Gain Of Function Research Articles

CTCF-dependent insulation of Hoxb13 and the heterochronic control of tail length

Mammalian tail length is controlled by several genetic determinants, among which are Hox13 genes, whose function is to terminate the body axis. Accordingly, the precise timing in the transcriptional activation of these genes may impact upon body length. Unlike other Hox clusters, HoxB lacks posterior genes between Hoxb9 and Hoxb13, two genes separated by a ca. 70 kb large DNA segment containing a high number of CTCF sites, potentially isolating Hoxb13 from the rest of the cluster and thereby delaying its negative impact on trunk extension. We deleted the spacer DNA to induce a potential heterochronic gain of function of Hoxb13 at physiological concentration and observed a shortening of the tail as well as other abnormal phenotypes. These defects were all rescued by inactivating Hoxb13 in-cis with the deletion. A comparable gain of function was observed in mutant Embryonic Stem (ES) cells grown as pseudoembryos in vitro, which allowed us to examine in detail the importance of both the number and the orientation of CTCF sites in the insulating activity of the DNA spacer. A short cassette containing all the CTCF sites was sufficient to insulate Hoxb13 from the rest of HoxB, and additional modifications of this CTCF cassette showed that two CTCF sites in convergent orientations were already capable of importantly delaying Hoxb13 activation in these conditions. We discuss the relative importance of genomic distance versus number and orientation of CTCF sites in preventing Hoxb13 to be activated too early during trunk extension and hence to modulate tail length.

The outcome of haematopoietic stem cell transplantation in a patient with STAT1 gain-of-function: a case report

Background: The human signal transducer and activator of transcription 1 (STAT1) is a latent cytoplasmic transcription factor and one of seven members of the STAT family. Autosomal dominant (AD) STAT1 gain-of-function (GOF), characterized by enhanced phosphorylation of the tyrosine-701 residue and STAT1 activation, is commonly associated with chronic mucocutaneous candidiasis (CMCC), immunodeficiency, aneurysms, malignancies, and autoimmune phenomena. The best management strategies for patients with STAT1 GOF remain unclear. Typically, the standard care includes supportive treatments such as antimicrobial prophylaxis, either alone or combined with immunoglobulin replacement therapy. Biological therapies such as JAK inhibitors have been shown to alleviate symptoms of infection and autoimmune disorders in patients with STAT1 GOF mutations. However, there is a lack of long-term outcome data for JAK inhibition. Hematopoietic stem cell transplantation (HSCT) is an alternative treatment option for a subset who experience a persistent disease course despite conventional therapy. However, the effectiveness of HSCT for this condition is not yet well established. Methods: Our patient’s medical records were analyzed retrospectively, including her medical history. Results: We present the outcome of HSCT performed on a 27-year-old female with STAT1 GOF, conducted as a treatment for acute myeloid leukemia (AML). Conclusion: HSCT may serve as an alternative and potentially curative treatment for certain STAT1 GOF patients with progressive, life-threatening conditions that do not respond to conventional therapies. Additional research is needed to improve the management of these patients. Statement of Novelty: We present a novel case study of HSCT as a treatment for AML in a 27-year-old patient with STAT1 GOF, highlighting the potential for curative outcomes in progressive, life-threatening conditions unresponsive to conventional therapies. This case contributes to the limited body of evidence supporting the efficacy and challenges of HSCT in STAT1 GOF patients.

