LinQURE: A novel AAV gene silencing platform that supports multi-transcript targeting for complex disorders.

Given that numerous genetic disorders, driven by diverse pathogenic mechanisms, may be amenable to recombinant adeno-associated virus (rAAV)-delivered gene therapies, the sustained innovation of rAAV-based therapeutic modalities is crucial. The progression and severity of genetic diseases can be reduced by targeting the toxic transcripts of a defective gene using microRNA (miRNA)-based miQURE technology delivered within an AAV vector. By adapting the delivered cassette, it may be possible to simultaneously regulate the expression profile of multiple genes involved in the pathogenesis of complex genetic diseases. The established miQURE gene silencing strategy was expanded by concatenating several miQURE molecules in a single construct, resulting in the novel linQURE platform. Here, a proof of mechanism is established by demonstrating that linQURE technology enables the concomitant expression of two synthetic miRNAs in vitro and in vivo, allowing more efficient downregulation of their disease-causing mRNA targets. This approach supports the development of multi-targeting therapeutic strategies, enabling gene therapy products to adapt to more complex multigenic indications, thus expanding the toolbox of readily available gene therapies.

Innate lymphoid cells are activated in HFRS, and their function can be modulated by hantavirus-induced type I interferons.

Hantaviruses cause the acute zoonotic diseases hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). Infected patients show strong systemic inflammation and immune cell activation. NK cells are highly activated in HFRS, suggesting that also other innate lymphoid cells (ILCs) might be responding to infection. Here, we characterized peripheral ILC responses, and measured plasma levels of soluble factors and plasma viral load, in 17 Puumala virus (PUUV)-infected HFRS patients. This revealed an increased frequency of ILC2 in patients, in particular the ILC2 lineage-committed c-Kitlo ILC2 subset. Patients' ILCs showed an activated profile with increased proliferation and displayed altered expression of several homing markers. How ILCs are activated during viral infection is largely unknown. When analyzing PUUV-mediated activation of ILCs in vitro we observed that this was dependent on type I interferons, suggesting a role for type I interferons-produced in response to virus infection-in the activation of ILCs. Further, stimulation of naïve ILC2s with IFN-ß affected ILC2 cytokine responses in vitro, causing decreased IL-5 and IL-13, and increased IL-10, CXCL10, and GM-CSF secretion. These results show that ILCs are activated in HFRS patients and suggest that the classical antiviral type I IFNs are involved in shaping ILC functions.

Optical coherence tomography biomarkers in MYO7A-inherited retinal dystrophy: longitudinal study in pediatric patients.

PURPOSE: This study aims to answer a key question: is MYO7A-inherited retinal dystrophy (MYO7A-IRD) a photoreceptor-first or retinal pigment epithelium-first disease? A second aim was to determine the most useful biomarkers to monitor disease progression in pediatric patients with Usher syndrome type 1B (USH1) secondary to MYO7A mutation. METHODS: Fifty-two eyes from 26 patients with genetically-confirmed MYO7A-IRD underwent swept-source optical coherence tomography (SS-OCT). Structural abnormalities were evaluated and correlated with follow-up time and best corrected visual acuity (BCVA). All patients were evaluated at baseline and after ≥ 40 months of follow-up. RESULTS: The mean (SD) patient age was 9.92 (± 4.1) years. Mean follow-up time was 43 (± 3.2) months. At the final evaluation, the most common qualitative abnormalities in the subfoveal area were alterations in the photoreceptor outer segments (76.9% of eyes) and in the interdigitation zone (IZ) (80.8%). The presence of cystoid macular edema at baseline was independently associated with worse BCVA at the final assessment (increase in LogMAR estimate = 0.142; t(45.00) = 2.78, p = 0.009). The mean width of the ellipsoid and interdigitation zones decreased significantly (by 668 µm and 278 µm, respectively; both p < 0.001). CONCLUSION: This study shows that disruption of the photoreceptor outer segments and the IZ are the first alterations detected by SS-OCT in the early phases of MYO7A-IRD. These data highlight the potential value of measuring the width of the ellipsoid and IZ to evaluate disease progression. These findings also demonstrate the utility of monitoring for the emergence of cystic lesions as biomarkers of worse visual prognosis in patients with MYO7A-IRD.

