Paired Control Research Articles

Exploring the influence of pre-analytical variables on gene expression measurements and relative expression orderings in cancer research

Gene expression profiling is an effective method for identifying predictive and prognostic biomarkers. However, measurements are prone to uncertainty and errors due to various pre-analytical variables. Systematic evaluating effects of these variables on gene expression measurements and relative expression orderings (REOs) of gene pairs, is necessary. A total of 18 datasets were collected, comprising over 800 paired samples. These paired samples were utilized to assess the impact of pre-analytical variables on gene expression measurements and REOs, including sampling methods, tumor sample heterogeneity, fixed time delays, preservation conditions, degradation levels, library preparation kits, amplification kits, RNA quantity, measuring platforms, and laboratory sites at single and multi-variable level. Low-quality samples served as the case group, while paired high-quality samples constituted the control group. In both single and multiple variable analyses, comparing each case sample to paired control sample revealed thousands of genes exhibited a twofold change in expression values. In contrast, on average, 82% and 76% of gene pairs keep consistent REO pattern between paired samples in single-variable and multi-variable analyses, respectively. Notably, the rate steadily increased after excluding gene pairs with the closest expression levels. Statistical analyses shown a higher proportion of differentially expressed genes (DEGs) than that of reversed gene pairs between case and control groups in both single-variable and multi-variable analyses. Furthermore, the proportion of reversal gene pairs among all gene pairs involving DEGs remained below 20% in the majority of comparisons. Our research demonstrates that REOs exhibit higher robustness under the influence of pre-analytical variables. These findings indicate the potential of the REOs-based approach in transcriptomics research and its applicability for biomarker studies.

The Anti-Inflammatory Effects of Liraglutide in Equine Inflammatory Joint Models.

This study investigates the anti-inflammatory properties of liraglutide, a glucagon-like peptide 1 receptor agonists, in equine in vitro models and in an in vivo acute synovitis model in Shetland ponies. The anti-inflammatory effect of liraglutide was assessed by measuring concentrations of inflammatory biomarker C-C Motif Chemokine Ligand 2 (CCL2) in culture media of equine whole blood, peripheral blood mononuclear cells (PBMCs), chondrocytes, and synoviocytes, with or without lipopolysaccharide (LPS) or interleukin-1β. In the in vivo experiment, acute synovitis was bilaterally induced with 0.25 ng LPS in the intercarpal joints of seven healthy Shetland ponies. The ponies were subsequently treated with either 6 mg liraglutide or a placebo as a paired control in each joint. The impact of liraglutide on biomarkers associated with inflammation (including white blood cell count, total protein, CCL2, and bradykinin) and cartilage metabolism (such as glycosaminoglycans, general matrix metalloproteinase activity, carboxypropeptide type II collagen, and collagen-cleavage neoepitope of type II collagen) was assessed across serial synovial fluid samples. Liraglutide was found to have an anti-inflammatory effect by reducing CCL2 concentrations in culture media of whole blood, PBMCs, chondrocytes, and synoviocytes. In contrast, no significant differences in synovial fluid inflammatory nor cartilage metabolism biomarker levels were found between joints treated with LPS and 6 mg liraglutide, versus LPS and placebo. In conclusion, liraglutide demonstrates the potential to attenuate inflammatory processes in joint cells. Additional research is necessary to validate its efficacy within the complex milieu of an inflamed joint.

Wood Chip Incorporation in Almond Orchard Planting Berms Increased Gravimetric Water Content, Soil CO2 Efflux, and Fine Root Length Density

Incorporation of chipped woody material into soil instead of burning or transporting wood chips (WCs) off-site is a practical solution to dispose of old orchard trees. In woody plants, the most external (fine) lateral roots are critical in acquiring water and nutrients and are the part of the root system that is most responsive to changes in the soil environment. Our objective was to measure the impact of pre-planting WC incorporation into the planting site on young almond tree root parameters such as root length density (root length per soil volume) and root architecture. We hypothesized that presence of decomposing WCs in soils in the years following planting leads to increased fine root length density and changes in root architecture that increase soil exploration per unit root biomass. Plum WCs were incorporated into the orchard soil 6 months before planting and three blocks were established with paired treatment and control (no WC) plots. Root traits, soil water content, soil bulk density, and soil CO2 efflux were measured in both the berm and alleyways twice in years 2 and 3 after planting. We found significant increases in standing root length density in planting berms with WC incorporation 2 and 3 years after trees were planted. This was due to an increase in root mass per soil volume rather than an increase in specific root length or a change in proportion of fine roots produced. Thus, although root foraging was increased in planting berms with WCs incorporated, this was not due to a change in root architectural traits.

