High ferritin is associated with liver and bone marrow iron accumulation: Effects of 1-year deferoxamine treatment in hemodialysis-associated iron overload.

BACKGROUND: Iron (Fe) supplementation is a critical component of anemia therapy for patients with chronic kidney disease (CKD). However, serum Fe, ferritin, and transferrin saturation, used to guide Fe replacement, are far from optimal, as they can be influenced by malnutrition and inflammation. Currently, there is a trend of increasing Fe supplementation to target high ferritin levels, although the long-term risk has been overlooked. METHODS: We prospectively enrolled 28 patients with CKD on hemodialysis with high serum ferritin (> 1000 ng/ml) and tested the effects of 1-year deferoxamine treatment, accompanied by withdrawal of Fe administration, on laboratory parameters (Fe status, inflammatory and CKD-MBD markers), heart, liver, and iliac crest Fe deposition (quantitative magnetic resonance imaging [MRI]), and bone biopsy (histomorphometry and counting of the number of Fe positive cells in the bone marrow). RESULTS: MRI parameters showed that none of the patients had heart iron overload, but they all presented iron overload in the liver and bone marrow, which was confirmed by bone histology. After therapy, ferritin levels decreased, although neither hemoglobin levels nor erythropoietin dose was changed. A significant decrease in hepcidin and FGF-23 levels was observed. Fe accumulation was improved in the liver and bone marrow, reaching normal values only in the bone marrow. No significant changes in turnover, mineralization or volume were observed. CONCLUSIONS: Our data suggest that treatment with deferoxamine was safe and could improve Fe accumulation, as measured by MRI and histomorphometry. Whether MRI is considered a standard tool for investigating bone marrow Fe accumulation requires further investigation. Registry and the registration number of clinical trial: ReBEC (Registro Brasileiro de Ensaios Clinicos) under the identification RBR-3rnskcj available at: https://ensaiosclinicos.gov.br/pesquisador.

Real-World Outcomes of the 0.19 mg Fluocinolone Acetonide Intravitreal Implant for Diabetic Macular Edema.

Purpose: To evaluate the real-world clinical and safety outcomes of a 0.19 mg fluocinolone acetonide intravitreal implant to treat diabetic macular edema (DME). Methods: This retrospective analysis comprised patients treated with a single fluocinolone acetonide intravitreal implant for the clinical indication of ME secondary to diabetic retinopathy. Primary outcomes included changes in best-corrected visual acuity (BCVA), central subfield thickness (CST), and the frequency of DME-related treatments 12 months before and up to 36 months after fluocinolone acetonide administration. Safety outcomes were also assessed. Results: One hundred forty-eight eyes (115 patients) were followed for a mean (±SD) of 12.3 ± 4.2 months before and 29.4 ± 14.5 months after fluocinolone acetonide administration. A 0.8-letter decrease (Early Treatment Diabetic Retinopathy Study) in the mean BCVA was observed at month 24. The BCVA was 70 letters (20/40 Snellen equivalent) or more in 20.6% of eyes at baseline and in 23.7% of eyes 24 months after implant administration. The mean CST was 379.9 µm and 323.7 µm, respectively. The CST was 300 µm or less in 58.7% of eyes at month 24 (P < .001). The mean frequency of intravitreal antivascular endothelial growth factor injections or laser photocoagulation decreased from 4.9 to 1.5 per year after fluocinolone acetonide administration (P < .001). Implant migration to the anterior chamber occurred in 3 eyes, 2 of which were vitrectomized and later required removal. Conclusions: The 0.19 mg fluocinolone acetonide intravitreal implant provided long-term stabilization of VA and macular anatomy in patients with DME, despite a significant reduction in treatment frequency.

Association of autoantibodies with the IFN signature and NETosis in patients with systemic lupus erythematosus.

