Perspectives on carboxylates generation from Ecuadorian agro-wastes.

Carboxylates generation from banana (peel and pulp), coffee, and cacao fermentation agro-waste, upon uncontrolled and controlled pHs of 6.6 (heat-driven methanogens inactivation) and 5.2 (pH inactivation), was studied. Regarding volatile fatty acids (VFAs), acetic was the highest for cocoa (96.2 g kg-1TVS) at pH 4.5. However, butyric was relevant for banana pulp (90.7 g kg-1TVS), at controlled pH 6.6. The highest medium chain fatty acid (MCFAs) level was hexanoic (cocoa, 3.5 g kg-1TVS), while octanoic reached a maximum of 2.8 g kg-1TVS for coffee at pH 6.6. At pH 5.2 MCFAs yield was relatively low. Uncontrolled pH conditions, using banana resulted in superior VFAs production compared to controlled conditions. Thus, pH became a determining variable when deciding the time and kind of carboxylic acid to be recovered. The bacterial community at the end of the chain elongation process was dominated by phyla Firmicutes, and Clostridium as the most common genera.

Incidence and management of the main serious adverse events reported after COVID-19 vaccination.

Coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2n first appeared in Wuhan, China in 2019. Soon after, it was declared a pandemic by the World Health Organization. The health crisis imposed by a new virus and its rapid spread worldwide prompted the fast development of vaccines. For the first time in human history, two vaccines based on recombinant genetic material technology were approved for human use. These mRNA vaccines were applied in massive immunization programs around the world, followed by other vaccines based on more traditional approaches. Even though all vaccines were tested in clinical trials prior to their general administration, serious adverse events, usually of very low incidence, were mostly identified after application of millions of doses. Establishing a direct correlation (the cause-effect paradigm) between vaccination and the appearance of adverse effects has proven challenging. This review focuses on the main adverse effects observed after vaccination, including anaphylaxis, myocarditis, vaccine-induced thrombotic thrombocytopenia, Guillain-Barré syndrome, and transverse myelitis reported in the context of COVID-19 vaccination. We highlight the symptoms, laboratory tests required for an adequate diagnosis, and briefly outline the recommended treatments for these adverse effects. The aim of this work is to increase awareness among healthcare personnel about the serious adverse events that may arise post-vaccination. Regardless of the ongoing discussion about the safety of COVID-19 vaccination, these adverse effects must be identified promptly and treated effectively to reduce the risk of complications.

NMDA receptor-mediated Ca2+ signaling: Impact on cell cycle regulation and the development of neurodegenerative diseases and cancer.

NMDA receptors are Ca2+-permeable ligand-gated ion channels that mediate fast excitatory transmission in the central nervous system. NMDA receptors regulate the proliferation and differentiation of neural progenitor cells and also play critical roles in neural plasticity, memory, and learning. In addition to their physiological role, NMDA receptors are also involved in glutamate-mediated excitotoxicity, which results from excessive glutamate stimulation, leading to Ca2+ overload, and ultimately to neuronal death. Thus, NMDA receptor-mediated excitotoxicity has been linked to several neurodegenerative diseases such as Alzheimer's, Parkinson's, Huntington's, dementia, and stroke. Interestingly, in addition to its effects on cell death, aberrant expression or activation of NMDA receptors is also involved in pathological cellular proliferation, and is implicated in the invasion and proliferation of various types of cancer. These disorders are thought to be related to the contribution of NMDA receptors to cell proliferation and cell death through cell cycle modulation. This review aims to discuss the evidence implicating NMDA receptor activity in cell cycle regulation and the link between aberrant NMDA receptor activity and the development of neurodegenerative diseases and cancer due to cell cycle dysregulation. The information presented here will provide insights into the signaling pathways and the contribution of NMDA receptors to these diseases, and suggests that NMDA receptors are promising targets for the prevention and treatment of these diseases, which are leading causes of death and disability worldwide.

SIDT2 Associates with Apolipoprotein A1 (ApoA1) and Facilitates ApoA1 Secretion in Hepatocytes.