P-1797. Implementation of a Handshake Stewardship Pilot for Inpatient General Medicine Teams at an Academic Medical Center

Abstract Background Handshake stewardship is an approach to antimicrobial stewardship that relies on in-person discussions between ASP teams and primary inpatient teams. This model has been implemented in pediatric and intensive care settings, and outcomes have been described in observational studies. In November 2023, UNCMC implemented a handshake stewardship pilot with crossover design for select inpatient general medicine teams. Methods Three pairs of matched inpatient general medicine teams at UNCMC were selected, with one team from each pair included in a three month intervention period. After three months, the teams were switched. The intervention consisted of twice weekly meetings of ASP physician and/or pharmacist and primary teams, during which antimicrobial regimens of all patients were reviewed except those with an ID consult. The intervention was delivered via in-person meetings for the two pairs of teams located at UNCMC, and via teleconference for a pair of teams located on a separate campus. The remote teams withdrew from the pilot during the intervention period. Recommendations made were catalogued as iVents in the Epic EMR and electronic surveys were circulated to primary team providers. Results 287 patients were reviewed across 95 meetings to date, yielding 73 recommendations. Common themes included narrowing coverage and transitioning to oral agents, although in some cases ASP teams recommended broadening therapy or a formal ID consult (Figure 1). 93% of recommendations were implemented. Survey data from 23 participants showed positive attitudes (Figure 2), with many valuing the opportunity to discuss patients that didn’t require a formal ID consult. Survey feedback from the virtual team included frustration with cessation of in-person ID consults at the site (unrelated to this pilot). Conclusion Handshake stewardship with in-person meetings has been well-received at UNCMC based on logged interventions and provider surveys, although the virtual component was unsuccessful. A planned crossover comparison between teams and paired controls will be helpful in assessing whether this has had an impact on antimicrobial use. Facilitating successful virtual handshake ASP will require improved coordination of meeting times and ensuring adequate access to in-person ID consults. Disclosures All Authors: No reported disclosures

CA-125 as a Biomarker in Renal Medullary Carcinoma: Integrated Molecular Profiling, Functional Characterization, and Prospective Clinical Validation.

Renal medullary carcinoma (RMC) is a highly aggressive malignancy defined by the loss of the SMARCB1 tumor suppressor. It mainly affects young individuals of African descent with sickle cell trait, and it is resistant to conventional therapies used for other renal cell carcinomas. This study aimed to identify potential biomarkers for early detection and disease monitoring of RMC. Integrated profiling of primary untreated RMC tumor tissues and paired adjacent kidney controls was performed using RNA-sequencing (RNA-seq) and histone Chromatin Immunoprecipitation Sequencing (ChIP-seq). The expression of serum cancer antigen 125 (CA-125), was prospectively evaluated in 47 patients with RMC. Functional studies were conducted in RMC cell lines to assess the effects of SMARCB1 re-expression. MUC16, encoding for CA-125, was identified as one of the top upregulated genes in RMC tissues, with concomitant enrichment of active histone marks H3K4me3 and H3K27ac at its promoter. Elevated serum CA-125 levels were found in 31 of 47 (66%) RMC patients and correlated significantly with metastatic tumor burden (p = 0.03). Functional studies in RMC cell lines demonstrated that SMARCB1 re-expression significantly reduced MUC16 expression. The correlation between serum CA-125 levels and metastatic burden suggests that CA-125 is a clinically relevant biomarker for RMC. These findings support further exploration of CA-125 for disease monitoring and targeted therapeutics in RMC.

The cortical high-flow sign in oligodendroglioma, IDH-mutant and 1p/19q-codeleted is correlated with histological cortical vascular density.

