A cost-effective method for the sensitive detection of levofloxacin using a 3D composite electrode composed of nail polish, graphite and aluminium oxide.

The potential impact on human health and the environment has spurred significant interest in detecting and quantifying pharmaceutical compounds across various matrices, from environmental to biological samples. Here, we present an electrochemical approach for determining levofloxacin in drug, synthetic urine, water, and breast milk samples. An affordable sensor was constructed using 3D printing and composite material based on nail polish, graphite, and aluminum oxide. The conductive composite material was characterized spectroscopically, electrochemically, and by imaging techniques. Subsequently, an electrochemical method based on square wave voltammetry was optimized and applied. The method exhibited good sensitivity (5.11 ± 0.0912 µA L µmol-1 cm-2) and enhanced stability (RSD = 7.2%), with electrochemical responses correlating with the concentration of levofloxacin in the samples tested, yielding recovery values in the range of 98 to 111%. The developed method demonstrated a robust linear working range from 2 to 100 µmol L-1 and a nanomolar detection limit of 128 nmol L-1, rendering it suitable for quantitative analysis. The sensor also shows promise as a platform for the sensitive detection of pharmaceutical compounds, contributing to greater safety and sustainability in these domains.

YAP1 status defines two intrinsic subtypes of LCNEC with distinct molecular features and therapeutic vulnerabilities.

PURPOSE: Large cell neuroendocrine carcinoma (LCNEC) is a high-grade neuroendocrine malignancy that, like small cell lung cancer (SCLC), is associated with an absence of druggable oncogenic drivers and dismal prognosis. In contrast to SCLC, however, there is little evidence to guide optimal treatment strategies which are often adapted from SCLC and non-small cell lung cancer (NSCLC) approaches. EXPERIMENTAL DESIGN: To better define the biology of LCNEC, we analyzed cell line and patient genomic data and performed immunohistochemistry and single-cell (sc)RNAseq of core needle biopsies from LCNEC patients and preclinical models. RESULTS: Here, we demonstrate that the presence or absence of YAP1 distinguishes two subsets of LCNEC. The YAP1-high subset is mesenchymal and inflamed and characterized, alongside TP53 mutations, by co-occurring alterations in CDKN2A/B and SMARCA4. Therapeutically, the YAP1-high subset demonstrates vulnerability to MEK and AXL targeting strategies, including a novel preclinical AXL CAR-T cell. Meanwhile, the YAP1-low subset is epithelial and immune-cold and more commonly features TP53 and RB1 co-mutations, similar to those observed in pure SCLC. Notably, the YAP1-low subset is also characterized by expression of SCLC subtype-defining transcription factors - especially ASCL1 and NEUROD1 - and, as expected given its transcriptional similarities to SCLC, exhibits putative vulnerabilities reminiscent of SCLC, including Delta-like ligand 3 (DLL3) and CD56 targeting, as with novel preclinical DLL3 and CD56 CAR T-cells, and DNA damage repair (DDR) inhibition. CONCLUSION: YAP1 defines distinct subsets of LCNEC with unique biology. These findings highlight the potential for YAP1 to guide personalized treatment strategies for LCNEC.

Patterns of kidney function and risk assessment in a nationwide laboratory database: the Brazilian CHECK-CKD study.

BACKGROUND: Chronic kidney disease (CKD) is a global health problem with rising prevalence, morbidity, mortality, and associated costs. Early identification and risk stratification are key to preventing progression to kidney failure. However, there is a paucity of data on practice patterns of kidney function assessment to guide the development of improvement strategies, particularly in lower-income countries. METHODS: A retrospective observational analysis was conducted in a nationwide laboratory database in Brazil. We included all adult patients with at least one serum creatinine assessment between June 2018 and May 2021. Our primary objective was to determine the proportion of patients with estimated glomerular filtration rate (eGFR) evaluations accompanied by predicted levels of urinary albumin-to-creatinine ratio (pACR) assessments within 12 months. RESULTS: Out of 4,5323,332 serum creatinine measurements, 42% lacked pACR measurements within 12 months. Approximately 10.8% of tests suggested CKD, mostly at stage 3a. The proportion of serum creatinine exams paired with pACR assessment varied according to the CKD stage. Internal Medicine, Cardiology, and Obstetrics/Gynecology were the specialties requesting most of the creatinine tests. Nephrology contributed with only 1.1% of serum creatinine requests for testing. CONCLUSION: Our findings reveal that a significant proportion of individuals with a creatinine test lack an accompanying urinary albuminuria measurement in Brazil, contrary to the recommendations of the international guidelines. Non-Nephrologists perform most kidney function evaluations, even among patients with presumable advanced CKD. This highlights the urge to incorporate in clinical practice the early detection of CKD and to encourage more collaborative multidisciplinary care to improve CKD management.