SIDT2 is a lysosomal protein involved in the degradation of nucleic acids and the transport of cholesterol between membranes. Previous studies identified two "cholesterol recognition/interaction amino acid consensus" (CRAC) motifs in SIDT1 and SIDT2 members. We have previously shown that the first CRAC motif (CRAC-1) is essential for protein translocation to the PM upon cholesterol depletion in the cell. In the present study, we show that SIDT2 and the apolipoprotein A1 (ApoA1) form a complex which requires the second CRAC-2 motif in SIDT2 to be established. The overexpression of SIDT2 and ApoA1 results in enhanced ApoA1 secretion by HepG2 cells. This is not observed when overexpressing the SIDT2 with the CRAC-2 domain mutated to render it unfunctional. All these results provide evidence of a novel role for SIDT2 as a protein forming a complex with ApoA1 and enhancing its secretion to the extracellular space.

The conducting state of TRPA1 modulates channel lateral mobility.

We have studied Danio rerio (Zebrafish) TRPA1 channel using a method that combines single channel electrophysiological and optical recordings to evaluate lateral mobility and channel gating simultaneously in single channels. TRPA1 channel activation by two distinct chemical ligands: allyl isothiocyanate (AITC) and TRPswitch B, results in substantial reduction of channel lateral mobility at the plasma membrane. Incubation with the cholesterol sequestering agent methyl-ß-cyclodextrin (MßCD), prevents the reduction on lateral mobility induced by the two chemical agonists. This results strongly suggest that the open conformation of TRPA1 modulates channel lateral mobility probably by facilitating the insertion of the channel into cholesterol-enriched domains at the plasma membrane.

STIM1: The lord of the rings?

STIM1 and Orai1 are the central core of the Store Operated Calcium Entry (SOCE). This calcium influx mechanism is triggered after the activation of Gq protein-coupled receptors at the plasma membrane (PM) that activate phospholipase C. The phospholipase C produces Inositol triphosphate (IP3) which rapidly diffuses throughout the cytosol, resulting in the binding and activation of IP3 receptors (IP3R) and the rapid efflux of calcium from the endoplasmic reticulum (ER) to the cytosol. The calcium depletion in the ER is sensed by the stromal interaction molecule 1 (STIM1) a single-pass transmembrane protein at the ER that binds intraluminal calcium through an EF-hand domain in its amino terminal region (Fig. 1A). The cytosolic portion of STIM1 contains multiple domains. The region that interacts and activates Orai channels is known as SOAR (the STIM1 Orai activating region) [1]. For SOAR be accessible to Orai1, STIM1 must get an extended conformation that unlocks SOAR from its coiled-coil 1 (CC1) region [2]. The extended conformation is triggered by calcium depletion at the ER that oligomerizes STIM1. The oligomers of STIM1 then translocate to a close distance between two opposing membranes, forming what is known as ER-PM junctions. STIM1 accumulates at ER-PM junctions conforming the denominated STIM1 puncta.

Roles of Cholesterol and PtdIns(4,5)P2 in the Regulation of STIM1-Orai1 Channel Function.

Calcium is one of the most prominent second messengers. It is involved in a wide range of functions at the single-cell level but also in modulating regulatory mechanisms in the entire organism. One process mediating calcium signaling involves hydrolysis of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) by the phospholipase-C (PLC). Thus, calcium and PtdIns(4,5)P2 are intimately intertwined two second-messenger cascades that often depend on each other. Another relevant lipid associated with calcium signaling is cholesterol. Both PtdIns(4,5)P2 and cholesterol play key roles in the formation and maintenance of specialized signaling nanodomains known as lipid rafts. Lipid rafts are particularly important in calcium signaling by concentrating and localizing calcium channels such as the Orai1 channel. Depletion of internal calcium stores is initiated by the production of inositol-1,4,5-trisphosphate (IP3). Calcium depletion from the ER induces the oligomerization of STIM1, which binds Orai1 and initiates calcium influx into the cell. In the present review, we analyzed the complex interactions between cholesterol, PtdIns(4,5)P2, and the complex formed by the Orai1 channel and the signaling molecule STIM1. We explore some of the complex mechanisms governing calcium homeostasis and phospholipid metabolism, as well as the interaction between these two apparently independent signaling cascades.