Optical Coherence Tomography in Inherited Macular Dystrophies: A Review.

Macular dystrophies (MDs) constitute a collection of hereditary retina disorders leading to notable visual impairment, primarily due to progressive macular atrophy. These conditions are distinguished by bilateral and relatively symmetrical abnormalities in the macula that significantly impair central visual function. Recent strides in fundus imaging, especially optical coherence tomography (OCT), have enhanced our comprehension and diagnostic capabilities for MD. OCT enables the identification of neurosensory retinal disorganization patterns and the extent of damage to retinal pigment epithelium (RPE) and photoreceptor cells in the dystrophies before visible macular pathology appears on fundus examinations. It not only helps us in diagnostic retinal and choroidal pathologies but also guides us in monitoring the progression of, staging of, and response to treatment. In this review, we summarize the key findings on OCT in some of the most common MD.

Synthesis of Cobalt-Based Nanoparticles as Catalysts for Methanol Synthesis from CO2 Hydrogenation.

The increasing emission of carbon dioxide into the atmosphere has urged the scientific community to investigate alternatives to alleviate such emissions, being that they are the principal contributor to the greenhouse gas effect. One major alternative is carbon capture and utilization (CCU) toward the production of value-added chemicals using diverse technologies. This work aims at the study of the catalytic potential of different cobalt-derived nanoparticles for methanol synthesis from carbon dioxide hydrogenation. Thanks to its abundance and cost efficacy, cobalt can serve as an economical catalyst compared to noble metal-based catalysts. In this work, we present a systematic comparison among different cobalt and cobalt oxide nanocomposites in terms of their efficiency as catalysts for carbon dioxide hydrogenation to methanol as well as how different supports, zeolites, MnO2, and CeO2, can enhance their catalytic capacity. The oxygen vacancies in the cerium oxide act as carbon dioxide adsorption and activation sites, which facilitates a higher methanol production yield.

Deciphering complexity: TULP1 variants linked to an atypical retinal dystrophy phenotype.

Introduction: TULP1 exemplifies the remarkable clinical and genetic heterogeneity observed in inherited retinal dystrophies. Our research describes the clinical and molecular characteristics of a patient manifesting an atypical retinal dystrophy pattern, marked by the identification of both a previously unreported and a rarely encountered TULP1 variant. Methods: Whole-exome sequencing was performed to identify potential causative variants. The pathogenicity of the identified TULP1 variants was evaluated through in silico predictors and a minigene splice assay, specifically designed to assess the effect of the unreported TULP1 variant. Results: We identified two TULP1 gene variants in a patient exhibiting unusual and symmetrical alterations in both retinas, characterized by an increase in autofluorescence along the distribution of retinal vessels. These variants included a known rare missense variant, c.1376T>C, and a novel splice site variant, c.822G>T. For the latter variant (c.822G>T), we conducted a minigene splice assay that demonstrated the incorporation of a premature stop codon. This finding suggests a likely activation of the nonsense-mediated mRNA decay mechanism, ultimately resulting in the absence of protein production from this allele. Segregation analysis confirmed that these variants were in trans. Discussion: Our data support that individuals with biallelic TULP1 variants may present with a unique pattern of macular degeneration and periarteriolar vascular pigmentation. This study highlights the importance of further clinical and molecular characterization of TULP1 variants to elucidate genotype-phenotype correlations in the context of inherited retinal dystrophies.

Kv1.3 in the spotlight for treating immune diseases.