The interplay between climate change and ageing: A systematic review of health indicators.

Climate change and rapid population ageing pose challenges for communities and public policies. This systematic review aims to gather data from studies that present health indicators establishing the connection between climate change and the physical and mental health of the older population (≥ 65 years), who experience a heightened vulnerability to the impacts of climate change when compared to other age cohorts. This review was conducted according to the PICO strategy and following Cochrane and PRISMA guidelines. Three databases (PubMed, Scopus and Greenfile) were searched for articles from 2015 to 2022. After applying inclusion and exclusion criteria,nineteen studies were included. The findings indicated that various climate change phenomena are associated with an elevated risk of mortality and morbidity outcomes in older adults. These included cardiovascular, respiratory, renal, and mental diseases, along with physical injuries. Notably, the impact of climate change was influenced by gender, socioeconomic status, education level, and age-vulnerability factors. Climate change directly affected the health of older adults through ambient temperature variability, extreme and abnormal temperatures, strong winds, sea temperature variability, extreme El Niño-southern Oscillation (ENSO) conditions and droughts, and indirectly by air pollution resulting from wildfires. This review presents further evidence confirming that climate change significantly impacts the health and well-being of older adults. It highlights the urgency for implementing effective strategies to facilitate adaptation and mitigation, enhancing the overall quality of life for all individuals.

Polycomb protein binding and looping in the ON transcriptional state.

Polycomb group (PcG) proteins mediate epigenetic silencing of important developmental genes by modifying histones and compacting chromatin through two major protein complexes, PRC1 and PRC2. These complexes are recruited to DNA by CpG islands (CGIs) in mammals and Polycomb response elements (PREs) in Drosophila. When PcG target genes are turned OFF, PcG proteins bind to PREs or CGIs, and PREs serve as anchors that loop together and stabilize gene silencing. Here, we address which PcG proteins bind to PREs and whether PREs mediate looping when their targets are in the ON transcriptional state. While the binding of most PcG proteins decreases at PREs in the ON state, one PRC1 component, Ph, remains bound. Further, PREs can loop to each other and with presumptive enhancers in the ON state and, like CGIs, may act as tethering elements between promoters and enhancers. Overall, our data suggest that PREs are important looping elements for developmental loci in both the ON and OFF states.

Non-bacterial thrombotic endocarditis in ROS1-rearranged lung cancer: a report of two cases.

Background: Lung cancer is one of the tumor types with highest incidence of thromboembolic events (TE), especially adenocarcinoma subtype. ROS1 rearrangements confer higher risk of TE. Non-bacterial thrombotic endocarditis (NBTE) is a rare event, frequently diagnosed on autopsy. Clinical suspicion is key to reach the diagnosis and start early treatment of the underlying cause and anticoagulation in order to improve patients' outcomes. Case Description: Here we present two cases of NBTE in patients with ROS1-rearranged lung cancer with different clinical debuts. A 42-year-old woman presented initial tetraplegia and impaired level of consciousness, and the other patient, a 54-year-old man, was diagnosed of stroke with sensitive loss of left body. Both were diagnosed of NBTE, confirmed by the finding of cardiac vegetation on echocardiogram and no microorganisms found on blood cultures. Both responded well to targeted therapy with lorlatinib and crizotinib and anticoagulation with heparin. Conclusions: NBTE is an infrequent disease which can cause severe neurological symptoms that impair quality of life, performance status and survival. Early clinical suspicion in patients with higher risk of TEs such as patients with rearrangements of ROS1 gene is of essence. Adequate management of underlying disease and anticoagulation may impact in the recovery of symptoms.

Loss of Ezh2 in the medial ganglionic eminence alters interneuron fate, cell morphology and gene expression profiles.