Objective: Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by a variety of disease symptoms and an unpredictable clinical course. To improve treatment outcome, stratification based on immunological manifestations commonly seen in patients with SLE such as autoantibodies, type I interferon (IFN) signature and neutrophil extracellular trap (NET) release may help. It is assumed that there is an association between these immunological phenomena, since NET release induces IFN production and IFN induces autoantibody formation via B-cell activation. Here we studied the association between autoantibodies, the IFN signature, NET release, and clinical manifestations in patients with SLE. Methods: We performed principal component analysis (PCA) and hierarchical clustering of 57 SLE-related autoantibodies in 25 patients with SLE. We correlated each autoantibody to the IFN signature and NET inducing capacity. Results: We observed two distinct clusters: one cluster contained mostly patients with a high IFN signature. Patients in this cluster often present with cutaneous lupus, and have higher anti-dsDNA concentrations. Another cluster contained a mix of patients with a high and low IFN signature. Patients with high and low NET inducing capacity were equally distributed between the clusters. Variance between the clusters is mainly driven by antibodies against histones, RibP2, RibP0, EphB2, RibP1, PCNA, dsDNA, and nucleosome. In addition, we found a trend towards increased concentrations of autoantibodies against EphB2, RibP1, and RNP70 in patients with an IFN signature. We found a negative correlation of NET inducing capacity with anti-FcER (r = -0.530; p = 0.007) and anti-PmScl100 (r = -0.445; p = 0.03). Conclusion: We identified a subgroup of patients with an IFN signature that express increased concentrations of antibodies against DNA and RNA-binding proteins, which can be useful for further patient stratification and a more targeted therapy. We did not find positive associations between autoantibodies and NET inducing capacity. Our study further strengthens the evidence of a correlation between RNA-binding autoantibodies and the IFN signature.

A Genome-Wide Association Study Suggests New Susceptibility Loci for Primary Antiphospholipid Syndrome.

OBJECTIVE: Primary antiphospholipid syndrome (PAPS) is a rare autoimmune disease characterized by the presence of antiphospholipid antibodies and the occurrence of thrombotic events and pregnancy complications. Our study aimed to identify novel genetic susceptibility loci associated with PAPS. METHODS: We performed a genome-wide association study comprising 5,485 individuals (482 affected individuals) of European ancestry. Significant and suggestive independent variants from a meta-analysis of approximately 7 million variants were evaluated for functional and biological process enrichment. The genetic risk variability for PAPS in different populations was also assessed. Hierarchical clustering, Mahalanobis distance, and Dirichlet Process Mixtures with uncertainty clustering methods were used to assess genetic similarities between PAPS and other immune-mediated diseases. RESULTS: We revealed genetic associations with PAPS in a regulatory locus within the HLA class II region near HLA-DRA and in STAT1-STAT4 with a genome-wide level of significance; 34 additional suggestive genetic susceptibility loci for PAPS were also identified. The disease risk allele near HLA-DRA is associated with overexpression of HLA-DRB6, HLA-DRB9, HLA-DQA2, and HLA-DQB2 in immune cells, vascular tissue, and nervous tissue. This association is independent of the association between PAPS and HLA-DRB1*1302. Functional analyses highlighted immune-related pathways in PAPS-associated loci. The comparison with other immune-mediated diseases revealed a close genetic relatedness to neuromyelitis optica, systemic sclerosis, and Sjögren syndrome, suggesting co-localized causal variations close to STAT1-STAT4, TNPO3, and BLK. CONCLUSION: This study represents a comprehensive large-scale genetic analysis for PAPS and provides new insights into the genetic basis and pathophysiology of this rare disease.

Remote neurological evaluation reliably identifies patients eligible to endovascular therapy while non-eligible to intravenous thrombolysis.