The cortical high-flow sign has been more commonly reported in oligodendroglioma, IDH-mutant and 1p/19q-codeleted (ODG IDHm-codel) compared to diffuse glioma with IDH-wildtype or astrocytoma, IDH-mutant. Besides tumor types, higher grades of glioma might also contribute to the cortical high flow. Therefore, we investigated whether the histological cortical vascular density or CNS WHO grade was associated with the cortical high-flow sign in patients with ODG IDHm-codel. This retrospective study consisted of pathologically confirmed 25 adult patients with ODG IDHm-codel. We implemented pseudo-continuous arterial spin labeling technique with background suppression. Subtraction images were generated from paired control and label images. Tumor-affecting cortices without intense contrast enhancement on conventional MR imaging were targeted for the determination of the cortical high-flow sign. Immunohistochemical staining of CD31 antibody was performed for the identification of vascular endothelial cells. A microscopic field of the most intense vascularization was captured in each specimen. The vessel number and the relative vascular density (%Vessel) were compared between the positive cortical high-flow sign (CHFS+) and the negative cortical high-flow sign (CHFS-) groups using the Mann-Whitney U test. Second, Fisher's exact test was used to compare the difference between the presence or absence of cortical high-flow sign and CNS WHO grades. Finally, the vessel number and %Vessel were compared between the CNS WHO grade 2 and grade 3 using the Mann-Whitney U test. The vessel number and %Vessel were higher in patients with the CHFS+ group than in patients with CHFS- group (p = 0.016 and p = 0.005, respectively). We observed no significant differences (p = 1.00) in the frequency of cortical high-flow sign between the CNS WHO grade 2 and grade 3. In addition, no significant differences are found in the vessel number and %Vessel between the CNS WHO grade 2 and grade 3 (p = 0.121 and p = 0.475, respectively). The cortical high-flow sign on ASL, which is more commonly found in ODG IDHm-codel than in diffuse glioma with IDH-wildtype or astrocytoma, is associated with the histological cortical vascular density in patients with ODG IDHm-codel.

Life‐history trade‐offs and stress resistance in Drosophila melanogaster populations adapted to pathogenic bacterial infection

AbstractEvolution of increased immune defence is often limited by costs: correlated changes in other traits (viz. life‐history traits) that otherwise reduce the fitness of the host organisms. Experimental evolution studies are useful for understanding the evolution of immune function and correlated changes in other traits. We experimentally evolved replicate Drosophila melanogaster populations to better survive infection challenges with an entomopathogenic bacteria, Enterococcus faecalis. Within 35 generations of directional selection, selected populations showed a marked increase in post‐infection survival than ancestrally paired controls. We next measured various life‐history traits of these populations. Our results show that the selected populations do not differ from control populations for larval development time and body weight at eclosion. No difference is also observed in case of fecundity and longevity (following the acute phase of infection), either when the flies were subjected to infection or when the flies were uninfected, although infected flies from all populations die much earlier than uninfected flies. Selected populations are either equally good or occasionally better as the control populations at surviving abiotic stressors (starvation and desiccation), although infected flies from all populations are more susceptible to stress than uninfected flies. Therefore, we conclude that (a) D. melanogaster populations can rapidly evolve to be more immune to infection with E. faecalis, (b) the evolution of increased defence against E. faecalis entails no life‐history cost for the hosts and (c) evolving defence against a biotic threat (pathogen) does not make flies more susceptible to abiotic stressors.

The influence of lightning on insect and fungal dynamics in a lowland tropical forest.

Lightning strikes are a common source of disturbance in tropical forests, and a typical strike generates large quantities of dead wood. Lightning-damaged trees are a consistent resource for tropical saproxylic (i.e., dead wood-dependent) organisms, but patterns of consumer colonization and succession following lightning strikes are not known. Here, we documented the occurrence of four common consumer taxa spanning multiple trophic levels-beetles, Azteca ants, termites, and fungi-in lightning strike sites and nearby undamaged control sites over time in a lowland forest of Panama. Beetle abundance was 10 times higher in lightning strike sites than in paired control sites, and beetle assemblages were compositionally distinct. Those in strike sites were initially dominated by bark and ambrosia beetles (Curculionidae: Platypodinae, Scolytinae); bark and ambrosia beetles, and predaceous taxa increased in abundance relatively synchronously. Beetle activity and fungal fruiting bodies, respectively, were 3.8 and 12.2 times more likely to be observed in lightning-damaged trees in strike sites versus undamaged trees in paired control sites, whereas the occurrence probabilities of Azteca ants and termites were similar between damaged trees in lightning strike sites and undamaged trees in control sites. Tree size also was important; larger dead trees in strike sites were more likely to support beetles, termites, and fungal fruiting bodies, and larger trees-regardless of mortality status-were more likely to host Azteca. Beetle presence was associated with higher rates of subsequent fungal presence, providing some evidence of beetle-associated priority effects on colonization patterns. These results suggest that lightning plays a key role in supporting tropical insect and fungal consumers by providing localized patches of suitable habitat. Any climate-driven changes in lightning frequency in tropical forests will likely affect a broad suite of consumer organisms, potentially altering ecosystem-level processes.

