Investigation of the role of VHL-HIF signaling in DNA repair and apoptosis in zebrafish

pVHL is a tumor suppressor. The lack of its function leads to various tumors, among which ccRCC (clear cell renal cell carcinoma) has the most serious outcome due to its resistance to chemotherapies and radiotherapies. Although HIF promotes the progression of ccRCC, the precise mechanism by which the loss of VHL leads to tumor initiation remains unclear. We exploited two zebrafish vhl mutants, vhl and vll, and Tg (phd3:: EGFP)i144 fish to identify crucial functions of Vhl in tumor initiation. Through the mutant analysis, we found that the role of pVHL in DNA repair is conserved in zebrafish Vll. Interestingly, we also discovered that Hif activation strongly suppressed genotoxic stress induced DNA repair defects and apoptosis in vll and brca2 mutants and in embryos lacking ATM activity. These results suggest the potential of HIF as a clinical modulator that can protect cells from accumulating DNA damage and apoptosis which can lead to cancers and neurodegenerative disorders

Enhancer trap lines with GFP driven by smad6b and frizzled1 regulatory sequences for the study of epithelial morphogenesis in the developing zebrafish inner ear

Live imaging in the zebrafish embryo using tissue-specific expression of fluorescent proteins can yield important insights into the mechanisms that drive sensory organ morphogenesis and cell differentiation. Morphogenesis of the semicircular canal ducts of the vertebrate inner ear requires a complex rearrangement of epithelial cells, including outgrowth, adhesion, fusion and perforation of epithelial projections to generate pillars of tissue that form the hubs of each canal. We report the insertion sites and expression patterns of two enhancer trap lines in the developing zebrafish embryo, each of which highlight different aspects of epithelial cell morphogenesis in the inner ear. A membrane-linked EGFP driven by smad6b regulatory sequences is expressed throughout the otic epithelium, most strongly on the lateral side of the ear and in the sensory cristae. A second enhancer trap line, with cytoplasmic EGFP driven by frizzled1 (fzd1) regulatory sequences, specifically marks cells of the ventral projection and pillar in the developing ear, and marginal cells in the sensory cristae, together with variable expression in the retina and epiphysis, and neurons elsewhere in the developing central nervous system. We have used a combination of methods to identify the insertion sites of these two transgenes, which were generated through random insertion, and show that Targeted Locus Amplification is a rapid and reliable method for the identification of insertion sites of randomly inserted transgenes

Lipoprotein(a) Genotype Influences the Clinical Diagnosis of Familial Hypercholesterolemia

Background Evidence suggests that LPA risk genotypes are a possible contributor to the clinical diagnosis of familial hypercholesterolemia (FH). This study aimed at determining the prevalence of LPA risk variants in adult individuals with FH enrolled in the Italian LIPIGEN (Lipid Transport Disorders Italian Genetic Network) study, with (FH/M+) or without (FH/M-) a causative genetic variant. Methods and ResultsAn lp(a) [lipoprotein(a)] genetic score was calculated by summing the number risk-increasing alleles inherited at rs3798220 and rs10455872 variants. Overall, in the 4.6% of 1695 patients with clinically diagnosed FH, the phenotype was not explained by a monogenic or polygenic cause but by genotype associated with high lp(a) levels. Among 765 subjects with FH/M- and 930 subjects with FH/M+, 133 (17.4%) and 95 (10.2%) were characterized by 1 copy of either rs10455872 or rs3798220 or 2 copies of either rs10455872 or rs3798220 (lp(a) score >= 1). Subjects with FH/M- also had lower mean levels of pretreatment low-density lipoprotein cholesterol than individuals with FH/M+ (t test for difference in means between FH/M- and FH/M+ groups <0.0001); however, subjects with FH/M- and lp(a) score >= 1 had higher mean (SD) pretreatment low-density lipoprotein cholesterol levels (223.47 [50.40] mg/dL) compared with subjects with FH/M- and lp(a) score=0 (219.38 [54.54] mg/dL for), although not statistically significant. The adjustment of low-density lipoprotein cholesterol levels based on lp(a) concentration reduced from 68% to 42% the proportion of subjects with low-density lipoprotein cholesterol level >= 190 mg/dL (or from 68% to 50%, considering a more conservative formula). ConclusionsOur study supports the importance of measuring lp(a) to perform the diagnosis of FH appropriately and to exclude that the observed phenotype is driven by elevated levels of lp(a) before performing the genetic test for FH

