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Mutations in the genes encoding for voltage-gated sodium channels cause profound sensory disturbances and other symptoms dependent on the distribution of a particular channel subtype in different organs. Humans with the gain-of-function mutation p.Leu811Pro in SCN11A (encoding for the voltage-gated Nav1.9 channel) exhibit congenital insensitivity to pain, pruritus, self-inflicted injuries, slow healing wounds, muscle weakness, Charcot-like arthropathies, and intestinal dysmotility. As already shown, knock-in mice (Scn11a+/L799P) carrying the orthologous mutation p.Leu799Pro replicate reduced pain sensitivity and show frequent tissue lesions. In the present study we explored whether Scn11a+/L799P mice develop also pruritus, muscle weakness, and changes in gastrointestinal transit time. Furthermore, we analyzed morphological and functional differences in nerves, skeletal muscle, joints and small intestine from Scn11a+/L799P and Scn11a+/+ wild type mice. Compared to Scn11a+/+ mice, Scn11a+/L799P mice showed enhanced scratching bouts before skin lesions developed, indicating pruritus. Scn11a+/L799P mice exhibited reduced grip strength, but no disturbances in motor coordination. Skeletal muscle fiber types and joint architecture were unaltered in Scn11a+/L799P mice. Their gastrointestinal transit time was unaltered. The small intestine from Scn11a+/L799P showed a small shift towards less frequent peristaltic movements. Similar proportions of lumbar dorsal root ganglion neurons from Scn11a+/L799P and Scn11a+/+ mice were calcitonin gene-related peptide (CGRP-) positive, but isolated sciatic nerves from Scn11a+/L799P mice exhibited a significant reduction of the capsaicin-evoked release of CGRP indicating reduced neurogenic inflammation. These data indicate important Nav1.9 channel functions in several organs in both humans and mice. They support the pathophysiological relevance of increased basal activity of Nav1.9 channels for sensory abnormalities (pain and itch) and suggest resulting malfunctions of the motor system and of the gastrointestinal tract. Scn11a+/L799P mice are suitable to investigate the role of Nav1.9, and to explore the pathophysiological changes and mechanisms which develop as a consequence of Nav1.9 hyperactivity. [ABSTRACT FROM AUTHOR]

Massive apoptosis of lymphocytes is a hallmark of sepsis. The resulting immunosuppression is associated with secondary infections, which are often lethal. Moreover, sepsis-survivors are burdened with increased morbidity and mortality for several years after the sepsis episode. The duration and clinical consequences of sepsis induced-immunosuppression are currently unknown. We have used the mouse model of peritoneal contamination and infection (PCI) to investigate the quantitative and qualitative recovery of T lymphocytes for 3.5 months after sepsis with or without IL-7 treatment. Thymic output and the numbers of naive and effector/memory CD4+ and CD8+ lymphocytes quickly recovered after sepsis. IL-7 treatment resulted in an accelerated recovery of CD8+ lymphocytes. Next generation sequencing revealed no significant narrowing of the T cell receptor repertoire 3.5 months after sepsis. In contrast, detailed functional analyses of T helper (Th)-cell responses towards a fungal antigen revealed a significant loss of Th cells. Whereas cytokine production was not impaired at the single cell level, the absolute number of Th cells specific for the fungal antigen was reduced. Our data indicate a clinically relevant loss of pathogen-specific T cell clones after sepsis. Given the small number of naive T lymphocytes specific for a given antigen, this decrement of T cell clones remains undetected even by sensitive methods such as deep sequencing. Taken together, our data are compatible with long lasting impairments in CD4+ T-cell responses after sepsis despite rapid recovery of T lymphocyte populations. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) may serve as elegant tool to improve risk stratification in bicuspid aortic valve (BAV)-associated aortopathy. However, the exact pathogenetic pathway by which miRNAs impact aortopathy progression is unknown. Herewith, we aimed to analyze the association between circulating miRNAs and rare variants of aortopathy-related genes. 