Fueling the heartbeat: Dynamic regulation of intracellular ATP during excitation-contraction coupling in ventricular myocytes.

The heart beats approximately 100,000 times per day in humans, imposing substantial energetic demands on cardiac muscle. Adenosine triphosphate (ATP) is an essential energy source for normal function of cardiac muscle during each beat, as it powers ion transport, intracellular Ca2+ handling, and actin-myosin cross-bridge cycling. Despite this, the impact of excitation-contraction coupling on the intracellular ATP concentration ([ATP]i) in myocytes is poorly understood. Here, we conducted real-time measurements of [ATP]i in ventricular myocytes using a genetically encoded ATP fluorescent reporter. Our data reveal rapid beat-to-beat variations in [ATP]i. Notably, diastolic [ATP]i was <1 mM, which is eightfold to 10-fold lower than previously estimated. Accordingly, ATP-sensitive K+ (KATP) channels were active at physiological [ATP]i. Cells exhibited two distinct types of ATP fluctuations during an action potential: net increases (Mode 1) or decreases (Mode 2) in [ATP]i. Mode 1 [ATP]i increases necessitated Ca2+ entry and release from the sarcoplasmic reticulum (SR) and were associated with increases in mitochondrial Ca2+. By contrast, decreases in mitochondrial Ca2+ accompanied Mode 2 [ATP]i decreases. Down-regulation of the protein mitofusin 2 reduced the magnitude of [ATP]i fluctuations, indicating that SR-mitochondrial coupling plays a crucial role in the dynamic control of ATP levels. Activation of ß-adrenergic receptors decreased [ATP]i, underscoring the energetic impact of this signaling pathway. Finally, our work suggests that cross-bridge cycling is the largest consumer of ATP in a ventricular myocyte during an action potential. These findings provide insights into the energetic demands of EC coupling and highlight the dynamic nature of ATP concentrations in cardiac muscle.

Cerebral organoids display dynamic clonal growth and tunable tissue replenishment.

During brain development, neural progenitors expand through symmetric divisions before giving rise to differentiating cell types via asymmetric divisions. Transition between those modes varies among individual neural stem cells, resulting in clones of different sizes. Imaging-based lineage tracing allows for lineage analysis at high cellular resolution but systematic approaches to analyse clonal behaviour of entire tissues are currently lacking. Here we implement whole-tissue lineage tracing by genomic DNA barcoding in 3D human cerebral organoids, to show that individual stem cell clones produce progeny on a vastly variable scale. By using stochastic modelling we find that variable lineage sizes arise because a subpopulation of lineages retains symmetrically dividing cells. We show that lineage sizes can adjust to tissue demands after growth perturbation via chemical ablation or genetic restriction of a subset of cells in chimeric organoids. Our data suggest that adaptive plasticity of stem cell populations ensures robustness of development in human brain organoids.

BIN1 knockdown rescues systolic dysfunction in aging male mouse hearts.

Cardiac dysfunction is a hallmark of aging in humans and mice. Here we report that a two-week treatment to restore youthful Bridging Integrator 1 (BIN1) levels in the hearts of 24-month-old mice rejuvenates cardiac function and substantially reverses the aging phenotype. Our data indicate that age-associated overexpression of BIN1 occurs alongside dysregulated endosomal recycling and disrupted trafficking of cardiac CaV1.2 and type 2 ryanodine receptors. These deficiencies affect channel function at rest and their upregulation during acute stress. In vivo echocardiography reveals reduced systolic function in old mice. BIN1 knockdown using an adeno-associated virus serotype 9 packaged shRNA-mBIN1 restores the nanoscale distribution and clustering plasticity of ryanodine receptors and recovers Ca2+ transient amplitudes and cardiac systolic function toward youthful levels. Enhanced systolic function correlates with increased phosphorylation of the myofilament protein cardiac myosin binding protein-C. These results reveal BIN1 knockdown as a novel therapeutic strategy to rejuvenate the aging myocardium.

Regional modulation of toll-like receptor signaling pathway genes in acute epididymitis in mice.