Refinement of the diagnostic approach for the identification of children and adolescents affected by familial hypercholesterolemia: Evidence from the LIPIGEN study

Background and aims: We aimed to describe the limitations of familiar hypercholesterolemia (FH) diagnosis in childhood based on the presence of the typical features of FH, such as physical sings of cholesterol accumulation and personal or family history of premature cardiovascular disease or hypercholesterolemia, comparing their prevalence in the adult and paediatric FH population, and to illustrate how additional information can lead to a more effective diagnosis of FH at a younger age. Methods: From the Italian LIPIGEN cohort, we selected 1188 (≥18 years) and 708 (<18 years) genetically-confirmed heterozygous FH, with no missing personal FH features. The prevalence of personal and familial FH features was compared between the two groups. For a sub-group of the paediatric cohort (N = 374), data about premature coronary heart disease (CHD) in second-degree family members were also included in the evaluation. Results: The lower prevalence of typical FH features in children/adolescents vs adults was confirmed: the prevalence of tendon xanthoma was 2.1% vs 13.1%, and arcus cornealis was present in 1.6% vs 11.2% of the cohorts, respectively. No children presented clinical history of premature CHD or cerebral/peripheral vascular disease compared to 8.8% and 5.6% of adults, respectively. The prevalence of premature CHD in first-degree relatives was significantly higher in adults compared to children/adolescents (38.9% vs 19.7%). In the sub-cohort analysis, a premature CHD event in parents was reported in 63 out of 374 subjects (16.8%), but the percentage increased to 54.0% extending the evaluation also to second-degree relatives. Conclusions: In children, the typical FH features are clearly less informative than in adults. A more thorough data collection, adding information about second-degree relatives, could improve the diagnosis of FH at younger age

Morphogenesis of the lateral semicircular canal in the zebrafish inner ear

The study of the morphogenetic processes underlying organogenesis has always proved to be challenging, as many aspects need to be considered (e.g. cell movement and genetic interactions). This is particularly interesting in the case of very complex organs, such as the inner ear, which hosts both the auditory and vestibular apparatus, the latter being the organ of balance. This is composed by three semicircular canals, which detect angular acceleration of the head and send information to the brain to allow the maintenance of posture and stabilisation of gaze. A failure in the activity of these canals has been linked to various kinds of disease, such as the Branchio-Oto-Renal syndrome and Menière’s disease. This project is focused on the development of the lateral (horizontal) canal, as it has been proposed to be the last to have evolved and to be the most affected by inner ear malformations. In the zebrafish, the lateral canal development requires the formation of an epithelial pillar, between 48 and 72 hours post fertilisation (hpf), which is the hub of the canal. Previous studies have described the ventral pillar formation as an epithelial fusion between two cell populations (ventral bulge and ventral projection), but the cell movements required to achieve that have never been described in detail. By taking advantage of light-sheet microscopy, I have demonstrated that, upon fusion, other cell rearrangements need to take place to develop the ventral pillar. In particular, after the fusion event, the cells exhibit complex movements and a dynamism that is not characteristic of an epithelial state. In addition, I built on previous knowledge regarding the inner ear phenotype of the otx1 and eya1 mutants to analyse a possible genetic interaction between these two genes and how their mutation affects the ventral pillar development. Here, I show that these two mutants exhibit reciprocal phenotypes, with respect to the ventral pillar, and that this correlates with changes in the expression pattern of specific markers of the ventral otic epithelium. In conclusion, this study sheds light on the cell movements and genetics underlying the development of the ventral pillar and, more broadly, provides new ways of analysing the morphogenetic processes that take place during organogenesis

Conservation and transformation of heritage : conditions of preservation of the values of a living heritage

