BRCA2 stabilises RAD51 and DMC1 nucleoprotein filaments through a conserved interaction mode.

BRCA2 is essential for DNA repair by homologous recombination in mitosis and meiosis. It interacts with recombinases RAD51 and DMC1 to facilitate the formation of nucleoprotein filaments on resected DNA ends that catalyse recombination-mediated repair. BRCA2's BRC repeats bind and disrupt RAD51 and DMC1 filaments, whereas its PhePP motifs bind recombinases and stabilise their nucleoprotein filaments. However, the mechanism of filament stabilisation has hitherto remained unknown. Here, we report the crystal structure of a BRCA2-DMC1 complex, revealing how core interaction sites of PhePP motifs bind to recombinases. The interaction mode is conserved for RAD51 and DMC1, which selectively bind to BRCA2's two distinct PhePP motifs via subtly divergent binding pockets. PhePP motif sequences surrounding their core interaction sites protect nucleoprotein filaments from BRC-mediated disruption. Hence, we report the structural basis of how BRCA2's PhePP motifs stabilise RAD51 and DMC1 nucleoprotein filaments for their essential roles in mitotic and meiotic recombination.

Analysis of the Structure of 14 Therapeutic Antibodies Using Circular Dichroism Spectroscopy.

Understanding the impact of the manufacturing environment on therapeutic monoclonal antibody (mAb) structures requires new process analytical technology. Here, we describe the creation of a new reference set for the circular dichroism (CD) spectra of mAbs. Data sets of the highest quality were collected by synchrotron radiation CD for 14 different mAbs in both native and acid-stressed states. Deconvolution of far-UV spectra for the mAb cohort identified two current reference sets (SP175 and SMP180) as assigning accurate secondary structures, irrespective of the analysis program employed. Scrutiny of spectra revealed significant variation in the far-UV and especially near-UV CD of the 14 mAbs. Two spectral features were found to be sensitive to changes in solution pH, i.e., the far-UV positive peak at 201-202 nm and the near-UV negative exciton couplet around 230-240 nm. The latter feature offers attractive possibilities for in-line CD-based monitoring of the mAb structure during manufacture.

Prevalence of cardiomyopathy and cardiac mortality in a colony of non-purebred cats in New Zealand.

AIMS: To evaluate the prevalence of subclinical cardiomyopathy and cardiac mortality in a research colony of non-purebred cats, established as a model of the wider cat population in New Zealand. METHODS: All apparently healthy, compliant, non-pregnant, non-neonatal cats in the colony at the Centre for Feline Nutrition (Massey University, Palmerston North, NZ) underwent physical examination and echocardiography using a 4.4-6.2-MHz probe by a board-certified veterinary cardiologist. Cardiac phenotype was classified following current guidelines. Hypertrophic cardiomyopathy (HCM) phenotype was defined as an end-diastolic left ventricular wall thickness ≥ 6 mm. Colony mortality data from February 2012 to February 2022 was reviewed to determine cardiac mortality. RESULTS: Cats (n = 132; 65 females and 67 males) included in the study had a median age of 4.1 (IQR 3.0-8.0) years. Thirty-two (24%) cats had a heart murmur, and three (2%) cats had an arrhythmia. Echocardiography revealed heart disease in 24 (18.2%) cats, including 23 with an HCM phenotype and one with a restrictive cardiomyopathy phenotype. Of the cats with the HCM phenotype, 3/23 had systemic hypertension or hyperthyroidism or both, and these cats were excluded from the final diagnosis of HCM (20/132; 15.2 (95% CI = 9.5-22.4)%).Between 2012 and 2022, 168 colony cats died, with 132 undergoing post-mortem examination. Heart disease was considered the cause of death in 7/132 (5.3%; 95% CI = 2.2-10.6%) cats; five had HCM, one a congenital heart defect, and one myocarditis. The overall prevalence of death related to HCM in the colony during this period was 3.8% (95% CI = 1.2-8.6%). Three cats with HCM and the cat with a congenital heart defect died unexpectedly without prior clinical signs, while congestive heart failure was observed prior to death in two cats with HCM and the cat with myocarditis. Additionally, 30/132 (22.7%) cats had cardiac abnormalities but died for non-cardiac reasons. CONCLUSIONS: Subclinical cardiomyopathy, specifically HCM, was common in cats in the colony. Given that the colony originated as a convenience selection of non-purebred cats in New Zealand, the true prevalence of HCM in the wider New Zealand population is likely to fall within the 95% CI (9.5-22%). The proportion of deaths of colony cats due to HCM was lower (3.8%) supporting the conclusion that subclinical cardiomyopathy may not progress to clinical disease causing death. CLINICAL RELEVANCE: Veterinarians should be aware of the high prevalence of subclinical HCM when treating cats. ABBREVIATIONS: CAM: Systolic anterior motion of the chordae tendineae; CFN: Centre for Feline Nutrition; HCM: Hypertrophic cardiomyopathy; LA/Ao: Left atrial to aortic ratio; LV FS: Left ventricular fractional shortening; LVIDd: Left ventricular internal diameters in end-diastole; LVIDs: Left ventricular internal diameter in end-systole; LVWT: Max Maximum left ventricular wall thickness; SAM: Systolic anterior motion of the mitral valve; 2D: Two-dimensional.