INTRODUCTION/BACKGROUND: Early identification of suspected stroke patients who might be eligible for a reperfusion strategy is a daily challenge in the management of patient referrals. The aim of this study was to evaluate the performance of a remote medical assessment in identifying patients eligible for endovascular therapy (EVT) while not eligible for intravenous thrombolysis (IVT), compared with a decision based on bedside clinico-radiological data. METHODS: Patients admitted to the emergency department for acute neurological symptoms lasting for less than 24h were prospectively included. Assessment of the clinical severity and medical history was performed simultaneously by two vascular neurologists (VNs), one remotely using a mobile telemedicine solution (NOMADEEC), the other at the bedside. RACE score was calculated from the NIHSS score. At the end of the evaluation, both VNs quoted their treatment convictions (IVT/EVT). Final therapeutic decision following brain and vascular imaging was recorded and compared to remote and bedside predictions. The performances of three different conditions were evaluated: complete medical evaluation (NIHSS+medical history), NIHSS score alone, and RACE score alone. Remote and bedside performances were also compared. Diagnostic accuracy parameters (sensitivity, specificity, positive and negative predictive values) of each condition were estimated, along with their two-sided 95% binomial confidence intervals. RESULTS: Out of 215 enrolled patients, 186 had a complete evaluation, 91 (54.3%) were diagnosed with an ischemic stroke or transient ischemic attack and 46 (24.7%) had an intracranial occlusion. Considering the three conditions evaluated remotely, RACE score-based decision provided the best sensitivity 54.6% [95% CI 23.4; 83.2]/specificity 80.6% [73.9; 86.2] combination. However, the complete medical evaluation had the best specificity (88.6% [82.9; 92.9] compared to RACE scores alone (P=0.038). Remote and bedside performances did not differ (κ=0.68 [0.59; 0.77]). DISCUSSION/CONCLUSION: This real-life study performed in the setting of emergency demonstrates that remote medical evaluations including recording of extensive medical information and NIHSS examination to address patient's eligibility to revascularization treatment is swiftly feasible and is as effective as bedside prediction to EVT and/or IVT. Remote standardized medical evaluation might improve the decision of patients' primary orientation and avoid overcrowding of comprehensive stroke centres.

Expanding the Chemical Space of Electrophilic ß-Glycosyl ß-Lactams through Photoinduced Diastereoselective Functionalization.

Herein, we present a photoinduced diastereoselective C-3 functionalization of electrophilic ß-glycosyl ß-lactams. The developed protocol is simple, mild, and scalable and explores the use of 3-exomethylene ß-lactams as reaction partners in a Giese type reaction. The key nucleophilic alkyl radical is generated by a photoinduced electron transfer process in the EDA complex formed by NHPI and Hantzsch esters. The diastereoselective hydrogen atom transfer to the ß-lactam radical intermediate enables the synthesis of various N-phenyl ß-glycosyl ß-lactams.

The effect of temporal context on memory for art.

Although considerable research has been done on memory for temporal information, as well as on the relationship between context and cognition, not much is known about the influence of temporal context on memory formation and retention. In this study, given that our sample comes from a largely Roman Catholic population, we used religious practices that occur throughout the calendar year to operationalize temporal context into two religious seasons (Lent and Ordinary Time). In addition, we used religious art to assess experience and memory as a function of whether there was temporal congruity or incongruity. This allowed us to explore different levels of memory representation; namely, memory for perceptual details of the art, memory for more inferential understanding of the art, and autobiographical memory for the initial experience of the art. Participants viewed 22 representational and abstract artworks during either Lent or Ordinary Time. After viewing, memory was tested at immediate, 1-day, and 7-day delays. We expected that the congruent temporal context (i.e., Lent) would lead to more activated semantic knowledge, which would then aid memory encoding and retention. This was the case only for perceptual details of the art. In addition, during Lent, forgetting followed a more linear pattern. These results suggest that priming semantic knowledge through temporal context leads encoding to focus on low-level information, as opposed to the processing of more complex information. Overall, these findings suggest that temporal context can influence cognition, but to a limited extent.

miR-100 and miR-125b Contribute to Enhanced 3D Growth and Invasiveness and can be Functionally Transferred to Silence Target Genes in Recipient Cells.