Baculovirus Display of Peptides and Proteins for Medical Applications.

Baculoviridae is a large family of arthropod-infective viruses. Recombinant baculoviruses have many applications, the best known is as a system for large scale protein production in combination with insect cell cultures. More recently recombinant baculoviruses have been utilized for the display of proteins of interest with applications in medicine. In the present review we analyze the different strategies for the display of proteins and peptides on the surface of recombinant baculoviruses and provide some examples of the different proteins displayed. We analyze briefly the commercially available systems for recombinant baculovirus production and display and discuss the future of this emerging and powerful technology.

Mielitis transversa aguda asociada con infección por SARS-CoV-2 / Acute Transverse Myelitis Associated to SARS-CoV-2 Infection

Resumen Además de provocar problemas respiratorios, la infección con el SARS-CoV-2 puede causar un espectro amplio de complicaciones neurológicas como son cefalea, ageusia, anosmia, encefalopatía, enfermedad cerebrovascular, síndrome de Guillain-Barré y otras polineuropatías, meningoencefalitis, encefalopatía hemorrágica necrotizante aguda y síndromes del sistema nervioso central asociados a inflamación. En esta revisión presentamos 39 casos de mielitis transversa aguda (MTA) asociados a infección por el SARS-CoV-2, confirmado por una prueba de reacción en cadena de la polimerasa (PCR) por hisopado nasofaríngeo y/o de antígeno en plasma. Los análisis bioquímicos de los pacientes confirmaron la ausencia de otros patógenos y de autoanticuerpos involucrados en inflamación del sistema nervioso, excepto por 2 casos, uno con autoanticuerpos contra la glicoproteína de la mielina de los oligodendrocitos (anti-MOG IgG) y otro con anti-MOG IgG y antidescarboxilasa del ácido glutámico (anti-GAD65), indicativos de una enfermedad autoinmune del sistema nervioso central. Los estudios de imagenología de resonancia magnética (RMN) confirmaron el diagnóstico de MTA. Aunque no se puede inferir causalidad, es muy probable que los casos aislados de MTA sean consecuencia de un proceso para o postinfeccioso en la COVID-19. La comprensión de los mecanismos que desencadenan estos trastornos neurológicos durante o al final de la infección viral, ayudarán a optimizar las estrategias terapéuticas para el manejo del paciente.

Abstract In addition to causing respiratory problems, SARS-CoV-2 can cause a wide spectrum of neurological complications such as headaches, ageusia, anosmia, encephalopathy, cerebrovascular disease, Guillain-Barré syndrome and other polyneuropathies, meningoencephalitis, acute necrotizing hemorrhagic encephalopathy, and central nervous system syndromes associated with inflammation. In this review we present 39 cases of acute transverse myelitis (ATM) associated to SARS-CoV-2 infection. All cases had a positive polymerase chain reaction (PCR) nasopharyngeal swab and/or antigen test. Biochemical analyses confirmed the absence of other pathogens and autoantibody-mediated neuroinflammatory disease, except for two cases, one with autoantibodies against the oligodendrocyte myelin glycoprotein (anti-MOG IgG) and another with anti-MOG IgG and anti-glutamic acid decarboxylase (anti-GAD65). Magnetic Resonance Imaging (MRI) studies confirmed the diagnosis of ATM. Although causality cannot be inferred, it is likely that isolated cases of ATM are the consequence of a para or post-infectious process in SARS-CoV-2. In this work, the probable causes of ATM associated with SARS-CoV-2 are discussed. The understanding of the mechanisms behind these neurological disorders triggered by a viral infection will help to optimize the therapeutic strategies for patient management.

An ambient-temperature stable nanoparticle-based vaccine for nasal application that confers long-lasting immunogenicity to carried antigens.