Mapping domains of early life determinants of future multimorbidity across three UK longitudinal cohort studies.

Many studies use a reductionist approach to isolate the influence of one factor in childhood on multimorbidity rather than consider the combined effect of wider determinants. We explored how potential multiple early life determinants of multimorbidity can be characterised across three UK cohort studies. We used the National Child Development Study (NCDS), the 1970 British Cohort Study (BCS70), and the Aberdeen Children of the 1950s Study (ACONF) to identified early life variables that fit into 12 conceptualised domains of early life determinants of multimorbidity. Variables were assigned into 12 domains; principal component analysis reduced the dimensionality of the data and structured variables into subgroups. The data audit identified 7 domains in ACONF, 10 domains in NCDS and 12 domains in BCS70. Dominant components included maternal fertility histories within the prenatal, antenatal and birth domain, long-term illnesses within the child health domain, educational ability within the child education and health literacy domain, ethnicity within the demography domain, parental health behaviours within the transgenerational domain, housing within the socioeconomic domain and parental-child interactions within the parental-family domain. We demonstrated that if multiple large scale longitudinal studies are used, there is enough data available for researchers to consider conceptualising early life risk factors of multimorbidity across groups or domains. Such conceptualisation can help challenge the existing understanding of disease aetiology and develop new ideas for prevention of multimorbidity.

Adiponectin restores cholinergic vasodilation in resistance vessels from adult offspring exposed to maternal obesity in utero.

Maternal obesity increases the risk of cardiovascular and metabolic disease in the offspring both during childhood and adult life. Pregnant women and mice with obesity have lower circulating levels of adiponectin (ADN) than controls with normal body mass index. ADN is an adipokine involved in regulating energy metabolism, vascular function, and placental function. We hypothesized that offspring of obese mice have impaired resistance artery function, which can be prevented by restoration of normal circulating ADN levels in obese dams during late pregnancy. Adult female mice were fed either control or obesogenic diet and mated with control diet-fed males. Control dams received a continuous infusion of phosphate saline buffer (PBS) during late pregnancy whereas obese females received either PBS or ADN. After weaning, offspring were fed a control diet. Mesenteric arteries (MsA) were dissected from adult offspring and mounted in a wire myograph or fixed for histology. MsA responses to vasoconstrictors (phenylephrine and endothelin-1) were not different between infusion groups. However, the vasodilatory responses to acetylcholine were reduced in offspring from obese dams as compared to control-fed dams. ADN supplementation during pregnancy restored the cholinergic vasodilatory responses of resistance vessels in offspring from obese dams. These observations suggest a target for the prevention of the long-lasting vascular effects of obesity during pregnancy in the offspring.

