RANKL, but Not R-Spondins, Is Involved in Vascular Smooth Muscle Cell Calcification through LGR4 Interaction.

Vascular calcification has a global health impact that is closely linked to bone loss. The Receptor Activator of Nuclear Factor Kappa B (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) system, fundamental for bone metabolism, also plays an important role in vascular calcification. The Leucine-rich repeat-containing G-protein-coupled receptor 4 (LGR4), a novel receptor for RANKL, regulates bone remodeling, and it appears to be involved in vascular calcification. Besides RANKL, LGR4 interacts with R-spondins (RSPOs), which are known for their roles in bone but are less understood in vascular calcification. Studies were conducted in rats with chronic renal failure fed normal or high phosphorus diets for 18 weeks, with and without control of circulating parathormone (PTH) levels, resulting in different degrees of aortic calcification. Additionally, vascular smooth muscle cells (VSMCs) were cultured under non-calcifying (1 mM phosphate) and calcifying (3 mM phosphate) media with different concentrations of PTH. To explore the role of RANKL in VSMC calcification, increasing concentrations of soluble RANKL were added to non-calcifying and calcifying media. The effects mediated by RANKL binding to its receptor LGR4 were investigated by silencing the LGR4 receptor in VSMCs. Furthermore, the gene expression of the RANK/RANKL/OPG system and the ligands of LGR4 was assessed in human epigastric arteries obtained from kidney transplant recipients with calcification scores (Kauppila Index). Increased aortic calcium in rats coincided with elevated systolic blood pressure, upregulated Lgr4 and Rankl gene expression, downregulated Opg gene expression, and higher serum RANKL/OPG ratio without changes in Rspos gene expression. Elevated phosphate in vitro increased calcium content and expression of Rankl and Lgr4 while reducing Opg. Elevated PTH in the presence of high phosphate exacerbated the increase in calcium content. No changes in Rspos were observed under the conditions employed. The addition of soluble RANKL to VSMCs induced genotypic differentiation and calcification, partly prevented by LGR4 silencing. In the epigastric arteries of individuals presenting vascular calcification, the gene expression of RANKL was higher. While RSPOs show minimal impact on VSMC calcification, RANKL, interacting with LGR4, drives osteogenic differentiation in VSMCs, unveiling a novel mechanism beyond RANKL-RANK binding.

Inflammatory and Cardiovascular Biomarkers to Monitor Fabry Disease Progression.

Fabry disease is an invalidating multisystemic disorder affecting α-Galactosidase, a rate-limiting hydrolase dedicated to lipid catabolism. Non-metabolized substrates, such as Globotriaosylceramide and its derivatives trigger the direct or indirect activation of inflammatory events and endothelial dysfunction. In spite of the efficacy demonstrated by enzyme replacement therapy or pharmacological chaperones in delaying disease progression, few studies have analyzed whether these treatments can improve the pro-inflammatory state of FD patients. Therefore, the aim of this work was to assess cytokines and cardiovascular risk-related proteins detectable in plasma from FD patients, whether treated or not with ERT, to evaluate the reliability of these markers in monitoring disease stage and treatment effects. We identified inflammatory and endothelial dysfunction markers (ADAMTS-13, TNF-α, GDF-15, MIP-1ß, VEGFA, MPO, and MIC-1) that cooperate in a common pathway and are increased in FD patients' plasma samples. As shown by the assessment of these proteins over time, they can help to evaluate the risk of higher severity in FD, as well as ERT effects. Even though the analyzed proteins cannot be considered as proper biomarkers due to their non-specificity to FD, taken together they can provide a signature of reference molecules with prognostic value for early diagnosis, and evaluation of disease progression and treatment efficacy, using blood samples.

Forecast Cosmological Constraints with the 1D Wavelet Scattering Transform and the Lyman-α Forest.