Introduction: Enhancer of zeste homolog 2 (Ezh2) is responsible for trimethylation of histone 3 at lysine 27 (H3K27me3), resulting in repression of gene expression. Here, we explore the role of Ezh2 in forebrain GABAergic interneuron development. Methods: We removed Ezh2 in the MGE by generating Nkx2-1Cre;Ezh2 conditional knockout mice. We then characterized changes in MGE-derived interneuron fate and electrophysiological properties in juvenile mice, as well as alterations in gene expression, chromatin accessibility and histone modifications in the MGE. Results: Loss of Ezh2 increases somatostatin-expressing (SST+) and decreases parvalbumin-expressing (PV+) interneurons in the forebrain. We observe fewer MGE-derived interneurons in the first postnatal week, indicating reduced interneuron production. Intrinsic electrophysiological properties in SST+ and PV+ interneurons are normal, but PV+ interneurons display increased axonal complexity in Ezh2 mutant mice. Single nuclei multiome analysis revealed differential gene expression patterns in the embryonic MGE that are predictive of these cell fate changes. Lastly, CUT&Tag analysis revealed that some genomic loci are particularly resistant or susceptible to shifts in H3K27me3 levels in the absence of Ezh2, indicating differential selectivity to epigenetic perturbation. Discussion: Thus, loss of Ezh2 in the MGE alters interneuron fate, morphology, and gene expression and regulation. These findings have important implications for both normal development and potentially in disease etiologies.

Tumor- and circulating-free DNA methylation identifies clinically relevant small cell lung cancer subtypes.

Small cell lung cancer (SCLC) is an aggressive malignancy composed of distinct transcriptional subtypes, but implementing subtyping in the clinic has remained challenging, particularly due to limited tissue availability. Given the known epigenetic regulation of critical SCLC transcriptional programs, we hypothesized that subtype-specific patterns of DNA methylation could be detected in tumor or blood from SCLC patients. Using genomic-wide reduced-representation bisulfite sequencing (RRBS) in two cohorts totaling 179 SCLC patients and using machine learning approaches, we report a highly accurate DNA methylation-based classifier (SCLC-DMC) that can distinguish SCLC subtypes. We further adjust the classifier for circulating-free DNA (cfDNA) to subtype SCLC from plasma. Using the cfDNA classifier (cfDMC), we demonstrate that SCLC phenotypes can evolve during disease progression, highlighting the need for longitudinal tracking of SCLC during clinical treatment. These data establish that tumor and cfDNA methylation can be used to identify SCLC subtypes and might guide precision SCLC therapy.

Consolidation Osimertinib Versus Durvalumab Versus Observation After Concurrent Chemoradiation in Unresectable EGFR-Mutant NSCLC: A Multicenter Retrospective Cohort Study.

INTRODUCTION: Durvalumab improves survival when used as consolidation therapy after chemoradiation (CRT) in patients with stage III NSCLC. The optimal consolidation therapy for patients with EGFR-mutant (EGFRmut) stage III NSCLC remains unknown. METHODS: In this multi-institutional, international retrospective analysis across 24 institutions, we evaluated outcomes in patients with stage III EGFRmut NSCLC treated with concurrent CRT followed by consolidation therapy with osimertinib, durvalumab, or observation between 2015 and 2022. Kaplan-Meier method was used to estimate real-world progression-free survival (rwPFS, primary end point) and overall survival (secondary end point). Treatment-related adverse events (trAEs) during consolidation treatment were defined using Common Terminology Criteria for Adverse Events version 5.0. Multivariable Cox regression analysis was used. RESULTS: Of 136 patients with stage III EGFRmut NSCLC treated with definitive concurrent CRT, 56 received consolidation durvalumab, 33 received consolidation osimertinib, and 47 was on observation alone. Baseline characteristics were similar across the three cohorts. With a median follow-up of 46 months for the entire cohort, the median duration of treatment was not reached (NR) for osimertinib (interquartile range: NR-NR) and was 5.5 (interquartile range: 2.4-10.8) months with durvalumab. After adjusting for nodal status, stage III A/B/C, and age, patients treated with consolidation osimertinib had significantly longer 24-month rwPFS compared to those treated with durvalumab or in the observation cohorts (osimertinib: 86%, durvalumab: 30%, observation: 27%, p < 0.001 for both comparisons). There was no difference in rwPFS between the durvalumab and the observation cohorts. No significant difference in overall survival across the three cohorts was detected, likely due to the limited follow-up. Any-grade trAE occurred in 52% (2 [6.1%] grade ≥3) and 48% (10 [18%] grade ≥3) of patients treated with osimertinib and durvalumab, respectively. Of 45 patients who progressed on consolidation durvalumab, 37 (82%) subsequently received EGFR tyrosine kinase inhibitors. Of these, 14 (38%) patients developed trAEs including five patients with pneumonitis (14%; 2 [5.4%] grade ≥3) and five patients with diarrhea (14%; 1 [2.7%] grade ≥3). CONCLUSIONS: This study suggests that among patients with stage III unresectable NSCLC with a sensitizing EGFR mutation, consolidation osimertinib was associated with a significantly longer rwPFS compared to durvalumab or observation. No unanticipated safety signals were observed with consolidation osimertinib.