INTRODUCTION: Kv1.3 is the main voltage-gated potassium channel of leukocytes from both the innate and adaptive immune systems. Channel function is required for common processes such as Ca2+ signaling but also for cell-specific events. In this context, alterations in Kv1.3 are associated with multiple immune disorders. Excessive channel activity correlates with numerous autoimmune diseases, while reduced currents result in increased cancer prevalence and immunodeficiencies. AREAS COVERED: This review offers a general view of the role of Kv1.3 in every type of leukocyte. Moreover, diseases stemming from dysregulations of the channel are detailed, as well as current advances in their therapeutic research. EXPERT OPINION: Kv1.3 arises as a potential immune target in a variety of diseases. Several lines of research focused on channel modulation have yielded positive results. However, among the great variety of specific channel blockers, only one has reached clinical trials. Future investigations should focus on developing simpler administration routes for channel inhibitors to facilitate their entrance into clinical trials. Prospective Kv1.3-based treatments will ensure powerful therapies while minimizing undesired side effects.

Linking structural and functional changes during aging using multilayer brain network analysis.

Brain structure and function are intimately linked, however this association remains poorly understood and the complexity of this relationship has remained understudied. Healthy aging is characterised by heterogenous levels of structural integrity changes that influence functional network dynamics. Here, we use the multilayer brain network analysis on structural (diffusion weighted imaging) and functional (magnetoencephalography) data from the Cam-CAN database. We found that the level of similarity of connectivity patterns between brain structure and function in the parietal and temporal regions (alpha frequency band) is associated with cognitive performance in healthy older individuals. These results highlight the impact of structural connectivity changes on the reorganisation of functional connectivity associated with the preservation of cognitive function, and provide a mechanistic understanding of the concepts of brain maintenance and compensation with aging. Investigation of the link between structure and function could thus represent a new marker of individual variability, and of pathological changes.

Dynamic multilayer functional connectivity detects preclinical and clinical Alzheimer's disease.

Increasing evidence suggests that patients with Alzheimer's disease present alterations in functional connectivity but previous results have not always been consistent. One of the reasons that may account for this inconsistency is the lack of consideration of temporal dynamics. To address this limitation, here we studied the dynamic modular organization on resting-state functional magnetic resonance imaging across different stages of Alzheimer's disease using a novel multilayer brain network approach. Participants from preclinical and clinical Alzheimer's disease stages were included. Temporal multilayer networks were used to assess time-varying modular organization. Logistic regression models were employed for disease stage discrimination, and partial least squares analyses examined associations between dynamic measures with cognition and pathology. Temporal multilayer functional measures distinguished all groups, particularly preclinical stages, overcoming the discriminatory power of risk factors such as age, sex, and APOE ϵ4 carriership. Dynamic multilayer functional measures exhibited strong associations with cognition as well as amyloid and tau pathology. Dynamic multilayer functional connectivity shows promise as a functional imaging biomarker for both early- and late-stage Alzheimer's disease diagnosis.

Vagus nerve dysfunction in the post-COVID-19 condition: a pilot cross-sectional study.

OBJECTIVES: The post-COVID-19 condition (PCC) is a disabling syndrome affecting at least 5%-10% of subjects who survive COVID-19. SARS-CoV-2 mediated vagus nerve dysfunction could explain some PCC symptoms, such as dysphonia, dysphagia, dyspnea, dizziness, tachycardia, orthostatic hypotension, gastrointestinal disturbances, or neurocognitive complaints. METHODS: We performed a cross-sectional pilot study in subjects with PCC with symptoms suggesting vagus nerve dysfunction (n = 30) and compared them with subjects fully recovered from acute COVID-19 (n = 14) and with individuals never infected (n = 16). We evaluated the structure and function of the vagus nerve and respiratory muscles. RESULTS: Participants were mostly women (24 of 30, 80%), and the median age was 44 years (interquartile range [IQR] 35-51 years). Their most prevalent symptoms were cognitive dysfunction 25 of 30 (83%), dyspnea 24 of 30 (80%), and tachycardia 24 of 30 (80%). Compared with COVID-19-recovered and uninfected controls, respectively, subjects with PCC were more likely to show thickening and hyperechogenic vagus nerve in neck ultrasounds (cross-sectional area [CSA] [mean ± standard deviation]: 2.4 ± 0.97mm2 vs. 2 ± 0.52mm2 vs. 1.9 ± 0.73 mm2; p 0.08), reduced esophageal-gastric-intestinal peristalsis (34% vs. 0% vs. 21%; p 0.02), gastroesophageal reflux (34% vs. 19% vs. 7%; p 0.13), and hiatal hernia (25% vs. 0% vs. 7%; p 0.05). Subjects with PCC showed flattening hemidiaphragms (47% vs. 6% vs. 14%; p 0.007), and reductions in maximum inspiratory pressure (62% vs. 6% vs. 17%; p ≤ 0.001), indicating respiratory muscle weakness. The latter findings suggest additional involvement of the phrenic nerve. DISCUSSION: Vagus and phrenic nerve dysfunction contribute to the complex and multifactorial pathophysiology of PCC.