BACKGROUND: Region-specific immune environments in the epididymis influence the immune responses to uropathogenic Escherichia coli (UPEC) infection, a relevant cause of epididymitis in men. Toll-like receptors (TLRs) are essential to orchestrate immune responses against bacterial infections. The epididymis displays region-specific inflammatory responses to bacterial-derived TLR agonists, such as lipopolysaccharide (LPS; TLR4 agonist) and lipoteichoic acid (LTA; TLR2/TLR6 agonist), suggesting that TLR-associated signaling pathways could influence the magnitude of inflammatory responses in epididymitis. OBJECTIVES: To investigate the expression and regulation of key genes associated with TLR4 and TLR2/TLR6 signaling pathways during epididymitis induced by UPEC, LPS, and LTA in mice. MATERIAL AND METHODS: Epididymitis was induced in mice using UPEC, ultrapure LPS, or LTA, injected into the interstitial space of the initial segment or the lumen of the vas deferens close to the cauda epididymidis. Samples were harvested after 1, 5, and 10 days for UPEC-treated animals and 6 and 24 h for LPS-/LTA-treated animals. Ex vivo epididymitis was induced by incubating epididymal regions from naive mice with LPS or LTA. RT-qPCR and Western blot assays were conducted. RESULTS: UPEC infection up-regulated Tlr2, Tlr4, and Tlr6 transcripts and their associated signaling molecules Cd14, Ticam1, and Traf6 in the cauda epididymidis but not in the initial segment. In these epididymal regions, LPS and LTA differentially modulated Tlr2, Tlr4, Tlr6, Cd14, Myd88, Ticam1, Traf3, and Traf6 expression levels. NFKB and AP1 activation was required for LPS- and LTA-induced up-regulation of TLR-associated signaling transcripts in the cauda epididymidis and initial segment, respectively. CONCLUSION: The dynamic modulation of TLR4 and TLR2/TLR6 signaling pathways gene expression during epididymitis indicates bacterial-derived antigens elicit an increased tissue sensitivity to combat microbial infection in a spatial manner in the epididymis. Differential activation of TLR-associated signaling pathways may contribute to fine-tuning inflammatory responses along the epididymis.

A cell fate decision map reveals abundant direct neurogenesis bypassing intermediate progenitors in the human developing neocortex.

The human neocortex has undergone strong evolutionary expansion, largely due to an increased progenitor population, the basal radial glial cells. These cells are responsible for the production of a diversity of cell types, but the successive cell fate decisions taken by individual progenitors remain unknown. Here we developed a semi-automated live/fixed correlative imaging method to map basal radial glial cell division modes in early fetal tissue and cerebral organoids. Through the live analysis of hundreds of dividing progenitors, we show that basal radial glial cells undergo abundant symmetric amplifying divisions, and frequent self-consuming direct neurogenic divisions, bypassing intermediate progenitors. These direct neurogenic divisions are more abundant in the upper part of the subventricular zone. We furthermore demonstrate asymmetric Notch activation in the self-renewing daughter cells, independently of basal fibre inheritance. Our results reveal a remarkable conservation of fate decisions in cerebral organoids, supporting their value as models of early human neurogenesis.

Fairness of Machine Learning Algorithms for Predicting Foregone Preventive Dental Care for Adults.