L’enjeu d’un patrimoine est sa transmission : celle-ci entre dans la définition même du mot patrimoine. La conservation n’est qu’un moyen au service de la transmission. Or il peut arriver que conservation et transmission entrent en conflit, en particulier pour ce qu’on appelle les patrimoines vivants – non seulement au sens biologique du terme, mais tout ce qui est défini dans le temps : paysage, ville, patrimoine immatériel, et, d’une manière ou d’une autre, presque tous les patrimoines. Le vivant est changeant et même mortel. On peut essayer de conserver un patrimoine en le rendant le plus solide possible sans qu’il y ait besoin de personne pour le gérer : on le conserve, mais on ne le transmet pas à de nouveaux gestionnaires. À l’inverse, la transmission implique le plus souvent une transformation. A partir d’exemples de patrimoine paysagers (vignobles, fleuve, routes, rivière urbaine), cette recherche examine quelles transformations conservent ce que l’on veut transmettre d’un patrimoine vivantThe stake of heritage is its transmission (inheritance from one generation to another). Conservation is a mere tool for transmission. But it may happen that conservation and transmission are in conflict, especially for the so-called living heritage – not only in the biological sense of the word, but all that is defined within time : landscape, city, intangible heritage, and, more or less, any property of heritage value. Living material is changing and even mortal. One can try and conserve a property by making it as strong as possible, without needing anybody to manage it : it is conserved, but not transmitted to new managers. Conversely, transmission often implies a transformation. From examples of heritage landscapes (vineyards, large river, roads, urban river), this research examines which transformations conserve what is intended to be transmitted of a living heritag

Temporal muscle thickness predicts mortality and disability in older adults diagnosed with mild dementia

ANTECEDENTES: La sarcopenia contribuye al aumento de las hospitalizaciones, deterioro cognitivo, caídas y mortalidad por todas las causas. Diagnóstico actual métodos, como la resonancia magnética corporal y la energía dual. La absorciometría de rayos X es costosa y poco práctica. Cabe destacar que no hay Enfoque estandarizado para evaluar la sarcopenia en clínicas de demencia. Nosotros estudió la asociación del grosor del músculo temporal (TMT) con clave Factores pronósticos en personas con enfermedad de Alzheimer (EA) y Lewy. Demencia corporal (DLB). MÉTODOS: Utilizamos datos del DemVest, una cohorte longitudinal estudio, e incluyó participantes clínicamente diagnosticados con EA leve o DLB. El TMT se midió mediante resonancias magnéticas iniciales. El resultado principal Las medidas fueron cognición, rendimiento funcional, desnutrición y mortalidad. Se consideraron varios factores demográficos y clínicos como posibles factores de confusión. RESULTADOS: La muestra de AD estuvo compuesta principalmente por mujeres (76,9%), 75,5 años (DE 6,95). La muestra DLB estuvo compuesta mayoritariamente por hombres (63,6%), edad 75,8 (DE 6,85). Al inicio del estudio, el TMT mostró asociación con el rendimiento cognitivo en el grupo DLB (Est.=0,593, valor p=0,049). El análisis longitudinal reveló importantes asociaciones entre TMT y deterioro funcional en DLB (Est. = -0,123, valor p 0,007) y aumento de la mortalidad en toda la muestra(HR=0,815, valor p 0,002), el grupo AD (HR=0,834 valor p=0,031) y el grupo DLB grupo (HR=0,767 valor p=0,019) respectivamente. Estas asociaciones siguió siendo significativo después de ajustar por factores de confusión. CONCLUSIONES: La medición TMT se asoció con la mortalidad. en ambos grupos de demencia, así como con la cognición y la función en DLB. TMT surge como una medida rentable de masa muscular que indica relevancia clínica y utilidad en entornos sanitarios. La implementación de la evaluación TMT podría mejorar la atención al paciente y ayudar a identificar a las personas en riesgo de resultados adversos en la demencia leve. Palabras clave: Sarcopenia, Demencia, Grosor del músculo temporal (TMT), músculo, funcionalidad, desnutrición.Q1Q1BACKGROUND: Sarcopenia contributes to increased hospitalizations, cognitive impairment, falls, and all-cause mortality. Current diagnostic methods, like body Magnetic Resonance Imaging and dual-energy X-ray absorptiometry, are costly and impractical. Notably, there is no standardized approach for assessing sarcopenia in dementia clinics. We studied the association of temporal muscle thickness (TMT) with key prognostic factors in people with Alzheimer’s disease (AD) and Lewy body dementia (DLB). METHODS: We utilized data from the DemVest, a longitudinal cohort study, and included participants clinically diagnosed with mild AD or DLB. TMT was measured using baseline MRI scans. The main outcome measures were cognition, functional performance, malnutrition, and mortality. Various demographic and clinical factors were considered as potential confounders. RESULTS: The AD sample was mainly composed by females(76.9%), age 75.5(SD 6.95). The DLB sample was mostly composed by men(63.6%), age 75.8(SD 6.85). At baseline TMT showed significant association with cognitive performance in the DLB group (Est.=0.593, p-value=0.049). The longitudinal analysis revealed significant associations between TMT and functional decline in DLB (Est.=-0.123, p-value 0.007) and increased mortality in the whole sample(HR=0.815, p-value 0.002), the AD group (HR=0.834 p-value=0.031), and the DLB group (HR=0.767 p-value=0.019) respectively. These associations remained significant after adjusting for confounders. CONCLUSIONS: The TMT measurement was associated with mortality in both dementia groups as well as with cognition and function in DLB. TMT emerges as a cost-efficient measure of muscle mass indicating clinical relevance and utility in healthcare settings. Implementing TMT assessment could improve patient care and aid in identifying individuals at risk of adverse outcomes in mild dementia. Key words: Sarcopenia, Dementia, Temporal muscle thickness (TMT), muscle, functionality, malnutrition.https://orcid.org/0000-0001-5832-0603https://scholar.google.com/citations?user=MrICwaMAAAAJ&hl=enhttps://scienti.minciencias.gov.co/cvlac/visualizador/generarCurriculoCv.do?cod_rh=0001429659Revista Internacional - IndexadaS