Novel intragenic deletion within the FXN gene in a patient with typical phenotype of Friedreich ataxia: may be more prevalent than we think?

BACKGROUND: Friedreich ataxia is the most common inherited ataxia in Europe and is mainly caused by biallelic pathogenic expansions of the GAA trinucleotide repeat in intron 1 of the FXN gene that lead to a decrease in frataxin protein levels. Rarely, affected individuals carry either a large intragenic deletion or whole-gene deletion of FXN on one allele and a full-penetrance expanded GAA repeat on the other allele. CASE PRESENTATION: We report here a patient that presented the typical clinical features of FRDA and genetic analysis of FXN intron 1 led to the assumption that the patient carried the common biallelic expansion. Subsequently, parental sample testing led to the identification of a novel intragenic deletion involving the 5'UTR upstream region and exons 1 and 2 of the FXN gene by MLPA. CONCLUSIONS: With this case, we want to raise awareness about the potentially higher prevalence of intragenic deletions and underline the essential role of parental sample testing in providing accurate genetic counselling.

Age-related differences in the functional topography of the locus coeruleus and their implications for cognitive and affective functions.

The locus coeruleus (LC) is an important noradrenergic nucleus that has recently attracted a lot of attention because of its emerging role in cognitive and psychiatric disorders. Although previous histological studies have shown that the LC has heterogeneous connections and cellular features, no studies have yet assessed its functional topography in vivo, how this heterogeneity changes over aging, and whether it is associated with cognition and mood. Here, we employ a gradient-based approach to characterize the functional heterogeneity in the organization of the LC over aging using 3T resting-state fMRI in a population-based cohort aged from 18 to 88 years of age (Cambridge Centre for Ageing and Neuroscience cohort, n=618). We show that the LC exhibits a rostro-caudal functional gradient along its longitudinal axis, which was replicated in an independent dataset (Human Connectome Project [HCP] 7T dataset, n=184). Although the main rostro-caudal direction of this gradient was consistent across age groups, its spatial features varied with increasing age, emotional memory, and emotion regulation. More specifically, a loss of rostral-like connectivity, more clustered functional topography, and greater asymmetry between right and left LC gradients was associated with higher age and worse behavioral performance. Furthermore, participants with higher-than-normal Hospital Anxiety and Depression Scale (HADS) ratings exhibited alterations in the gradient as well, which manifested in greater asymmetry. These results provide an in vivo account of how the functional topography of the LC changes over aging, and imply that spatial features of this organization are relevant markers of LC-related behavioral measures and psychopathology.

Age-related differences in the functional topography of the locus coeruleus: implications for cognitive and affective functions.

The locus coeruleus (LC) is an important noradrenergic nucleus that has recently attracted a lot of attention because of its emerging role in cognitive and psychiatric disorders. Although previous histological studies have shown that the LC has heterogeneous connections and cellular features, no studies have yet assessed its functional topography in vivo, how this heterogeneity changes over aging and whether it is associated with cognition and mood. Here we employ a gradient-based approach to characterize the functional heterogeneity in the organization of the LC over aging using 3T resting-state fMRI in a population-based cohort aged from 18 to 88 years old (Cambridge Centre for Ageing and Neuroscience cohort, n=618). We show that the LC exhibits a rostro-caudal functional gradient along its longitudinal axis, which was replicated in an independent dataset (Human Connectome Project 7T dataset, n=184). Although the main rostro-caudal direction of this gradient was consistent across age groups, its spatial features varied with increasing age, emotional memory and emotion regulation. More specifically, a loss of rostral-like connectivity, more clustered functional topography and greater asymmetry between right and left LC gradients was associated with higher age and worse behavioral performance. Furthermore, participants with higher-than-normal Hospital Anxiety and Depression Scale ratings exhibited alterations in the gradient as well, which manifested in greater asymmetry. These results provide an in vivo account of how the functional topography of the LC changes over aging, and imply that spatial features of this organization are relevant markers of LC-related behavioral measures and psychopathology.