Importance: Access to routine dental care prevents advanced dental disease and improves oral and overall health. Identifying individuals at risk of foregoing preventive dental care can direct prevention efforts toward high-risk populations. Objective: To predict foregone preventive dental care among adults overall and in sociodemographic subgroups and to assess the algorithmic fairness. Design, Setting, and Participants: This prognostic study was a secondary analyses of longitudinal data from the US Medical Expenditure Panel Survey (MEPS) from 2016 to 2019, each with 2 years of follow-up. Participants included adults aged 18 years and older. Data analysis was performed from December 2022 to June 2023. Exposure: A total of 50 predictors, including demographic and socioeconomic characteristics, health conditions, behaviors, and health services use, were assessed. Main Outcomes and Measures: The outcome of interest was foregoing preventive dental care, defined as either cleaning, general examination, or an appointment with the dental hygienist, in the past year. Results: Among 32 234 participants, the mean (SD) age was 48.5 (18.2) years and 17 386 participants (53.9%) were female; 1935 participants (6.0%) were Asian, 5138 participants (15.9%) were Black, 7681 participants (23.8%) were Hispanic, 16 503 participants (51.2%) were White, and 977 participants (3.0%) identified as other (eg, American Indian and Alaska Native) or multiple racial or ethnic groups. There were 21 083 (65.4%) individuals who missed preventive dental care in the past year. The algorithms demonstrated high performance, achieving an area under the receiver operating characteristic curve (AUC) of 0.84 (95% CI, 0.84-0.85) in the overall population. While the full sample model performed similarly when applied to White individuals and older adults (AUC, 0.88; 95% CI, 0.87-0.90), there was a loss of performance for other subgroups. Removing the subgroup-sensitive predictors (ie, race and ethnicity, age, and income) did not impact model performance. Models stratified by race and ethnicity performed similarly or worse than the full model for all groups, with the lowest performance for individuals who identified as other or multiple racial groups (AUC, 0.76; 95% CI, 0.70-0.81). Previous pattern of dental visits, health care utilization, dental benefits, and sociodemographic characteristics were the highest contributing predictors to the models' performance. Conclusions and Relevance: Findings of this prognostic study using cohort data suggest that tree-based ensemble machine learning models could accurately predict adults at risk of foregoing preventive dental care and demonstrated bias against underrepresented sociodemographic groups. These results highlight the importance of evaluating model fairness during development and testing to avoid exacerbating existing biases.

Proteomic analysis of Cryptostegia grandiflora latex, purification, characterization, and biological activity of two osmotin isoforms.

Although latex fluids are found in >20,000 plant species, the biochemical composition and biological function of their proteins are still poorly explored. Thus, this work aimed to conduct a proteomic analysis of Cryptostegia grandiflora latex (CgLP) for subsequent purification and characterization of an antifungal protein. After 2D-SDS-PAGE and mass spectrometry, 27 proteins were identified in CgLP, including a polygalacturonase inhibitor, cysteine peptidases, pathogenesis-related proteins (PR-4), and osmotins. Then, two osmotin isoforms (CgOsm) were purified, and a unique N-terminal sequence was determined (1ATFDIRSNCPYTVWAAAVPGGGRRLDRGQTWTINVAPGTA40). The PCR products revealed a cDNA sequence of 609 nucleotides for CgOsm, which encoded a polypeptide with 203 amino acid residues. The structure of CgOsm has features of typical osmotin or thaumatin-like proteins (TLPs), such as 16 conserved Cys residues, REDDD and FF motifs, an acidic cleft, and three main domains. Atomic force microscopy (AFM) and bioinformatics suggested that CgOsm is associated with three chain units. This result was interesting since the literature describes osmotins and TLPs as monomers. AFM also showed that Fusarium falciforme spores treated with CgOsm were drastically damaged. Therefore, it is speculated that CgOsm forms pores in the membrane of these cells, causing the leakage of cytoplasmic content.

Prescription and Effects of Cardiorespiratory Training in Individuals with Intellectual Disability: A Systematic Review.

This study aims to systematize effects of cardiorespiratory training (CT) programs in individuals with intellectual disability (ID) and identifying the fundamental and structuring aspects for the prescription of CT. This systematic review was carried out through four databases (Pubmed, Web of Science, Scopus, and SPORTDiscus), considering data from the period between 2013 and 2022. From 257 studies, 12 studies were included in this systematic review. Three studies used interval CT, while seven used continuous CT. Seven were carried out in the population with Down syndrome, while only three were carried out with participants with ID. The CT programs had the following characteristics: duration of 8 to 12 weeks, weekly frequency of three sessions, for 20 to 60 min, the intensity of 50% to 80% of maximal heart rate or 70% to 80% of peak oxygen consumption, using an ergometer cycle or an outdoor walking. The studies reported improvements in cardiorespiratory function, lipid, hemodynamic and metabolic profile, body composition, and neuromuscular and cognitive capacity. This review presents characteristics and recommendations that technicians can follow when structuring, prescribing, and implementing CT programs to individuals with ID.

Diagnosing and treating pericarditis and myocarditis in the emergency department.