Extracellular communication via the transfer of vesicles and nanoparticles is now recognized to play an important role in tumor microenvironment interactions. Cancer cells upregulate and secrete abundant levels of miR-100 and miR-125b that can alter gene expression by both cell- and non-cell-autonomous mechanisms. We previously showed that these miRNAs activate Wnt signaling in colorectal cancer (CRC) through noncanonical pairing with 5 negative regulators of Wnt signaling. To identify additional targets of miR-100 and miR-125b , we used bioinformatic approaches comparing multiple CRC cell lines, including knockout lines lacking one or both of these miRNAs. From an initial list of 96 potential mRNA targets, we tested 15 targets with 8 showing significant downregulation in the presence of miR-100 and miR-125b . Among these, Cingulin (CGN) and Protein tyrosine phosphatase receptor type-R (PTPRR) are downregulated in multiple cancers, consistent with regulation by increased levels of miR-100 and miR-125b. We also show that increased cellular levels of miR-100 and miR-125b enhance 3D growth and invasiveness in CRC and glioblastoma cell lines. Lastly, we demonstrate that extracellular transfer of miR-100 and miR-125b can silence both reporter and endogenous mRNA targets in recipient cells and also increase the invasiveness of recipient spheroid colonies when grown under 3D conditions in type I collagen.

Satellite-enabled enviromics to enhance crop improvement.

Enviromics refers to the characterization of micro- and macroenvironments based on large-scale environmental datasets. By providing genotypic recommendations with predictive extrapolation at a site-specific level, enviromics could inform plant breeding decisions across varying conditions and anticipate productivity in a changing climate. Enviromics-based integration of statistics, envirotyping (i.e., determining environmental factors), and remote sensing could help unravel the complex interplay of genetics, environment, and management. To support this goal, exhaustive envirotyping to generate precise environmental profiles would significantly improve predictions of genotype performance and genetic gain in crops. Already, informatics management platforms aggregate diverse environmental datasets obtained using optical, thermal, radar, and light detection and ranging (LiDAR)sensors that capture detailed information about vegetation, surface structure, and terrain. This wealth of information, coupled with freely available climate data, fuels innovative enviromics research. While enviromics holds immense potential for breeding, a few obstacles remain, such as the need for (1) integrative methodologies to systematically collect field data to scale and expand observations across the landscape with satellite data; (2) state-of-the-art AI models for data integration, simulation, and prediction; (3) cyberinfrastructure for processing big data across scales and providing seamless interfaces to deliver forecasts to stakeholders; and (4) collaboration and data sharing among farmers, breeders, physiologists, geoinformatics experts, and programmers across research institutions. Overcoming these challenges is essential for leveraging the full potential of big data captured by satellites to transform 21st century agriculture and crop improvement through enviromics.

Differences in DNA methylation status explain phenotypic variability in patients with 5p- syndrome.

Cri Du Chat syndrome, or 5p- syndrome, is characterized by a terminal or interstitial deletion on the short arm of chromosome 5 that causes variable clinical manifestations, including high-pitched cry in newborns, delayed growth, and global development. Different cytogenomic rearrangements, family history, and environmental factors may hinder the genotype-phenotype association. Thus, the phenotypic variability of this syndrome may not be limited only to variations in gene structure, such as deletions and duplications. It is possible that other mechanisms related to the activation or inactivation of promoters and/or exons of actively transcribed genes, such as DNA methylation are involved. Therefore, we studied the genome-wide methylation status profile of peripheral blood samples from fifteen patients with Cri du Chat Syndrome and nine control samples through the array method to look for Differentially Methylated Regions. We found that Differentially Methylated Regions outside the 5p region are mainly associated with regulating gene transcription, splicing, and chromatin remodeling. Most biological pathways are related to transcription, histone and chromatin binding, spliceosome and ribosomal complex, and RNA processing. Our results suggest that changes in the 5p region can cause an imbalance in other chromosomal regions capable of affecting gene modulation and thus explain the phenotypic differences in patients with 5p-.

Patterns of concomitant traumatic brain injury and ocular trauma in US service members.

