Modulation of the K/BxN arthritis mouse model and the effector functions of human fibroblast-like synoviocytes by liver X receptors.

The role of liver X receptors (LXR) in rheumatoid arthritis (RA) remains controversial. We studied the effect of LXR agonists on fibroblast-like synoviocytes (FLS) from RA patients and the K/BxN arthritis model in LXRα and ß double-deficient (Nr1h2/3-/-) mice. Two synthetic LXR agonists, GW3965 and T0901317, were used to activate LXRs and investigate their effects on cell growth, proliferation and matrix metalloproteinases, and chemokine production in cultured FLS from RA patients. The murine model K/BxN serum transfer of inflammatory arthritis in Nr1h2/3-/- animals was used to investigate the role of LXRs on joint inflammation in vivo. LXR agonists inhibited the FLS proliferative capacity in response to TNF, the chemokine-induced migration, the collagenase activity in FLS supernatant and FLS CXCL12 production. In the K/BxN mouse model, Nr1h2/3-/- animals showed aggravated arthritis, histological inflammation, and joint destruction, as well as an increase in synovial metalloproteases and expression of proinflammatory mediators such as IL-1ß and CCL2 in joints compared with wild type animals. Taken together, these data underscore the importance of LXRs in modulating the joint inflammatory response and highlight them as potential therapeutic targets in RA.

Olaparib maintenance versus placebo in platinum-sensitive non-small cell lung cancer: the Phase 2 randomized PIPSeN trial.

BACKGROUND: Platinum-sensitivity is a phenotypic biomarker of Poly (ADP-ribose) polymerase inhibitors (PARPi) sensitivity in histotypes where PARPi are approved. Approximately one-third of non-small cell lung cancers (NSCLC) are platinum-sensitive. The double-blind, randomized phase II PIPSeN (NCT02679963) study evaluated olaparib, a PARPi, as maintenance therapy for patients with platinum-sensitive advanced NSCLC. METHODS: Chemonaïve patients with ECOG performance status of 0-1, platinum-sensitive, EGFR- and ALK-wild-type, stage IIIB-IV NSCLC were randomized (R) to receive either olaparib (O) maintenance or a placebo (P). The primary objective was progression-free survival (PFS) from R. Secondary objectives included overall survival (OS) and safety. With an anticipated hazard ratio of 0.65, 144 patients were required to be randomized, and approximately 500 patients enrolled. RESULTS: The trial was prematurely terminated because anti-PD(L)1 therapy was approved during the trial recruitment. A total of 182 patients were enrolled, with 60 patients randomized: 33 and 27 in the O and P arms, respectively. Patient and tumor characteristics were well-balanced between arms, except for alcohol intake (33% vs 11% in the O and P arms, respectively, p = 0.043). The median PFS was 2.9 and 2.0 months in the O and P arms, respectively (logrank p = 0.99). The median OS was 9.4 and 9.5 months in the O and P arms, respectively (p = 0.28). Grade ≥3 toxicities occurred in 15 and 8 patients in O and P arms, with no new safety concerns. CONCLUSION: PIPSeN was terminated early after enrollment of only 50% of the pre-planned population, thus being statistically underpowered. Olaparib maintenance did neither improve median PFS nor OS in this patient population.

Effect of combined physical-cognitive training on the functional and cognitive capacity of older people with mild cognitive impairment: a randomized controlled trial.

BACKGROUND: The increase in population aging highlights the growing prevalence of mild cognitive impairment, prompting the adoption of interventions that combine physical exercise and cognitive training to improve health and cognitive performance in older adults. The aim of this study was to analyze the efficacy of a combined program on physical and cognitive health in older people with cognitive impairment. METHODS: A 12-week randomized controlled clinical trial involving 95 participants (aged 72.12 ± 4.25 years), 47 individuals participated in a control group (CG) that only underwent cognitive stimulation, while 48 individuals were in an experimental group (EG) that participated in a combined program. Balance was measured using the Tinetti scale, upper body strength was assessed with the arm curl test, lower body strength was evaluated with the 30-s chair stand test, flexibility was tested using the back scratch test and chair sit-and-reach test, physical function was measured with the Timed Up and Go test, cognitive function was assessed using the Mini Mental State Examination, cognitive impairment was evaluated with the Montreal Cognitive Assessment, verbal fluency was tested with the Isaac test, and executive functions were assessed using the Trail Making Test. RESULTS: The results of the study show significant improvements in both physical and cognitive aspects, such as balance, gait, upper and lower body strength, flexibility, physical function, cognitive function, cognitive impairment, verbal fluency, and executive functions in the group that carried out the intervention compared to the control group. CONCLUSION: A combined program for older individuals with mild cognitive impairment leads to enhancements in physical and cognitive health. These improvements underscore the importance of integrating physical exercise with cognitive training as an effective strategy for enhancing overall health and quality of life in older adults. TRIAL REGISTRATION: NCT05503641.

