and Trichodesmium cyanobacteria. Humans can be exposed to these toxins by different routes with a series of adverse effects but the most severe risk is associated with poisonings by the consumption of edible marine organisms accumulating these toxins, as occurs in (sub)-tropical areas. In temperate areas, adverse effects ascribed to PLTXs have been recorded after inhalation of marine aerosols and/or cutaneous contact with seawater during Ostreopsis blooms, as well as during cleaning procedures of Palythoa-containing home aquaria. Besides instrumental analytical methods, in the last years a series of alternative or complementary methods based on biological/biochemical tools have been developed for the rapid and specific PLTX detection required for risk assessment. These methods are usually sensitive, cost- and time-effective, and do not require highly specialized operators. Among them, structural immunoassays and functional cellbased assays are reviewed. The availability of specific anti-PLTX antibodies allowed the development of different sensitive structural assays, suitable for its detection also in complex matrices, such as mussels. In addition, knowing the mechanism of PLTX action, a series of functional identification methods has been developed. Despite some of them being limited by matrix effects and specificity issues, biological methods for PLTX detection represent a feasible tool, suitable for rapid screening

Narcolepsy risk loci outline role of T cell autoimmunity and infectious triggers in narcolepsy

Narcolepsy has genetic and environmental risk factors, but the specific genetic risk loci and interaction with environmental triggers are not well understood. Here, the authors identify genetic loci for narcolepsy, suggesting infection as a trigger and dendritic and helper T cell involvement. Narcolepsy type 1 (NT1) is caused by a loss of hypocretin/orexin transmission. Risk factors include pandemic 2009 H1N1 influenza A infection and immunization with Pandemrix (R). Here, we dissect disease mechanisms and interactions with environmental triggers in a multi-ethnic sample of 6,073 cases and 84,856 controls. We fine-mapped GWAS signals within HLA (DQ0602, DQB1*03:01 and DPB1*04:02) and discovered seven novel associations (CD207, NAB1, IKZF4-ERBB3, CTSC, DENND1B, SIRPG, PRF1). Significant signals at TRA and DQB1*06:02 loci were found in 245 vaccination-related cases, who also shared polygenic risk. T cell receptor associations in NT1 modulated TRAJ*24, TRAJ*28 and TRBV*4-2 chain-usage. Partitioned heritability and immune cell enrichment analyses found genetic signals to be driven by dendritic and helper T cells. Lastly comorbidity analysis using data from FinnGen, suggests shared effects between NT1 and other autoimmune diseases. NT1 genetic variants shape autoimmunity and response to environmental triggers, including influenza A infection and immunization with Pandemrix (R)

Self-assembly, nematic phase formation and organocatalytic behaviour of a proline-functionalized lipopeptide

The self-assembly of the amphiphilic lipopeptide PAEPKI-C16 (P = proline, A = alanine, E = glutamic acid, K = lysine, I = isoleucine, C16 = hexadecyl) was investigated using a combination of spectroscopic, microscopic and scattering methods and compared to C16-IKPEAP with the same (reversed) peptide sequence and the alkyl chain positioned N-terminally and which lacks a free N-terminal proline residue. The catalytic activity of these peptides were then compared using a model aldol reaction system. For PAEPKI-C16, Cryo-TEM images showed the formation of micrometer length fibers, which by Small-angle X-ray scattering (SAXS) were found to have a radius of 2.5 - 2.6 nm. Spectroscopic analysis shows these fibers are built from -sheets. This behaviour is in complete contrast to that of C16-IKPEAP which forms spherical micelles with peptides in a disordered conformation [Hutchinson, J. A. et al. J. Phys. Chem. B 2019, 123, 613]. For PAEPKI-C16, the spontaneous alignment of fibers was observed upon increasing pH, which was accompanied by observed birefringence and anisotropy of SAXS patterns. This shows the formation of a nematic liquids and unprecedented nematic hydrogel formation was also observed these lipopeptides at sufficiently high concentrations. SAXS shows retention of an ultrafine (1.7 nm core radius) fibrillar network within the hydrogel. PAEPKI-C16 with free N-terminal proline shows enhanced anti:syn diastereoselectivity and better conversion compared to C16-IKPEAP. The cytotoxicity of PAEPKI-C16 was also lower than C16-IKPEAP for both fibroblast and cancer cell lines. These results highlight the sensitivity of lipopeptide properties to the presence of a free proline residue. The spontaneous nematic phase formation by PAEPKI-C16 points to the highly anisotropy of its ultrafine fibrillar structure and the formation of such a phase at low concentration in aqueous solution may be valuable for future applications

