Endpoint Measures Research Articles

Coupling Exponential to Linear Amplification for Endpoint Quantitative Analysis.

Exponential DNA amplification techniques are fundamental in ultrasensitive molecular diagnostics. These systems offer a wide dynamic range, but the quantification requires real-time monitoring of the amplification reaction. Linear amplification schemes, despite their limited sensitivity, can achieve quantitative measurement from a single end-point readout, suitable for low-cost, point-of-care, or massive testing. Reconciling the sensitivity of exponential amplification with the simplicity of end-point readout would thus break through a major design dilemma and open a route to a new generation of massively scalable quantitative bioassays. Here a hybrid nucleic acid-based circuit design is introduced to compute a logarithmic function, therefore providing a wide dynamic range based on a single end-point measurement. CELIA (Coupling Exponential amplification reaction to LInear Amplification) exploits a versatile biochemical circuit architecture to couple a tunable linear amplification stage - optionally embedding an inverter function - downstream of an exponential module in a one-pot format. Applied to the detection of microRNAs, CELIA provides a limit of detection in the femtomolar range and a dynamic range of six decades. This isothermal approach bypasses thermocyclers without compromising sensitivity, thereby opening the way to applications in various diagnostic assays, and providing a simplified, cost-efficient, and high throughput solution for quantitative nucleic acid analysis.

Outcome prediction of cardiac arrest with automatically computed gray-white matter ratio on computed tomography images

BackgroundThis study aimed to develop an automated method to measure the gray-white matter ratio (GWR) from brain computed tomography (CT) scans of patients with out-of-hospital cardiac arrest (OHCA) and assess its significance in predicting early-stage neurological outcomes.MethodsPatients with OHCA who underwent brain CT imaging within 12 h of return of spontaneous circulation were enrolled in this retrospective study. The primary outcome endpoint measure was a favorable neurological outcome, defined as cerebral performance category 1 or 2 at hospital discharge. We proposed an automated method comprising image registration, K-means segmentation, segmentation refinement, and GWR calculation to measure the GWR for each CT scan. The K-means segmentation and segmentation refinement was employed to refine the segmentations within regions of interest (ROIs), consequently enhancing GWR calculation accuracy through more precise segmentations.ResultsOverall, 443 patients were divided into derivation N=265, 60% and validation N=178, 40% sets, based on age and sex. The ROI Hounsfield unit values derived from the automated method showed a strong correlation with those obtained from the manual method. Regarding outcome prediction, the automated method significantly outperformed the manual method in GWR calculation (AUC 0.79 vs. 0.70) across the entire dataset. The automated method also demonstrated superior performance across sensitivity, specificity, and positive and negative predictive values using the cutoff value determined from the derivation set. Moreover, GWR was an independent predictor of outcomes in logistic regression analysis. Incorporating the GWR with other clinical and resuscitation variables significantly enhanced the performance of prediction models compared to those without the GWR.ConclusionsAutomated measurement of the GWR from non-contrast brain CT images offers valuable insights for predicting neurological outcomes during the early post-cardiac arrest period.

Retinal Imaging Findings in Inherited Retinal Diseases.

Inherited retinal diseases (IRDs) represent one of the major causes of progressive and irreversible vision loss in the working-age population. Over the last few decades, advances in retinal imaging have allowed for an improvement in the phenotypic characterization of this group of diseases and have facilitated phenotype-to-genotype correlation studies. As a result, the number of clinical trials targeting IRDs has steadily increased, and commensurate to this, the need for novel reproducible outcome measures and endpoints has grown. This review aims to summarize and describe the clinical presentation, characteristic imaging findings, and imaging endpoint measures that are being used in clinical research on IRDs. For the purpose of this review, IRDs have been divided into four categories: (1) panretinal pigmentary retinopathies affecting rods or cones; (2) macular dystrophies; (3) stationary conditions; (4) hereditary vitreoretinopathies.

Physical Activity with Sports Scientist (PASS) programme to promote physical activity among patients with non-communicable diseases: a pragmatic randomised controlled trial protocol

Physical activity (PA) effectively prevents and treats non-communicable diseases in clinical settings. PA promotion needs to be more consistent, especially in busy primary care. Sports scientists have the potential to...