Unlocking bacterial defense: Exploring the potent inhibition of NorA efflux pump by coumarin derivatives in Staphylococcus aureus.

The occurrence of bacterial resistance has been increasing, compromising the treatment of various infections. The high virulence of Staphylococcus aureus allows for the maintenance of the infectious process, causing many deaths and hospitalizations. The MepA and NorA efflux pumps are transporter proteins responsible for expelling antimicrobial agents such as fluoroquinolones from the bacterial cell. Coumarins are phenolic compounds that have been studied for their diverse biological actions, including against bacteria. A pharmacokinetic in silico characterization of compounds C10, C11, C13, and C14 was carried out according to the principles of Lipinski's Rule of Five, in addition to searching for similarity in ChemBL and subsequent search for publications in CAS SciFinder. All compounds were evaluated for their in vitro antibacterial and modulatory activity against standard and multidrug-resistant Gram-positive and Gram-negative strains. The effect of coumarins C9, C10, C11, C13, and C14 as efflux pump inhibitors in Staphylococcus aureus strains was evaluated using the microdilution method (MepA or NorA) and fluorimetry (NorA). The behavior of coumarins regarding the efflux pump was determined from their interaction properties with the membrane and coumarin-protein using molecular docking and molecular dynamics simulations. Only the isolated coumarin compound C13 showed antibacterial activity against standard strains of Staphylococcus aureus and Escherichia coli. However, the other tested coumarins showed modulatory capacity for fluoroquinolone and aminoglycoside antibacterials. Compounds C10, C13, and C14 were effective in reducing the MIC of both antibiotics for both multidrug-resistant strains, while C11 potentiated the effect of norfloxacin and gentamicin for Gram-positive and Gram-negative bacteria and only norfloxacin for Gram-negative. Only coumarin C14 produced synergistic effects when associated with ciprofloxacin in MepA-carrying strains. All tested coumarins have the ability to inhibit the NorA efflux pump present in Staphylococcus aureus, both in reducing the MIC and inducing increased ethidium bromide fluorescence emission in fluorimetry. The findings of this study offer an atomistic perspective on the potential of coumarins as active inhibitors of the NorA pump, highlighting their specific mode of action mainly targeting protein inhibition. In molecular docking, it was observed that coumarins are capable of interacting with various amino acid residues of the NorA pump. The simulation showed that coumarin C10 can cross the bilayer; however, the other coumarins interacted with the membrane but were unable to cross it. Coumarins demonstrated their potentiating role in the effect of norfloxacin through a dual mechanism: efflux pump inhibition through direct interaction with the protein (C9, C10, C11, and C13) and increased interaction with the membrane (C10 and C13). In the context of pharmacokinetic prediction studies, the studied structures have a suitable chemical profile for possible oral use. We suggest that coumarin derivatives may be an interesting alternative in the future for the treatment of resistant bacterial infections, with the possibility of a synergistic effect with other antibacterials, although further studies are needed to characterize their therapeutic effects and toxicity.

Treatment of post-allogeneic hematopoietic stem cell transplant cytopenias with sequential doses of multipotent mesenchymal stromal/stem cells.