Sex differences in multilayer functional network topology over the course of aging in 37543 UK Biobank participants.

Aging is a major risk factor for cardiovascular and neurodegenerative disorders, with considerable societal and economic implications. Healthy aging is accompanied by changes in functional connectivity between and within resting-state functional networks, which have been associated with cognitive decline. However, there is no consensus on the impact of sex on these age-related functional trajectories. Here, we show that multilayer measures provide crucial information on the interaction between sex and age on network topology, allowing for better assessment of cognitive, structural, and cardiovascular risk factors that have been shown to differ between men and women, as well as providing additional insights into the genetic influences on changes in functional connectivity that occur during aging. In a large cross-sectional sample of 37,543 individuals from the UK Biobank cohort, we demonstrate that such multilayer measures that capture the relationship between positive and negative connections are more sensitive to sex-related changes in the whole-brain connectivity patterns and their topological architecture throughout aging, when compared to standard connectivity and topological measures. Our findings indicate that multilayer measures contain previously unknown information on the relationship between sex and age, which opens up new avenues for research into functional brain connectivity in aging.

Nonlinear changes in delayed functional network topology in Alzheimer's disease: relationship with amyloid and tau pathology.

BACKGROUND: Alzheimer's disease is a neurodegenerative disorder associated with the abnormal deposition of pathological processes, such as amyloid-ß and tau, which produces nonlinear changes in the functional connectivity patterns between different brain regions across the Alzheimer's disease continuum. However, the mechanisms underlying these nonlinear changes remain largely unknown. Here, we address this question using a novel method based on temporal or delayed correlations and calculate new whole-brain functional networks to tackle these mechanisms. METHODS: To assess our method, we evaluated 166 individuals from the ADNI database, including amyloid-beta negative and positive cognitively normal subjects, patients with mild cognitive impairment, and patients with Alzheimer's disease dementia. We used the clustering coefficient and the global efficiency to measure the functional network topology and assessed their relationship with amyloid and tau pathology measured by positron emission tomography, as well as cognitive performance using tests measuring memory, executive function, attention, and global cognition. RESULTS: Our study found nonlinear changes in the global efficiency, but not in the clustering coefficient, showing that the nonlinear changes in functional connectivity are due to an altered ability of brain regions to communicate with each other through direct paths. These changes in global efficiency were most prominent in early disease stages. However, later stages of Alzheimer's disease were associated with widespread network disruptions characterized by changes in both network measures. The temporal delays required for the detection of these changes varied across the Alzheimer's disease continuum, with shorter delays necessary to detect changes in early stages and longer delays necessary to detect changes in late stages. Both global efficiency and clustering coefficient showed quadratic associations with pathological amyloid and tau burden as well as cognitive decline. CONCLUSIONS: This study suggests that global efficiency is a more sensitive indicator of network changes in Alzheimer's disease when compared to clustering coefficient. Both network properties were associated with pathology and cognitive performance, demonstrating their relevance in clinical settings. Our findings provide an insight into the mechanisms underlying nonlinear changes in functional network organization in Alzheimer's disease, suggesting that it is the lack of direct connections that drives these functional changes.

The Phosphorylation of Kv1.3: A Modulatory Mechanism for a Multifunctional Ion Channel.