We make forecasts for the constraining power of the 1D wavelet scattering transform when used with a Lyman-α forest cosmology survey. Using mock simulations and a Fisher matrix, we show that there is considerable cosmological information in the scattering transform coefficients not captured by the flux power spectrum. We estimate mock covariance matrices assuming uncorrelated Gaussian pixel noise for each quasar at a level drawn from a simple log-normal model. The extra information comes from a smaller estimated covariance in the first-order wavelet power and from second-order wavelet coefficients that probe non-Gaussian information in the forest. Forecast constraints on cosmological parameters from the wavelet scattering transform are more than an order of magnitude tighter than for the power spectrum, shrinking a 4D parameter space by a factor of 10^{6}. Should these improvements be realized with the Dark Energy Spectroscopic Instrument, inflationary running would be constrained to test common inflationary models predicting α_{s}=-6×10^{-4} and neutrino mass constraints would be improved enough for a 5-σ detection of the minimal neutrino mass.

INF2 formin variants linked to human inherited kidney disease reprogram the transcriptome, causing mitotic chaos and cell death.

Mutations in the human INF2 gene cause autosomal dominant focal segmental glomerulosclerosis (FSGS)-a condition characterized by podocyte loss, scarring, and subsequent kidney degeneration. To understand INF2-linked pathogenicity, we examined the effect of pathogenic INF2 on renal epithelial cell lines and human primary podocytes. Our study revealed an increased incidence of mitotic cells with surplus microtubule-organizing centers fostering multipolar spindle assembly, leading to nuclear abnormalities, particularly multi-micronucleation. The levels of expression of exogenous pathogenic INF2 were similar to those of endogenous INF2. The aberrant nuclear phenotypes were observed regardless of the expression method used (retrovirus infection or plasmid transfection) or the promoter (LTR or CMV) used, and were absent with exogenous wild type INF2 expression. This indicates that the effect of pathogenic INF2 is not due to overexpression or experimental cell manipulation, but instead to the intrinsic properties of pathogenic INF2. Inactivation of the INF2 catalytic domain prevented aberrant nuclei formation. Pathogenic INF2 triggered the translocation of the transcriptional cofactor MRTF into the nucleus. RNA sequencing revealed a profound alteration in the transcriptome that could be primarily attributed to the sustained activation of the MRTF-SRF transcriptional complex. Cells eventually underwent mitotic catastrophe and death. Reducing MRTF-SRF activation mitigated multi-micronucleation, reducing the extent of cell death. Our results, if validated in animal models, could provide insights into the mechanism driving glomerular degeneration in INF2-linked FSGS and may suggest potential therapeutic strategies for impeding FSGS progression.

Opposing effects of the purinergic P2X7 receptor on seizures in neurons and microglia in male mice.

BACKGROUND: The purinergic ATP-gated P2X7 receptor (P2X7R) is increasingly recognized to contribute to pathological neuroinflammation and brain hyperexcitability. P2X7R expression has been shown to be increased in the brain, including both microglia and neurons, in experimental models of epilepsy and patients. To date, the cell type-specific downstream effects of P2X7Rs during seizures remain, however, incompletely understood. METHODS: Effects of P2X7R signaling on seizures and epilepsy were analyzed in induced seizure models using male mice including the kainic acid model of status epilepticus and pentylenetetrazole model and in male and female mice in a genetic model of Dravet syndrome. RNA sequencing was used to analyze P2X7R downstream signaling during seizures. To investigate the cell type-specific role of the P2X7R during seizures and epilepsy, we generated mice lacking exon 2 of the P2rx7 gene in either microglia (P2rx7:Cx3cr1-Cre) or neurons (P2rx7:Thy-1-Cre). To investigate the protective potential of overexpressing P2X7R in GABAergic interneurons, P2X7Rs were overexpressed using adeno-associated virus transduction under the mDlx promoter. RESULTS: RNA sequencing of hippocampal tissue from wild-type and P2X7R knock-out mice identified both glial and neuronal genes, in particular genes involved in GABAergic signaling, under the control of the P2X7R following seizures. Mice with deleted P2rx7 in microglia displayed less severe acute seizures and developed a milder form of epilepsy, and microglia displayed an anti-inflammatory molecular profile. In contrast, mice lacking P2rx7 in neurons showed a more severe seizure phenotype when compared to epileptic wild-type mice. Analysis of single-cell expression data revealed that human P2RX7 expression is elevated in the hippocampus of patients with temporal lobe epilepsy in excitatory and inhibitory neurons. Functional studies determined that GABAergic interneurons display increased responses to P2X7R activation in experimental epilepsy. Finally, we show that viral transduction of P2X7R in GABAergic interneurons protects against evoked and spontaneous seizures in experimental temporal lobe epilepsy and in mice lacking Scn1a, a model of Dravet syndrome. CONCLUSIONS: Our results suggest a dual and opposing action of P2X7R in epilepsy and suggest P2X7R overexpression in GABAergic interneurons as a novel therapeutic strategy for acquired and, possibly, genetic forms of epilepsy.