Age related changes in skin sensitivity assessed with smartphone vibration testing.

The capacity to perceive tactile input at the fingertips, referred to as tactile sensitivity, is known to diminish with age due to regressive changes to mechanoreceptor density and morphology. Sensitivity is measured as perceptual responses to stimuli of varying intensity. Contrary to traditional sensitivity monitoring instruments, smartphones are uniquely suited for remote assessment and have shown to deliver highly calibrated stimuli along a broad spectrum of intensity, which may improve test reliability. The aim of this study was to evaluate a vibration-emitting smartphone application, the Vibratus App, as a mode of estimating tactile sensory thresholds in the aging adult. The peripheral nerve function of 40 neurologically healthy volunteers (ages 18-71) was measured using monofilaments, a 128-Hz tuning fork, the Vibratus App, and nerve conduction studies (NCS). Between group differences were analyzed to determine each measurement's sensitivity to age. Spearman correlation coefficients depicted the associative strength between hand-held measurements and sensory nerve action potential (SNAP) amplitude. Inter-rater reliability of traditional instruments and the software-operated smartphone were assessed by intraclass correlation coefficient (ICC2,k). Measurements taken with Vibratus App were significantly different between age groups (p < 0.001). The inter-rater reliability of monofilament, smartphone vibration, and tuning fork testing was moderate to good (ICC2,k = 0.65, 0.69, and 0.79, respectively). The findings of this study support further investigation of smartphones as sensitivity monitoring devices for at home monitoring of skin sensitivity.

MEILB2-BRME1 forms a V-shaped DNA clamp upon BRCA2-binding in meiotic recombination.

DNA double-strand break repair by homologous recombination has a specialised role in meiosis by generating crossovers that enable the formation of haploid germ cells. This requires meiosis-specific MEILB2-BRME1, which interacts with BRCA2 to facilitate loading of recombinases onto resected DNA ends. Here, we report the crystal structure of the MEILB2-BRME1 2:2 core complex, revealing a parallel four-helical assembly that recruits BRME1 to meiotic double-strand breaks in vivo. It forms an N-terminal ß-cap that binds to DNA, and a MEILB2 coiled-coil that bridges to C-terminal ARM domains. Upon BRCA2-binding, MEILB2-BRME1 2:2 complexes dimerize into a V-shaped 2:4:4 complex, with rod-like MEILB2-BRME1 components arranged at right-angles. The ß-caps located at the tips of the MEILB2-BRME1 limbs are separated by 25 nm, allowing them to bridge between DNA molecules. Thus, we propose that BRCA2 induces MEILB2-BRME1 to function as a DNA clamp, connecting resected DNA ends or homologous chromosomes to facilitate meiotic recombination.

Life histories are not just fast or slow.

Life history strategies, which combine schedules of survival, development, and reproduction, shape how natural selection acts on species' heritable traits and organismal fitness. Comparative analyses have historically ranked life histories along a fast-slow continuum, describing a negative association between time allocation to reproduction and development versus survival. However, higher-quality, more representative data and analyses have revealed that life history variation cannot be fully accounted for by this single continuum. Moreover, studies often do not test predictions from existing theories and instead operate as exploratory exercises. To move forward, we offer three recommendations for future investigations: standardizing life history traits, overcoming taxonomic siloes, and using theory to move from describing to understanding life history variation across the Tree of Life.

Integrated system for temperature-controlled fast protein liquid chromatography. IV. Continuous 'one-column' 'low-salt' hydrophobic interaction chromatography.