63 BAV patients (mean age 47.3±11.3 years, 92% male) with a root dilatation phenotype, who underwent aortic valve+/-proximal aortic surgery at a single institution (mean post-AVR follow-up 10.3±6.9 years) were analyzed. A custom-made HaloPlex HS panel including 20 aortopathy-related genes was used for the genetic testing. miRNAs were extracted from whole blood and miRNA analysis was performed using miRNA-specific assay. Study endpoint was the association between circulating miRNAs and rare genetic variants in the aortopathy gene panel. The study cohort was divided into a subgroup with rare variants vs. a subgroup without rare variants based on the presence of rare variants in the respective genes (i.e., at least one variant present). The genetic analysis yielded n = 6 potentially and likely pathogenic rare variants within the NOTCH1 gene as being the most common finding. Univariate analysis between blood miRNAs and NOTCH1 variants revealed a significantly lower expression of miR-145 in the subgroup of patients with NOTCH1 variants vs. those without NOTCH1 variants (i.e., delta Ct 4.95±0.74 vs. delta Ct 5.57±0.78, p = 0.04). Our preliminary data demonstrate a significant association between blood miR-145 expression and the presence of rare NOTCH1 variants. This association may be indicative of a specific pathogenetic pathway in the development of genetically-triggered bicuspid aortopathy. [ABSTRACT FROM AUTHOR]

Pain is necessary to alert us to actual or potential tissue damage. Specialized nerve cells in the body periphery, so called nociceptors, are fundamental to mediate pain perception and humans without pain perception are at permanent risk for injuries, burns and mutilations. Pain insensitivity can be caused by sensory neurodegeneration which is a hallmark of hereditary sensory and autonomic neuropathies (HSANs). Although mutations in several genes were previously associated with sensory neurodegeneration, the etiology of many cases remains unknown. Using next generation sequencing in patients with congenital loss of pain perception, we here identify bi-allelic mutations in the FLVCR1 (Feline Leukemia Virus subgroup C Receptor 1) gene, which encodes a broadly expressed heme exporter. Different FLVCR1 isoforms control the size of the cytosolic heme pool required to sustain metabolic activity of different cell types. Mutations in FLVCR1 have previously been linked to vision impairment and posterior column ataxia in humans, but not to HSAN. Using fibroblasts and lymphoblastoid cell lines from patients with sensory neurodegeneration, we here show that the FLVCR1-mutations reduce heme export activity, enhance oxidative stress and increase sensitivity to programmed cell death. Our data link heme metabolism to sensory neuron maintenance and suggest that intracellular heme overload causes early-onset degeneration of pain-sensing neurons in humans. [ABSTRACT FROM AUTHOR]

Hereditary spastic paraplegia (HSP) is characterized by a dying back degeneration of corticospinal axons which leads to progressive weakness and spasticity of the legs. SPG11 is the most common autosomal-recessive form of HSPs and is caused by mutations in SPG11. A recent in vitro study suggested that Spatacsin, the respective gene product, is needed for the recycling of lysosomes from autolysosomes, a process known as autophagic lysosome reformation. The relevance of this observation for hereditary spastic paraplegia, however, has remained unclear. Here, we report that disruption of Spatacsin in mice indeed causes hereditary spastic paraplegia-like phenotypes with loss of cortical neurons and Purkinje cells. Degenerating neurons accumulate autofluorescent material, which stains for the lysosomal protein Lamp1 and for p62, a marker of substrate destined to be degraded by autophagy, and hence appears to be related to autolysosomes. Supporting a more generalized defect of autophagy, levels of lipidated LC3 are increased in Spatacsin knockout mouse embryonic fibrobasts (MEFs). Though distinct parameters of lysosomal function like processing of cathepsin D and lysosomal pH are preserved, lysosome numbers are reduced in knockout MEFs and the recovery of lysosomes during sustained starvation impaired consistent with a defect of autophagic lysosome reformation. Because lysosomes are reduced in cortical neurons and Purkinje cells in vivo, we propose that the decreased number of lysosomes available for fusion with autophagosomes impairs autolysosomal clearance, results in the accumulation of undegraded material and finally causes death of particularly sensitive neurons like cortical motoneurons and Purkinje cells in knockout mice. [ABSTRACT FROM AUTHOR]