Características clínicas y evolución del paciente anciano mayor de 80 años en una Unidad de Cuidados Intensivos / Clinical features and disease progression of elderlypatients at the ICU setting

Objetivo Describir y caracterizar una cohorte de pacientes octogenarios ingresados en la UCI del Hospital Universitario Central de Asturias (HUCA). Diseño Estudio retrospectivo, observacional y descriptivo de 14 meses de duración. Ámbito Unidad de Cuidados Intensivos (UCI) Cardiaca y UCI Polivalente del Servicio de Medicina Intensiva del HUCA (Oviedo). Participantes Pacientes mayores de 80 años que ingresaron en la UCI durante más de 24 horas.Intervenciones Ninguna. Variables de interés principales Edad, sexo, comorbilidad, capacidad funcional, tratamiento, complicaciones, evolución, mortalidad. Resultados Los motivos de ingreso más frecuentes fueron la cirugía cardiaca y la neumonía. La estancia media de ingreso fue significativamente mayor en pacientes menores de 85 años (p=0,037). El 84,3% de estos últimos se benefició de ventilación mecánica invasiva (VMI) vs. 46,2% de los pacientes más mayores (p=<0,001). Los pacientes mayores de 85 años presentaron mayor fragilidad. El ingreso por intervención quirúrgica cardiaca se asoció con menor riesgo de mortalidad (hazard ratio [HR]=0,18; intervalo de confianza [IC] 95%, 0,062-0,527; p=0,002). Conclusiones Los resultados muestran una asociación entre el motivo de ingreso en UCI y el riesgo de mortalidad en pacientes octogenarios. La cirugía cardiaca se asoció con mejor pronóstico frente a la patología médica, donde la neumonía se asoció con mayor riesgo de mortalidad. Además, se observó una relación positiva significativa entre edad y fragilidad. (AU)

ObjectiveTo describe and characterize a cohort of octogenarian patients admitted to the ICU of the University Central Hospital of Asturias (HUCA). Design Retrospective, observational and descriptive study of 14 months’ duration. Setting Cardiac and Medical Intensive Care Units (ICU) of the HUCA (Oviedo). Participants Patients over 80 years old who were admitted to the ICU for more than 24hours. Interventions None. Main variables of interest Age, sex, comorbidity, functional dependence, treatment, complications, evolution, mortality. Results The most frequent reasons for admission were cardiac surgery and pneumonia. The average admission stay was significantly longer in patients under 85 years of age (p=0,037). 84,3% of the latter benefited from invasive mechanical ventilation compared to 46,2% of older patients (p=<0,001). Patients over 85 years of age presented greater fragility. Admission for cardiac surgery was associated with a lower risk of mortality (HR=0,18; 95% CI (0,062-0,527; p=0,002). Conclusions The results have shown an association between the reason for admission to the ICU and the risk of mortality in octogenarian patients. Cardiac surgery was associated with a better prognosis compared to medical pathology, where pneumonia was associated with a higher risk of mortality. Furthermore, a significant positive association was observed between age and frailty. (AU)

Neutrophils secrete exosome-associated DNA to resolve sterile acute inflammation.