Proteomic insights from extracellular vesicles into the molecular mechanisms of health effects induced by Per- and polyfluoroalkyl substances

Per- and polyfluoroalkyl substances (PFASs) can induce a range of adverse health effects, with the precise molecular mechanisms remaining elusive. Extracellular vesicles (EVs) have demonstrated their potential to elucidate unknown molecular mechanisms. Building upon the close alignment of their biological functions with the observed health effects of PFASs, this study innovatively focuses on proteomic insights from EVs into the molecular mechanisms underlying the systemic health effects of PFASs. Through rat exposure experiments and proteomics technology, it not only demonstrated the occurrence of PFASs in EVs but also revealed the alterations in the serum EVs and the expression of their protein cargos following mixed exposure to PFASs, leading to changes in related pathways. These changes encompass various biological processes, including proteasome activity, immune response, cytoskeletal organization, oxidative stress, cell signaling, and nervous system function. Particularly noteworthy is the uncovering of the activation of the proteasome pathway, highlighting significant key contributing proteins. These novel findings provide a new perspective for exploring the molecular mechanism underlying the systemic health effects of PFASs and offer reliable screening for potential biomarkers. Additionally, comparisons with serum confirmed the potential of serum EVs as biological responders and measurable endpoints for evaluating PFASs-induced toxicity.

Pneumococcal conjugate vaccination schedules in infants—acquisition, immunogenicity, and pneumococcal conjugate and yellow fever vaccine co-administration study: statistical analysis plan

RationaleThe effectiveness of immunisation with pneumococcal conjugate vaccine (PCV) has been demonstrated in many countries. However, the global impact of PCV is limited by its cost, which has prevented its introduction in some countries. Reducing the cost of PCV programmes will facilitate further vaccine introductions and improve the sustainability of PCV in low-income countries when they transition from subsidised vaccine supply. We are conducting a large, population-level, cluster-randomised field trial (PVS) of an alternative reduced-dose schedule of PCV compared to the standard schedule. We are also conducting a nested sub-study at the individual level to investigate the immunogenicity of the two schedules and their effects on pneumococcal carriage acquisition (PVS-AcqImm).Methods and designPVS-AcqImm is a prospective, cluster-randomised trial of an alternative schedule of one dose of PCV scheduled at age 6 weeks with a booster dose at age 9 months compared to the standard of three primary doses scheduled at 6, 10, and 14 weeks of age. Sub-groups within the alternative schedule group receive yellow fever vaccine separately or co-administered with PCV at 9 months of age. The primary endpoints are (a) concentrations of vaccine-type anti-pneumococcal IgG at 18 months of age, (b) proportions with yellow fever neutralising antibody titre ≥ 1:8 4 weeks after separate or co-administration of PCV and yellow fever vaccines, and (c) rate of nasopharyngeal vaccine-type pneumococcal acquisition from 10–14 months of age. Participants and field staff are not masked to group allocation while measurement of the laboratory endpoints is masked. Approximately equal numbers of participants are resident in each of 28 randomly allocated geographic clusters (14 clusters in each group); 784 enrolled for acquisition measurements and 336 for immunogenicity measurements.PurposeThis statistical analysis plan (SAP) describes the PVS-AcqImm cohort and follow-up criteria to be used in different analyses. The SAP defines the endpoints and describes how adherence to the interventions will be presented. We describe the approach to analyses and how we will account for the effect of clustering. Defining the SAP prior to the conduct of analysis will avoid bias in analyses that may arise from prior knowledge of trial findings.Trial registrationISRCTN, ISRCTN7282161328. Registered on 28 November 2019. https://www.isrctn.com/ISRCTN72821613.Protocol: MRCG SCC number 1670, LSHTM Ref 17683.Current protocol version: 6.0, 24 May 2021.Version: 1.0 (5 April 2023); SAP revisions—none.

Abstract 2048: Integrated tumor models for immune oncology: Using live cell imaging for prediction of treatment efficacy in vitro and in vivo