Neoangiogenesis and Blood-brain Barrier Dysfunction in Human TSC Brain Lesions

Introduction: Tuberous sclerosis complex (TSC) is a genetic disorder characterized by the presence of multiple benign tumors throughout the body and brain. Patients with TSC experience severe cognitive dysfunction and therapy-resistant seizures, which can be associated with refractory epilepsy and poor developmental outcomes. We hypothesize that neoangiogenesis, disruption of the blood-brain barrier, and leakage of serum proteins into the brain parenchyma play vital roles in the pathogenesis of TSC. Methods: In order to assess blood-brain barrier integrity, cortical tissue samples from TSC patients with intractable seizures, non-TSC patients with therapy-resistant epilepsy, and control subjects were immunolabeled for the serum protein fibrinogen, the adherens junction protein V-cadherin, and the tight junction protein occludin. Lectin was used to visualize blood vessels. Quantification was performed to assess average blood vessel segment length and branching. The fraction of membrane-associated V-cadherin and occludin, relative to the blood vessel surface area represented by lectin, was also analyzed. Results: The average length of blood vessel segments and the average number of branch nodes were significantly increased in TSC compared to epilepsy and control. The average surface area fraction of V-cadherin and occludin was significantly decreased in TSC compared to control. In addition, fibrinogen staining outside of the blood vessels was extensive in both TSC and epilepsy. These results confirm our hypothesis, suggesting blood-brain barrier dysfunction in TSC, with disease-specific neoangiogenic mechanisms in TSC. Discussion: Our results show increased blood-brain barrier permeability and increased vascular proliferation in TSC. These findings are likely due to decreased expression of tight junctions and adherens junctions in TSC cortical tissue. These results suggest that antiangiogenic therapies targeting the blood-brain barrier may offer a novel approach to preventing epileptogenesis in patients with TSC

Synthesis and characterization of mixed ligand chiral nanoclusters

Chiral mixed ligand silver nanoclusters were synthesized in the presence of a chiral and an achiral ligand. The ratio of the ligands was changed to track the formation of these clusters. While the chiral ligand lead to nanoparticles, Presence of the achiral ligand induced the formation of nanoclusters with chiral properties

Phylogeny-aware identification and correction of taxonomically mislabeled sequences

Molecular sequences in public databases are mostly annotated by the submitting authors without further validation. This procedure can generate erroneous taxonomic sequence labels. Mislabeled sequences are hard to identify, and they can induce downstream errors because new sequences are typically annotated using existing ones. Furthermore, taxonomic mislabelings in reference sequence databases can bias metagenetic studies which rely on the taxonomy. Despite significant efforts to improve the quality of taxonomic annotations, the curation rate is low because of the labor-intensive manual curation process. Here, we present SATIVA, a phylogeny-aware method to automatically identify taxonomically mislabeled sequences (‘mislabels’) using statistical models of evolution. We use the Evolutionary Placement Algorithm (EPA) to detect and score sequences whose taxonomic annotation is not supported by the underlying phylogenetic signal, and automatically propose a corrected taxonomic classification for those. Using simulated data, we show that our method attains high accuracy for identification (96.9% sensitivity/91.7% precision) as well as correction (94.9% sensitivity/89.9% precision) of mislabels. Furthermore, an analysis of four widely used microbial 16S reference databases (Greengenes, LTP, RDP and SILVA) indicates that they currently contain between 0.2% and 2.5% mislabels. Finally, we use SATIVA to perform an in-depth evaluation of alternative taxonomies for Cyanobacteria. SATIVA is freely available at https://github.com/amkozlov/sativa