Pericarditis and myocarditis represent a challenging set of diseases to diagnose and treat. These diseases typically present with chest pain and dyspnea in previously healthy young people, often in the weeks following a viral illness, including COVID-19. Nonetheless, the etiologies can be very diverse, including infectious, noninfectious, drug-induced, and autoimmune causes. This review focuses on the evaluation, diagnosis, and management of emergency department patients presenting with pericarditis and myocarditis and summarizes current guidelines and best-practice medical management strategies in order to avoid potential life-threatening cardiac complications.

The microtubule cytoskeleton of radial glial progenitor cells.

A high number of genetic mutations associated with cortical malformations are found in genes coding for microtubule-related factors. This has stimulated research to understand how the various microtubule-based processes are regulated to build a functional cerebral cortex. Here, we focus our review on the radial glial progenitor cells, the stem cells of the developing neocortex, summarizing research mostly performed in rodents and humans. We highlight how the centrosomal and acentrosomal microtubule networks are organized during interphase to support polarized transport and proper attachment of the apical and basal processes. We describe the molecular mechanism for interkinetic nuclear migration (INM), a microtubule-dependent oscillation of the nucleus. Finally, we describe how the mitotic spindle is built to ensure proper chromosome segregation, with a strong focus on factors mutated in microcephaly.

A Machine Learning Prediction Model for Immediate Graft Function After Deceased Donor Kidney Transplantation.

BACKGROUND: After kidney transplantation (KTx), the graft can evolve from excellent immediate graft function (IGF) to total absence of function requiring dialysis. Recipients with IGF do not seem to benefit from using machine perfusion, an expensive procedure, in the long term when compared with cold storage. This study proposes to develop a prediction model for IGF in KTx deceased donor patients using machine learning algorithms. METHODS: Unsensitized recipients who received their first KTx deceased donor between January 1, 2010, and December 31, 2019, were classified according to the conduct of renal function after transplantation. Variables related to the donor, recipient, kidney preservation, and immunology were used. The patients were randomly divided into 2 groups: 70% were assigned to the training and 30% to the test group. Popular machine learning algorithms were used: eXtreme Gradient Boosting (XGBoost), Light Gradient Boosting Machine, Gradient Boosting classifier, Logistic Regression, CatBoost classifier, AdaBoost classifier, and Random Forest classifier. Comparative performance analysis on the test dataset was performed using the results of the AUC values, sensitivity, specificity, positive predictive value, negative predictive value, and F1 score. RESULTS: Of the 859 patients, 21.7% (n = 186) had IGF. The best predictive performance resulted from the eXtreme Gradient Boosting model (AUC, 0.78; 95% CI, 0.71-0.84; sensitivity, 0.64; specificity, 0.78). Five variables with the highest predictive value were identified. CONCLUSIONS: Our results indicated the possibility of creating a model for the prediction of IGF, enhancing the selection of patients who would benefit from an expensive treatment, as in the case of machine perfusion preservation.

Coating of SPIONs with a Cysteine-Decorated Copolyester: A Possible Novel Nanoplatform for Enzymatic Release.

Superparamagnetic iron oxide nanoparticles (SPIONs) have their use approved for the diagnosis/treatment of malignant tumors and can be metabolized by the organism. To prevent embolism caused by these nanoparticles, they need to be coated with biocompatible and non-cytotoxic materials. Here, we synthesized an unsaturated and biocompatible copolyester, poly (globalide-co-ε-caprolactone) (PGlCL), and modified it with the amino acid cysteine (Cys) via a thiol-ene reaction (PGlCLCys). The Cys-modified copolymer presented reduced crystallinity and increased hydrophilicity in comparison to PGlCL, thus being used for the coating of SPIONS (SPION@PGlCLCys). Additionally, cysteine pendant groups at the particle's surface allowed the direct conjugation of (bio)molecules that establish specific interactions with tumor cells (MDA-MB 231). The conjugation of either folic acid (FA) or the anti-cancer drug methotrexate (MTX) was carried out directly on the amine groups of cysteine molecules present in the SPION@PGlCLCys surface (SPION@PGlCLCys_FA and SPION@PGlCLCys_MTX) by carbodiimide-mediated coupling, leading to the formation of amide bonds, with conjugation efficiencies of 62% for FA and 60% for MTX. Then, the release of MTX from the nanoparticle surface was evaluated using a protease at 37 °C in phosphate buffer pH~5.3. It was found that 45% of MTX conjugated to the SPIONs were released after 72 h. Cell viability was measured by MTT assay, and after 72 h, 25% reduction in cell viability of tumor cells was observed. Thus, after a successful conjugation and subsequent triggered release of MTX, we understand that SPION@PGlCLCys has a strong potential to be treated as a model nanoplatform for the development of treatments and diagnosis techniques (or theranostic applications) that can be less aggressive to patients.