Abstract Background: The preclinical evaluation of novel cancer treatments demands comprehensive model systems in vitro that provide meaningful data before entering in vivo studies. Here we evaluate the capabilities of live cell imaging systems to evaluate novel immune therapies. Using integrated immune and tumor cell models in vitro we demonstrate, that these model systems can generate reliable data of pharmacodynamic activity of biologicals, small molecules or combinatorial approaches for further preclinical in vivo characterization. Methods: Target tumor cell killing was assessed in vitro with immune cells (T- and NK-cells) and engagers. Tumor cells were transduced with a fluorescent marker to discriminate tumor cells from immune cells. The technology was used to determine inhibition of cell motility (re-invasion) after scratching of tumor cell monolayers. Cells were monitored using the IncuCyte. Dose-response-curves of single treatments and all combinations were generated in parallel. Active therapies were selected for further in vivo validation of immune cell killing. Humanized mice were generated by injection of CD34+ HSC or human immune cell subsets. Immune cell engraftment was monitored by FACS. To analyze the effect biologicals or small molecules, tumor cells were transplanted into these humanized mice. Tumor development and therapeutic effects were monitored by BLI measurements. Results and conclusion: Tumor cell killing by immune cells and monolayer scratch assay in 96 well format were successfully monitored in the IncuCyte. Here, data can be generated over time without the need of new samples at every time point compared with conventional end point measurements. Using antibodies directing immune cells to attack target cells extensive cell killing was observed over time. These data predicted in vivo treatment outcome in mice co-engrafted with human immune cells. After successful humanization of mice, immune cells can be directed to kill target tumor cells. Small molecule combinations were tested in vitro utilizing the metastasis/2D scratch assay. After setup of dose-response curves for two molecules combinatorial treatments were tested. Here we found a synergistic increase in efficacy. These combinations were tested in vivo to evaluate their abilities to inhibit cell motility and distant metastasis. Here we show that the in vitro assays predicted correctly the highest efficacy of combined treatments compared to mono treatments.The IncuCyte System provides data that translate our integrated model systems into in vivo studies. We have shown that activated immune cells can kill target tumor cells in vitro. These data have been validated in vivo using immune cell humanized mice. Further, immune cells, biologicals and small molecule based treatments can be tested either alone or in combination, allowing the preselection of active combinations for further development. Citation Format: Dennis Kobelt, Maria Stecklum, Simone Rhein, Wolfgang Walther, Jens Hoffmann. Integrated tumor models for immune oncology: Using live cell imaging for prediction of treatment efficacy in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2048.

Neoadjuvant immunotherapy based on PD-1/L1 inhibitors for gastrointestinal tumors: a review of the rationale and clinical advances.

The landscape of current tumor treatment has been revolutionized by the advent of immunotherapy based on PD-1/PD-L1 inhibitors. Leveraging its capacity to mobilize systemic antitumor immunity, which is primarily mediated by T cells, there is growing exploration and expansion of its potential value in various stages of clinical tumor treatment. Neoadjuvant immunotherapy induces a robust immune response against tumors prior to surgery, effectively facilitating tumor volume reduction, early eradication or suppression of tumor cell activity, and control of potential metastatic spread, to improve curative surgical resection rates, and prevent tumor recurrence. This review delineates the theoretical basis of neoadjuvant immunotherapy from preclinical research evidence, discusses specific challenges in clinical application, and provides a comprehensive overview of clinical research progress in neoadjuvant immunotherapy for gastrointestinal tumors. These findings suggest that neoadjuvant immunotherapy has the potential to ameliorate immunosuppressive states and enhance cytotoxic T cell function while preserving lymphatic drainage in the preoperative period. However, further investigations are needed on specific treatment regimens, suitable patient populations, and measurable endpoints. Despite numerous studies demonstrating the promising efficacy and manageable adverse events of neoadjuvant immunotherapy in gastrointestinal tumors, the availability of high-quality randomized controlled trials is limited, which highlights the necessity for further research.

Evaluation of molecular inhibitors of loop-mediated isothermal amplification (LAMP)

Loop-mediated isothermal amplification (LAMP) is a cost-effective and easy-to-perform assay that enables the direct detection of DNA. Its use in point-of-care diagnostic tests is growing, while it has the potential to be used in presumptive on-the-field forensic tests. Samples are often collected from complex matrices that contain high levels of contaminants. Herein, we evaluate the effect of seven common DNA amplification inhibitors on LAMP – bile salts, calcium chloride, hematin, humic acid, immunoglobulin G, tannic acid and urea. We study the effect of each inhibitor individually in real-time detection systems coupled with end-point measurements to delineate their inhibitory effects from the matrix in which they may be found. Our studies show LAMP inhibitors generally delay the onset of amplicon formation and quench fluorescence at similar or higher concentrations compared to PCR, but that end-point measurements of LAMP amplicons are unaffected. This is important as LAMP amplicons can be detected in non-fluorometric ways thus contributing to the assertions that LAMP is more robust to inhibitors than PCR.