Background: Concomitant traumatic brain injury (TBI) and ocular trauma (OT) are caused by the same physical mechanisms, which may complicate therapeutic intervention if screening and evaluation of each condition are not promptly initiated. The aim of this study is to identify concomitant TBI in OT patients and characterize the pattern of those injured service members (SMs) in non-combat environments to assist in the early detection and treatment of both TBI and OT. Methods: Encounters matching the case definitions of TBI and OT for injured SMs were extracted from the Military Health System. Concomitant TBI and OT was identified as patients who were diagnosed with both medical conditions within 30 days. Incidence rates of concomitance were analyzed using a Poisson regression model. The odds of mechanisms and types of OT with concomitant TBI were analyzed using logistic regression models. Results: From 2017 to 2021, there were 71 689 SMs diagnosed with TBI, and 69 358 patients diagnosed with OT. There were 3251 concomitant cases identified. The overall concomitance rate in OT patients was 4.7%. Clinical presentations of concomitant OT had a higher rate of complications. Blast, transport accidents, assaults, alcohol, falls, and sports-related injuries (in decreasing order) were significantly associated with concomitance rates. Compared with closed globe injuries, OT with orbital fractures, rupture, laceration, adnexal periocular injury, and penetrating injury had higher risks of concomitant TBI. For patients with orbital fractures, nearly half (44.1%) sustained a concomitant TBI. Conclusions: A practical approach using temporal proximity of diagnostic data was developed to identify concomitant cases of TBI and OT which presented with more severe injury types than non-concomitant cases. These results indicate OT patients with orbital or open globe injuries sustained from high-impact mechanisms warrant further TBI screening to prompt early detection and treatment. Level of evidence: IV.

Evaluating antibiotic occurrence, degradation, and environmental risks in poultry litter within Argentina's agricultural hub.

This study explores the relationship between poultry farming's antibiotic administration practices and residual antibiotic levels in the litter before its application onto agricultural soils. Twenty-three antibiotics were performed across 19 Argentinean farms representing diverse antibiotic management practices. Analysis revealed up to 20 antibiotics from eight chemical classes in poultry litter samples, with tylosin, enrofloxacin, and salinomycin being the most relevant drugs. Farms with restricted antibiotic use in feed exhibited lower residual concentrations. A self-heating treatment was tested to reduce litter antibiotic levels. Although a 60 % reduction of antibiotics was found after treatment, prevalent compounds persisted at residual levels. Regulatory measures and comprehensive litter treatments pre-application are crucial to mitigate environmental risks. This is the first study that provides insight on the occurrence of >20 drugs in real poultry production scenarios from Latin America and demonstrates how relatively simple treatments can be readily applied to decrease the associated environmental risks.

Agreement between Handheld and Standard Echocardiography for Diagnosis of Latent Rheumatic Heart Disease in Brazilian Schoolchildren from High-Prevalence Settings (Agreement between Screening and Standard Echo for RHD).

Introduction: Handheld echocardiography (echo) is the tool of choice for rheumatic heart disease (RHD) screening. We aimed to assess the agreement between screening and standard echo for latent RHD diagnosis in schoolchildren from an endemic setting. Methods: Over 14 months, 3 nonphysicians used handheld machines and the 2012 WHF Criteria to determine RHD prevalence in consented schoolchildren from Brazilian low-income public schools. Studies were interpreted by telemedicine by 3 experts (Brazil, US). RHD-positive children (borderline/definite) and those with congenital heart disease (CHD) were referred for standard echo, acquired and interpreted by a cardiologist. Agreement between screening and standard echo, by WHF subgroups, was assessed. Results: 1390 students were screened in 6 schools, with 110 (7.9%, 95% CI 6.5-9.5) being screen positive (14 ± 2 years, 72% women). Among 16 cases initially diagnosed as definite RHD, 11 (69%) were confirmed, 4 (25%) reclassified to borderline, and 1 to normal. Among 79 cases flagged as borderline RHD, 19 (24%) were confirmed, 50 (63%) reclassified to normal, 8 (10%) reclassified as definite RHD, and 2 had mild CHD. Considering the 4 diagnostic categories, kappa was 0.18. In patients with borderline RHD reclassified to non-RHD, the most frequent WHF criterion was B (isolated mitral regurgitation, 64%), followed by A (2 mitral valve morphological features, 31%). In 1 patient with definite RHD reclassified to normal, the WHF criterion was D (borderline RHD in aortic and mitral valves). After standard echo, RHD prevalence was 3.2% (95% CI 2.3-4.2). Conclusions: Although practical, RHD screening with handheld devices tends to overestimate prevalence.