Polycomb protein binding and looping mediated by Polycomb Response Elements in the ON transcriptional state.

Polycomb group proteins (PcG) mediate epigenetic silencing of important developmental genes and other targets. In Drosophila, canonical PcG-target genes contain Polycomb Response Elements (PREs) that recruit PcG protein complexes including PRC2 that trimethylates H3K27 forming large H3K27me3 domains. In the OFF transcriptional state, PREs loop with each other and this looping strengthens silencing. Here we address the question of what PcG proteins bind to PREs when canonical PcG target genes are expressed, and whether PREs loop when these genes are ON. Our data show that the answer to this question is PRE-specific but general conclusions can be made. First, within a PcG-target gene, some regulatory DNA can remain covered with H3K27me3 and PcG proteins remain bound to PREs in these regions. Second, when PREs are within H3K27ac domains, PcG-binding decreases, however, this depends on the protein and PRE. The DNA binding protein GAF, and the PcG protein Ph remain at PREs even when other PcG proteins are greatly depleted. In the ON state, PREs can still loop with each other, but also form loops with presumptive enhancers. These data support the model that, in addition to their role in PcG silencing, PREs can act as "promoter-tethering elements" mediating interactions between promoter proximal PREs and distant enhancers.

Comprehensive NGS profiling to enable detection of ALK gene rearrangements and MET amplifications in non-small cell lung cancer.

Introduction: Next-generation sequencing (NGS) is currently widely used for biomarker studies and molecular profiling to identify concurrent alterations that can lead to the better characterization of a tumor's molecular landscape. However, further evaluation of technical aspects related to the detection of gene rearrangements and copy number alterations is warranted. Methods: There were 12 ALK rearrangement-positive tumor specimens from patients with non-small cell lung cancer (NSCLC) previously detected via fluorescence in situ hybridization (FISH), immunohistochemistry (IHC), and an RNA-based NGS assay, and 26 MET high gene copy number (GCN) cases detected by FISH, selected for this retrospective study. All 38 pre-characterized cases were reassessed utilizing the PGDx™ elio™ tissue complete assay, a 505 gene targeted NGS panel, to evaluate concordance with these conventional diagnostic techniques. Results: The detection of ALK rearrangements using the DNA-based NGS assay demonstrated excellent sensitivity with the added benefit of characterizing gene fusion partners and genomic breakpoints. MET copy number alterations were also detected; however, some discordances were observed likely attributed to differences in algorithm, reporting thresholds and gene copy number state. TMB was also assessed by the assay and correlated to the presence of NSCLC driver alterations and was found to be significantly lower in cases with NGS-confirmed canonical driver mutations compared with those without (p=0.0019). Discussion: Overall, this study validates NGS as an accurate approach for detecting structural variants while also highlighting the need for further optimization to enable harmonization across methodologies for amplifications.

Hospital mortality from covid-19 in children and adolescents in Brazil in 2020-2021.