Acute inflammation, characterized by a rapid influx of neutrophils, is a protective response that can lead to chronic inflammatory diseases when left unresolved. Secretion of LTB 4 -containing exosomes is required for effective neutrophil infiltration during inflammation. In this study, we show that neutrophils release nuclear DNA in a non-lytic, rapid, and repetitive manner, via a mechanism distinct from suicidal NET release and cell death. The packaging of nuclear DNA occurs in the lumen of nuclear envelope (NE)-derived multivesicular bodies (MVBs) that harbor the LTB 4 synthesizing machinery and is mediated by the lamin B receptor (LBR) and chromatin decondensation. Disruption of secreted exosome-associated DNA (SEAD) in a model of sterile inflammation in mouse skin amplifies and prolongs the presence of neutrophils, impeding the onset of resolution. Together, these findings advance our understanding of neutrophil functions during inflammation and the physiological significance of NETs, with implications for novel treatments for inflammatory disorders.

Thrombus discrimination with electron density: potential implications for non-contrast computed tomography imaging.

The potential clinical usefulness of electron density (ED) imaging, that can be directly estimated using dual-layer spectral computed tomography (CT), has been poorly investigated. We explored whether ED imaging might improve thrombus identification compared to conventional imaging in vitro. We evaluated mechanical thrombectomy material obtained from patients with acute ischemic stroke (AIS) treated in a tertiary level stroke center and immediately fixed in 10% neutral buffered formalin and stored in polystyrene test tubes. The test tubes were immersed in a bucket of water for evaluation by spectral CT, along with scattered control tubes. All images were obtained using a dual-layer detector CT scanner. Each tube was assessed using multiparametric side-by-side view of conventional CT (120 kVp), low monoenergetic imaging (40 keV), and ED images. Fifty-eight polystyrene tubes were analyzed, comprising 52 tubes with thrombectomy material of at least 1 mm2 size obtained from 52 AIS patients, and six control tubes filled with formalin. ED imaging identified accurately the presence of material in all tubes, whereas 2 (3%) of the tubes containing thrombus were not identified by conventional CT, leading to a very good agreement between observers for the presence of material using conventional CT and ED imaging (kappa =0.84, P<0.001). Using ED imaging, thrombus material showed a mean density of 108.8±2.9 percent ED relative to water (%EDW), water had a mean density of 100.0±0.3 %EDW, and formalin a mean density of 103.5±1.2 %EDW. Compared to conventional imaging and 40 keV monoenergetic, ED imaging had a significantly higher signal-to-noise ratio (conventional 10.4±7.0, vs. 40 keV 11.5±8.4, vs. ED 490.0±304.5, P<0.001) and contrast-to-noise ratio (CNR) (conventional 4.3±4.3, vs. 40 keV 5.7±11.2, vs. ED 37.8±29.1, P<0.001). In this in-vitro study, we demonstrated improved visualization of thrombus with ED imaging compared to conventional imaging and low monoenergetic imaging, with a significant increase in CNR.

H2AX promotes replication fork degradation and chemosensitivity in BRCA-deficient tumours.

Histone H2AX plays a key role in DNA damage signalling in the surrounding regions of DNA double-strand breaks (DSBs). In response to DNA damage, H2AX becomes phosphorylated on serine residue 139 (known as γH2AX), resulting in the recruitment of the DNA repair effectors 53BP1 and BRCA1. Here, by studying resistance to poly(ADP-ribose) polymerase (PARP) inhibitors in BRCA1/2-deficient mammary tumours, we identify a function for γH2AX in orchestrating drug-induced replication fork degradation. Mechanistically, γH2AX-driven replication fork degradation is elicited by suppressing CtIP-mediated fork protection. As a result, H2AX loss restores replication fork stability and increases chemoresistance in BRCA1/2-deficient tumour cells without restoring homology-directed DNA repair, as highlighted by the lack of DNA damage-induced RAD51 foci. Furthermore, in the attempt to discover acquired genetic vulnerabilities, we find that ATM but not ATR inhibition overcomes PARP inhibitor (PARPi) resistance in H2AX-deficient tumours by interfering with CtIP-mediated fork protection. In summary, our results demonstrate a role for H2AX in replication fork biology in BRCA-deficient tumours and establish a function of H2AX separable from its classical role in DNA damage signalling and DSB repair.