Systematic development of a temperature-controlled isocratic process for one-column low-salt hydrophobic interaction chromatography (HIC) of proteins employing a travelling cooling zone reactor (TCZR) system, is described. Batch binding and confocal scanning microscopy were employed to define process conditions for temperature-reversible binding of bovine serum albumin (BSA) which were validated in pulse-response temperature switching HIC experiments, before transferring to TCZR-HIC. A thin-walled stainless-steel column mounted with a movable assembly of copper blocks and Peltier elements (travelling cooling zone, TCZ) was used for TCZR-HIC. In pulse-response TCZR-HIC, 12 TCZ movements along the column desorbed 86.3% of the applied BSA monomers in 95.3% purity depleted >6-fold in 2-4 mers and nearly 260-fold in higher molecular weight (HMW) species. For continuous TCZR-HIC, the TCZ was moved 49-58 times during uninterrupted loading of BSA feeds at 0.25, 0.5 or 1 mg·mL-1. Each TCZ movement generated a sharp symmetrical elution peak. In the best case, (condition 1: 0.25 mg·mL-1 BSA; >17 mg BSA applied per mL of bed) the height of TCZ elution peaks approached pseudo-steady midway through the loading phase with no rise in baseline UV280 signal between peaks. Peak composition remained constant averaging 94.4% monomer, 5.6% 2-4 mers and <0.05% HMW. Monomers were recovered in quantitative yield depleted >3.1 fold in 2-4 mers and 92-fold in HMW species cf. the feed (63.6% monomers, 21.8% 2-4 mers, 14.6% HMW). However, increasing the BSA concentration to 1 mg·mL-1 (condition 2) or employing a fouled HIC column with 0.5 mg·mL-1 BSA (condition 3) compromised monomer purification performance.

Structural basis for Mis18 complex assembly and its implications for centromere maintenance.

The centromere, defined by the enrichment of CENP-A (a Histone H3 variant) containing nucleosomes, is a specialised chromosomal locus that acts as a microtubule attachment site. To preserve centromere identity, CENP-A levels must be maintained through active CENP-A loading during the cell cycle. A central player mediating this process is the Mis18 complex (Mis18α, Mis18ß and Mis18BP1), which recruits the CENP-A-specific chaperone HJURP to centromeres for CENP-A deposition. Here, using a multi-pronged approach, we characterise the structure of the Mis18 complex and show that multiple hetero- and homo-oligomeric interfaces facilitate the hetero-octameric Mis18 complex assembly composed of 4 Mis18α, 2 Mis18ß and 2 Mis18BP1. Evaluation of structure-guided/separation-of-function mutants reveals structural determinants essential for cell cycle controlled Mis18 complex assembly and centromere maintenance. Our results provide new mechanistic insights on centromere maintenance, highlighting that while Mis18α can associate with centromeres and deposit CENP-A independently of Mis18ß, the latter is indispensable for the optimal level of CENP-A loading required for preserving the centromere identity.

High resolution assessment of commercial fisheries activity along the US West Coast using Vessel Monitoring System data with a case study using California groundfish fisheries.

Commercial fisheries along the US West Coast are important components of local and regional economies. They use various fishing gear, target a high diversity of species, and are highly spatially heterogeneous, making it challenging to generate a synoptic picture of fisheries activity in the region. Still, understanding the spatial and temporal dynamics of US West Coast fisheries is critical to meet the US legal mandate to manage fisheries sustainably and to better coordinate activities among a growing number of users of ocean space, including offshore renewable energy, aquaculture, shipping, and interactions with habitats and key non-fishery species such as seabirds and marine mammals. We analyzed vessel tracking data from Vessel Monitoring System (VMS) from 2010 to 2017 to generate high-resolution spatio-temporal estimates of contemporary fishing effort across a wide range of commercial fisheries along the entire US West Coast. We identified over 247,000 fishing trips across the entire VMS data, covering over 25 different fisheries. We validated the spatial accuracy of our analyses using independent estimates of spatial groundfish fisheries effort generated through the NOAA's National Marine Fisheries Service Observer Program. Additionally, for commercial groundfish fisheries operating in federal waters in California, we combined the VMS data with landings and ex-vessel value data from California commercial fisheries landings receipts to generate highly resolved estimates of landings and ex-vessel value, matching over 38,000 fish tickets with VMS data that included 87% of the landings and 76% of the ex-vessel value for groundfish. We highlight fisheries-specific and spatially-resolved patterns of effort, landings, and ex-vessel value, a bimodal distribution of fishing effort with respect to depth, and variable and generally declining effort over eight years. The information generated by our study can help inform future sustainable spatial fisheries management and other activities in the marine environment including offshore renewable energy planning.