Mural Cells Initiate Endothelial-to-Mesenchymal Transition in Adjacent Endothelial Cells in Extracranial AVMs.

Extracranial arteriovenous malformations (eAVMs) are complex vascular lesions characterized by anomalous arteriovenous connections, vascular instability, and disruptions in endothelial cell (EC)-to-mural cell (MC) interactions. This study sought to determine whether eAVM-MCs could induce endothelial-to-mesenchymal transition (EndMT), a process known to disrupt vascular integrity, in the eAVM microenvironment. eAVM and paired control tissues were analyzed using RT-PCR for EC (CD31, CD34, and CDH5) and EndMT-specific markers (SNAI1, SNAI2, ACTA2/α-SMA, N-cadherin/CDH2, VIM). Immunohistochemistry (IHC) was also performed to analyze MC- (PDGFR-β and α-SMA), EC (CD31, CD34, and CDH5), and EndMT-specific markers (CDH2 and SNAI1) in sequential paraffin-embedded sections of eAVM patient biopsies and in adjacent normal tissue biopsies from the same patients. Furthermore, eAVM-MCs and MCs from normal paired tissues (NMCs) were then isolated from fresh human surgical samples using flow cytometry and co-cultured with normal human umbilical vein vascular endothelial cells (HUVECs), followed by analysis of CD31 by immunofluorescence. RT-PCR analysis did not show a significant difference in the expression of EC markers between eAVM tissues and controls, whereas expression of EndMT-specific markers was upregulated in eAVM tissues compared to controls. IHC of eAVMs and paired control tissues demonstrated regions of significant perivascular eAVM-MC expansion (PDGFR-β+, and α-SMA+) surrounding dilated, morphologically abnormal vessels. These regions contained endothelium undergoing EndMT as evidenced by loss of CD31, CD34, and CDH5 expression and upregulation of the EndMT-associated genes CDH2 and SNAI1. Isolated eAVM-MCs induced loss of CD31 in HUVECs when grown in co-culture, while NMCs did not. This study suggests that the eAVM endothelium surrounded by expanded eAVM-MCs undergoes EndMT, potentially leading to the formation of dilated and fragile vessels, and implicates the eAVM-MCs in EndMT initiation and eAVM pathology.

Steering spin fluctuations in lattice systems via two-tone Floquet engineering

Abstract We report on the controlled creation and destruction of antiferromagnetic dimers using two-tone Floquet engineering. We consider a one-dimensional spin-1/2 lattice with periodically modulated bonds using parametric resonances. The stroboscopic dynamics generated from distributed bond modulations lead to pair correlation between spins. Consequently, subharmonic response in local observables breaks discrete time translational symmetry and leads to the emergence of Floquet dynamical dimerisation. We present a protocol allowing the control of local spin-correlated pairs driven by one-period evolution operators, providing significant insight into new nonequilibrium states of matter that can be feasibly implemented in current quantum simulator platforms.

In vivo reduction of biofilm seeded on orthopaedic implants.

Electromagnetic induction heating has demonstrated in vitro antibacterial efficacy over biofilms on metallic biomaterials, although no in vivo studies have been published. Assessment of side effects, including thermal necrosis of adjacent tissue, would determine transferability into clinical practice. Our goal was to assess bone necrosis and antibacterial efficacy of induction heating on biofilm-infected implants in an in vivo setting. Titanium-aluminium-vanadium (Ti6Al4V) screws were implanted in medial condyle of New Zealand giant rabbit knee. Study intervention consisted of induction heating of the screw head up to 70°C for 3.5 minutes after implantation using a portable device. Both knees were implanted, and induction heating was applied unilaterally keeping contralateral knee as paired control. Sterile screws were implanted in six rabbits, while the other six received screws coated with Staphylococcus aureus biofilm. Sacrifice and sample collection were performed 24, 48, or 96 hours postoperatively. Retrieved screws were sonicated, and adhered bacteria were estimated via drop-plate. Width of bone necrosis in retrieved femora was assessed through microscopic examination. Analysis was performed using non-parametric tests with significance fixed at p ≤ 0.05. The width of necrosis margin in induction heating-treated knees ranged from 0 to 650 μm in the sterile-screw group, and 0 to 517 μm in the biofilm-infected group. No significant differences were found between paired knees. In rabbits implanted with sterile screws, no bacteria were detected. In rabbits implanted with infected screws, a significant bacterial load reduction with median 0.75 Log10 colony-forming units/ml was observed (p = 0.016). Induction heating was not associated with any demonstrable thermal bone necrosis in our rabbit knee model, and might reduce bacterial load in S. aureus biofilms on Ti6Al4V implants.