Frustraevo: a web server to localize and quantify the conservation of local energetic frustration in protein families.

According to the Principle of Minimal Frustration, folded proteins can only have a minimal number of strong energetic conflicts in their native states. However, not all interactions are energetically optimized for folding but some remain in energetic conflict, i.e. they are highly frustrated. This remaining local energetic frustration has been shown to be statistically correlated with distinct functional aspects such as protein-protein interaction sites, allosterism and catalysis. Fuelled by the recent breakthroughs in efficient protein structure prediction that have made available good quality models for most proteins, we have developed a strategy to calculate local energetic frustration within large protein families and quantify its conservation over evolutionary time. Based on this evolutionary information we can identify how stability and functional constraints have appeared at the common ancestor of the family and have been maintained over the course of evolution. Here, we present FrustraEvo, a web server tool to calculate and quantify the conservation of local energetic frustration in protein families.

Market analysis of the presence of endocrine disrupting chemicals in cosmetic products intended for oncological patients and other vulnerable groups.

There is growing concern about the presence of endocrine disrupting chemicals (EDCs) in cosmetics. We aimed to identify the main cosmetic ingredients with suspected endocrine-disrupting properties, and analyse their presence in current marketed products. Particular attention was given to products intended for susceptible (due to physiological status) and vulnerable (due to specific pathologies) groups with a view to informing cosmetologists and related health professionals of the scientific basis and current status of any concerns. Suspected EDCs used as cosmetic ingredients, included in lists published by regulatory agencies, were documented and investigated by weight of evidence analysis based on endocrine-related toxicity studies. In total, 49 suspected EDCs were identified from a sample of over a thousand cosmetic products marketed in the European Union. Suspected EDCs were found in approximately one third of products, with a similar frequency in products intended for susceptible and vulnerable groups. Avobenzone (CAS number:70356-09-1), octisalate (CAS number: 118-60-5), and butylated hydroxytoluene (CAS number: 128-37-0) were mostly commonly identified. The presence of EDCs was particularly high for sun care cosmetic products. Our results highlight potentially significant exposure through cosmetics to substances currently studied by regulatory institutions as suspected endocrine disrupters. EDCs are not yet universally regulated, and informing health professionals and educating the population as a precaution are options to reduce individual exposure levels, especially in vulnerable and susceptible groups. Special recommendations are needed for products intended for oncological patients.

Barriers and enablers to people-centred viral hepatitis care in Vietnam and the Philippines: Results of a patient journey mapping study.

In Vietnam and the Philippines, viral hepatitis is the leading cause of cirrhosis and liver cancer. This study aims to understand the barriers and enablers of people receiving care for hepatitis B and C to support both countries' efforts to eliminate viral hepatitis as a public health threat by 2030. Retrospective, semi-structured interviews were conducted with a purposive, quota-based sample of 63 people living with hepatitis B or C in one province of Vietnam and one region of the Philippines. A rapid deductive approach to thematic analysis produced key findings among the three phases of care: (1) pre-awareness and testing, (2) linkage and treatment initiation and (3) ongoing treatment and recovery. The research found that participants followed five typical journeys, from a variety of entry points. Barriers during the pre-awareness and testing phase included limited awareness about hepatitis and its management, stigma and psychological impacts. Enablers included being familiar with the health system and/or patients benefiting from social connections within the health systems. During the linkage and treatment initiation phase, barriers included difficult physical access, complex navigation and inadequate counselling. In this phase, family support emerged as a critical enabler. During the ongoing treatment and recovery phase, the cost of care and socially and culturally informed perceptions of the disease and medication use were both barriers and enablers. Exploring peoples' journeys with hepatitis B and C in Vietnam and the Philippines revealed many similarities despite the different cultural and health system contexts. Insights from this study may help generate a contextualized, people-centred evidence base to inform the design and improvement of primary care services for hepatitis in both research sites.