Harmonising outcome measurement for child focused domestic abuse interventions. Reflections on the development and implementation of a core outcome set.

There is appetite in the UK to better measure the impact of domestic violence and abuse (DVA) interventions on children. The spread of outcomes-based commissioning means outcome measurement is no longer just the territory of academic researchers but is now firmly within the purview of practitioners and policy makers. However, outcomes measured in trials only partially represent the views of those delivering and using services with respect to how success should be defined and captured. Even within trials there is huge inconsistency in the definition and measurement of important endpoints. This yields a body of evidence that is difficult to make sense of, defeating the ends for which it was produced - to improve the response to children and families who have experienced abuse. Development of Core Outcome Sets (COS) is seen as a solution to this problem, by establishing consensus across key stakeholder groups regarding a minimum standard for outcome measurement in trials, and increasingly in service delivery contexts. To date COS development has addressed outcomes relating to health conditions or interventions, with limited application to public health challenges. We reflect on our efforts to develop a COS to evaluate psychosocial interventions for children and families experiencing DVA. We highlight the value of COS development as a mechanism for improving evidence quality and the response to families experiencing abuse. Finally, we make recommendations to researchers and COS guideline developers to support this broader application of COS methodology.

Phase II Study of Allogeneic Hematopoietic Stem Cell Transplantation for Children with High-Risk Neuroblastoma Using a Reduced-Intensity Conditioning Regimen: Results from the AIEOP Trial

Despite aggressive multimodal treatment, the outcomes of pediatric patients with high-risk (HR) neuroblastoma (NB) remain poor. The rationale for allogeneic hematopoietic stem cell transplantation (allo-HCT) to treat NB was based on the possible graft-versus-tumor effect; however, toxicity limits its efficacy. We sought to prospectively assess the feasibility and efficacy of allo-HCT using a reduced-intensity conditioning regimen in pediatric patients with HR NB in a multicenter phase II trial. Primary endpoints were the rate of neutrophil and platelet engraftment, 5-year transplantation-related mortality (TRM), and disease-free survival (DFS). Secondary endpoint measures included the incidence of acute graft-versus-host disease (aGVHD) and chronic GVHD. Fifty-one patients were enrolled in the study. The 5-year cumulative incidence (CuI) of TRM was 29.4 ± 6.4%, and that of DFS was 11.8 ± 4.5%. Patients undergoing allo-HCT within 1 year of diagnosis or with bone marrow as their stem cell source had a higher DFS probability. The CuI of neutrophil engraftment, platelet engraftment, and grade II-IV aGVHD was 97.9 ± 2.1%, 93.8 ± 3.5%, and 47.1 ± 7.0%, respectively. The development of new therapeutic strategies could further improve disease control.

Quantifiable and Inexpensive Cell-Free Fluorescent Method to Confirm the Ability of Novel Compounds to Chelate Iron.

Cancer cells require large amounts of iron to maintain their proliferation. Iron metabolism is considered a hallmark of cancer, making iron a valid target for anti-cancer approaches. The development of novel compounds and the identification of leads for further modification requires that proof of mechanism assays be carried out. There are many assays to evaluate the impact on proliferation; however, the ability to chelate iron is an important and sometimes overlooked end-point measure due to the high costs of equipment and the challenge to quickly and reproducibly quantify the strength of chelation. Here, we describe a quantifiable and inexpensive cell-free fluorescent method to confirm the ability of novel compounds to chelate iron. Our assay relies on the commercially available inexpensive fluorescent dye Calcein, whose fluorescence can be quantified on most fluorescent microtiter plate readers. Calcein is a weak iron chelator, and its fluorescence is quenched when it binds Fe2+/3+; fluorescence is restored when a novel chelator outcompetes Calcein for bound Fe2+/3+. The removal of fluorescent quenching and the resulting increase in fluorescence allows the chelation ability of a novel putative chelator to be determined. Therefore, we offer an inexpensive, high-throughput assay that allows the rapid screening of novel candidate chelator compounds.

Comparing the Analgesic Effects Between the Pre- and Post-costal Cartilage Harvest Cohorts Using Ultrasound-Guided Deep Serratus Anterior Plane Block in Children with Microtia Undergoing Auricular Reconstruction: A Randomized Clinical Trial.