Acute phosphatidylinositol 4,5 bisphosphate depletion destabilizes sarcolemmal expression of cardiac L-type Ca2+ channel CaV1.2.

CaV1.2 channels are critical players in cardiac excitation-contraction coupling, yet we do not understand how they are affected by an important therapeutic target of heart failure drugs and regulator of blood pressure, angiotensin II. Signaling through Gq-coupled AT1 receptors, angiotensin II triggers a decrease in PIP2, a phosphoinositide component of the plasma membrane (PM) and known regulator of many ion channels. PIP2 depletion suppresses CaV1.2 currents in heterologous expression systems but the mechanism of this regulation and whether a similar phenomenon occurs in cardiomyocytes is unknown. Previous studies have shown that CaV1.2 currents are also suppressed by angiotensin II. We hypothesized that these two observations are linked and that PIP2 stabilizes CaV1.2 expression at the PM and angiotensin II depresses cardiac excitability by stimulating PIP2 depletion and destabilization of CaV1.2 expression. We tested this hypothesis and report that CaV1.2 channels in tsA201 cells are destabilized after AT1 receptor-triggered PIP2 depletion, leading to their dynamin-dependent endocytosis. Likewise, in cardiomyocytes, angiotensin II decreased t-tubular CaV1.2 expression and cluster size by inducing their dynamic removal from the sarcolemma. These effects were abrogated by PIP2 supplementation. Functional data revealed acute angiotensin II reduced CaV1.2 currents and Ca2+ transient amplitudes thus diminishing excitation-contraction coupling. Finally, mass spectrometry results indicated whole-heart levels of PIP2 are decreased by acute angiotensin II treatment. Based on these observations, we propose a model wherein PIP2 stabilizes CaV1.2 membrane lifetimes, and angiotensin II-induced PIP2 depletion destabilizes sarcolemmal CaV1.2, triggering their removal, and the acute reduction of CaV1.2 currents and contractility.

Magnetic Bacterial Cellulose Biopolymers: Production and Potential Applications in the Electronics Sector.

Bacterial cellulose (BC) is a biopolymer that has been widely investigated due to its useful characteristics, such as nanometric structure, simple production and biocompatibility, enabling the creation of novel materials made from additive BC in situ and/or ex situ. The literature also describes the magnetization of BC biopolymers by the addition of particles such as magnetite and ferrites. The processing of BC with these materials can be performed in different ways to adapt to the availability of materials and the objectives of a given application. There is considerable interest in the electronics field for novel materials and devices as well as non-polluting, sustainable solutions. This sector influences the development of others, including the production and optimization of new equipment, medical devices, sensors, transformers and motors. Thus, magnetic BC has considerable potential in applied research, such as the production of materials for biotechnological electronic devices. Magnetic BC also enables a reduction in the use of polluting materials commonly found in electronic devices. This review article highlights the production of this biomaterial and its applications in the field of electronics.

Photophysical Rate Constants and Oxygen Dependence for Si and Ge Rhodamine Zwitterions.