Hereditary spastic paraplegias (HSPs) are characterized by progressive weakness and spasticity of the legs because of the degeneration of cortical motoneuron axons. SPG15 is a recessively inherited HSP variant caused by mutations in the ZFYVE26 gene and is additionally characterized by cerebellar ataxia, mental decline, and progressive thinning of the corpus callosum. ZFYVE26 encodes the FYVE domain-containing protein ZFYVE26/SPASTIZIN, which has been suggested to be associated with the newly discovered adaptor protein 5 (AP5) complex. We show that Zfyve26 is broadly expressed in neurons, associates with intracellular vesicles immunopositive for the early endosomal marker EEA1, and co-fractionates with a component of the AP5 complex. As the function of ZFYVE26 in neurons was largely unknown, we disrupted Zfyve26 in mice. Zfyve26 knockout mice do not show developmental defects but develop late-onset spastic paraplegia with cerebellar ataxia confirming that SPG15 is caused by ZFYVE26 deficiency. The morphological analysis reveals axon degeneration and progressive loss of both cortical motoneurons and Purkinje cells in the cerebellum. Importantly, neuron loss is preceded by accumulation of large intraneuronal deposits of membrane-surrounded material, which co-stains with the lysosomal marker Lamp1. A density gradient analysis of brain lysates shows an increase of Lamp1-positive membrane compartments with higher densities in Zfyve26 knockout mice. Increased levels of lysosomal enzymes in brains of aged knockout mice further support an alteration of the lysosomal compartment upon disruption of Zfyve26. We propose that SPG15 is caused by an endolysosomal membrane trafficking defect, which results in endolysosomal dysfunction. This appears to be particularly relevant in neurons with highly specialized neurites such as cortical motoneurons and Purkinje cells. [ABSTRACT FROM AUTHOR]

Courses of SARS-CoV-2 infections are highly variable, ranging from asymptomatic to lethal COVID-19. Though research has shown that host genetic factors contribute to this variability, cohort-based joint analyses of variants from the entire allelic spectrum in individuals with confirmed SARS-CoV-2 infections are still lacking. Here, we present the results of whole genome sequencing in 1,220 mainly vaccine-naïve individuals with confirmed SARS-CoV-2 infection, including 827 hospitalized COVID-19 cases. We observed the presence of autosomal-recessive or likely compound heterozygous monogenic disorders in six individuals, all of which were hospitalized and significantly younger than the rest of the cohort. We did not observe any suggestive causal variants in or around the established risk gene TLR7. Burden testing in the largest population subgroup (i.e., Europeans) suggested nominal enrichments of rare variants in coding and non-coding regions of interferon immune response genes in the overall analysis and male subgroup. Case-control analyses of more common variants confirmed associations with previously reported risk loci, with the key locus at 3p21 reaching genome-wide significance. Polygenic scores accurately captured risk in an age-dependent manner. By enabling joint analyses of different types of variation across the entire frequency spectrum, this data will continue to contribute to the elucidation of COVID-19 etiology., Competing Interests: JBR is CEO and founder of the company 5 Prime Sciences Inc, which offers analyses of human genetic data for the selection of drug targets. PK received grants or personal fees from Ambu GmbH, Gilead Sciences, Mundipharma Resarch Limited, Noxxon N.V. and Pfizer Pharma; he received honoraria for lectures from Akademie für Infektionsmedizin e.V., Ambu GmbH, Astellas Pharma, BioRad Laboratories Inc., Datamed GmbH, European Confederation of Medical Mycology, Gilead Sciences, GPR Academy Ruesselsheim, HELIOS Kliniken GmbH, Lahn-Dill-Kliniken GmbH, medupdate GmbH, MedMedia GmbH, MSD Sharp & Dohme GmbH, Pfizer Pharma GmbH, Scilink Comunicación Científica SC, streamedup! GmbH and University Hospital and LMU Munich and he participates in an Advisory Board from Ambu GmbH, Gilead Sciences, Mundipharma Resarch Limited and Pfizer Pharma. CDS received grants or personal fees from AstraZeneca, BBraun Melsungen, BioNtech, Gilead Sciences, Janssen-Cilag, Eli