OA25 Not so “savy” afterall

Abstract Introduction Interferonopathies (IFN’s) are a constellation of diseases that cause recurrent pathogenic inflammation, arising primarily through antigen-independent hyperactivation of immune pathways¹. It is primarily due to abnormal production or signalling of type I IFNs². A disorder of both the innate and adaptive immune system, this condition is relatively new with the term only coined in 2011. Currently there are more than 20 interferonopathies recognized, with new interferonopathies being identified like Sting associated vasculopathy with onset in infancy (SAVI). Case description Our patient first presented with a severe panniculitis rash, Raynaud’s, swelling of her limbs and hepatosplenomegaly at five weeks of age. She developed significant respiratory distress requiring non-invasive ventilation. A CT Thorax revealed diffuse groundglass opacification suggestive of interstitial lung disease. She had elevated inflammatory markers (CRP 24 mg/L, ferritin 1448 ng/ml, ESR 88 mm/hr), a transaminitis (ALT 193 U/ml, AST 356 U/L). She had multiple positive auto antibodies (Table 1) Clinically SAVI was suspected as the most likely diagnosis. Her interferon signature gene assay demonstrated exaggerated IFI 27 and IFI44L levels. A Trio exome sequencing and IKBKG sequencing however have not been indicative of any currently recognised interferonopathy. Further genetic analysis has been requested to explore possible novel pathogenic mutations. A reasonable clinical response was seen with corticosteroid therapy in combination with Jak Inhibition. With worsening of her cutaneous disease and an increasing need for corticosteroid use a trial of anti IL-6 therapy was initiated. This was subsequently discontinued when she presented with severe acute digital ischemia. It was managed with escalating doses of ruxolitinib, pulse high dose corticosteroids and epoprostenol infusion. Her interferon signature gene assay demonstrated exaggerated IFI 27 and IFI44L levels. A Trio exome sequencing and Inhibitor of nuclear factor kappa B gene (IKBKG) sequencing however have not been indicative of any currently recognised interferonopathy. Further genetic analysis has been requested to explore possible novel pathogenic mutations. A reasonable clinical response was seen with corticosteroid therapy in combination with Jak Inhibition. With worsening of her cutaneous disease and an increasing need for corticosteroid use a trial of anti IL-6 therapy was initiated. This was subsequently discontinued when she presented with severe acute digital ischemia. It was managed with escalating doses of ruxolitinib, pulse high dose corticosteroids and epoprostenol infusion. Discussion This case enhances our current understanding of interferonopathies. Although the phenotype and course of the disease is compatible with SAVI³, genetic testing is negative to date. Presently there are eleven heterozygous gain-of-function (GoF) variants for SAVI4. With JAK inhibition as the most successful therapeutic option to date5. Our case possibly represents an as yet unreported genetic variant, further studies are awaited.This case enhances our current understanding of interferonopathies. Although the phenotype and course of the disease is compatible with SAVI³, genetic testing is negative to date. Presently there are eleven heterozygous gain-of-function (GoF) variants for SAVI4. With JAK inhibition as the most successful therapeutic option to date5. Our case possibly represents an as yet unreported genetic variant, further studies are awaited.This case enhances our current understanding of interferonopathies. Although the phenotype and course of the disease is compatible with SAVI³, genetic testing is negative to date. Presently there are eleven heterozygous gain-of-function (GoF) variants for SAVI4. With JAK inhibition as the most successful therapeutic option to date5. Our case possibly represents an as yet unreported genetic variant, further studies are awaited. Key learning points • Early suspicion for an interferonopathy in very young infants with interstitial lung disease • Expanding the potentially new genetic variants of this condition • Challenge of treatment management

Polo-like kinase 2 targeting as novel strategy to sensitize mutant p53-expressing tumor cells to anticancer treatments.

Polo-like kinase 2 (Plk2) belongs to a family of serine/threonine kinases, and it is involved in tumorigenesis of diverse kind of tissues. We previously reported that Plk2 gene was a transcriptional target of the mutant p53/NF-Y oncogenic complex. Plk2 protein can bind to and phosphorylate mutant p53 triggering an oncogenic autoregulatory feedback loop involved in cancer cell proliferation and chemoresistance. In this study, we aimed to assess whether the specific inhibition of Plk2 kinase activity by the selective TC-S 7005 inhibitor could decrease cell proliferation and migration inhibiting mutant p53 phosphorylation, thus disarming its oncogenic potential. We found that the Plk2 inhibitor treatment sensitized the cells to the irradiation and chemotherapy drugs, thereby overcoming the mutant p53-dependent chemoresistance. Taken together, we provided results that Plk2 could be considered a tractable pharmacological target for cancers expressing mutant p53 proteins. The combined treatment with conventional chemotherapeutic drugs and Plk2 inhibitors may represent a new candidate intervention approach, which may be considered for improving tumor cell sensitivity to DNA damaging drugs. KEY MESSAGES : Missense mutations are present in the TP53 gene in about half of all human cancers and correlate with poor patient outcome. Mutant p53 proteins exert gain of function (GOF) activities in tumor cells such as increased proliferation, genomic instability and resistance to therapies. Polo-like kinase 2 (PLK2) binds and phosphorylates mutant p53 protein strengthening its GOF activities. Pharmacologically targeting PLK2 weakens mutant p53 proteins and sensitizes tumor cells to therapeutic treatments.