The family of group XIV rhodamine zwitterions are fluorescence probes with carbon, silicon, germanium, or tin substituted in the 10-position of the xanthene ring. Because of their inherent near-infrared fluorescence, photostability and high quantum yields in aqueous solutions, the Si and Ge containing fluorophores in this class have become increasingly important for fluorescent labeling of proteins and biological molecules. This study fully characterizes photophysical rates derived from a model consisting of a singlet ground state, the lowest singlet excited state, and the lowest triplet excited state for two exemplar group XIV rhodamine zwitterions, one containing Si and the other Ge. Within a simple Jablonski diagram, all radiative and non-radiative rates, including intersystem crossing and triplet depopulation rates, were measured as a function of oxygen concentration. It was shown that the triplet depopulation rates are intrinsically fast in comparison with traditional xanthene containing fluorophores, probably due to the increased spin-obit coupling from the Si and Ge substitution in the xanthene ring. Dissolved oxygen increases both the intersystem crossing and triplet depopulation rates. Stern-Volmer analysis was conducted to estimate rates of quenching by oxygen. The experimental data was used to estimate the initial rates for reactive oxygen production by Si and Ge containing fluorophores in aqueous solutions containing different concentrations of dissolved O2. These estimates showed a significantly slower initial rate of reactive oxygen production in comparison with rhodamine 6G. This goes a long way to explaining their inherent photostability. Spectroscopic experiments were also conducted in 77 K viscous aqueous glasses where it was observed that the fluorescence spectra remained unchanged, and the quantum yields increased from 0.53 to 0.84 and from 0.52 to 0.89 for the Si and Ge containing fluorophores respectively; no phosphorescence was observed. All intersystem crossing and triplet depopulation rates were measured using fluorescence correlation spectroscopy (FCS) and analyzed using a new method that extrapolated the power dependence of the FCS curves to optical saturation. This method was verified using published data.

Natural and artificial pollination of white-fleshed pitaya.

To produce pitaya (Hylocereus spp.), it is essential that pollination occur, either artificially or by pollinating agents. This study evaluated the viability of self-pollination, cross-pollination, and natural pollination, as well as pollen viability, stigma receptivity and ideal pollination window in pitaya flowers (Hylocereus spp.). An experiment was conducted with a randomized block design consisting of four treatments: T1: manual self-pollination; T2: nocturnal open pollination; T3: diurnal open pollination; and T4: manual cross-pollination - red-fleshed pitaya [ Hylocereus polyrhizus] pollen placed on white-fleshed pitaya [ Hylocereus undatus] stigma. The experiment had four replicates, with two plants per replicate (two flowers per plant), totaling 16 flowers per treatment. The analyzed variables were fruit weight, length, diameter, titratable acidity, pH, soluble solids, and SS/TA ratio; the germination percentage of pollen grains; and the receptivity of stigmas. Artificial pollination (self- and cross-pollination) of white-fleshed pitaya (Hylocereus undatus) is viable, resulting in larger fruits than natural pollination, with cross-pollination being the most recommended. Throughout the floral opening period, the pitaya flower (Hylocereus spp.) has receptive stigma capable of receiving pollen. The flowers exhibit the highest pollen germination rate at 7 p.m, which is the ideal pollination window for pitaya.

"Playing the beat": Occurrence of Bio-duck calls in Santos Basin (Brazil) reveals a complex acoustic behaviour for the Antarctic minke whale (Balaenoptera bonaerensis).

The Antarctic minke whale (Balaenoptera bonaerensis) (AMW) is one of the smallest species among baleen whales, occurring in the southern hemisphere from Antarctica to near the equator, and performing seasonal migrations from polar to tropical waters. Information about (AMW) occurrence in the winter breeding grounds is scarce, mostly coming from old records from whaling stations before the 1960's international moratorium, such as Costinha Station in Northeastern Brazil (6° S / 34° W) and some sightings from few dedicated visual surveys. Acoustic methods can provide important data on the occurrence and distribution of migratory species. This work describes the occurrence of the Antarctic minke whale through acoustic detections of their "Bioduck" vocalisations in the Santos Basin, South-Southeastern Brazil (22° and 28° S / 42° and 48° W). Data was recorded between November 12 and December 19, 2015. AMW calls were detected for 12 days. We detected and classified 9 different Bio-duck calls in Brazilian coastal waters, evidencing a highly diverse acoustic behavior for the minke whale breeding ground. This is the first attempt to describe the acoustic diversity of AMW vocalizations in lower latitudes, contributing important information for future conservation efforts and management of AMW populations and their habitat. Therefore, our study presents the foremost acoustic evidence of the Antarctic minke whale in Brazilian coastal waters.