OBJECTIVE: To describe cases, deaths, and hospital mortality from covid-19 in children and adolescents in Brazil, according to age group, during the evolving phases of the pandemic in 2020 and 2021. METHODS: Census of patients aged up to 19 committed with severe acute respiratory syndrome, due to covid-19 or unspecified, notified to the Brazilian Influenza Epidemiological Surveillance Information System, from January 1, 2020, to December 31, 2021. The two years were divided into six phases, covering the spread of the disease-first, second and third wave-as well as the impact of vaccination. The pediatric population was categorized into infants, preschoolers, schoolchildren, and adolescents. Hospital mortality was assessed by pandemic phase and age group. RESULTS: A total of 144,041 patients were recorded in the two years, 18.2% of whom had confirmed cases of covid-19. Children under 5 years old (infants and preschoolers) accounted for 62.8% of those hospitalized. A total of 4,471 patients died, representing about 6.1 deaths per day. Infants were the ones who most progressed to the intensive care unit (24.7%) and had the highest gross number of deaths (n = 2,012), but mortality was higher among adolescents (5.7%), reaching 9.8% in phase 1. The first peak of deaths occurred in phase 1 (May/2020), and two other peaks occurred in phase 4 (March/2021 and May/2021). There was an increase in cases and deaths for younger ages since phase 4. Hospital mortality in the pediatric population was higher in phases 1, 4, and 6, following the phenomena of dissemination/interiorization of the virus in the country, beginning of the second wave and beginning of the third wave, respectively. CONCLUSION: The absolute number of cases of covid-19 in children and adolescents is significant. Although complete vaccination in descending order of age provided a natural deviation in age range, there was a greater gap between the curve of new hospitalized cases and the curve of deaths, indicating the positive impact of immunization.

Code Integrity and Confidentiality: An Active Data Approach for Active and Healthy Ageing.

Internet of Things cybersecurity is gaining attention as the number of devices installed in IoT environments is exponentially increasing while the number of attacks successfully addressed to these devices are also proliferating. Security concerns have, however, been mainly addressed to service availability and information integrity and confidentiality. Code integrity, on the other hand, is not receiving proper attention, mainly because of the limited resources of these devices, thus preventing the implementation of advanced protection mechanisms. This situation calls for further research on how traditional mechanisms for code integrity can be adapted to IoT devices. This work presents a mechanism for code integrity in IoT devices based on a virtual-machine approach. A proof-of-concept virtual machine is presented, specially designed for providing code integrity during firmware updates. The proposed approach has been experimentally validated in terms of resource consumption among the most-widespread micro-controller units. The obtained results demonstrate the feasibility of this robust mechanism for code integrity.

Pre-existing tumor host immunity characterization in resected non-small cell lung cancer.

INTRODUCTION: Neoadjuvant and adjuvant immune checkpoint blockade (ICB) have recently become standard of care in resectable non-small cell lung cancer (NSCLC). Yet, biomarkers that inform patients who benefit from this approach remain largely unknown. Here, we interrogated the tumor immune microenvironment (TIME) in early-stage NSCLC patients that underwent up-front surgery. METHODS: A total of 185 treatment-naïve patients with early-stage NSCLC, that underwent up-front surgical treatment between 2006 and 2018 at Hospital del Mar were included. 124 lung adenocarcinomas (LUADs), and 61 squamous cell carcinoma (LUSCs) were included in a tissue microarray. Immunohistochemistry for CD3, CD4, CD8, CD68, CD80, CD103, FOXP3, PD-1, PD-L1, PD-L2 and HLA class II were evaluated by digital image analysis (QuPath software). TIME was categorized into four groups using PD-L1 expression in tumor cells (<1 % or ≥1 %) and tumor resident memory (CD103+) immune cells (using the median as cut-off). We explored the association between different TIME dimensions and patient's clinicopathological features and outcomes. RESULTS: We found increased levels of T cell markers (CD3+, CD4+, CD8+ cells), functional immune markers (FOXP3+ cells) as well as, higher HLA-II tumor membrane expression in LUADs compared to LUSCs (p < 0.05 for all). In contrast, LUSCs displayed higher percentage of intratumor macrophages (CD68+ cells) as well as, higher PD-L1 and PD-L2 tumor membrane expression (p < 0.05 for all). Unsupervised analysis revealed three different tumor subsets characterized by membrane tumor expression of PD-L1, PD-L2 and HLA-class II. Enrichment of T cells (CD3+, CD8+ cells), regulatory T cells (FOXP3+ cells) and macrophages (CD68+ cells) was observed in the CD103+/PD-L1+ group (p < 0.05 for all). Multivariate analysis showed that infiltration by CD103+ immune cells was associated with improved OS (p = 0.009). CONCLUSIONS: TIME analysis in resected NSCLC highlighted differences by histology, PD-L1 expression and molecular subgroups. Biomarker studies using IHC might aid to individually tailor adjuvant treatment in early-stage NSCLC.