BACKGROUND AIMS: Cytopenias after allogeneic stem cell transplantation (allo-SCT) are a common complication, the underlying pathogenic mechanisms of which remain incompletely understood. Multipotent mesenchymal stromal/stem cell (MSC) therapy has been successfully employed in the treatment of immune-related disorders and can aid in the restoration of the hematopoietic niche. METHODS: A phase II clinical trial to assess the efficacy and safety of administering four sequential doses of ex-vivo expanded bone marrow MSCs from a third-party donor to patients with persistent severe cytopenias after allo-SCT was performed. RESULTS: The overall response rate on day 90 was 75% among the 27 evaluable patients (comprising 12 complete responses, 8 partial responses, and 7 with no response). The median time to respond was 14.5 days. Responses were observed across different profiles, including single or multiple affected lineages, primary or secondary timing, and potential immune-mediated or post-infectious pathophysiology versus idiopathic origin. With a median follow-up for surviving patients of 85 months after MSC infusion, 53% of patients are alive. Notably, no adverse events related to MSC therapy were reported. CONCLUSIONS: In summary, the sequential infusion of third-party MSCs emerges as a viable and safe therapeutic option, exhibiting potential benefits for patients experiencing cytopenias following allo-SCT.

Clinical impact of the TPSAB1 genotype in mast cell diseases: A REMA study in a cohort of 959 individuals.

BACKGROUND: A close association between hereditary alpha-tryptasemia (HAT) and mast cell (MC) disorders has been previously reported. However, the relationship between HAT and the diagnostic subtypes and clinical features of MC disorders still remains to be established. OBJECTIVE: To determine the prevalence of HAT in healthy donors (HD) vs patients with different diagnostic subtypes of MC activation syndromes (MCAS) and mastocytosis, and its relationship with the clinical behavior of the disease. METHODS: A total of 959 subjects were studied including 346 healthy donors (HD), 464 mastocytosis, and 149 non-clonal MCAS patients. Molecular studies to assess the TPSAB1 genotype were performed, and data on serum baseline tryptase (sBT) and basal MC-mediator release episodes and triggers of anaphylaxis were collected. RESULTS: HAT was detected in 15/346 (4%) HD versus 43/149 (29%) non-clonal MCAS and 84/464 (18%) mastocytosis cases. Among mastocytosis, HAT was more frequently found in patients with MC-restricted KITD816V (21% vs. 10% among multilineage KITD816V patients; p = .008). Overall, median sBT was higher in cases presenting with HAT (28.9 vs. 24.5 ng/mL; p = .008), while no significant differences in sBT were observed among HAT+ mastocytosis patients depending on the presence of 1 vs. ≥2 extra copies of the α-tryptase gene (44.1 vs. 35.2 ng/mL, p > .05). In turn, anaphylaxis was more frequently observed in HAT+ versus HAT- mastocytosis patients (76% vs. 65%; p = .018), while HAT+ and HAT- patients who did not refer anaphylaxis as the presenting symptom (n = 308) showed a similar prevalence of subsequent anaphylaxis (35% vs. 36%, respectively). CONCLUSION: The frequency of HAT in MC disorders varies according to the diagnostic subtype of the disease. HAT does not imply a higher risk (and severity) of anaphylaxis in mastocytosis patients in whom anaphylaxis is not part of the presenting symptoms of the disease.

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.

Risk factors of death or chronic renal replacement therapy requirements in patients with thrombotic microangiopathies without ADAMTS-13 deficiency.

Thrombotic microangiopathy (TMA), characterized by microangiopathic hemolytic anemia, thrombocytopenia, and multisystem organ dysfunction, is a life-threatening disease. Patients with TMA who do not exhibit a severe ADAMTS-13 deficiency (defined as a disintegrin-like and metalloprotease with thrombospondin type 1 motif no. 13 activity ≥10%: TMA-13n) continue to experience elevated mortality rates. This study explores the prognostic indicators for augmented mortality risk or necessitating chronic renal replacement therapy (composite outcome: CO) in TMA-13n patients. We included 42 TMA-13n patients from January 2008 to May 2018. Median age of 41 years and 60% were female. At presentation, 62% required dialysis, and 57% warranted intensive care unit admission. CO was observed in 45% of patients, including a 9-patient mortality subset. Multivariate logistic regression revealed three independent prognostic factors for CO: early administration of eculizumab (median time from hospitalization to eculizumab initiation: 5 days, range 0-19 days; odds ratio [OR], 0.14; 95% confidence interval [CI], 0.02-0.94), presence of neuroradiological lesions (OR, 6.67; 95% CI, 1.12-39.80), and a PLASMIC score ≤4 (OR, 7.39; 95% CI, 1.18-46.11). In conclusion, TMA-13n patients exhibit a heightened risk of CO in the presence of low PLASMIC scores and neuroradiological lesions, while early eculizumab therapy was the only protective factor.