Integrating single-cell sequencing and genetic lineage tracing reveals endothelial plasticity during atrial remodeling

Abstract Background Atrial fibrillation (AF), as the most burdensome cardiac arrhythmia, could lead to the significant mortality and morbidity. Atrial fibrosis is a well-recognized pathophysiological factor of the development and progression of AF, which also hampers its treatment. However, the underlying mechanisms of this intricate process remain largely unknown. Purpose This study sought to reveal the cellular composition of AF substrate and investigate the vital contributors to atrial structural remodeling. Methods Left atrial appendages from three patients with AF undergoing surgical ablation and three individuals with sinus rhythm undergoing heart transplantation were subjected to 10X single-cell RNA sequencing (scRNA-seq). Another five public scRNA-seq or snRNA-seq datasets of potentially remodeled right or left appendages were retrieved across three publications. The vital contributors of atrial structural remodeling and their characteristics were identified by intercellular communication analysis and trajectory inference. Transverse aortic constriction (TAC) was used to induce atrial remodeling in Cdh5-CreERT2;RFP mice, followed by scRNA-seq of endothelial cells (ECs). Human primary atrial ECs and fibroblasts were isolated for cell-cell interaction experiments. ECs-specific mouse models of Transforming growth factor beta 1 (TGFB1) (knockout and overexpression) were generated. Electrophysiological and histology analyses were performed to determine the consequences of ECs-derived TGFB1 gain and loss of function in the atrium. Results The comprehensive analysis of different scRNA-seq datasets revealed that ECs played an important regulatory role in the homeostasis of atrial substrate and displayed remarkable mesenchymal activation during atrial remodeling, which was further confirmed by TAC mice. Immunofluorescence staining and high-content screening suggested that the complete transition from ECs to mesenchymal cells slightly contributed to atrial remodeling. However, TGFB1 secreted from mesenchymal activated ECs significantly promoted activation, proliferation, and collagen secretion of fibroblasts. We discovered that overexpression of TGFB1 in ECs of mice significantly increased atrial fibrosis, and thus prolonging AF duration. Besides, knockout of TGFB1 in ECs of mice underwent TAC significantly attenuated the atrial fibrosis and decreased the rate of AF. Conclusion Our work demonstrates that mesenchymal ECs-derived Tgfb1 plays an important role in atrial remodeling by regulating the fibroblast function. EC-specific interventions were suggested to facilitate the development of novel strategies for mitigating AF substrate remodeling.

Foxi1 regulates multiple steps of mucociliary development and ionocyte specification through transcriptional and epigenetic mechanisms.

Foxi1 is a master regulator of ionocytes (ISCs / INCs) across species and organs. Two subtypes of ISCs exist, and both α- and β-ISCs regulate pH- and ion-homeostasis in epithelia. Gain and loss of FOXI1 function are associated with human diseases, including Pendred syndrome, male infertility, renal acidosis and cancers. Foxi1 functions were predominantly studied in the context of ISC specification, however, reports indicate additional functions in early and ectodermal development. Here, we re-investigated the functions of Foxi1 in Xenopus laevis embryonic mucociliary epidermis development and found a novel function for Foxi1 in the generation of Notch-ligand expressing mucociliary multipotent progenitors (MPPs). We demonstrate that Foxi1 has multiple concentration-dependent functions: At low levels, Foxi1 confers ectodermal competence through transcriptional and epigenetic mechanisms, while at high levels, Foxi1 induces a multi-step process of ISC specification and differentiation. We further describe how foxi1 expression is affected through auto- and Notch-regulation, how Ubp1 and Dmrt2 regulate ISC subtype differentiation, and how this developmental program affects Notch signaling as well as mucociliary patterning. Together, we reveal novel functions for Foxi1 in Xenopus mucociliary epidermis formation, relevant to our understanding of vertebrate development and human disease.