Polyhedrins are viral proteins present in a large family of baculoviruses that form occlusion bodies (polyhedra). These structures protect the virus particles from the outside environment until they are ingested by susceptible insects. Occluded viruses can sustain inclement weather for long periods of time. Therefore, the polyhedra is a natural preservative that keeps the viral structure intact at ambient temperature for years. In a previous study we identified the first 110 amino acids from polyhedrin (PH(1-110)) as a good candidate to carry antigens of interest. As a proof of concept, we produced a fusion protein with PH(1-110) and the green fluorescent protein (PH(1-110)GFP). The fusion protein associates spontaneously during its synthesis resulting in the formation of nanoparticles. Nasal immunization with these nanoparticles and in the absence of any adjuvant, results in a robust immune response with the production of IgG immunoglobulins that remained elevated for months and that selectively recognize the GFP but not PH(1-110). These results indicate that PH(1-110) is poorly immunogenic but capable of enhancing the immune response to GFP.

Design of Hydrogel Silk-Based Microarrays and Molecular Beacons for Reagentless Point-of-Care Diagnostics.

We have developed a novel microarray system based on three technologies: 1) molecular beacons designed to interact with DNA targets at room temperature (25-27°C), 2) tridimensional silk-based microarrays containing the molecular beacons immersed in the silk hydrogel, and 3) shallow angle illumination, which uses separated optical pathways for excitation and emission. Unlike conventional microarrays that exhibit reduced signal-to-background ratio, require several stages of incubation, rinsing, and stringency control, and measure only end-point results, our microarray technology provides enhanced signal-to-background ratio (achieved by separating the optical pathways for excitation and emission, resulting in reduced stray light), performs analysis rapidly in one step without the need for labeling DNA targets, and measures the entire course of association kinetics between target DNA and the molecular beacons. To illustrate the benefits of our technology, we conducted microarray assays designed for the identification of influenza viruses. We show that in a single microarray slide, we can identify the virus subtype according to the molecular beacons designed for hemagglutinin (H1, H2, and H3) and neuraminidase (N1, N2). We also show the identification of human and swine influenza using sequence-specific molecular beacons. This microarray technology can be easily implemented for reagentless point-of-care diagnostics of several contagious diseases, including coronavirus variants responsible for the current pandemic.

LIPID transfer proteins regulate store-operated calcium entry via control of plasma membrane phosphoinositides.

The ER-resident proteins STIM1 together with the plasma membrane (PM)-localized Orai1 channels constitute the molecular components of the store-operated Ca2+ entry (SOCE) pathway. Prepositioning of STIM1 to the peripheral ER close to the PM ensures its efficient interaction with Orai1 upon a decrease in the ER luminal Ca2+ concentration. The C-terminal polybasic domain of STIM1 has been identified as mediating the interaction with PM phosphoinositides and hence positions the molecule to ER-PM contact sites. Here we show that STIM1 requires PM phosphatidylinositol 4-phosphate (PI4P) for efficient PM interaction. Accordingly, oxysterol binding protein related proteins (ORPs) that work at ER-PM junctions and consume PI4P gradients exert important control over the Ca2+ entry process. These studies reveal an important connection between non-vesicular lipid transport at ER-PM contact sites and regulation of ER Ca2+store refilling.

Super-resolution microscopy for the study of store-operated calcium entry.

The use of a variety of techniques based on super-resolution (SR) microscopy unveiled a close and complex relationship between cytoskeleton reorganization and SOCE. By using SR microscopy many new proteins involved in SOCE regulation have been identified over the last few years. Many enigmas remain unsolved in this highly dynamic field, however, recent developments in SR microscopy promise new answers soon. In the present review, we describe the most relevant findings in SOCE components and SOCE modulation using different methods derived from SR microscopy.