Comparison of ICSI, IVF, and in vivo derived embryos to produce CRISPR-Cas9 gene-edited pigs for xenotransplantation.

Genome editing in pigs for xenotransplantation has seen significant advances in recent years. This study compared three methodologies to generate gene-edited embryos, including co-injection of sperm together with the CRISPR-Cas9 system into oocytes, named ICSI-MGE (mediated gene editing); microinjection of CRISPR-Cas9 components into oocytes followed by in vitro fertilization (IVF), and microinjection of in vivo fertilized zygotes with the CRISPR-Cas9 system. Our goal was to knock-out (KO) porcine genes involved in the biosynthesis of xenoantigens responsible for the hyperacute rejection of interspecific xenografts, namely GGTA1, CMAH, and ß4GalNT2. Additionally, we attempted to KO the growth hormone receptor (GHR) gene with the aim of limiting the growth of porcine organs to a size that is physiologically suitable for human transplantation. Embryo development, pregnancy, and gene editing rates were evaluated. We found an efficient mutation of the GGTA1 gene following ICSI-MGE, comparable to the results obtained through the microinjection of oocytes followed by IVF. ICSI-MGE also showed higher rates of biallelic mutations compared to the other techniques. Five healthy piglets were born from in vivo-derived embryos, all of them exhibiting biallelic mutations in the GGTA1 gene, with three displaying mutations in the GHR gene. No mutations were observed in the CMAH and ß4GalNT2 genes. In conclusion, in vitro methodologies showed high rates of gene-edited embryos. Specifically, ICSI-MGE proved to be an efficient technique for obtaining homozygous biallelic mutated embryos. Lastly, only live births were obtained from in vivo-derived embryos showing efficient multiple gene editing for GGTA1 and GHR.

Household mixed plastic waste derived adsorbents for CO2 capture: A feasibility study.

The feasibility of producing activated carbon (AC) from real Household Mixed Plastic Waste (HMPW) comprising of LDPE, HDPE, PP, PS, and PET for carbon capture via direct carbonisation followed by microwave-assisted or conventional thermally assisted chemical activation was investigated. A microwave-assisted activation procedure was adopted to assess the impact on the CO2 capture capacity of the resulting AC using both a lower temperature (400 °C vs. 700 °C) and a shorter duration (5 vs. 120 mins) than that required for conventional activation. The results obtained showed that the AC yield was 71 and 78% for the conventional and microwave-assisted samples, respectively. Microwave activation consumed five-fold less energy (0.19 kWh) than the conventional activation (0.98 kWh). Thermal stability results indicated total weight loss of 10.0 and 8.3 wt%, respectively, for conventional and microwave-activated samples over the temperature range of 25-1000 °C, with ACs from both activation routes displaying a type 1 nitrogen isotherm. The dynamic CO2 uptake capacity at 1 bar and 25 °C was 1.53 mmol/g, with maximum equilibrium uptake ranging between 1.32 and 2.39 mmol/g at temperatures (0-50 °C) and 1 bar for the conventionally activated AC. The analogous microwave-activated sample showed a higher dynamic CO2 uptake of 1.62 mmol/g and equilibrium uptake in the range 1.58-2.88 mmol/g under equivalent conditions. The results therefore indicate that microwave activation results in enhanced carbon capture potential. To the best of our knowledge, this is the first-time microwave heating has been employed to convert household mixed plastic wastes directly into ACs for carbon capture applications. This report therefore demonstrates that the management of mixed plastics could lead to the development of a circular economy through the conversion of waste into value-added materials.

Novel Biomarkers of Bone Metabolism.