Over-the-Counter Hearing Aids Versus Traditional Hearing Aids in Patients With Mild-to-Moderate Hearing Loss: Protocol for a Noninferiority Randomized Controlled Trial.

With the aging of society, the prevalence of hearing loss (HL) is increasing. Currently, approximately 5% of the global population has HL, and this number is projected to reach 7 million by 2050. Although hearing aids (HAs) are the primary treatment for HL, their use is limited by barriers such as high costs and social stigma. To address these limitations, over-the-counter (OTC) HAs have been introduced, but their effectiveness and drawbacks require further investigation. This study aims to conduct a noninferiority randomized controlled trial comparing OTC HAs with traditional HAs to assess the clinical effectiveness of OTC HAs. We designed a noninferiority randomized controlled trial comparing OTC HAs and traditional HAs in adults with mild-to-moderate HL. A total of 64 participants (32 per group) will be recruited. Randomization will be performed using block randomization (block sizes of 2 or 4) with an equal allocation ratio. The study will include 2 types of HAs: an OTC HA (Jabra Enhance Pro) and a traditional HA (LiNX Quattro LE561-DRW) by GN ReSound A/S. OTC HAs will be self-fitted using a smartphone app, while traditional HAs will be fitted by a licensed audiologist using the National Acoustics Laboratories-Non-Linear Prescription, second generation. Assessments, including functional gain, real-ear measurement, speech audiometry, and questionnaires, will be conducted at 6-month intervals over the course of 3 visits. Statistical analysis will compare the 2 outcomes, focusing on functional gain, to determine noninferiority. This study is scheduled to begin in August 2024 and has not yet recruited any participants. The study will be conducted over 2 years, from August 2024 to July 2026. Each participant will have 2 follow-up visits at 6-month intervals, making the total follow-up period 1 year. Since 2022, the introduction of OTC HAs has revolutionized access to these devices. Researchers, clinicians, and the general public are keen to evaluate the clinical effectiveness of OTC HAs, as more individuals will likely use them for HL. This increased usage will provide valuable real-world data to understand the benefits and limitations of OTC HAs. Monitoring the outcomes and user feedback will provide insights into their effectiveness and impact on hearing rehabilitation. PRR1-10.2196/59894.

Discovery of Potent and Selective Blockers Targeting the Epilepsy-Associated KNa1.1 Channel.

Gain-of-function (GOF) mutations of the sodium-activated potassium channel KNa1.1 (Slack, Slo2.2, or KCa4.1) induce severe, drug-resistant forms of epilepsy in infants and children. Although quinidine has shown promise in treating KCNT1-related epilepsies compared to other drugs, its limited efficacy and substantial side effects necessitate the development of new KNa1.1 channel inhibitors. In this study, we developed a novel class of KNa1.1 inhibitors using combined silico approaches and structural optimization. Among these inhibitors, compound Z05 was identified as a selective potential KNa1.1 inhibitor, especially against the hERG channel. Moreover, its binding site and potential counteraction to a GOF mutant Y796H were identified by the mutation studies. Our data also showed that Z05 had significant pharmacological profiles, including high brain penetration and moderate oral bioavailability, offering a valuable in vitro tool compound for further drug development in treating KCNT1-related epilepsies.

MiR-30c-5p Gain and Loss of Function Modulate Sciatic Nerve Injury-Induced Nucleolar Stress Response in Dorsal Root Ganglia Neurons

miR-30c-5p Gain and Loss of Function Modulate Sciatic Nerve Injury-Induced Nucleolar Stress Response in Dorsal Root Ganglia Neurons

Comparison Between Total Hip and Knee Arthroplasty on Short-Term Performance-Based Outcomes and Factors Associated with the Improvement of Gait Function During Post-Acute Inpatient Rehabilitation