Lilly, Formycon, Pfizer, Roche, Apeiron, MSD, Cepheid, GSK, Molecular partners, SOBI, AbbVie, Synairgen, Shionogi and ViiV Healthcare. None of the funders did have any influence on study design or execution., (Copyright: © 2024 Schmidt et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Mutations in the genes encoding for voltage-gated sodium channels cause profound sensory disturbances and other symptoms dependent on the distribution of a particular channel subtype in different organs. Humans with the gain-of-function mutation p.Leu811Pro in SCN11A (encoding for the voltage-gated Nav1.9 channel) exhibit congenital insensitivity to pain, pruritus, self-inflicted injuries, slow healing wounds, muscle weakness, Charcot-like arthropathies, and intestinal dysmotility. As already shown, knock-in mice (Scn11a+/L799P) carrying the orthologous mutation p.Leu799Pro replicate reduced pain sensitivity and show frequent tissue lesions. In the present study we explored whether Scn11a+/L799P mice develop also pruritus, muscle weakness, and changes in gastrointestinal transit time. Furthermore, we analyzed morphological and functional differences in nerves, skeletal muscle, joints and small intestine from Scn11a+/L799P and Scn11a+/+ wild type mice. Compared to Scn11a+/+ mice, Scn11a+/L799P mice showed enhanced scratching bouts before skin lesions developed, indicating pruritus. Scn11a+/L799P mice exhibited reduced grip strength, but no disturbances in motor coordination. Skeletal muscle fiber types and joint architecture were unaltered in Scn11a+/L799P mice. Their gastrointestinal transit time was unaltered. The small intestine from Scn11a+/L799P showed a small shift towards less frequent peristaltic movements. Similar proportions of lumbar dorsal root ganglion neurons from Scn11a+/L799P and Scn11a+/+ mice were calcitonin gene-related peptide (CGRP-) positive, but isolated sciatic nerves from Scn11a+/L799P mice exhibited a significant reduction of the capsaicin-evoked release of CGRP indicating reduced neurogenic inflammation. These data indicate important Nav1.9 channel functions in several organs in both humans and mice. They support the pathophysiological relevance of increased basal activity of Nav1.9 channels for sensory abnormalities (pain and itch) and suggest resulting malfunctions of the motor system and of the gastrointestinal tract. Scn11a+/L799P mice are suitable to investigate the role of Nav1.9, and to explore the pathophysiological changes and mechanisms which develop as a consequence of Nav1.9 hyperactivity., Competing Interests: Matthias Ebbinghaus was funded by Charles River Discovery Research Services Germany GmbH after the experimental part of this study was finished. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Massive apoptosis of lymphocytes is a hallmark of sepsis. The resulting immunosuppression is associated with secondary infections, which are often lethal. Moreover, sepsis-survivors are burdened with increased morbidity and mortality for several years after the sepsis episode. The duration and clinical consequences of sepsis induced-immunosuppression are currently unknown. We have used the mouse model of peritoneal contamination and infection (PCI) to investigate the quantitative and qualitative recovery of T lymphocytes for 3.5 months after sepsis with or without IL-7 treatment. Thymic output and the numbers of naive and effector/memory CD4+ and CD8+ lymphocytes quickly recovered after sepsis. IL-7 treatment resulted in an accelerated recovery of CD8+ lymphocytes. Next generation sequencing revealed no significant narrowing of the T cell receptor repertoire 3.5 months after sepsis. In contrast, detailed functional analyses of T helper (Th)-cell responses towards a fungal antigen revealed a significant loss of Th cells. Whereas cytokine production was not impaired at the single cell level, the absolute number of Th cells specific for the fungal antigen was reduced. Our data indicate a clinically relevant loss of pathogen-specific T cell clones after sepsis. Given the small number of naive T lymphocytes specific for a given antigen, this decrement of T cell clones remains undetected even by sensitive methods such as deep sequencing. Taken together, our data are compatible with long lasting impairments in CD4+ T-cell responses after sepsis despite rapid recovery of T lymphocyte populations., Competing Interests: The authors have declared that no competing interests exist.