A growing disconnection from nature is evident in cultural products

Human connection with nature is widely believed to be in decline, even though empirical evidence on the magnitude and temporal pattern of the change is scarce. Studying works of popular culture in English throughout the 20th century and later, we document a cultural shift away from nature, beginning in the 1950s. Since then, references to nature have been decreasing steadily in fiction, song lyrics, and film storylines. No parallel decline is observed in references to the human-made environment. These findings are cause for concern, not only because they imply foregone benefits from engagement with nature, but also because cultural products are agents of socialization that can evoke curiosity, respect, and concern for the natural world

Induced pluripotent stem cells for therapy personalization in pediatric patients: Focus on drug-induced adverse events

Adverse drug reactions (ADRs) are major clinical problems, particularly in special populations such as pediatric patients. Indeed, ADRs may be caused by a plethora of different drugs leading, in some cases, to hospitalization, disability or even death. In addition, pediatric patients may respond differently to drugs with respect to adults and may be prone to developing different kinds of ADRs, leading, in some cases, to more severe consequences. To improve the comprehension, and thus the prevention, of ADRs, the set-up of sensitive and personalized assays is urgently needed. Important progress is represented by the possibility of setting up groundbreaking patient-specific assays. This goal has been powerfully achieved using induced pluripotent stem cells (iPSCs). Due to their genetic and physiological species-specific differences and their ability to be differentiated ideally into all tissues of the human body, this model may be accurate in predicting drug toxicity, especially when this toxicity is related to individual genetic differences. This review is an up-to-date summary of the employment of iPSCs as a model to study ADRs, with particular attention to drugs used in the pediatric field. We especially focused on the intestinal, hepatic, pancreatic, renal, cardiac, and neuronal levels, also discussing progress in organoids creation. The latter are three-dimensional in vitro culture systems derived from pluripotent or adult stem cells simulating the architecture and functionality of native organs such as the intestine, liver, pancreas, kidney, heart, and brain. Based on the existing knowledge, these models are powerful and promising tools in multiple clinical applications including toxicity screening, disease modeling, personalized and regenerative medicine

Toxic equivalency factors (TEFs) after acute oral exposure of azaspiracid 1,-2 and-3 in mice

Azaspiracids (AZAs) are marine algal toxins that can be accumulated by edible shellfish to cause a foodborne gastrointestinal poisoning in humans. In the European Union, only AZA1, -2 and -3 are currently regulated and their concentration in shellfish is determined through their toxic equivalency factors (TEFs) derived from the intraperitoneal lethal potency in mice. Nevertheless, considering the potential human exposure by oral route, AZAs TEFs should be calculated by comparative oral toxicity data. Thus, the acute oral toxicity of AZA1, -2 and -3 was investigated in female CD-1 mice treated with different doses (AZA1: 135-1100 mu g/kg; AZA2 and AZA3: 300-1100 mu g/kg) and sacrificed after 24 h or 14 days. TEFs derived from the median lethal doses (LD50) were 1.0, 0.7 and 0.5, respectively for AZA1, -2 and -3. In fact, after 24 h from gavage administration, LD(50)s were 443 mu g/kg (AZA1; 95% CL: 350-561 mu g/kg), 626 mu g/kg (AZA2; 95% CL: 430-911 mu g/kg) and 875 mu g/kg (AZA3; 95% CL: 757-1010 mu g/kg). Mice dead more than 5 h after the treatment or those sacrificed after 24 h (doses: = 175 mu g AZA1/kg, >= 500 mu g AZA2/kg and >= 600 mu g AZA3/kg) showed enlarged pale liver, while increased serum markers of liver alteration were recorded even at the lowest doses. Blood chemistry revealed significantly increased serum levels of K+ ions (>= 500 mg/kg), whereas light microscopy showed tissue changes in the gastrointestinal tract, liver and spleen. No lethality, macroscopic, tissue or haematological changes were recorded two weeks post exposure, indicating reversible toxic effects. LC-MS/MS analysis of the main organs showed a dose-dependency in gastrointestinal absorption of these toxins: at 24 h, the highest levels were detected in the stomach and, in descending order, in the intestinal content, liver, small intestine, kidneys, lungs, large intestine, heart as well as detectable traces in the brain. After 14 days, AZA1 and AZA2 were still detectable in almost all the organs and intestinal content