Chemically induced cone degeneration in the 13-lined ground squirrel.

Animal models of retinal degeneration are critical for understanding disease and testing potential therapies. Inducing degeneration commonly involves the administration of chemicals that kill photoreceptors by disrupting metabolic pathways, signaling pathways, or protein synthesis. While chemically induced degeneration has been demonstrated in a variety of animals (mice, rats, rabbits, felines, 13-lined ground squirrels (13-LGS), pigs, chicks), few studies have used noninvasive high-resolution retinal imaging to monitor the in vivo cellular effects. Here, we used longitudinal scanning light ophthalmoscopy (SLO), optical coherence tomography, and adaptive optics SLO imaging in the euthermic, cone-dominant 13-LGS (46 animals, 52 eyes) to examine retinal structure following intravitreal injections of chemicals, which were previously shown to induce photoreceptor degeneration, throughout the active season of 2019 and 2020. We found that iodoacetic acid induced severe pan-retinal damage in all but one eye, which received the lowest concentration. While sodium nitroprusside successfully induced degeneration of the outer retinal layers, the results were variable, and damage was also observed in 50% of contralateral control eyes. Adenosine triphosphate and tunicamycin induced outer retinal specific damage with varying results, while eyes injected with thapsigargin did not show signs of degeneration. Given the variability of damage we observed, follow-up studies examining the possible physiological origins of this variability are critical. These additional studies should further advance the utility of chemically induced photoreceptor degeneration models in the cone-dominant 13-LGS.

Mortality of Patients With Sepsis Administered Piperacillin-Tazobactam vs Cefepime.

Importance: Experimental and observational studies have suggested that empirical treatment for bacterial sepsis with antianaerobic antibiotics (eg, piperacillin-tazobactam) is associated with adverse outcomes compared with anaerobe-sparing antibiotics (eg, cefepime). However, a recent pragmatic clinical trial of piperacillin-tazobactam and cefepime showed no difference in short-term outcomes at 14 days. Further studies are needed to help clarify the empirical use of these agents. Objective: To examine the use of piperacillin-tazobactam compared with cefepime in 90-day mortality in patients treated empirically for sepsis, using instrumental variable analysis of a 15-month piperacillin-tazobactam shortage. Design, Setting, and Participants: In a retrospective cohort study, hospital admissions at the University of Michigan from July 1, 2014, to December 31, 2018, including a piperacillin-tazobactam shortage period from June 12, 2015, to September 18, 2016, were examined. Adult patients with suspected sepsis treated with vancomycin and either piperacillin-tazobactam or cefepime for conditions with presumed equipoise between piperacillin-tazobactam and cefepime were included in the study. Data analysis was conducted from December 17, 2022, to April 11, 2023. Main Outcomes and Measures: The primary outcome was 90-day mortality. Secondary outcomes included organ failure-free, ventilator-free, and vasopressor-free days. The 15-month piperacillin-tazobactam shortage period was used as an instrumental variable for unmeasured confounding in antibiotic selection. Results: Among 7569 patients (4174 men [55%]; median age, 63 [IQR 52-73] years) with sepsis meeting study eligibility, 4523 were treated with vancomycin and piperacillin-tazobactam and 3046 were treated with vancomycin and cefepime. Of patients who received piperacillin-tazobactam, only 152 (3%) received it during the shortage. Treatment groups did not differ significantly in age, Charlson Comorbidity Index score, Sequential Organ Failure Assessment score, or time to antibiotic administration. In an instrumental variable analysis, piperacillin-tazobactam was associated with an absolute mortality increase of 5.0% at 90 days (95% CI, 1.9%-8.1%) and 2.1 (95% CI, 1.4-2.7) fewer organ failure-free days, 1.1 (95% CI, 0.57-1.62) fewer ventilator-free days, and 1.5 (95% CI, 1.01-2.01) fewer vasopressor-free days. Conclusions and Relevance: Among patients with suspected sepsis and no clear indication for antianaerobic coverage, administration of piperacillin-tazobactam was associated with higher mortality and increased duration of organ dysfunction compared with cefepime. These findings suggest that the widespread use of empirical antianaerobic antibiotics in sepsis may be harmful.