Exploring dysfunctional barrier phenotypes associated with glaucoma using a human pluripotent stem cell-based model of the neurovascular unit

Glaucoma is a neurodegenerative disease that results in the degeneration of retinal ganglion cells (RGCs) and subsequent loss of vision. While RGCs are the primary cell type affected in glaucoma, neighboring cell types selectively modulate RGCs to maintain overall homeostasis. Among these neighboring cell types, astrocytes, microvascular endothelial cells (MVECs), and pericytes coordinate with neurons to form the neurovascular unit that provides a physical barrier to limit the passage of toxic materials from the blood into neural tissue. Previous studies have demonstrated that these barrier properties may be compromised in the progression of glaucoma, yet mechanisms by which this happens have remained incompletely understood. Thus, the goals of this study were to adapt a human pluripotent stem cell (hPSC)-based model of the neurovascular unit to the study of barrier integrity relevant to glaucoma. To achieve this, hPSCs were differentiated into the cell types that contribute to this barrier, including RGCs, astrocytes, and MVECs, then assembled into an established Transwell®-insert model. The ability of these cell types to contribute to an in vitro barrier model was tested for their ability to recapitulate characteristic barrier properties. Results revealed that barrier properties of MVECs were enhanced when cultured in the presence of RGCs and astrocytes compared to MVECs cultured alone. Conversely, the versatility of this system to model aspects of barrier dysfunction relevant to glaucoma was tested using an hPSC line with a glaucoma-specific Optineurin (E50K) mutation as well as a paired isogenic control, where MVECs then exhibited reduced barrier integrity. To identify factors that could result in barrier dysfunction, results revealed an increased expression of TGFβ2 in glaucoma-associated OPTN(E50K) astrocytes, indicating a potential role for TGFβ2 in disease manifestation. To test this hypothesis, we explored the ability to modulate exogenous TGFβ2 in both isogenic control and OPTN(E50K) experimental conditions. Collectively, the results of this study indicated that the repurposing of this in vitro barrier model for glaucoma reliably mimicked some aspects of barrier dysfunction, and may serve as a platform for drug discovery, as well as a powerful in vitro model to test the consequences of barrier dysfunction upon RGCs in glaucoma.

DNAR-10. PHASE I CLINICAL TRIAL OF PEPOSERTIB PLUS RADIOTHERAPY IN ADULTS WITH NEWLY-DIAGNOSED MGMT-UNMETHYLATED GLIOBLASTOMA

Abstract BACKGROUND DNA-dependent protein kinase (DNA-PK) is an attractive target in GBM because it promotes unwanted DNA repair and renders cancer cells less vulnerable to DNA damaging agents, such as radiotherapy (RT). Peposertib, a small molecule inhibitor of DNA-PK, has demonstrated pre-clinical efficacy in sensitizing GBM to RT, resulting in tumor regression. METHODS 21 patients with newly-diagnosed MGMT-unmethylated GBM received peposertib plus 6-weeks of RT to determine the maximum-tolerated dose (MTD) based on the Bayesian Optimal Interval design. The dose-limiting toxicity (DLT) period was 10-weeks. Patients received at least 6-cycles of adjuvant temozolomide (NCT04555577). Single-nuclei RNA-sequencing (snRNA-Seq) and spatial transcriptomics (Visium, 10x Genomics) were performed on matched pair primary-recurrent tumors and controls, incorporating tumor cell state and cell type definitions by automated cluster classification and differential expression of marker genes. RESULTS The MTD of peposertib in combination with RT was 300mg per RT fraction day. One DLT (G3 radiation necrosis @300mg) was observed. After a median follow-up of 15.4 months (interquartile-range 8.9-18.5), mPFS was 10.8 months (95%CI 8.33-NR) and mOS was 22.9 months (95%CI 16-NR). 6/21 (29%) patients have undergone re-resection to date. snRNA-Seq on 102,709 cells from 12 primary and 3 recurrent tumors revealed that pre-treatment tumor cell states were progenitor-like, while treatment with peposertib plus RT was associated with differentiated and inflammatory malignant cell states, along with macrophage infiltration. Spatial transcriptomics of 8 matched primary-recurrent tumors (20,793 transcriptomes) confirmed malignant cell differentiation and enrichment of pro-inflammatory macrophages after inhibition of DNA-PK. 5 patients will be enrolled into a window-of-opportunity expansion cohort to evaluate intratumoral drug concentration. CONCLUSION The combination of peposertib and intracranial RT was safe. Survival data for this MGMT-unmethylated GBM population was promising. Single nuclei and spatial transcriptomic analyses revealed enrichment of pro-inflammatory macrophages in the tumor microenvironmentafter DNA-PK inhibition. The study was supported by EMD-Serono (CrossRef Funder ID: 10.13039/100004755).