The Natural Products Discovery Center: Release of the First 8490 Sequenced Strains for Exploring Actinobacteria Biosynthetic Diversity.

Actinobacteria, the bacterial phylum most renowned for natural product discovery, has been established as a valuable source for drug discovery and biotechnology but is underrepresented within accessible genome and strain collections. Herein, we introduce the Natural Products Discovery Center (NPDC), featuring 122,449 strains assembled over eight decades, the genomes of the first 8490 NPDC strains (7142 Actinobacteria), and the online NPDC Portal making both strains and genomes publicly available. A comparative survey of RefSeq and NPDC Actinobacteria highlights the taxonomic and biosynthetic diversity within the NPDC collection, including three new genera, hundreds of new species, and ~7000 new gene cluster families. Selected examples demonstrate how the NPDC Portal's strain metadata, genomes, and biosynthetic gene clusters can be leveraged using genome mining approaches. Our findings underscore the ongoing significance of Actinobacteria in natural product discovery, and the NPDC serves as an unparalleled resource for both Actinobacteria strains and genomes.

Visible light-driven CO2 photoreduction by a Re(I) complex immobilized onto CuO/Nb2O5 heterojunctions.

The immobilization of Re(I) complexes onto metal oxide surfaces presents an elegant strategy to enhance their stability and reusability toward photocatalytic CO2 reduction. In this study, the photocatalytic performance of fac-[ClRe(CO)3(dcbH2)], where dcbH2 = 4,4'-dicarboxylic acid-2,2'-bipyridine, anchored onto the surface of 1%m/m CuO/Nb2O5 was investigated. Following adsorption, the turnover number for CO production (TONCO) in DMF/TEOA increased significantly, from ten in solution to 370 under visible light irradiation, surpassing the TONCO observed for the complex onto pristine Nb2O5 or CuO surfaces. The CuO/Nb2O5 heterostructure allows for efficient electron injection by the Re(I) center, promoting efficient charge separation. At same time CuO clusters introduce a new absorption band above 550 nm that contributes for the photoreduction of the reaction intermediates, leading to a more efficient CO evolution and minimization of side reactions.

Development of platensimycin, platencin, and platensilin overproducers by biosynthetic pathway engineering and fermentation medium optimization.

The platensimycin (PTM), platencin (PTN), and platensilin (PTL) family of natural products continues to inspire the discovery of new chemistry, enzymology, and medicine. Engineered production of this emerging family of natural products, however, remains laborious due to the lack of practical systems to manipulate their biosynthesis in the native-producing Streptomyces platensis species. Here we report solving this technology gap by implementing a CRISPR-Cas9 system in S. platensis CB00739 to develop an expedient method to manipulate the PTM, PTN, and PTL biosynthetic machinery in vivo. We showcase the utility of this technology by constructing designer recombinant strains S. platensis SB12051, SB12052, and SB12053, which, upon fermentation in the optimized PTM-MS medium, produced PTM, PTN, and PTL with the highest titers at 836 mg L-1, 791 mg L-1, and 40 mg L-1, respectively. Comparative analysis of these resultant recombinant strains also revealed distinct chemistries, catalyzed by PtmT1 and PtmT3, two diterpene synthases that nature has evolved for PTM, PTN, and PTL biosynthesis. The ΔptmR1/ΔptmT1/ΔptmT3 triple mutant strain S. platensis SB12054 could be envisaged as a platform strain to engineer diterpenoid biosynthesis by introducing varying ent-copalyl diphosphate-acting diterpene synthases, taking advantage of its clean metabolite background, ability to support diterpene biosynthesis in high titers, and the promiscuous tailoring biosynthetic machinery. ONE-SENTENCE SUMMARY: Implementation of a CRISPR-Cas9 system in Streptomyces platensis CB00739 enabled the construction of a suite of designer recombinant strains for the overproduction of platensimycin, platencin, and platensilin, discovery of new diterpene synthase chemistries, and development of platform strains for future diterpenoid biosynthesis engineering.