Naproxen chemoprevention induces proliferation of cytotoxic lymphocytes in Lynch Syndrome colorectal mucosa.

Background: Recent clinical trial data from Lynch Syndrome (LS) carriers demonstrated that naproxen administered for 6-months is a safe primary chemoprevention that promotes activation of different resident immune cell types without increasing lymphoid cellularity. While intriguing, the precise immune cell types enriched by naproxen remained unanswered. Here, we have utilized cutting-edge technology to elucidate the immune cell types activated by naproxen in mucosal tissue of LS patients. Methods: Normal colorectal mucosa samples (pre- and post-treatment) from a subset of patients enrolled in the randomized and placebo-controlled 'Naproxen Study' were obtained and subjected to a tissue microarray for image mass cytometry (IMC) analysis. IMC data was processed using tissue segmentation and functional markers to ascertain cell type abundance. Computational outputs were then used to quantitatively compare immune cell abundance in pre- and post-naproxen specimens. Results: Using data-driven exploration, unsupervised clustering identified four populations of immune cell types with statistically significant changes between treatment and control groups. These four populations collectively describe a unique cell population of proliferating lymphocytes within mucosal samples from LS patients exposed to naproxen. Conclusions: Our findings show that daily exposure of naproxen promotes T-cell proliferation in the colonic mucosa, which paves way for developing combination of immunoprevention strategies including naproxen for LS patients.

ALTAI Tool for Assessing AI-Based Technologies: Lessons Learned and Recommendations from SHAPES Pilots.

Across European countries, the SHAPES Project is piloting AI-based technologies that could improve healthcare delivery for older people over 60 years old. This article aims to present a study developed inside the SHAPES Project to find a theoretical framework focused on AI-assisted technology in healthcare for older people living in the home, to assess the SHAPES AI-based technologies using the ALTAI tool, and to derive ethical recommendations regarding AI-based technologies for ageing and healthcare. The study has highlighted concerns and reservations about AI-based technologies, namely dealing with living at home, mobility, accessibility, data exchange procedures in cross-board cases, interoperability, and security. A list of recommendations is built not only for the healthcare sector, but also for other pilot studies.

MLL3/MLL4 methyltransferase activities control early embryonic development and embryonic stem cell differentiation in a lineage-selective manner.

H3K4me1 methyltransferases MLL3 (KMT2C) and MLL4 (KMT2D) are critical for enhancer activation, cell differentiation and development. However, roles of MLL3/4 enzymatic activities and MLL3/4-mediated enhancer H3K4me1 in these processes remain unclear. Here we report that constitutive elimination of both MLL3 and MLL4 enzymatic activities prevents initiation of gastrulation and leads to early embryonic lethality in mice. However, selective elimination of MLL3/4 enzymatic activities in embryonic, but not extraembryonic, lineages leaves gastrulation largely intact. Consistent with this, embryonic stem cells (ESCs) lacking MLL3/4 enzymatic activities can differentiate toward the three embryonic germ layers but show aberrant differentiation to extraembryonic endoderm (ExEn) and trophectoderm. The failure in ExEn differentiation can be attributed to markedly reduced enhancer-binding of the lineage-determining transcription factor GATA6. Furthermore, we show that MLL3/4-catalyzed H3K4me1 is largely dispensable for enhancer activation during ESC differentiation. Together, our findings suggest a lineage-selective, but enhancer activation-independent, role of MLL3/4 methyltransferase activities in early embryonic development and ESC differentiation.

Study of the nerve endings and mechanoreceptors of the medial meniscotibial ligament of the knee: A structural and distribution analysis.