Telemedicine Beyond Borders: A Mixed-Methods Approach to Understanding Cross-National Usage Intention.

Introduction: Drawing on the Unified Theory of Acceptance and Use of Technology 2 and the Diffusion of Innovation Theory, this article investigates the adoption of telemedicine services from a patient perspective in Germany, Spain, and the United States using a mixed-methods approach. Digital health technologies have the potential to improve access to care and to alleviate the burden on traditional health care systems and are becoming more integrated into everyday medicine. Therefore, understanding the factors that impact patients' intentions to use telemedicine is crucial to ensure successful development. Methods: Based on 1,200 surveys collected in Germany, Spain, and the United States, structural equation modeling (IBM SPSS Amos 24) is employed to test the hypotheses. The article also explores how age and gender moderate the proposed relationships. Results: Seven out of the 10 hypotheses (performance expectancy, hedonic motivation, habit, relative advantage, and perceived security) are found to be positive, direct, and statistically significant. Furthermore, findings suggest stronger effects for telemedicine usage intention for younger female users than their male counterparts. Discussion: With digital health technologies becoming more prevalent, the outcomes of this study can endorse the development of effective strategies to promote the adoption of telemedicine, ultimately improving access to care and contributing to the advancement of and modern health care.

The differential illusion memory for high-associated abstract concepts (DIM-HA) effect.

A vast body of evidence has shown that concrete concepts are processed faster and more accurately than abstract concepts in a variety of cognitive tasks. This phenomenon is widely known as the concreteness effect, and explanations for its occurrence seem to reflect differences in processing and organization for both types of representations. While there is considerable evidence to support this concreteness effect, the nature of these differences is still controversial. In developing an explanation, we have proposed a relatively different approach from a false memory perspective using the Deese-Roediger-McDermott paradigm. To explore the implications of the association in creating false memories, we explore behavioral and electrophysiologically the false memory effect, where targets were manipulated according to their association strength and their concreteness. Results showed that false recognition rates differed significantly between concrete and abstract critical words when they were associated strongly with their respective lists, which led to a higher proportion of abstract false alarms both in behavioral and electrophysiological experiments. The principal outcome, which was called the DIM-HA effect, was discussed in terms of theories of associative activation and qualitatively different representation.

In vitro and in silico antibacterial evaluation of coumarin derivatives against MDR strains of Staphylococcus aureus and Escherichia coli.

The increase in antibiotic resistance rates has attracted the interest of researchers for antibacterial compounds capable of potentiating the activity of conventional antibiotics. Coumarin derivatives have been reported to develop effective antibacterials with possible new mechanisms of action for treating infectious diseases caused by bacteria with a profile of drug resistance. In this context, the aim of the present study we have now prepared one variety of new synthetic coumarins evaluating the pharmacokinetic and chemical similarity in silico, their antimicrobial activity against Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922), and potential for the modulation of antibiotic resistance against Staphylococcus aureus (SA10) and Escherichia coli (EC06) clinical isolate bacteria by in vitro assay. The antibacterial activity and antibiotic-enhancing properties were evaluated by the broth microdilution method and pharmacokinetically characterized according to the Lipinsk rule of 5 and had their similarity analyzed in databases such as ChemBL and CAS SciFinder. The results demonstrated that only compound C13 showed significant antibacterial activity (MIC ≤256 µg/mL), and all other coumarins did not display relevant antibacterial activity (MIC ≥1024 µg/mL). However, they did modulate the antibiotics activities to norfloxacin and gentamicin, except, compound C11 to norfloxacin against Staphylococcus aureus (SA10). The in silico properties prediction and drug-likeness results demonstrated that all coumarins presented a good drug-likeness score with no violations and promising in silico pharmacokinetic profiles showing that they have the potential to be developed into an oral drug. The results indicate that the coumarin derivatives showed good in vitro antibacterial activity. These new coumarin derivatives also demonstrated the capacity to modulate antibiotic resistance with potential synergy action for current antimicrobials assayed, as antibiotic adjuvants, to reduce the emergence of antimicrobial resistance.

In vitro and in silico evidences about the inhibition of MepA efflux pump by coumarin derivatives.