Kv1.3 blockade by ShK186 modulates CD4+ effector memory T-cell activity of patients with granulomatosis with polyangiitis.

OBJECTIVES: Granulomatosis with polyangiitis (GPA) is a chronic relapsing systemic autoimmune vasculitis. Current treatment of GPA is unsatisfactory, as it relies on strong immunosuppressive regimens, with either CYC or rituximab, which reduce the immunogenicity of several vaccines and are risk factors for a severe form of COVID-19. This emphasizes the need to identify new drug targets and to develop treatment strategies with less harmful side effects. Since CD4+ effector memory T cells (TEM) play a key role in the pathogenesis of GPA, we aimed in this study to modulate CD4+TEM cell activity via Kv1.3 blockade using the specific peptide inhibiter, ShK-186. METHODS: Peripheral blood samples from 27 patients with GPA in remission and 16 age- and sex-matched healthy controls (HCs) were pre-incubated in vitro in the presence or absence of ShK-186, followed by stimulation with phorbol myristate acetate, calcium ionophore and brefeldin-A. The effect of ShK-186 on the cytokine production (IFNγ, TNFα, IL-4, IL-17, IL-21) within total and subsets of CD4+ T helper (CD4+TH) cells were assessed using flow cytometry. RESULTS: ShK-186 reduced the expression level of IFNγ, TNFα, IL-4, IL-17 and IL-21 in CD4+TH cells from patients with GPA in vitro. Further analysis performed on sorted CD4+T cell subsets, revealed that ShK-186 predominantly inhibited the cytokine production of CD4+TEM cells. ShK-186 treatment reduced the production of the pro-inflammatory cytokines to the level seen in CD4+ TH cells from HCs. CONCLUSIONS: Modulation of cellular effector function by ShK-186 may constitute a novel treatment strategy for GPA with high specificity and less harmful side effects.

Epilepsy-linked kinase CDKL5 phosphorylates voltage-gated calcium channel Cav2.3, altering inactivation kinetics and neuronal excitability.

Developmental and epileptic encephalopathies (DEEs) are a group of rare childhood disorders characterized by severe epilepsy and cognitive deficits. Numerous DEE genes have been discovered thanks to advances in genomic diagnosis, yet putative molecular links between these disorders are unknown. CDKL5 deficiency disorder (CDD, DEE2), one of the most common genetic epilepsies, is caused by loss-of-function mutations in the brain-enriched kinase CDKL5. To elucidate CDKL5 function, we looked for CDKL5 substrates using a SILAC-based phosphoproteomic screen. We identified the voltage-gated Ca2+ channel Cav2.3 (encoded by CACNA1E) as a physiological target of CDKL5 in mice and humans. Recombinant channel electrophysiology and interdisciplinary characterization of Cav2.3 phosphomutant mice revealed that loss of Cav2.3 phosphorylation leads to channel gain-of-function via slower inactivation and enhanced cholinergic stimulation, resulting in increased neuronal excitability. Our results thus show that CDD is partly a channelopathy. The properties of unphosphorylated Cav2.3 closely resemble those described for CACNA1E gain-of-function mutations causing DEE69, a disorder sharing clinical features with CDD. We show that these two single-gene diseases are mechanistically related and could be ameliorated with Cav2.3 inhibitors.