BACKGROUND: The aim of the present study is to describe the morphology and distribution of the nerve endings of the meniscotibial ligament (MTL) of the knee, in order to understand the interaction between the proprioceptive system and knee mechanics. METHODS: Twenty medial MTLs were obtained from deceased organ donors. The ligaments were measured, weighed and cut. Sections (10 mm) were prepared on hematoxylin and eosin-stained slides for analysis of tissue integrity, and 50 mm sections were submitted to immunofluorescence with the protein gene product (PGP) 9.5 as primary antibody and Alexa Fluor 488 as secondary antibody, followed by microscopic analysis. RESULTS: The medial MTL was identified in 100% of the dissections, with average length, width, thickness and weight of 7.07 ± 1.34 mm, 32.25 ± 3.09 mm, 3.53 ± 0.27 mm and 0.67 ± 0.13 g, respectively. The hematoxylin and eosin-stained histological sections exhibited typical ligament structure, with dense well-organized collagen fibers and vascular tissue. All the specimens analyzed contained type I (Ruffini) mechanoreceptors and free (type IV) nerve endings, varying from parallel to intertwined fibers. Nerve endings not classified with different irregular shapes were also found. Most type I mechanoreceptors were found close to the MTL insertions on the tibial plateau, while the free nerve endings were found adjacent to the capsule. CONCLUSION: The medial MTL showed a peripheral nerve structure, primarily type I and IV mechanoreceptors. These findings suggest that the medial MTL is important for proprioception and medial knee stabilization.

Developing a Compost Quality Index (CQI) Based on the Electrochemical Quantification of Cd (HA) Reactivity.

The present work demonstrates the use of Cd2+ as a reactivity probe of the fulvic acids (FAs), humic acids (HAs) and dissolved organic matter (DOM) compost extracts. Significant differences were observed between the extracts, with the HA extract showing the highest reactivity. Comparing the different composts, the largest reactivity variation was again observed for HA then FA and finally DOM extracts. The Cd2+ binding extent was used to calculate the quality of composts and compared with a reference of uncomposted organic fertiliser (FLW), leading to the definition of an operational scale of compost quality. The parameter equivalent mass of fertiliser (mEF) was used for this scale sorted the seven composts from 0.353 to 1.09 kg FLW, for compost of sewage sludge (CSS) and vermicompost of domestic waste (CVDW), respectively. The significance of this parameter was verified through a correlation analysis between binding extent and the effect of compost application on lettuce crop growth in a field trial. The results demonstrate the potentiality of FA and HA extracts as markers of compost bioactivity and the use of Cd2+ as a reactivity probe.

Analysis of Immune Intratumor Heterogeneity Highlights Immunoregulatory and Coinhibitory Lymphocytes as Hallmarks of Recurrence in Stage I Non-Small Cell Lung Cancer.

Our understanding of the molecular mechanisms underlying postsurgical recurrence of non-small cell lung cancer (NSCLC) is rudimentary. Molecular and T cell repertoire intratumor heterogeneity (ITH) have been reported to be associated with postsurgical relapse; however, how ITH at the cellular level impacts survival is largely unknown. Here we report the analysis of 2880 multispectral images representing 14.2% to 27% of tumor areas from 33 patients with stage I NSCLC, including 17 cases (relapsed within 3 years after surgery) and 16 controls (without recurrence ≥5 years after surgery) using multiplex immunofluorescence. Spatial analysis was conducted to quantify the minimum distance between different cell types and immune cell infiltration around malignant cells. Immune ITH was defined as the variance of immune cells from 3 intratumor regions. We found that tumors from patients having relapsed display different immune biology compared with nonrecurrent tumors, with a higher percentage of tumor cells and macrophages expressing PD-L1 (P =.031 and P =.024, respectively), along with an increase in regulatory T cells (Treg) (P =.018), antigen-experienced T cells (P =.025), and effector-memory T cells (P =.041). Spatial analysis revealed that a higher level of infiltration of PD-L1+ macrophages (CD68+PD-L1+) or antigen-experienced cytotoxic T cells (CD3+CD8+PD-1+) in the tumor was associated with poor overall survival (P =.021 and P =.006, respectively). A higher degree of Treg ITH was associated with inferior recurrence-free survival regardless of tumor mutational burden (P =.022), neoantigen burden (P =.021), genomic ITH (P =.012) and T cell repertoire ITH (P =.001). Using multiregion multiplex immunofluorescence, we characterized ITH at the immune cell level along with whole exome and T cell repertoire sequencing from the same tumor regions. This approach highlights the role of immunoregulatory and coinhibitory signals as well as their spatial distribution and ITH that define the hallmarks of tumor relapse of stage I NSCLC.