Background: Data on the differences in functional recovery between inpatient rehabilitation for total hip arthroplasty (THA) and total knee arthroplasty (TKA) are lacking, and the factors influencing the improvement of short-term functional mobility remain unknown. In this study, we compared the short-term functional outcomes of both procedures and identified early postoperative predictors of physical function gain during post-acute rehabilitation. Methods: A total of 435 patients who underwent THA and TKA were included. The main outcomes were knee extension strength, the motor component of the Functional Independence Measure, Numerical Rating Scale, 10 Meter Walk Test, Timed Up and Go (TUG) test, and the Berg Balance Scale. The recovery process and rehabilitation outcomes were compared between patients with THA and TKA. Additionally, predictors related to physical performance improvement were examined for each procedure. Results: Patients with THA and TKA achieved significant short-term functional recovery after multidisciplinary rehabilitation (time; p < 0.001). However, the pain score was higher at discharge in patients with TKA (p < 0.001). Age (β: −0.264, p = 0.009) and TUG test (β: −0.884, p < 0.001) in THA, and non-operated knee extension strength (β: 0.234, p = 0.016) and TUG test (β: −0.783, p < 0.001) in TKA were significant early postoperative predictors of functional mobility. Conclusions: Multidisciplinary rehabilitation was beneficial for functional improvement in patients with THA and TKA despite persistent pain at discharge after TKA. Baseline functional levels in both groups and non-operated knee extension strength in TKA can be useful performance-based predictors of short-term gait function improvement.

Two-faced OsNAS3 influences disease resistance via nicotianamine and ethylene.

The loss and gain of OsNAS3 function both positively influence plant disease resistance. Overexpression of OsNAS3 boosts blast resistance by promoting nicotianamine accumulation, thereby enhancing blast resistance. Conversely, knockout of OsNAS3 increases ethylene biosynthesis, also contributing to improved blast resistance.

The Inhibitory Effects of a Factor B-Binding DNA Aptamer Family Supersede the Gain of Function of Factor B Variants Associated with Atypical Hemolytic Uremic Syndrome.

Aptamers are short, single-stranded oligonucleotides that selectively bind to target biomolecules. Although they generally exhibit good binding specificity, their affinities are often limited because of the relative lack of hydrophobic groups in nucleic acids. Chemically modified nucleotides incorporating hydrophobic structures into uracil have been synthesized to address this obstacle. Modified DNA aptamers containing such nonstandard nucleotides have been developed for >20 different complement proteins. These modified aptamers show increased affinity and enhanced serum stability and have potential value as therapeutic agents. We recently conducted a structure/function study on a family of modified DNA aptamers that bind specifically to complement Factor B (FB). This work revealed that these aptamers selectively inhibit the complement alternative pathway (AP) by preventing the formation of the AP complement component C3 (C3) proconvertase complex, C3bB. Certain patients with atypical hemolytic uremic syndrome express gain-of-function variants of FB that enhance the formation of the proconvertase complex and/or decrease the efficacy of endogenous regulators against the C3 convertases they form. To investigate whether these FB-binding aptamers could override the effects of disease-causing mutations in FB, we examined how they interacted with several FB variants, including D279G, F286L, K323E, and K350N, in various assays of complement function. We found that the inhibitory effect of the FB-binding aptamers superseded the gain-of-function mutations in FB, although the aptamers could not dissociate preformed C3 convertases. These findings suggest that FB-binding aptamers could be further developed as a potential treatment for certain atypical hemolytic uremic syndrome patients or those with other diseases characterized by excessive complement activity.

Recent Progress of Antisense Oligonucleotide Therapy for Superoxide-Dismutase-1-Mutated Amyotrophic Lateral Sclerosis: Focus on Tofersen.