Pain following costal cartilage harvest surgery is the most common complaint of auricular reconstruction (AR). Anesthesiologists are continuously searching for an effective postoperative pain control method. This study was conducted from 10 April 2022 to 10 June 2022. Sixty children undergoing AR using costal cartilage were randomly assigned to either a serratus anterior plane block performed before costal cartilage harvest (SAPB-pre-cohort; n=30) or the SAPB-post-cohort (Post-costal cartilage Harvest Cohort: n=30). The primary endpoint measures were the Numerical Rating Scale (NRS) scores of the chest and ear pain degrees recorded at 1-, 6-, 12-, 24-, and 48-h after surgery. Intraoperative anesthetic and analgesic dosages, sufentanil consumption and rescue analgesia consumption during the first 24h post-operation, cough score during extubation, extubation agitation score, length of stay, the extubation time, first ambulatory time, analgesia duration, and opioid-related adverse effects and SAPB-related adverse effects were the secondary endpoints. The rest and coughing NRS scores were significantly reduced in the SAPB-pre-cohort 6 and 12h post-operation in comparison with the SAPB-post-cohort (rest 6h p=0.002, others p<0.001). No significant difference in the NRS ear scores existed between the two cohorts (p>0.05). The use of propofol and remifentanil for general anesthesia during the SAPB-pre-procedure was significantly reduced compared to the SAPB-post-group, with statistical significance (p<0.001). Sufentanil consumption and rescue analgesia consumption were significantly reduced in the SAPB-pre-cohort (p=0.001, p=0.033). The extubation time and first ambulatory time were markedly shorter in the SAPB-pre-cohort (all p<0.001). Analgesia duration was markedly longer in the SAPB-pre-cohort (p<0.001). No significant differences were noted in the cough score during extubation, extubation agitation score, length of stay between the two cohorts (all p>0.05). Opioid-related adverse effects occurred more in the SAPB-post-cohort, while there was no statistical significance (16.7 vs. 36.7%; p=0.082). There were no blockade-related complications observed in either cohort. The analgesic effect of the SAPB-pre-cohort was better than the SAPB-post-cohort suggesting both efficacy and feasibility of preemptive analgesia. This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Measuring differential fitness costs and interactions between genetic cassettes using fluorescent spectrophotometry.

In this article, we present a method for designing, executing, and analyzing data from a microbial competition experiment. We use fluorescent reporters to label different competing strains and resolve individual growth curves using a fluorescent spectrophotometer. Our comprehensive data analysis pipeline integrates multiple experiments to simultaneously infer sources of variation, extract selection coefficients, and estimate the genetic contributions to fitness for various synthetic genetic cassettes (SGCs). To demonstrate the method, we employ a synthetic biological system based on Escherichia coli. Strains carry 1 of 10 different plasmids and one of three genomically integrated fluorescent markers. All strains are co-cultured to obtain real-time measurements of optical density (total population density) and fluorescence (sub-population densities). We identify challenges in calibrating between fluorescence and density and of fluorescent proteins maturing at different rates. To resolve these issues, we compare two methods of fluorescence calibration and correct for maturation by measuring in vivo maturation times. We provide evidence of genetic interactions occurring between our SGCs and further show how to use our statistical model to test some hypotheses about microbial growth and the costs of protein expression.IMPORTANCEFluorescently labeled co-cultures are becoming increasingly popular. The approach proposed here offers a high standard for experimental design and data analysis to measure selection coefficients and growth rates in competition. Measuring competitive differences is useful in many laboratory studies, allowing for fitness cost-correction of growth rates and ecological interactions and testing hypotheses in synthetic biology. Using time-resolved growth curves, rather than endpoint measurements, for competition assays allows us to construct a detailed scientific model that can be used to ask questions about fine-grained phenomena, such as bacterial growth dynamics, as well as higher-level phenomena, such as the interactions between synthetic cassette expression.

Prostate-Specific Membrane Antigen: Gateway to Management of Advanced Prostate Cancer.