Bone represents a metabolically active tissue subject to continuous remodeling orchestrated by the dynamic interplay between osteoblasts and osteoclasts. These cellular processes are modulated by a complex interplay of biochemical and mechanical factors, which are instrumental in assessing bone remodeling. This comprehensive evaluation aids in detecting disorders arising from imbalances between bone formation and reabsorption. Osteoporosis, characterized by a reduction in bone mass and strength leading to heightened bone fragility and susceptibility to fractures, is one of the more prevalent chronic diseases. Some epidemiological studies, especially in patients with chronic kidney disease (CKD), have identified an association between osteoporosis and vascular calcification. Notably, low bone mineral density has been linked to an increased incidence of aortic calcification, with shared molecules, mechanisms, and pathways between the two processes. Certain molecules emerging from these shared pathways can serve as biomarkers for bone and mineral metabolism. Detecting and evaluating these alterations early is crucial, requiring the identification of biomarkers that are reliable for early intervention. While traditional biomarkers for bone remodeling and vascular calcification exist, they suffer from limitations such as low specificity, low sensitivity, and conflicting results across studies. In response, efforts are underway to explore new, more specific biomarkers that can detect alterations at earlier stages. The aim of this review is to comprehensively examine some of the emerging biomarkers in mineral metabolism and their correlation with bone mineral density, fracture risk, and vascular calcification as well as their potential use in clinical practice.

Clinical features and disease progression of elderly patients at the ICU setting.

OBJECTIVE: To describe and characterize a cohort of octogenarian patients admitted to the ICU of the University Central Hospital of Asturias (HUCA). DESIGN: Retrospective, observational and descriptive study of 14 months' duration. SETTING: Cardiac and Medical intensive care units (ICU) of the HUCA (Oviedo). PARTICIPANTS: Patients over 80 years old who were admitted to the ICU for more than 24 h. INTERVENTIONS: None. MAIN VARIABLES OF INTEREST: Age, sex, comorbidity, functional dependence, treatment, complications, evolution, mortality. RESULTS: The most frequent reasons for admission were cardiac surgery and pneumonia. The average admission stay was significantly longer in patients under 85 years of age (p = 0,037). 84,3% of the latter benefited from invasive mechanical ventilation compared to 46,2% of older patients (p = <0,001). Patients over 85 years of age presented greater fragility. Admission for cardiac surgery was associated with a lower risk of mortality (HR = 0,18; 95% CI (0,062-0,527; p = 0,002). CONCLUSIONS: The results have shown an association between the reason for admission to the ICU and the risk of mortality in octogenarian patients. Cardiac surgery was associated with a better prognosis compared to medical pathology, where pneumonia was associated with a higher risk of mortality. Furthermore, a significant positive association was observed between age and frailty.

Soluble Klotho, a Potential Biomarker of Chronic Kidney Disease-Mineral Bone Disorders Involved in Healthy Ageing: Lights and Shadows.

Shortly after the discovery of Klotho, interest grew in its potential role in chronic kidney disease (CKD). There are three isoforms of the Klotho protein: αKlotho, ßKlotho and γKlotho. This review will focus on αKlotho due to its relevance as a biomarker in CKD. αKlotho is synthesized mainly in the kidneys, but it can be released into the bloodstream and urine as soluble Klotho (sKlotho), which undertakes systemic actions, independently or in combination with FGF23. It is usually accepted that sKlotho levels are reduced early in CKD and that lower levels of sKlotho might be associated with the main chronic kidney disease-mineral bone disorders (CKD-MBDs): cardiovascular and bone disease. However, as results are inconsistent, the applicability of sKlotho as a CKD-MBD biomarker is still a matter of controversy. Much of the inconsistency can be explained due to low sample numbers, the low quality of clinical studies, the lack of standardized assays to assess sKlotho and a lack of consensus on sample processing, especially in urine. In recent decades, because of our longer life expectancies, the prevalence of accelerated-ageing diseases, such as CKD, has increased. Exercise, social interaction and caloric restriction are considered key factors for healthy ageing. While exercise and social interaction seem to be related to higher serum sKlotho levels, it is not clear whether serum sKlotho might be influenced by caloric restriction. This review focuses on the possible role of sKlotho as a biomarker in CKD-MBD, highlighting the difference between solid knowledge and areas requiring further research, including the role of sKlotho in healthy ageing.

Oocyte electroporation prior to in vitro fertilization is an efficient method to generate single, double, and multiple knockout porcine embryos of interest in biomedicine and animal production.