Amyotrophic lateral sclerosis (ALS) is a refractory neurodegenerative disease characterized by the degeneration and loss of motor neurons, typically resulting in death within five years of onset. There have been few effective treatments, making the development of robust therapies an urgent challenge. Genetic mutations have been identified as contributors to ALS, with mutations in superoxide dismutase 1 (SOD1), which neutralizes the harmful reactive oxygen species superoxide, accounting for approximately 2% of all ALS cases. To counteract the toxic gain of function caused by SOD1 mutations, therapeutic strategies aimed at suppressing SOD1 gene expression have shown promise. Antisense oligonucleotide (ASO) is an artificially synthesized, short, single-stranded DNA/RNA molecule that binds to target RNA to alter gene expression, representing a next-generation therapeutic approach. In 2023, tofersen became the first ASO drug approved by the FDA for ALS. Administered intrathecally, tofersen specifically binds to SOD1 mRNA, inhibiting the production of toxic SOD1 protein, thereby improving biomarkers of ALS. The long-term efficacy and safety of tofersen require further validation, and the development of more optimized treatment protocols is essential. A series of studies and therapeutic developments related to SOD1 mutations have advanced the understanding of ALS pathophysiology and significantly contributed to treatment strategies for central nervous system disorders. This review focuses on an overview of SOD1 mutations and the development process of tofersen, aiming to deepen the understanding of advancements in ALS research and discuss future challenges and directions for ASO therapy.

STAT1 and STAT3 gain of function: clinically heterogenous immune regulatory disorders.

The identification of STAT1 gain-of-function (GOF) in 2011 and STAT3 GOF in 2014 has advanced our understanding of the host immunity along the JAK/STAT pathway and allowed targeted treatment approaches. We review the clinical features and pathogenesis of STAT1 and STAT3 GOF and how this has shaped new approaches to therapy. STAT1 GOF, initially described in patients with chronic mucocutaneous candidiasis (CMC) and autoimmune thyroid disease, is now recognized to cause early-onset multisystem autoimmunity and a range of infections. STAT3 GOF comprises mostly lymphoproliferation and autoimmunity but also with varying severity, including some with life threatening organ dysfunction. Treatment has evolved along with the understanding of the pathogenesis, with patients now receiving JAK inhibition to block upstream of the STAT defect with good response in autoimmunity and CMC in STAT1 GOF. Blockade of IL-6 signaling has also been used in STAT3 GOF. Hematopoietic cell transplantation had initial poor outcomes, but outcomes are now improving with focus on the control of inflammation pretransplant. Understanding the pathogenesis of STAT1 and STAT3 GOF has allowed great recent advancements in therapy, but many questions remain as to the best approach to therapy for each patient's clinical presentation as well as the durability of these therapies.

Value of Functional Protein S Activity Assays for Detection of Type II Protein S Deficiency

Abstract Background Protein S deficiency is a hereditary thrombophilia which occurs in three forms: Type I (quantitative), type II (qualitative) and type III (qualitative due to a gain of function). Literature suggests that of these, type II is the rarest and most difficult to diagnose. Type II protein S deficiency is only detectable by assaying protein S activity with a functional, clot-based test. However, this test is susceptible to interference and has a lower specificity when compared to antigen assays used to detect free and total protein S levels. Subsequently, the Choosing Wisely Guidelines released by the ABIM Foundation recommends against using protein S activity assays entirely. Instead, they suggest using only free protein S antigen assays. Although the rationale is to avoid false positive results from the protein S activity assay, it ultimately may result in a failure to detect type II protein S deficiency. Objective This study, performed at a tertiary medical center in the Midwestern region of the United States, aims to determine the proportion of false positives in functional assays as well as the percent of true type II protein S deficiency cases within this testing population. Methods All cases with abnormal protein S function within the institution were evaluated from July 2018 through July 2023 and classified as acquired or hereditary deficiencies. Cases were classified as acquired if there was evidence of liver disease, vitamin K deficiency, ongoing thrombosis, consumption, disseminated intravascular coagulopathy, or pregnancy. Possible hereditary deficiency cases were classified as type I, type II and type III: Type I if all assays were low, type III if both active and free protein S assays were low with normal total protein S, and type II if only protein S activity was low. The cutoff defined for normal total and free protein S levels was greater than 70% activity, and the low protein S activity cutoff was defined as less than 50%. Patients with low protein S activity but normal immunologic protein S and an elevated factor VIII were considered falsely low values. Chart review will be performed for remaining cases with low protein S activity to rule out confounders such as vitamin K deficiency, warfarin use, acute phase reaction, or pregnancy. Results From a total of 8805 panels performed to assess for possible thrombophilia, 1580 (18%) were found to have decreased protein S activity. Of these cases, 599 (38%) were considered falsely decreased due to elevated factor VIII levels. 322 cases (3.7%) were considered possible hereditary deficiencies: 217 type III (2.5%), 27 type II (0.3%), and 80 type I (0.9%). Discussion Our data indicates the prevalence of subtypes may be differently distributed than previously believed, which may call for current guidelines to be reevaluated.