A comparison of strategies for identifying patients at risk for carbapenem-resistant or extended ß-lactam-resistant Pseudomonas aeruginosa.

OBJECTIVES: To assess risk factors for carbapenem-resistant Pseudomonas aeruginosa (CR) and extended-ß-lactam-resistant P. aeruginosa (EBR) infection/colonization, and to develop and compare tools for predicting isolation of CR and EBR from clinical cultures. METHODS: This retrospective study analysed hospitalized patients with positive P. aeruginosa cultures between 2015 and 2021. Two case-control analyses were performed to identify risk factors and develop scoring tools for distinguishing patients with CR versus carbapenem-susceptible (CS) P. aeruginosa and EBR versus CS P. aeruginosa. The performance of institutionally derived scores, externally derived scores and the presence/absence of key risk factors to predict CR and EBR were then compared. RESULTS: A total of 2379 patients were included. Of these, 8.3% had a positive culture for CR, 5.0% for EBR and 86.7% for CS P. aeruginosa. There was substantial overlap in risk factors for CR and EBR. Institutional risk scores demonstrated modestly higher area under the ROC curve values than external scores for predicting CR (0.67 versus 0.58) and EBR (0.76 versus 0.70). Assessing the presence/absence of ≥1 of the two strongest predictors (prior carbapenem use or CR isolation within 90 days) was slightly inferior to scoring tools for predicting CR, and comparable for predicting EBR. CONCLUSIONS: Clinicians concerned about CR in P. aeruginosa should consider the likelihood of EBR when making treatment decisions. A simple approach of assessing recent history of CR isolation or carbapenem usage performed similarly to more complex scoring tools and offers a more pragmatic way of identifying patients who require coverage for resistant P. aeruginosa.

Outpatient COVID-19 convalescent plasma recipient antibody thresholds correlated to reduced hospitalizations within a randomized trial.

BACKGROUNDCOVID-19 convalescent plasma (CCP) virus-specific antibody levels that translate into recipient posttransfusion antibody levels sufficient to prevent disease progression are not defined.METHODSThis secondary analysis correlated donor and recipient antibody levels to hospitalization risk among unvaccinated, seronegative CCP recipients within the outpatient, double-blind, randomized clinical trial that compared CCP to control plasma. The majority of COVID-19 CCP arm hospitalizations (15/17, 88%) occurred in this unvaccinated, seronegative subgroup. A functional cutoff to delineate recipient high versus low posttransfusion antibody levels was established by 2 methods: (i) analyzing virus neutralization-equivalent anti-Spike receptor-binding domain immunoglobulin G (anti-S-RBD IgG) responses in donors or (ii) receiver operating characteristic (ROC) curve analysis.RESULTSSARS-CoV-2 anti-S-RBD IgG antibody was volume diluted 21.3-fold into posttransfusion seronegative recipients from matched donor units. Virus-specific antibody delivered was approximately 1.2 mg. The high-antibody recipients transfused early (symptom onset within 5 days) had no hospitalizations. A CCP-recipient analysis for antibody thresholds correlated to reduced hospitalizations found a statistical significant association between early transfusion and high antibodies versus all other CCP recipients (or control plasma), with antibody cutoffs established by both methods-donor-based virus neutralization cutoffs in posttransfusion recipients (0/85 [0%] versus 15/276 [5.6%]; P = 0.03) or ROC-based cutoff (0/94 [0%] versus 15/267 [5.4%]; P = 0.01).CONCLUSIONIn unvaccinated, seronegative CCP recipients, early transfusion of plasma units in the upper 30% of study donors' antibody levels reduced outpatient hospitalizations. High antibody level plasma units, given early, should be reserved for therapeutic use.TRIAL REGISTRATIONClinicalTrials.gov NCT04373460.FUNDINGDepartment of Defense (W911QY2090012); Defense Health Agency; Bloomberg Philanthropies; the State of Maryland; NIH (3R01AI152078-01S1, U24TR001609-S3, 1K23HL151826NIH); the Mental Wellness Foundation; the Moriah Fund; Octapharma; the Healthnetwork Foundation; the Shear Family Foundation; the NorthShore Research Institute; and the Rice Foundation.