Prostate-specific membrane antigen (PSMA) as a transmembrane protein is overexpressed by prostate cancer (PC) cells and is accessible for binding antibodies or low-molecular-weight radioligands due to its extracellular portion. Successful targeting of PSMA began with the development of humanized J591 antibody. Due to their faster clearance compared to antibodies, small-molecule radioligands for targeted imaging and therapy of PC have been favored in recent development efforts. PSMA positron emission tomography (PET) imaging has higher diagnostic performance than conventional imaging for initial staging of high-risk PC and biochemical recurrence detection/localization. However, it remains to be demonstrated how to integrate PSMA PET imaging for therapy response assessment and as an outcome endpoint measure in clinical trials. With the recent approval of 177Lu-PSMA-617 by the US Food and Drug Administration for metastatic castration-resistant PC progressing after chemotherapy, the high value of PSMA-targeted therapy was confirmed. Compared to standard of care, PSMA-based radioligand therapy led to a better outcome and a higher quality of life. This review, focusing on the advanced PC setting, provides an overview of different approved and nonapproved PSMA-targeted imaging and therapeutic modalities and discusses the future of PSMA-targeted theranostics, also with an outlook on non-radiopharmaceutical-based PSMA-targeted therapies.

Metabolomic Response of Thalassiosira weissflogii to Erythromycin Stress: Detoxification Systems, Steroidal Metabolites, and Energy Metabolism

Erythromycin, a macrolide antibiotic, is a prioritized pollutant that poses a high risk to environmental health. It has been detected in different environmental matrices and can cause undesired effects in aquatic organisms, particularly freshwater algae, which are primary producers. However, the impact of erythromycin on marine algae remains largely unexplored. Erythromycin has been reported to induce hormetic effects in the marine diatom Thalassiosira weissflogii (T. weissflogii). These effects are associated with the molecular pathways and biological processes of ribosome assembly, protein translation, photosynthesis, and oxidative stress. However, the alterations in the global gene expression have yet to be validated at the metabolic level. The present study used non-targeted metabolomic analysis to reveal the altered metabolic profiles of T. weissflogii under erythromycin stress. The results showed that the increased cell density was possibly attributed to the accumulation of steroidal compounds with potential hormonic action at the metabolic level. Additionally, slight increases in the mitochondrial membrane potential (MMP) and viable cells were observed in the treatment of 0.001 mg/L of erythromycin (an environmentally realistic level). Contrarily, the 0.75 and 2.5 mg/L erythromycin treatments (corresponding to EC20 and EC50, respectively) showed decreases in the MMP, cell density, and viable algal cells, which were associated with modified metabolic pathways involving ATP-binding cassette (ABC) transporters, the metabolism of hydrocarbons and lipids, thiamine metabolism, and the metabolism of porphyrin and chlorophyll. These findings suggest that metabolomic analysis, as a complement to the measurement of apical endpoints, could provide novel insights into the molecular mechanisms of hormesis induced by antibiotic agents in algae.

Vitamin D Status and Physical Functioning in Nursing Home Residents after Improved Adherence to the Vitamin D and Calcium Recommendation-A Quasiexperimental Study.

Dietary supplements with vitamin and calcium are recommended to nursing home residents in Denmark, but adherence to the recommendation is low. In a previous part of this study, we reported improved adherence by means of The Model for Improvement leading to increased awareness and change of workflows at two nursing homes. However, potential effects of this improved adherence are unknown. The objective of this substudy was to investigate if the improved adherence to the recommendation affected vitamin D status, muscle strength, and physical functioning of the residents. This was a 20-week quasiexperimental study involving 40 residents from two Danish nursing homes. Baseline and endpoint measurements took place in October 2021 and March 2022, respectively. Outcomes were number of residents taking vitamin D and calcium supplements; vitamin D status; handgrip strength; and physical functioning with timed-up-and-go test and 30-second chair stand test. Prevalence of vitamin D supplement users increased from 45 to 78% (mean dose 41 μg) and of calcium supplement users from 40 to 72% (mean dose 769 mg) (both P=0.002). Among those having blood sampled at both baseline and endpoint (n = 30), mean vitamin D status increased from 66.6 ± 31.7 nmol/L to 82.8 ± 26.3 nmol/L (P < 0.001), and more residents were vitamin D sufficient at endpoint (90 vs. 63%, P=0.021). Endpoint vitamin D status among supplement users was 88.2 ± 22.2 nmol/L, which was higher compared to nonsupplement users (55.3 ± 30.4 nmol/L, P < 0.01). No effects were seen on muscle strength or physical functioning. Increased supplementation with vitamin D using The Model for Improvement positively affected vitamin D status and prevalence of vitamin D sufficiency but did not affect muscle strength or physical functioning. Longer-term studies involving more residents are needed to investigate effects of improved adherence on these outcomes. This trial is registered with NCT04956705.