A functionally augmented carbohydrate utilization locus from herbivore gut microbiota fueled by dietary β-glucans

Gut microbiota members from the Bacteroidota phylum play a pivotal role in mammalian health and metabolism. They thrive in this diverse ecosystem due to their notable ability to cope with distinct recalcitrant dietary glycans via polysaccharide utilization loci (PULs). Our study reveals that a PUL from an herbivore gut bacterium belonging to the Bacteroidota phylum, with a gene composition similar to that in the human gut, exhibits extended functionality. While the human gut PUL targets mixed-linkage β-glucans specifically, the herbivore gut PUL also efficiently processes linear and substituted β-1,3-glucans. This gain of function emerges from molecular adaptations in recognition proteins and carbohydrate-active enzymes, including a β-glucosidase specialized for β(1,6)-glucosyl linkages, a typical substitution in β(1,3)-glucans. These findings broaden the existing model for non-cellulosic β-glucans utilization by gut bacteria, revealing an additional layer of functional and evolutionary complexity within the gut microbiota, beyond conventional gene insertions/deletions to intricate biochemical interactions.

Impact of Mask Type as Training Target for Speech Intelligibility and Quality in Cochlear-Implant Noise Reduction

The selection of a target when training deep neural networks for speech enhancement is an important consideration. Different masks have been shown to exhibit different performance characteristics depending on the application and the conditions. This paper presents a comprehensive comparison of several different masks for noise reduction in cochlear implants. The study incorporated three well-known masks, namely the Ideal Binary Mask (IBM), Ideal Ratio Mask (IRM) and the Fast Fourier Transform Mask (FFTM), as well as two newly proposed masks, based on existing masks, called the Quantized Mask (QM) and the Phase-Sensitive plus Ideal Ratio Mask (PSM+). These five masks are used to train networks to estimate masks for the purpose of separating speech from noisy mixtures. A vocoder was used to simulate the behavior of a cochlear implant. Short-time Objective Intelligibility (STOI) and Perceptual Evaluation of Speech Quality (PESQ) scores indicate that the two new masks proposed in this study (QM and PSM+) perform best for normal speech intelligibility and quality in the presence of stationary and non-stationary noise over a range of signal-to-noise ratios (SNRs). The Normalized Covariance Measure (NCM) and similarity scores indicate that they also perform best for speech intelligibility/gauging the similarity of vocoded speech. The Quantized Mask performs better than the Ideal Binary Mask due to its better resolution as it approximates the Wiener Gain Function. The PSM+ performs better than the three existing benchmark masks (IBM, IRM, and FFTM) as it incorporates both magnitude and phase information.

Persistent Na + current couples spreading depolarization to seizures in Scn8a gain of function mice.