Physical seed dormancy in pea is genetically separable from seed coat thickness and roughness.

Introduction: The seeds of wild pea (Pisum) exhibit marked physical dormancy due to impermeability of the seed coat to water, and the loss of this dormancy is thought to have been critical for domestication. Wild pea seed coats are also notably thick and rough, traits that have also reduced during domestication and are anecdotally linked to increased permeability. However, how these traits specifically interact with permeability is unclear. Methods: To investigate this, we examined the genetic control of differences in seed coat characteristics between wild P. sativum ssp. humile and a non-dormant domesticated P. s. sativum accession in a recombinant inbred population. QTL effects were confirmed and their locations refined in segregating F4/5 populations. Results: In this population we found a moderate correlation between testa thickness and permeability, and identified loci that affect them independently, suggesting no close functional association. However, the major loci affecting both testa thickness and permeability collocated closely with Mendel's pigmentation locus A, suggesting flavonoid compounds under its control might contribute significantly to both traits. We also show that seed coat roughness is oligogenic in this population, with the major locus independent of both testa thickness and permeability, suggesting selection for smooth seed was unlikely to be due to effects on either of these traits. Discussion: Results indicate loss of seed coat dormancy during domestication was not primarily driven by reduced testa thickness or smooth seededness. The close association between major permeability and thickness QTL and Mendel's 'A' warrant further study, particularly regarding the role of flavonoids.

Temporary increase in circulating replication-competent latent HIV-infected resting CD4+ T cells after switch to an integrase inhibitor based antiretroviral regimen.

BACKGROUND: The principal barrier to an HIV cure is the presence of the latent viral reservoir (LVR), which has been understudied in African populations. From 2018 to 2019, Uganda instituted a nationwide rollout of ART consisting of Dolutegravir (DTG) with two NRTI, which replaced the previous regimen of one NNRTI and the same two NRTI. METHODS: Changes in the inducible replication-competent LVR (RC-LVR) of ART-suppressed Ugandans with HIV (n = 88) from 2015 to 2020 were examined using the quantitative viral outgrowth assay. Outgrowth viruses were examined for viral evolution. Changes in the RC-LVR were analyzed using three versions of a Bayesian model that estimated the decay rate over time as a single, linear rate (model A), or allowing for a change at time of DTG initiation (model B&C). FINDINGS: Model A estimated the slope of RC-LVR change as a non-significant positive increase, which was due to a temporary spike in the RC-LVR that occurred 0-12 months post-DTG initiation (p < 0.005). This was confirmed with models B and C; for instance, model B estimated a significant decay pre-DTG initiation with a half-life of 6.9 years, and an ∼1.7-fold increase in the size of the RC-LVR post-DTG initiation. There was no evidence of viral failure or consistent evolution in the cohort. INTERPRETATION: These data suggest that the change from NNRTI- to DTG-based ART is associated with a significant temporary increase in the circulating RC-LVR. FUNDING: Supported by the NIH (grant 1-UM1AI164565); Gilead HIV Cure Grants Program (90072171); Canadian Institutes of Health Research (PJT-155990); and Ontario Genomics-Canadian Statistical Sciences Institute.