The voltage-gated potassium channel Kv1.3 plays a pivotal role in a myriad of biological processes, including cell proliferation, differentiation, and apoptosis. Kv1.3 undergoes fine-tuned regulation, and its altered expression or function correlates with tumorigenesis and cancer progression. Moreover, posttranslational modifications (PTMs), such as phosphorylation, have evolved as rapid switch-like moieties that tightly modulate channel activity. In addition, kinases are promising targets in anticancer therapies. The diverse serine/threonine and tyrosine kinases function on Kv1.3 and the effects of its phosphorylation vary depending on multiple factors. For instance, Kv1.3 regulatory subunits (KCNE4 and Kvß) can be phosphorylated, increasing the complexity of channel modulation. Scaffold proteins allow the Kv1.3 channelosome and kinase to form protein complexes, thereby favoring the attachment of phosphate groups. This review compiles the network triggers and signaling pathways that culminate in Kv1.3 phosphorylation. Alterations to Kv1.3 expression and its phosphorylation are detailed, emphasizing the importance of this channel as an anticancer target. Overall, further research on Kv1.3 kinase-dependent effects should be addressed to develop effective antineoplastic drugs while minimizing side effects. This promising field encourages basic cancer research while inspiring new therapy development.

Functional gradients of the medial parietal cortex in a healthy cohort with family history of sporadic Alzheimer's disease.

BACKGROUND: The medial parietal cortex is an early site of pathological protein deposition in Alzheimer's disease (AD). Previous studies have identified different subregions within this area; however, these subregions are often heterogeneous and disregard individual differences or subtle pathological alterations in the underlying functional architecture. To address this limitation, here we measured the continuous connectivity gradients of the medial parietal cortex and assessed their relationship with cerebrospinal fluid (CSF) biomarkers, ApoE ε4 carriership and memory in asymptomatic individuals at risk to develop AD. METHODS: Two hundred sixty-three cognitively normal participants with a family history of sporadic AD who underwent resting-state and task-based functional MRI using encoding and retrieval tasks were included from the PREVENT-AD cohort. A novel method for characterizing spatially continuous patterns of functional connectivity was applied to estimate functional gradients in the medial parietal cortex during the resting-state and task-based conditions. This resulted in a set of nine parameters that described the appearance of the gradient across different spatial directions. We performed correlation analyses to assess whether these parameters were associated with CSF biomarkers of phosphorylated tau181 (p-tau), total tau (t-tau), and amyloid-ß1-42 (Aß). Then, we compared the spatial parameters between ApoE ε4 carriers and noncarriers, and evaluated the relationship between these parameters and memory. RESULTS: Alterations involving the superior part of the medial parietal cortex, which was connected to regions of the default mode network, were associated with higher p-tau, t-tau levels as well as lower Aß/p-tau levels during the resting-state condition (p < 0.01). Similar alterations were found in ApoE ε4 carriers compared to non-carriers (p < 0.003). In contrast, lower immediate memory scores were associated with changes in the middle part of the medial parietal cortex, which was connected to inferior temporal and posterior parietal regions, during the encoding task (p = 0.001). No results were found when using conventional connectivity measures. CONCLUSIONS: Functional alterations in the medial parietal gradients are associated with CSF AD biomarkers, ApoE ε4 carriership, and lower memory in an asymptomatic cohort with a family history of sporadic AD, suggesting that functional gradients are sensitive to subtle changes associated with early AD stages.

Response Variability to Drug Testing in Two Models of Chemically Induced Colitis.

The lack of knowledge regarding the pathogenesis of IBD is a challenge for the development of more effective and safer therapies. Although in vivo preclinical approaches are critical for drug testing, none of the existing models accurately reproduce human IBD. Factors that influence the intra-individual response to drugs have barely been described. With this in mind, our aim was to compare the anti-inflammatory efficacy of a new molecule (MTADV) to that of corticosteroids in TNBS and DSS-induced colitis mice of both sexes in order to clarify further the response mechanism involved and the variability between sexes. The drugs were administered preventively and therapeutically, and real-time bioluminescence was performed for the in vivo time-course colitis monitoring. Morphometric data were also collected, and colonic cytokines and acute plasma phase proteins were analyzed by qRT-PCR and ELISA, respectively-bioluminescence images correlated with inflammatory markers. In the TNBS model, dexamethasone worked better in females, while MTADV improved inflammation in males. In DSS-colitis, both therapies worked similarly. Based on the molecular profiles, interaction networks were constructed to pinpoint the drivers of therapeutic response that were highly dependent on the sex. In conclusion, our results suggest the importance of considering sex in IBD preclinical drug screening.