Spreading depolarization (SD) is a slowly propagating wave of massive cellular depolarization that transiently impairs the function of affected brain regions. While SD typically arises as an isolated hemispheric event, we previously reported that reducing M-type potassium current (I KM ) by ablation of Kcnq2 in forebrain excitatory neurons results in tightly coupled spontaneous bilateral seizure-SD complexes in the awake mouse cortex. Here we find that enhanced persistent Na + current due to gain-of-function (GOF) mutations in Scn8a (N1768D/+, hereafter D/+) produces a similar compound cortical excitability phenotype. Chronic DC-band EEG recording detected spontaneous bilateral seizure-SD complexes accompanied by seizures with a profound tonic component, which occurs predominantly during the light phase and were detected in the mutant mice across ages between P40-100. Laser speckle contrast imaging of cerebral blood flow dynamics resolved SD as bilateral wave of hypoperfusion and subsequent hour-lasting hypoperfusion in Scn8a D/+ cortex in awake head-restrained mice subjected to a subconvulsive PTZ. Subcortical recordings in freely moving mice revealed that approximately half of the spontaneous cortical seizure-SD complexes arose with concurrent SD-like depolarization in the thalamus and delayed depolarization in the striatum. In contrast, SD-like DC potential shifts were rarely detected in the hippocampus or upper pons. Consistent with the high spontaneous incidence in vivo , cortical slices from Scn8a D/+ mice showed a raised SD susceptibility, and pharmacological inhibition of persistent Na + current (I NaP ), which is enhanced in Scn8a D/+ neurons, inhibited SD generation in cortical slices ex vivo , indicating that I NaP contributes to SD susceptibility. Ex vivo Ca 2+ imaging studies using acute brain slices expressing genetic Ca 2+ sensor (Thy1-GCAMP6s) demonstrated that pharmacological activation of I KM suppressed Ca 2+ spikes and SD, whereas I KM inhibitor drastically increased the frequency of Ca 2+ spikes in the hippocampus of Scn8a D/+ mice, but not in WT, suggesting that I KM restrains the hyperexcitability resulting from Scn8a GOF mutation. Together, our study identifies a cortical SD phenotype in Scn8a GOF mice shared with the Kcnq2 - cKO model of developmental epileptic encephalopathy and reveals that an imbalance of non-inactivating inward and outward membrane currents bidirectionally modulates spatiotemporal SD susceptibility.

Disturbance observer-based fuzzy adaptive switched tuning control of uncertain nonlinear systems with full state constraints

This article examines the issue of disturbance observer-based fuzzy adaptive switched tuning output feedback control for a family of uncertain nonlinear systems subjected to full state constraints. By introducing monotonically bounded positive time-varying gain functions (TVGFs), a set of improved intermediate-variable-based disturbance observers (IVBDOs) is devised. Applying the convex combination technique and the LMI toolbox, the solution of the observation gain matrix is realized. Combining the switched tuning control function and the composite integral barrier Lyapunov method, a new switching nature controller is constructed. Particularly, the developed framework overcomes the conservatism of conventional constraints. Through theoretical analysis, it is exhibited that for any given initial value of the system, all states of the system are enclosed within predetermined compact sets while all signals are bounded. Lastly, simulation and experimental results under unknown disturbances attest the validity and superiority of the suggested strategy.

Heterozygous expression of a Kcnt1 gain-of-function variant has differential effects on somatostatin- and parvalbumin-expressing cortical GABAergic neurons

More than 20 recurrent missense gain-of-function (GOF) mutations have been identified in the sodium-activated potassium (KNa) channel gene KCNT1 in patients with severe developmental and epileptic encephalopathies (DEEs), most of which are resistant to current therapies. Defining the neuron types most vulnerable to KCNT1 GOF will advance our understanding of disease mechanisms and provide refined targets for precision therapy efforts. Here, we assessed the effects of heterozygous expression of a Kcnt1 GOF variant (Kcnt1Y777H) on KNa currents and neuronal physiology among cortical glutamatergic and GABAergic neurons in mice, including those expressing vasoactive intestinal polypeptide (VIP), somatostatin (SST), and parvalbumin (PV), to identify and model the pathogenic mechanisms of autosomal dominant KCNT1 GOF variants in DEEs. Although the Kcnt1Y777H variant had no effects on glutamatergic or VIP neuron function, it increased subthreshold KNa currents in both SST and PV neurons but with opposite effects on neuronal output; SST neurons became hypoexcitable with a higher rheobase current and lower action potential (AP) firing frequency, whereas PV neurons became hyperexcitable with a lower rheobase current and higher AP firing frequency. Further neurophysiological and computational modeling experiments showed that the differential effects of the Kcnt1Y777H variant on SST and PV neurons are not likely due to inherent differences in these neuron types, but to an increased persistent sodium current in PV, but not SST, neurons. The Kcnt1Y777H variant also increased excitatory input onto, and chemical and electrical synaptic connectivity between, SST neurons. Together, these data suggest differential pathogenic mechanisms, both direct and compensatory, contribute to disease phenotypes, and provide a salient example of how a pathogenic ion channel variant can cause opposite functional effects in closely related neuron subtypes due to interactions with other ionic conductances.