A ‘Tuba Drain’ incorporated in sink drains reduces counts of antibiotic-resistant bacterial species at the plughole: a blinded, randomized trial in 36 sinks in a hospital outpatient department with a low prevalence of sink colonization by antibiotic-resistant species

Multi-resistant Gram-negative bacteria (GNB) survive in hospital drains in traps that contain water and may ascend into the sink because of splashes, or biofilm growth. To investigate whether the 'Tuba Drain' (TD) a long, bent, continually descending copper tube between the sink outlet and the trap prevents the ascent of bacteria. After initial laboratory tests confirmed that the TD prevented bacteria in the U-bend from splashing upwards into the sink outlet, TDs were assessed in a randomized, blinded trial in a hospital outpatient department built in 2019. Sinks were paired into those with a similar clinical exposure and each member of each pair was randomized to receive either new, standard plumbing up to and including the trap (18 sinks) or the same new standard plumbing but including the TD inserted between the sink outlet and trap. Bacterial counts in swabs from the sink outlets were determined blindly before and monthly after the plumbing change for a year. GNB that are associated with clinical infection and carriage of resistance genes, Pseudomonas aeruginosa, Acinetobacter baumanii, Stenotrophomonas maltophilia and all Enterobacterales were the organisms of primary interest and termed target bacteria. The TDs fitted into the required spaces and functioned without problems. The geometric means (over months) of the counts of target bacteria in TD-plumbed sinks was lower than those in their paired controls, P=0.012 (sign test, two-tailed). Prevalence of target bacteria in sinks was low. TDs were effective for reducing target bacteria in sinks.

Digoxin reduces senescence and restores dysfunctional endothelial-subcutaneous adipocyte cross-talk and dysregulated glucose homeostasis in patients with type 2 diabetes mellitus

Abstract Background Normal subcutaneous adipose tissue (SAT) function is dependent on SAT microvascular expansion. Heart failure with reduced ejection fraction (HF) and type 2 diabetes mellitus (DM) have a synergistic deleterious relationship and, of relevance to this, metabolic dysregulation may drive HF progression in DM. Chronic hyperinsulinemia is associated with adipocyte senescence, however any contributing role of SAT microvascular endothelial cell (SATMVEC) senescence on adverse microvascular expansion in type 2 diabetes mellitus (DM) remains unknown. Digoxin has been shown to have senolytic activity in murine cancer models. However, the potential therapeutic role of digoxin on SAT metabolic function is unexplored. Purpose We establish the presence of SATMVEC senescence and its effects on adipocyte function in people with HF and DM (HFDM). Moreover, we establish a therapeutic role for digoxin in targeting SATMVEC senescence, modulating SATMVEC-adipocyte crosstalk and restoring metabolic homeostasis. Methods We took pectoral SAT biopsies from 59 patients undergoing pacemaker implantation of whom 25 (42%) had HF and DM (HFDM). SATMVEC were isolated from SAT and were treated with 1ng/mL digoxin vs control for 24 hours before characterisation (cell function, expression of senescence-associated ß-galactosidase (ß-gal) and senescence associated secretory phenotype, SASP). Crosstalk between SATMVEC and subcutaneous white adipocytes were examined using co-culture techniques (Fig 1A). Paired digoxin vs control treated SATMVEC were seeded co-cultured with adipocytes for 24 hours, before adipocyte health was quantified, through 2-NBD-glucose uptake and expression of key adipogenic genes and proteins. Results Compared to HF alone, isolated SATMVEC from patients with HFDM had a senescent phenotype with increased SASP and ß-gal expression (p<0.05), reduced metabolic activity, ATP production, proliferative capacity, and impaired angiogenesis (p<0.05). In co-culture, SATMVEC from patients with HFDM had deleterious effects on adipocyte function: increasing expression of IL-6 (Fig 1C) and reducing 2-NBD-glucose uptake (p<0.05). Treatment of SATMVEC with digoxin reduced the SATMVEC senescent phenotype (Fig 1B) and increased 2-NBD-glucose adipocyte uptake (Figure 1D) (p<0.001). Conclusions SATMVEC from patients with HFDM have a senescent phenotype, and when in co-culture induce a proinflammatory adipocyte phenotype, resistant to glucose uptake. Digoxin reduces SATMVEC senescence, restores healthy adipocyte glucose homeostasis, and thus has the potential to repair dysfunctional SAT in patients with HFDM.