Implications of the Immune Polymorphisms of the Host and the Genetic Variability of SARS-CoV-2 in the Development of COVID-19.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for the current pandemic affecting almost all countries in the world. SARS-CoV-2 is the agent responsible for coronavirus disease 19 (COVID-19), which has claimed millions of lives around the world. In most patients, SARS-CoV-2 infection does not cause clinical signs. However, some infected people develop symptoms, which include loss of smell or taste, fever, dry cough, headache, severe pneumonia, as well as coagulation disorders. The aim of this work is to report genetic factors of SARS-CoV-2 and host-associated to severe COVID-19, placing special emphasis on the viral entry and molecules of the immune system involved with viral infection. Besides this, we analyze SARS-CoV-2 variants and their structural characteristics related to the binding to polymorphic angiotensin-converting enzyme type 2 (ACE2). Additionally, we also review other polymorphisms as well as some epigenetic factors involved in the immunopathogenesis of COVID-19. These factors and viral variability could explain the increment of infection rate and/or in the development of severe COVID-19.

An artificial virus-like triblock protein shows low in vivo humoral immune response and high stability.

The use of viral vectors for in vivo gene therapy can be severely limited by their immunogenicity. Non-viral vectors may represent an alternative, however, reports analyzing their immunogenicity are still lacking. Here, we studied the humoral immune response in a murine model triggered by artificial virus-like particles (AVLPs) carrying plasmid or antisense DNA. The AVLPs were assembled using a family of modular proteins based on bioinspired collagen-like and silk-like sequences that produce virus-like particles. We compared our AVLPs against an Adeno Associated Virus 1 (AAV), a widely used viral vector for in vivo gene delivery that has been approved by the FDA and EMA for gene therapy. We found that a 1000-fold higher mass of AVLPs than AAV are necessary to obtain similar specific antibody titters. Furthermore, we studied the stability of AVLPs against relevant biological reagents such as heparin and fetal bovine serum to ensure nucleic acid protection in biological media. Our study demonstrates that the AVLPs are stable in physiological conditions and can overcome safety limitations such as immunogenicity. The scarce humoral immunogenicity and high stability found with AVLPs suggest that they have potential to be used as stealth non-viral gene delivery systems for in vivo studies or gene therapy.

SARS-CoV-2 Vaccines Based on the Spike Glycoprotein and Implications of New Viral Variants.

Coronavirus 19 Disease (COVID-19) originating in the province of Wuhan, China in 2019, is caused by the severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), whose infection in humans causes mild or severe clinical manifestations that mainly affect the respiratory system. So far, the COVID-19 has caused more than 2 million deaths worldwide. SARS-CoV-2 contains the Spike (S) glycoprotein on its surface, which is the main target for current vaccine development because antibodies directed against this protein can neutralize the infection. Companies and academic institutions have developed vaccines based on the S glycoprotein, as well as its antigenic domains and epitopes, which have been proven effective in generating neutralizing antibodies. However, the emergence of new SARS-CoV-2 variants could affect the effectiveness of vaccines. Here, we review the different types of vaccines designed and developed against SARS-CoV-2, placing emphasis on whether they are based on the complete S glycoprotein, its antigenic domains such as the receptor-binding domain (RBD) or short epitopes within the S glycoprotein. We also review and discuss the possible effectiveness of these vaccines against emerging SARS-CoV-2 variants.

Genome-Wide Association Study Identifies a Functional SIDT2 Variant Associated With HDL-C (High-Density Lipoprotein Cholesterol) Levels and Premature Coronary Artery Disease.