The discovery of antibiotics has significantly transformed the outcomes of bacterial infections in the last decades. However, the development of antibiotic resistance mechanisms has allowed an increasing number of bacterial strains to overcome the action of antibiotics, decreasing their effectiveness against infections they were developed to treat. This study aimed to evaluate the antibacterial activity of synthetic coumarins Staphylococcus aureus in vitro and analyze their interaction with the MepA efflux pump in silico. The Minimum Inhibitory Concentration (MIC) determination showed that none of the test compounds have antibacterial activity. However, all coumarin derivatives decreased the MIC of the standard efflux inhibitor ethidium bromide, indicating antibacterial synergism. On the other hand, the C14 derivative potentiated the antibacterial activity of ciprofloxacin against the resistant strain. In silico analysis showed that C9, C11, and C13 coumarins showed the most favorable interaction with the MepA efflux pump. Nevertheless, due to the present in silico and in vitro investigation limitations, further experimental research is required to confirm the therapeutic potential of these compounds in vivo.

Preantral follicle numbers and size in heifers carrying Trio, a bovine high fecundity allele.

In brief: The bovine high fecundity allele, Trio, results in the occurrence of multiple ovulations and is characterized by antral follicles that develop slower and acquire ovulatory capacity at smaller sizes. This study provides novel information on the effect of the Trio allele on early folliculogenesis. Abstract: The bovine high fecundity allele, Trio, causes overexpression in granulosa cells (GCs) of SMAD6, an inhibitor of BMP15-activated SMAD signalling. Furthermore, the Trio allele results in antral follicles that develop slower, acquire ovulatory capacity at smaller sizes, and have three-fold greater ovulation rate compared to half-sib non-carriers. The present study was designed to determine preantral follicle numbers and size in Trio carrier and non-carrier cattle testing the hypothesis that inhibition of SMAD signalling would alter preantral follicle activation and/or growth. Ovarian tissues from Trio carrier (n = 12) and non-carrier (n = 12) heifers were obtained by laparotomy after follicle wave synchronization. Follicle numbers and dimensions were determined for each stage of development (primordial, transitional, primary, and secondary) from paraffin-embedded sections. There were no differences in the number of primordial, transitional, or secondary follicles or in antral follicle count, circulating AMH, or ovarian volume between carriers and non-carriers. Trio carriers had ~2.5-fold greater (P < 0.01) number of primary follicles than non-carriers, and transitional and primary follicles were larger (~1.2-fold; P < 0.1) in Trio carriers. Oocyte volume of primordial and transitional follicles was not different between genotypes; however, oocytes were larger (P < 0.05) in primary (~1.3-fold) and secondary (~1.8-fold) follicles for Trio carriers. Granulosa cell numbers were not different (P > 0.3) between carriers and non-carriers, irrespective of the stage of development. These results suggest that, after primordial follicle activation, follicles in Trio carrier cattle have slower progression through the primary stage, hence the larger oocyte and greater number of primary follicles.

Prediction of blood volume to be processed to achieve a target number of CD34+ cells: Development, validation and implementation of a formula.

BACKGROUND AND OBJECTIVES: Calculation of blood volume (BV) to be processed to achieve the target number of CD34+ cells can be accomplished by using collection efficiency 2 (CE2) formula. Our aim was to develop a BV web formula. MATERIALS AND METHODS: We calculated CE2 from aphereses performed between January 2015 and March 2020 in allogeneic donors and patients. From May 2020 to May 2021, we validated a formula: BV = ((Target CD34+ cells in the product)/(CD34+ pre-apheresis cells × CE2)) × 100. Subsequently, we compared the outcome of the procedures carried out before formula implementation (pre-formula), when standard three total BV collection was performed. RESULTS: CE2 was assessed in 384 apheresis procedures before formula implementation. CE2 was higher in allogeneic donors than in patients (53% ± 17% vs. 48% ± 15%, p = 0.008). CE2 was higher in multiple myeloma and non-Hodgkin lymphoma than Hodgkin's lymphoma (48% ± 15%, 48% ± 15% and 42% ± 13%, respectively; p = 0.008). Our formula (available on a website: Publisheet) was prospectively used in 54 individuals. The formula was very accurate: predicted versus observed CD34 + cells/kg collected had an r-value of 0.89 (p < 0.0001). We compared their results with 78 pre-formula individuals. In the post-formula group, a greater BV was processed in patients and less BV in allogeneic donors. Among individuals under 60 years of age, it was significantly less frequent than the need for more than one apheresis in the post-formula group. CONCLUSION: Formula calculations were accurate. Formula implementation allowed the optimization of the procedures and reduced the rate of individuals in need of apheresis for more than 1 day.