An Exploratory Study of Contextual Control Modes in Teamwork.

This study investigated the relationship between human team member contextual control and team performance under time constraints. Contextual control modes, which describe different strategies for action selection in dynamic environments, characterize how humans maintain performance under variable demands. Control modes have not yet been studied in teamwork settings. Modeling of the cross-level interaction between team members' control modes and emerging team behaviors can improve understanding of effective teamwork in dynamic environments. A human-subjects study explored the relationship between individual contextual control and team performance. Questionnaires about contextual control were used to elicit individual control modes. Analysis compared team members' control modes and investigated how control modes changed under varying time pressures. Participant's control modes differed in their look ahead horizon, the extensiveness of prior action evaluation, and their prior experience. Many team members shifted control modes during trials, resulting in both convergence and divergence of paired control modes. No effects on communication rate were found due to changes in team members' control modes, but partially significant findings may suggest that the control mode divergence affects performance. Teams can operate in multiple control mode configurations that change dynamically according to context. Further research with an increased sample size is warranted to analyze how time constraints influence team members' control modes and overall teaming processes and whether divergence of team control mode is favorable under time pressures. Further study of contextual control in teams may help improve team design to better support teams in coping with time constraints in dynamic environments.

Vascular endothelial growth factor A inhibition remodels the transcriptional signature of lipid metabolism in psoriasis non-lesional skin in 12 h ex vivo culture.

Vascular endothelial growth factor A (VEGF-A)-mediated angiogenesis is involved in the pathogenesis of psoriasis. VEGF-A inhibitors are widely used to treat oncological and ophthalmological diseases but have not been used in psoriasis management. The molecular mechanisms underlying the effects of VEGF-A inhibition in psoriatic skin remain unknown. To identify the genes and canonical pathways affected by VEGF-A inhibition in non-lesional and plaque skin ex vivo. Total RNA sequencing was performed on skin biopsies from patients with psoriasis (n=6; plaque and non-lesional skin) and healthy controls (n=6) incubated with anti-VEGF-A monoclonal antibody (bevacizumab, Avastin®) or human IgG1 isotype control for 12h in serum-free organ culture. Differentially expressed genes between paired control and treated samples with adjusted p-values <0.1 were considered significant. Gene ontology and ingenuity pathway analysis was used to identify enriched biological processes, canonical pathways and upstream regulators. VEGF-A inhibition upregulated the expression of genes involved in lipid metabolism. Pathway enrichment analysis identified the activation of pathways involved in fatty acids and lipid biosynthesis and degradation in non-lesional skin and ferroptosis in plaque skin. VEGF-A inhibition downregulated endothelial cell apoptosis in non-lesional psoriasis skin and members of the interferon family were identified as potential regulators of the effects of VEGF-A inhibition in non-lesional skin. Early response to VEGF-A inhibition is associated with changes in lipid metabolism in non-lesional psoriasis skin and cellular stress in psoriasis plaque. More investigation is needed to validate these findings.