Lacticaseibacillus paracasei PS23 increases ghrelin levels and modulates microbiota composition: a post-hoc analysis of a randomized controlled study.

Muscle damage can occur due to excessive, high-intensity, or inappropriate exercise. It is crucial for athletes and sports enthusiasts to have access to ways that expedite their recovery and alleviate discomfort. Our previous clinical trial demonstrated the anti-inflammatory and muscle damage-ameliorating properties of Lacticaseibacillus paracasei PS23 (PS23), prompting us to further explore the role of this probiotic in muscle damage recovery. This post-hoc analysis of a randomized controlled study investigated potential mediators between the intake of PS23 and the prevention of strength loss after muscle damage. We recruited 105 students from a sports university who had participated in the previously published clinical trial. These participants were randomly allocated to three groups, receiving capsuled live PS23 (L-PS23), heat-treated PS23 (HT-PS23), or a placebo over a period of six weeks. Baseline and endpoint measurements were taken for the levels of circulating ghrelin and other blood markers, stress, mood, quality of life, and the fecal microbiota. A significant increase in ghrelin levels was recorded in the L-PS23 group compared to the other groups. Additionally, both L-PS23 and HT-PS23 interventions led to positive shifts in the gut microbiota composition, particularly in elevated Lacticaseibacillus, Blautia, and Lactobacillus populations. The abundance of these bacteria was positively correlated with exercise performance and inversely correlated with inflammatory markers. In conclusion, dietary supplementation with PS23 may enhance exercise performance and influence muscle damage by increasing ghrelin levels and modulating the gut microbiota composition. Further clarification of the possible mechanisms and clinical implications is required.

Direct fluoride monitoring using a fluorogenic RNA-based biosensor.

Fluorinated compounds, whether naturally occurring or from anthropogenic origin, have been extensively exploited in the last century. Degradation of these compounds by physical or biochemical processes is expected to result in the release of fluoride. Several fluoride detection mechanisms have been previously described. However, most of these methods are not compatible with high- and ultrahigh-throughput screening technologies, lack the ability to real-time monitor the increase of fluoride concentration in solution, or rely on costly reagents (such as cell-free expression systems). Our group recently developed "FluorMango" as the first completely RNA-based and direct fluoride-specific fluorogenic biosensor. To do so, we merged and engineered the Mango-III light-up RNA aptamer and the fluoride-specific aptamer derived from a riboswitch, crcB. In this chapter, we explain how this RNA-based biosensor can be produced in large scale before providing examples of how it can be used to quantitatively detect (end-point measurement) or monitor in real-time fluoride release in complex biological systems by translating it into measurable fluorescent signal.

A prospective, blinded, randomized controlled clinical trial evaluating the effect of the synthetic electrospun fiber matrix in the treatment of chronic diabetic foot ulcers

The goal of this randomized, controlled, single-blind clinical trial was to compare the efficacy of synthetic electrospun fiber matrix (SEFM) with standard of care (SOC) in the treatment of diabetic foot ulcers (DFUs). Patients with DFUs ≤30 cm2 were randomized to receive either SEFM (RESTRATA®, Acera Surgical, Inc., St. Louis, MO) or SOC weekly for up to 12 weeks. SOC included foam or alginate dressing changes. The primary endpoint measure was the percentage of wounds achieving 100 % re-epithelialization at 12 weeks. Secondary endpoints included decrease in wound area, time to closure, and number of applications. Forty-six subjects were enrolled and randomized into two groups. In the per protocol (PP) population, 14/19 wounds (74 %) in the SEFM Group demonstrated 100 % re-epithelialization, compared with 6/18 wounds (33 %) in the SOC Group. The proportion of wounds closed and the persistence of wound closure in the SEFM Group was statistically superior to the SOC Group (90 % CI: 0.14, 0.62 / 0.12, 0.65) (Farrington-Manning) in the PP population. Time to complete healing in the SEFM Group (6.6 ± 3.0 weeks) was significantly reduced compared to the SOC Group (p = 0.046, 0.026) (Cox regression) in both the ITT and PP populations. This study represents the first randomized controlled trial to evaluate SEFM in the treatment of chronic DFUs and demonstrates superiority to SOC in achieving 100 % re-epithelialization within 12 weeks. Clinical trial registryNCT04918784.