Cost-effectiveness of omalizumab for the treatment of severe pediatric allergic asthma-Results of a real-life study in Spain.

BACKGROUND: Severe pediatric allergic asthma (SPAA) induces a huge economic burden in terms of direct, indirect, and intangible costs. The use of omalizumab for the treatment of these patients has produced a significant improvement in several clinical outcomes, but at the same time, the cost for the management of the disease has also increased. The aim of this report was to evaluate whether the use of omalizumab is cost-effective. METHODS: A sample of 426 children with SPAA from the ANCHORS (Asthma iN CHildren: Omalizumab in Real-life in Spain) study was used to calculate the incremental cost-effectiveness ratio (ICER) for the avoidance of moderate-to-severe exacerbations (MSE) and also for the improvement in childhood Asthma Control Test (c-ACT) or the Asthma Control Questionnaire (ACQ5). We retrospectively collected data on health encounters and drug consumption before and up to 6 years after the beginning of the treatment with omalizumab. RESULTS: The ICER per avoided MSE was €2107 after 1 year, and it consistently decreased to €656 in those followed up to 6 years. Similarly, the ICER for the minimally important difference in control tests showed a decrease from €2059 to €380 per each 0.5 points of improvement in ACQ5 and from €3141 to €2322 per each 3 points improvement in c-ACT, at years 1 and 6, respectively. CONCLUSION: The use of OMZ is a cost-effective option for most children with uncontrolled SPAA, especially those who have frequent exacerbations; the costs are progressively reduced in successive years of treatment.

A pharmacometrics approach to assess the feasibility of capillary microsampling to replace venous sampling in clinical studies: Tafenoquine case study.

AIM: Microsampling has the advantage of smaller blood sampling volume and suitability in vulnerable populations compared to venous sampling in clinical pharmacokinetics studies. Current regulatory guidance requires correlative studies to enable microsampling as a technique. A post hoc population pharmacokinetic (POPPK) approach was utilized to investigate blood capillary microsampling as an alternative to venous sampling. METHODS: Pharmacokinetic data from microsampling and venous sampling techniques during a paediatric study evaluating tafenoquine, a single-dose antimalarial for P. vivax, were used. Separate POPPK models were developed and validated based on goodness of fit and visual predictive checks, with pharmacokinetic data obtained via each sampling technique. RESULTS: Each POPPK model adequately described tafenoquine pharmacokinetics using a two-compartment model with body weight based on allometric scaling of clearance and volume of distribution. Tafenoquine pharmacokinetic parameter estimates including clearance (3.4 vs 3.7 L/h) were comparable across models with slightly higher interindividual variability (38.3% vs 27%) in capillary microsampling-based data. A bioavailability/bioequivalence comparison demonstrated that the point estimate (90% CI) of capillary microsample versus venous sample model-based individual post hoc estimates for area under the concentration-time curve from time zero to infinity (AUC0-inf ) (100.7%, 98.0-103.5%) and Cmax (79.7%, 76.9-82.5%) met the 80-125% and 70-143% criteria, respectively. Overall, both POPPK models led to the same dose regimen recommendations across weight bins based on achieving target AUC. CONCLUSIONS: This analysis demonstrated that a POPPK approach can be employed to assess the performance of alternative pharmacokinetic sampling techniques. This approach provides a robust solution in scenarios where variability in pharmacokinetic data collected via venous sampling and microsampling may not result in a strong linear relationship. The findings also established that microsampling techniques may replace conventional venous sampling methods.

One health surveillance strategy for coronaviruses in Italian wildlife.

The recent reinforcement of CoV surveillance in animals fuelled by the COVID-19 pandemic provided increasing evidence that mammals other than bats might hide further diversity and play critical roles in human infectious diseases. This work describes the results of a two-year survey carried out in Italy with the double objective of uncovering CoV diversity associated with wildlife and of excluding the establishment of a reservoir for SARS-CoV-2 in particularly susceptible or exposed species. The survey targeted hosts from five different orders and was harmonised across the country in terms of sample size, target tissues, and molecular test. Results showed the circulation of 8 CoV species in 13 hosts out of the 42 screened. Coronaviruses were either typical of the host species/genus or normally associated with their domestic counterpart. Two novel viruses likely belonging to a novel CoV genus were found in mustelids. All samples were negative for SARS-CoV-2, with minimum detectable prevalence ranging between 0.49% and 4.78% in the 13 species reaching our threshold sample size of 59 individuals. Considering that within-species transmission in white-tailed deer resulted in raising the prevalence from 5% to 81% within a few months, this result would exclude a sustained cycle after spillback in the tested species.