Reindeer carcasses modulate vegetation composition and greenness in High-Arctic tundra

Vertebrate carrion is an integral part of foodwebs in ecosystems and can impact biodiversity at the local as well as the landscape scale. However, very little knowledge currently exists about the ecological role of carrion in the Arctic ecosystems. We conducted a ground survey on the cover of five plant functional groups at paired reindeer carcass and control sites and analysed the relationship between cover and carcass presence in the Arctic tundra of Svalbard. Vegetation indices from Red–Green–Blue (RGB) imagery captured by drones complemented this, assessing plant productivity in terms of “spectral greening” and modelling the relationship between vegetation index values and carcass distance. We show that graminoids capitalised most from carcass presence, whereas bryophytes and lichen showed decreases in cover. Woody plant and forb covers were not significantly impacted by carcass presence. The Red Green Blue Vegetation Index decreased locally at fresh carcasses (i.e., &lt;1 year old) but showed an increase at more established carcass sites (i.e., &gt;1 year). We show that carcasses have differential impacts on the plant functional groups of Svalbard's tundra and induce a local “green-up” through secondary succession within 2 m of the carcass. Given their non-random distribution, carcasses may contribute to vegetation heterogeneity at landscape scales. This is relevant for understanding how climate change-induced reindeer mortalities will impact tundra plant community composition in the future.

EFFECTS OF LOCAL HYPOTHERMIA ON LIMB VIABILITY IN A SWINE MODEL OF ACUTE LIMB ISCHEMIA DURING PROLONGED DAMAGE-CONTROL RESUSCITATION.

Background : New strategies are needed to mitigate further tissue injury during traumatic limb ischemia in cases requiring damage control resuscitation (DCR). Little is known about the pathophysiology and injury course in acute limb ischemia (ALI) with DCR in polytraumatized casualties. We therefore investigated the effects of therapeutic limb hypothermia in a swine model of ALI and DCR. Methods : Fifteen swine underwent a published 6-h DCR protocol of hemorrhage and then resuscitation. After hemorrhage, animals were randomized to 5°C or 15°C cooling of one hindlimb; the contralateral limb serving as an uncooled control. Physiologic variables, limb temperature, and limb tissue metabolites (glucose, lactate, and pyruvate) were measured throughout the DCR protocol. Muscle and nerve biopsies were obtained after the 6-h protocol. Results : Lactate and pyruvate levels were significantly lower in the cooled limbs than in the uncooled control limbs but did not differ between the 5°C and 15°C groups. Tissue glucose levels did not differ between the 5°C group, the 15°C group, and controls. Mean histologic muscle score was significantly higher in the 5°C group than in controls ( P = 0.03). Mean nerve histology scores did not differ between the 5°C and paired control limbs, or between the mean muscle and nerve histology scores of the 15°C and paired control limbs. Conclusion : Cooling to 15°C significantly reduced local tissue metabolites compared to paired controls, while producing no significant increase in histologic damage, whereas cooling to 5°C increased histologic muscle damage. These results suggest an approach to prevention of ischemic injury through local hypothermia but warrant further functional testing.

Impact of triflumezopyrim (insecticide) on blood and serum biochemistry of freshwater fish, subadult Labeo rohita (Hamilton, 1822)

The impact of insecticides on water bodies can be evaluated by studying the physiology and health of fish. These chemicals pose a significant and direct risk to aquatic ecosystems and hence aquatic animals like fish. Therefore, this study was designed to investigate the toxic effects of sub-lethal concentrations of triflumezopyrim, a novel synthetic insecticide commonly used in crop protection, on the biochemical and hematological parameters of freshwater sub-adult Labeo rohita L. Here, fish were subjected to sub-lethal doses of triflumezopyrim at various concentrations (1.41 ppm, 3.27 ppm, and 4.97 ppm) for 21 days. In addition, fish were grown in an insecticide-free control group to determine the toxic effects of the insecticides. The obtained data on various biochemical and hematological parameters of the fish were analyzed using state-of-the-art statistics. Our findings suggest that triflumezopyrim exposure has a significant effect on blood variables. However, variable response, such as mean corpuscular volume, hemoglobin concentration, neutrophils, and eosinophils were unaffected by insecticide treatment, regardless of the dose. Our interhematological parameter analysis revealed varying interrelationships at various insecticide doses. Similarly, triflumezopyrim exposure significantly affected the response of serum biomarker profiles i.e., the difference in mean treatment pairs (control and three doses of triflumezopyrim) had a significant effect on the response of most serum profile parameters.For example, cholesterol response was inversely related to serological markers, including total protein, alanine aminotransferase, and aspartate aminotransferase, under triflumezopyrim exposure. The relationship pattern and grouping dynamics of the blood and serum parameters changed drastically under various insecticide doses according to correlation and cluster analyses. Our results showed that repeated exposure of fish to various sub-lethal concentrations of triflumezopyrim negatively affected their physiology and serology, which is highly harmful to the organism.