Directed Functional Brain Connectivity is Altered in Sub-threshold Amyloid-ß Accumulation in Cognitively Normal Individuals.

Several studies have shown that amyloid-ß (Aß) deposition below the clinically relevant cut-off levels is associated with subtle changes in cognitive function and increases the risk of developing future Alzheimer's disease (AD). Although functional MRI is sensitive to early alterations occurring during AD, sub-threshold changes in Aß levels have not been linked to functional connectivity measures. This study aimed to apply directed functional connectivity to identify early changes in network function in cognitively unimpaired participants who, at baseline, exhibit Aß accumulation below the clinically relevant threshold. To this end, we analyzed baseline functional MRI data from 113 cognitively unimpaired participants of the Alzheimer's Disease Neuroimaging Initiative cohort who underwent at least one 18F-florbetapir-PET after the baseline scan. Using the longitudinal PET data, we classified these participants as Aß negative (Aß-) non-accumulators (n = 46) and Aß- accumulators (n = 31). We also included 36 individuals who were amyloid-positive (Aß+) at baseline and continued to accumulate Aß (Aß+ accumulators). For each participant, we calculated whole-brain directed functional connectivity networks using our own anti-symmetric correlation method and evaluated their global and nodal properties using measures of network segregation (clustering coefficient) and integration (global efficiency). When compared to Aß- non-accumulators, the Aß- accumulators showed lower global clustering coefficient. Moreover, the Aß+ accumulator group exhibited reduced global efficiency and clustering coefficient, which at the nodal level mainly affected the superior frontal gyrus, anterior cingulate cortex, and caudate nucleus. In Aß- accumulators, global measures were associated with lower baseline regional PET uptake values, as well as higher scores on the Modified Preclinical Alzheimer Cognitive Composite. Our findings indicate that directed connectivity network properties are sensitive to subtle changes occurring in individuals who have not yet reached the threshold for Aß positivity, which makes them a potentially viable marker to detect negative downstream effects of very early Aß pathology.

Improving Hereditary Hemorrhagic Telangiectasia Molecular Diagnosis: A Referral Center Experience.

BACKGROUND: Hereditary hemorrhagic telangiectasia (HHT) is a rare vascular disease inherited in an autosomal dominant manner. Disease-causing variants in endoglin (ENG) and activin A receptor type II-like 1 (ACVRL1) genes are detected in more than 90% of the patients undergoing molecular testing. The identification of variants of unknown significance is often seen as a challenge in clinical practice that makes family screening and genetic counseling difficult. Here, we show that the implementation of cDNA analysis to assess the effect of splice site variants on mRNA splicing is a powerful tool. METHODS: Gene panel sequencing of genes associated with HHT and other arteriovenous malformation-related syndromes was performed. To evaluate the effect of the splice site variants, cDNA analysis of ENG and ACVRL1 genes was carried out. RESULTS: three novel splice site variants were identified in ENG (c.68-2A > T and c.1311+4_1311+8del) and ACVLR1 (c.526-6C > G) genes correspondingly in three individuals with HHT that met ≥ 3 Curaçao criteria. All three variants led to an aberrant splicing inducing exon skipping (ENG:c.68-2A > T and ACVRL1:c.526-6C > G) or intron retention (ENG:c.1311+4_1311+8del) allowing the confirmation of the predicted effect on splicing and the reclassification from unknown significance to pathogenic/likely pathogenic of two of them. CONCLUSIONS: RNA analysis should be performed to assess and/or confirm the impact of variants on splicing. The molecular diagnosis of HHT patients is crucial to allow family screening and accurate genetic counseling. A multidisciplinary approach including clinicians and geneticists is crucial when dealing with patients with rare diseases.