Objective: Low HDL-C (high-density lipoprotein cholesterol) is the most frequent dyslipidemia in Mexicans, but few studies have examined the underlying genetic basis. Our purpose was to identify genetic variants associated with HDL-C levels and cardiovascular risk in the Mexican population. Approach and Results: A genome-wide association studies for HDL-C levels in 2335 Mexicans, identified four loci associated with genome-wide significance: CETP, ABCA1, LIPC, and SIDT2. The SIDT2 missense Val636Ile variant was associated with HDL-C levels and was replicated in 3 independent cohorts (P=5.9×10−18 in the conjoint analysis). The SIDT2/Val636Ile variant is more frequent in Native American and derived populations than in other ethnic groups. This variant was also associated with increased ApoA1 and glycerophospholipid serum levels, decreased LDL-C (low-density lipoprotein cholesterol) and ApoB levels, and a lower risk of premature CAD. Because SIDT2 was previously identified as a protein involved in sterol transport, we tested whether the SIDT2/Ile636 protein affected this function using an in vitro site-directed mutagenesis approach. The SIDT2/Ile636 protein showed increased uptake of the cholesterol analog dehydroergosterol, suggesting this variant affects function. Finally, liver transcriptome data from humans and the Hybrid Mouse Diversity Panel are consistent with the involvement of SIDT2 in lipid and lipoprotein metabolism. Conclusions: This is the first genome-wide association study for HDL-C levels seeking associations with coronary artery disease in the Mexican population. Our findings provide new insight into the genetic architecture of HDL-C and highlight SIDT2 as a new player in cholesterol and lipoprotein metabolism in humans.

Palmitic acid induces insulin resistance by a mechanism associated with energy metabolism and calcium entry in neuronal cells.

Palmitic acid (PA) is a saturated fatty acid whose high consumption has been largely associated with the development of different metabolic alterations, such as insulin resistance, metabolic syndrome, and type 2 diabetes. Particularly in the brain, insulin signaling disruption has been linked to cognitive decline and is considered a risk factor for Alzheimer's disease. Cumulative evidence has demonstrated the participation of PA in the molecular cascade underlying cellular insulin resistance in peripheral tissues, but its role in the development of neuronal insulin resistance and the mechanisms involved are not fully understood. It has generally been accepted that the brain does not utilize fatty acids as a primary energy source, but recent evidence shows that neurons possess the machinery for fatty acid ß-oxidation. However, it is still unclear under what conditions neurons use fatty acids as energy substrates and the implications of their oxidative metabolism in modifying insulin-stimulated effects. In the present work, we have found that neurons differentiated from human neuroblastoma MSN exposed to high but nontoxic concentrations of PA generate ATP through mitochondrial metabolism, which is associated with an increase in the cytosolic Ca2+ and diminished insulin signaling in neurons. These findings reveal a novel mechanism by which saturated fatty acids produce Ca2+ entry and insulin resistance that may play a causal role in increasing neuronal vulnerability associated with metabolic diseases.

Developing a Portable Device for the Identification of miRNAs in Fluids.

In the present work we describe a novel system for the identification of microRNAs (miRNAs) in fluids. The method is based on combined novel 3D microarray technology using silk as scaffold and total internal reflection fluorescence microscopy (TIRFM), which allows for the rapid identification of miRNAs using a portable device.

Phosphorylation of NMDA receptors by cyclin B/CDK1 modulates calcium dynamics and mitosis.

N-methyl-D-aspartate receptors (NMDAR) are glutamate-gated calcium channels named after their artificial agonist. NMDAR are implicated in cell proliferation under normal and pathophysiological conditions. However, the role of NMDAR during mitosis has not yet been explored in individual cells. We found that neurotransmitter-evoked calcium entry via endogenous NMDAR in cortical astrocytes was transient during mitosis. The same occurred in HEK293 cells transfected with the NR1/NR2A subunits of NMDAR. This transient calcium entry during mitosis was due to phosphorylation of the first intracellular loop of NMDAR (S584 of NR1 and S580 of NR2A) by cyclin B/CDK1. Expression of phosphomimetic mutants resulted in transient calcium influx and enhanced NMDAR inactivation independent of the cell cycle phase. Phosphomimetic mutants increased entry of calcium in interphase and generated several alterations during mitosis: increased mitotic index, increased number of cells with lagging chromosomes and fragmentation of pericentriolar material. In summary, by controlling cytosolic calcium, NMDAR modulate mitosis and probably cell differentiation/proliferation. Our results suggest that phosphorylation of NMDAR by cyclin B/CDK1 during mitosis is required to preserve mitotic fidelity. Altering the modulation of the NMDAR by cyclin B/CDK1 may conduct to aneuploidy and cancer.