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.

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.

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.

Vasopressin acts as a synapse organizer in limbic regions by boosting PSD95 and GluA1 expression.

Hypothalamic arginine vasopressin (AVP)-containing magnocellular neurosecretory neurons (AVPMNN) emit collaterals to synaptically innervate limbic regions influencing learning, motivational behaviour, and fear responses. Here, we characterize the dynamics of expression changes of two key determinants for synaptic strength, the postsynaptic density (PSD) proteins AMPAR subunit GluA1 and PSD scaffolding protein 95 (PSD95), in response to in vivo manipulations of AVPMNN neuronal activation state, or exposure to exogenous AVP ex vivo. Both long-term water deprivation in vivo, which powerfully upregulates AVPMNN metabolic activity, and exogenous AVP application ex vivo, in brain slices, significantly increased GluA1 and PSD95 expression as measured by western blotting, in brain regions reportedly receiving direct ascending innervations from AVPMNN (i.e., ventral hippocampus, amygdala and lateral habenula). By contrast, the visual cortex, a region not observed to receive AVPMNN projections, showed no such changes. Ex vivo application of V1a and V1b antagonists to ventral hippocampal slices ablated the AVP stimulated increase in postsynaptic protein expression measured by western blotting. Using a modified expansion microscopy technique, we were able to quantitatively assess the significant augmentation of PSD95 and GLUA1 densities in subcellular compartments in locus coeruleus tyrosine hydroxylase immunopositive fibres, adjacent to AVP axon terminals. Our data strongly suggest that the AVPMNN ascending system plays a role in the regulation of the excitability of targeted neuronal circuits through upregulation of key postsynaptic density proteins corresponding to excitatory synapses.

Metabolic acidosis and hyperkalemia differentially regulate cation HCN3 channel in the rat nephron.

The kidney controls body fluids, electrolyte and acid-base balance. Previously, we demonstrated that hyperpolarization-activated and cyclic nucleotide-gated (HCN) cation channels participate in ammonium excretion in the rat kidney. Since acid-base balance is closely linked to potassium metabolism, in the present work we aim to determine the effect of chronic metabolic acidosis (CMA) and hyperkalemia (HK) on protein abundance and localization of HCN3 in the rat kidney. CMA increased HCN3 protein level only in the outer medulla (2.74 ± 0.31) according to immunoblot analysis. However, immunofluorescence assays showed that HCN3 augmented in cortical proximal tubules (1.45 ± 0.11) and medullary thick ascending limb of Henle's loop (4.48 ± 0.45) from the inner stripe of outer medulla. HCN3 was detected in brush border membranes (BBM) and mitochondria of the proximal tubule by immunogold electron and confocal microscopy in control conditions. Acidosis did not alter HCN3 levels in BBM and mitochondria but augmented them in lysosomes. HCN3 was also immuno-detected in mitoautophagosomes. In the distal nephron, HCN3 was expressed in principal and intercalated cells from cortical to medullary collecting ducts. CMA did not change HCN3 abundance in these nephron segments. In contrast, HK doubled HCN3 level in cortical collecting ducts and favored its basolateral localization in principal cells from the inner medullary collecting ducts. These findings further support HCN channels contribution to renal acid-base and potassium balance.

"Funny" channels in cardiac mitochondria modulate membrane potential and oxygen consumption.

The hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are encoded by a family of four genes (HCN1-4). All isoforms are expressed in the heart, HCN4 being the most abundant in the sinoatrial node (SAN). HCN channels are responsible for the "funny" current (If) associated with the generation and autonomic control of the diastolic depolarization phase of cardiac action potential. In this work we performed a proteomic analysis of HCN4 transfected in HEK293 cells. Most of the identified proteins in the HCN4 network belonged to mitochondria. The subcellular localization of HCN channels was predicted in plasma membrane, mitochondria and nucleus. Experimentally, HCN2 (full-length, truncated), HCN3 (full-length, truncated) and HCN4 (truncated) were detected in rat heart mitochondria by immunoblotting. If sensitive to ZD7288, was recorded by patch-clamp in mitoplasts from cardiomyocytes. Mitochondrial membrane potential (ΔΨm) assessment in H9c2 cells revealed that ZD7288 induced almost 50% higher hyperpolarization respect to control at 30 min. Furthermore, ZD7288 reduced oxygen consumption attributed to ATP synthesis in H9c2 cells. In conclusion, we identify for the first time functional HCN channels in mammalian cardiac mitochondria and demonstrate their impact on ΔΨm and respiration.

Novel Potassium Channels in Kidney Mitochondria: The Hyperpolarization-Activated and Cyclic Nucleotide-Gated HCN Channels.

Hyperpolarization-activated cationic HCN channels comprise four members (HCN1-4) that control dendritic integration, synaptic transmission and action potential firing. In the kidney, HCN1, HCN2 and HCN3 are differentially expressed and contribute to the transport of sodium, potassium (K+) and ammonium into the nephrons. HCN3 is regulated by K+ diets in the kidney. In this work we performed a proteomic analysis of HCN3 expressed in human embryonic kidney cells (HEK293 cells). More than 50% of the interacting proteins belonged to mitochondria. Therefore, we explored the presence of HCN channels in kidney mitochondria. By immunoblotting and immunogold electron microscopy HCN3 protein expression was found in rat kidney mitochondria; it was also confirmed in human kidney. Patch-clamp recordings of renal mitochondria and mitochondria from HEK293 cells overexpressing HCN1, HCN2 and HCN3 channels, stained with MitoTracker Green FM, indicated that only HCN3 could produce inwardly K+ currents that were inhibited by ZD7288, a specific blocker of HCN channels. Furthermore, ZD7288 caused inhibition of the oxygen consumption coupled to ATP synthesis and hyperpolarization of the inner mitochondrial membrane. In conclusion, we show for the first time that pacemaker HCN channels contribute to K+ transport in mitochondria facilitating the activity of the respiratory chain and ATP synthesis by controlling the inner mitochondrial membrane potential.

Insights in cullin 3/WNK4 and its relationship to blood pressure regulation and electrolyte homeostasis.

One of the most important systems for protein degradation is the ubiquitin-proteasome system (UPS). The highly specific process called ubiquitination is provided by the E3 ubiquitin ligases, which mediates degradation via the proteasome system. The ubiquitin ligases based on cullins are the type of ubiquitin ligases known so far. The complex based on cullin 3 (Cul3) requires that its target protein has a bric-a-brac/tram-track/broad-complex (BTB) domain to recognize it. Cul3 has been widely associated with Kelch-like erythroid cell-derived protein with CNC homology (ECH)-associated protein 1 (Keap1) and the cytoprotective nuclear factor erythroid 2 related factor 2 (Nrf2) pathway and the proper control of cell cycle progression. Recently, Cul3 has been linked to the development of type II pseudohypoaldosteronism (PHAII or Gordon's syndrome) due to the fact that Cul3 has the ability to bind to Kelch-like 3 protein (KLHL3) and therefore mediating the degradation of some members of the WNK kinases. In this work we focused on highlighting how Cul3 system is involved in the regulation of electrolyte homeostasis and blood pressure.