Peripheral blood mononuclear cells reactivity in recent-onset type I diabetes patients is directed against the leader peptide of preproinsulin, GAD65271-285 and GAD65431-450

IntroductionT cell reactivity against pancreatic autoantigens is considered one of the main contributors to the destruction of insulin-producing cells in type 1 diabetes (T1D). Over the years, peptide epitopes derived from these autoantigens have been described in NOD mice and in both HLA class II transgenic mice and humans. However, which ones are involved in the early onset or in the progressive phases of the disease is still unclear. MethodsIn this work we have investigated, in early-onset T1D pediatric patients and HLA-matched controls from Sardinia, the potential of preproinsulin (PPI) and glutamate decarboxylase 65 (GAD65)-derived peptides to induce spontaneous T cell proliferation responses of peripheral blood mononuclear cells (PBMCs). ResultsSignificant T cell responses against PPI1-18, PPI7-19 and PPI31-49, the first two belonging to the leader sequence of PPI, and GAD65271-285 and GAD65431-450, were found in HLA-DR4, -DQ8 and -DR3, -DQ2 T1D children. ConclusionsThese data show that cryptic epitopes from the leader sequence of the PPI and GAD65271-285 and GAD65431-450 peptides might be among the critical antigenic epitopes eliciting the primary autoreactive responses in the early phases of the disease. These results may have implications in the design of immunogenic PPI and GAD65 peptides for peptide-based immunotherapy

Asymmetric synthesis of a tertiary arsine by nucleophilic addition to a chiral phosphine-stabilized arsenium salt

A new method for the asymmetric synthesis of As-chiral tertiary arsines is described. The addition at -95 °C of n-butyllithium to a dichloromethane solution of a phosphine-stabilized arsenium salt of the type (±)-[R P→AsMePh]PF, where RP is an enantiomerically pure, atropisomeric phosphepine derived from lithiated (aR)-2,2'-dimethyl-1,1'-binaphthalene, furnishes (S)-(+)-(n-butyl) methylphenylarsine in 85% enantioselectivity (70% enantiomeric excess) with displacement of the (aR)-phosphepine. The enantioselectivity of the synthesis is lower than the diastereoselectivity of coordination of the (aR )-phosphepine to the prochiral, six-electron methylphenylarsenium ion with which it is in equilibrium in solution by P-As bond dissociation, 94% diastereomeric excess, as determined by NMR spectroscopy at -95 °C. The excess of the S enantiomer of the arsine, however, is consistent with the S 2 mechanism proposed for the reaction and the solution and solid-state structures of the predominant diastereomer of the phosphepine-arsenium complex. A feature of the design of the phosphepine auxiliary is the attachment of a 2-(methoxymethyl)phenyl substituent at phosphorus, the oxygen of which interacts with the arsenic of the arsenium ion, in solution and in the solid state, and facilitates Stereodifferentiation by the chiral phosphepine of the enantiotopic faces of the prochiral, six-electron methylphenylarsenium ion; the significance of this anchimeric interaction is borne out by DFT calculations on a closely related model complex