Genetically modified pigs play a critical role in mimicking human diseases, xenotransplantation, and the development of pigs resistant to viral diseases. The use of programmable endonucleases, including the CRISPR/Cas9 system, has revolutionized the generation of genetically modified pigs. This study evaluates the efficiency of electroporation of oocytes prior to fertilization in generating edited gene embryos for different models. For single gene editing, phospholipase C zeta (PLC ζ) and fused in sarcoma (FUS) genes were used, and the concentration of sgRNA and Cas9 complexes was optimized. The results showed that increasing the concentration resulted in higher mutation rates without affecting the blastocyst rate. Electroporation produced double knockouts for the TPC1/TPC2 genes with high efficiency (79 %). In addition, resistance to viral diseases such as PRRS and swine influenza was achieved by electroporation, allowing the generation of double knockout embryo pigs (63 %). The study also demonstrated the potential for multiple gene editing in a single step using electroporation, which is relevant for xenotransplantation. The technique resulted in the simultaneous mutation of 5 genes (GGTA1, B4GALNT2, pseudo B4GALNT2, CMAH and GHR). Overall, electroporation proved to be an efficient and versatile method to generate genetically modified embryonic pigs, offering significant advances in biomedical and agricultural research, xenotransplantation, and disease resistance. Electroporation led to the processing of numerous oocytes in a single session using less expensive equipment. We confirmed the generation of gene-edited porcine embryos for single, double, or quintuple genes simultaneously without altering embryo development to the blastocyst stage. The results provide valuable insights into the optimization of gene editing protocols for different models, opening new avenues for research and applications in this field.

Uninformed Teacher-Student for hard-samples distillation in weakly supervised mitosis localization.

BACKGROUND AND OBJECTIVE: Mitotic activity is a crucial biomarker for diagnosing and predicting outcomes for different types of cancers, particularly breast cancer. However, manual mitosis counting is challenging and time-consuming for pathologists, with moderate reproducibility due to biopsy slide size, low mitotic cell density, and pattern heterogeneity. In recent years, deep learning methods based on convolutional neural networks (CNNs) have been proposed to address these limitations. Nonetheless, these methods have been hampered by the available data labels, which usually consist only of the centroids of mitosis, and by the incoming noise from annotated hard negatives. As a result, complex algorithms with multiple stages are often required to refine the labels at the pixel level and reduce the number of false positives. METHODS: This article presents a novel weakly supervised approach for mitosis detection that utilizes only image-level labels on histological hematoxylin and eosin (H&E) images, avoiding the need for complex labeling scenarios. Also, an Uninformed Teacher-Student (UTS) pipeline is introduced to detect and distill hard samples by comparing weakly supervised localizations and the annotated centroids, using strong augmentations to enhance uncertainty. Additionally, an automatic proliferation score is proposed that mimicks the pathologist-annotated mitotic activity index (MAI). The proposed approach is evaluated on three publicly available datasets for mitosis detection on breast histology samples, and two datasets for mitotic activity counting in whole-slide images. RESULTS: The proposed framework achieves competitive performance with relevant prior literature in all the datasets used for evaluation without explicitly using the mitosis location information during training. This approach challenges previous methods that rely on strong mitosis location information and multiple stages to refine false positives. Furthermore, the proposed pipeline for hard-sample distillation demonstrates promising dataset-specific improvements. Concretely, when the annotation has not been thoroughly refined by multiple pathologists, the UTS model offers improvements of up to ∼4% in mitosis localization, thanks to the detection and distillation of uncertain cases. Concerning the mitosis counting task, the proposed automatic proliferation score shows a moderate positive correlation with the MAI annotated by pathologists at the biopsy level on two external datasets. CONCLUSIONS: The proposed Uninformed Teacher-Student pipeline leverages strong augmentations to distill uncertain samples and measure dissimilarities between predicted and annotated mitosis. Results demonstrate the feasibility of the weakly supervised approach and highlight its potential as an objective evaluation tool for tumor proliferation.