Obesogenic diet in pregnancy disrupts placental iron handling and ferroptosis and stress signalling in association with fetal growth alterations.

Obesity and gestational diabetes (GDM) impact fetal growth during pregnancy. Iron is an essential micronutrient needed for energy-intense feto-placental development, but if mis-handled can lead to oxidative stress and ferroptosis (iron-dependent cell death). In a mouse model showing maternal obesity and glucose intolerance, we investigated the association of materno-fetal iron handling and placental ferroptosis, oxidative damage and stress signalling activation with fetal growth. Female mice were fed a standard chow or high fat, high sugar (HFHS) diet during pregnancy and outcomes were measured at day (d)16 or d19 of pregnancy. In HFHS-fed mice, maternal hepcidin was reduced and iron status maintained (tissue iron levels) at both d16 and d19. However, fetal weight, placental iron transfer capacity, iron deposition, TFR1 expression and ERK2-mediated signalling were reduced and oxidative damage-related lipofuscin accumulation in the placenta was increased in HFHS-fed mice. At d19, whilst TFR1 remained decreased, fetal weight was normal and placental weight, iron content and iron transporter genes (Dmt1, Zip14, and Fpn1) were reduced in HFHS-fed mice. Furthermore, there was stress kinase activation (increased phosphorylated p38MAPK, total ERK and JNK) in the placenta from HFHS-fed mice at d19. In summary, a maternal HFHS diet during pregnancy impacts fetal growth trajectory in association with changes in placental iron handling, ferroptosis and stress signalling. Downregulation of placental iron transporters in HFHS mice may protect the fetus from excessive oxidative iron. These findings suggest a role for alterations in placental iron homeostasis in determining perinatal outcomes of pregnancies associated with GDM and/or maternal obesity.

Laboratory Automated Interrogation of Data: an interactive web application for visualization of multilevel data from biological experiments.

A key step in understanding the results of biological experiments is visualization of the data. Many laboratory experiments contain a range of measurements that exist within a hierarchy of interdependence. An automated and facile way to visualize and interrogate such multilevel data, across many experimental variables, would (i) lead to improved understanding of the results, (ii) help to avoid misleading interpretation of statistics and (iii) easily identify outliers and sources of batch and confounding effects. While many excellent graphing solutions already exist, they are often geared towards the production of publication-ready plots and the analysis of a single variable at a time, require programming expertise or are unnecessarily complex for the task at hand. Here, we present Laboratory Automated Interrogation of Data (LAB-AID), an interactive tool specifically designed to automatically visualize and query hierarchical data resulting from biological experiments.

Progression free survival of myeloma patients who become IFE-negative correlates with the detection of residual monoclonal free light chain (FLC) by mass spectrometry.

Deeper responses are associated with improved survival in patients being treated for myeloma. However, the sensitivity of the current blood-based assays is limited. Historical studies suggested that normalisation of the serum free light chain (FLC) ratio in patients who were negative by immunofixation electrophoresis (IFE) was associated with improved outcomes. However, recently this has been called into question. Mass spectrometry (MS)-based FLC assessments may offer a superior methodology for the detection of monoclonal FLC due to greater sensitivity. To test this hypothesis, all available samples from patients who were IFE negative after treatment with carfilzomib and lenalidomide-based induction and autologous stem cell transplantation (ASCT) in the Myeloma XI trial underwent FLC-MS testing. FLC-MS response assessments from post-induction, day+100 post-ASCT and six months post-maintenance randomisation were compared to serum FLC assay results. Almost 40% of patients had discordant results and 28.7% of patients with a normal FLC ratio had residual monoclonal FLC detectable by FLC-MS. FLC-MS positivity was associated with reduced progression-free survival (PFS) but an abnormal FLC ratio was not. This study demonstrates that FLC-MS provides a superior methodology for the detection of residual monoclonal FLC with FLC-MS positivity identifying IFE-negative patients who are at higher risk of early progression.