Multisensory System for Long-Term Activity Monitoring to Facilitate Aging-in-Place.

Demographic changes and an ageing population require more effective methods to confront the increased prevalence of chronic diseases which generate dependence in older adults as well as an important rise in social expenditure. The challenge is not only to increase life expectancy, but also to ensure that the older adults can fully enjoy that moment in their lives, living where they wish to (private home, nursing home, …). Physical activity (PA) is a representative parameter of a person's state of health, especially when we are getting older, because it plays an important role in the prevention of diseases, and that is the reason why it is promoted in older adults. One of the goals of this work is to assess the feasibility of objectively measuring the PA levels of older adults wherever they live. In addition, this work proposes long-term monitoring that helps to gather daily activity patterns. We fuse inertial measurements with other technologies (WiFi- and ultrasonic-based location) in order to provide not only PA, but also information about the place where the activities are carried out, including both room-level location and precise positioning (depending on the technology used). With this information, we would be able to generate information about the person's daily routines which can be very useful for the early detection of physical or cognitive impairment.

Time-Dependent Changes in Muscle IGF1-IGFBP5-PAPP System after Sciatic Denervation.

Denervation-induced muscle atrophy is a frequent cause of skeletal muscle diseases. However, the role of the most important muscle growth factor, insulin-like growth factor (IGF-1), in this process is poorly understood. IGF-1 activity is controlled by six IGF-1 binding proteins (IGFBPs). In skeletal muscle, IGFBP-5 seems to have an important role in atrophic processes. Furthermore, pappalysins (PAPP-A) modulate muscle growth by increasing IGF-1 bioavailability through IGFBP cleavage. We aimed to study the time-dependent changes in the IGF1-IGFBP5-PAPP system and its regulators in gastrocnemius muscle after sciatic denervation. Gastrocnemius atrophy and overexpression of IGF-1 was observed from day 3 post-denervation. The proteolytic factors measured were elevated from day 1 post-denervation onwards. Expression of both IGFBP-5 and pappalysins were increased on days 1 and 3. Subsequently, on days 7 to 14 pappalysins returned to control levels while IGFBP-5 remained elevated. The ratio IGFBP-5/PAPP-A was correlated with the main proteolytic markers. All data suggest that the initial increase of pappalysins could facilitate the IGF-1 action on muscle growth, whereas their subsequent decrease could lead to further muscle wasting.

Enhancing the Hydrolytic Activity of a Lipase towards Larger Triglycerides through Lid Domain Engineering.

Lipases have valuable potential for industrial use, particularly those mostly active against water-insoluble substrates, such as triglycerides composed of long-carbon chain fatty acids. However, in most cases, engineered variants often need to be constructed to achieve optimal performance for such substrates. Protein engineering techniques have been reported as strategies for improving lipase characteristics by introducing specific mutations in the cap domain of esterases or in the lid domain of lipases or through lid domain swapping. Here, we improved the lipase activity of a lipase (WP_075743487.1, or LipMRD) retrieved from the Marine Metagenomics MarRef Database and assigned to the Actinoalloteichus genus. The improvement was achieved through site-directed mutagenesis and by substituting its lid domain (FRGTEITQIKDWLTDA) with that of Rhizopus delemar lipase (previously R. oryzae; UniProt accession number, I1BGQ3) (FRGTNSFRSAITDIVF). The results demonstrated that the redesigned mutants gain activity against bulkier triglycerides, such as glyceryl tridecanoate and tridodecanoate, olive oil, coconut oil, and palm oil. Residue W89 (LipMRD numbering) appears to be key to the increase in lipase activity, an increase that was also achieved with lid swapping. This study reinforces the importance of the lid domains and their amino acid compositions in determining the substrate specificity of lipases, but the generalization of the lid domain swapping between lipases or the introduction of specific mutations in the lid domain to improve lipase activity may require further investigation.