Phosphoproteomic analysis of the adaption of epididymal epithelial cells to corticosterone challenge.

BACKGROUND: The epididymis has long been of interest owing to its role in promoting the functional maturation of the male germline. More recent evidence has also implicated the epididymis as an important sensory tissue responsible for remodeling of the sperm epigenome, both under physiological conditions and in response to diverse forms of environmental stress. Despite this knowledge, the intricacies of the molecular pathways involved in regulating the adaptation of epididymal tissue to paternal stressors remains to be fully resolved. OBJECTIVE: The overall objective of this study was to investigate the direct impact of corticosterone challenge on a tractable epididymal epithelial cell line (i.e., mECap18 cells), in terms of driving adaptation of the cellular proteome and phosphoproteome signaling networks. MATERIALS AND METHODS: The newly developed phosphoproteomic platform EasyPhos coupled with sequencing via an Orbitrap Exploris 480 mass spectrometer, was applied to survey global changes in the mECap18 cell (phospho)proteome resulting from sub-chronic (10-day) corticosterone challenge. RESULTS: The imposed corticosterone exposure regimen elicited relatively subtle modifications of the global mECap18 proteome (i.e., only 73 out of 4171 [∼1.8%] proteins displayed altered abundance). By contrast, ∼15% of the mECap18 phosphoproteome was substantially altered following corticosterone challenge. In silico analysis of the corresponding parent proteins revealed an activation of pathways linked to DNA damage repair and oxidative stress responses as well as a reciprocal inhibition of pathways associated with organismal death. Corticosterone challenge also induced the phosphorylation of several proteins linked to the biogenesis of microRNAs. Accordingly, orthogonal validation strategies confirmed an increase in DNA damage, which was ameliorated upon selective kinase inhibition, and an altered abundance profile of a subset of microRNAs in corticosterone-treated cells. CONCLUSIONS: Together, these data confirm that epididymal epithelial cells are reactive to corticosterone challenge, and that their response is tightly coupled to the opposing action of cellular kinases and phosphatases.

Validation of three-dimensional thoracic electrical impedance tomography of horses during normal and increased tidal volumes.

Objective. Data from two-plane electrical impedance tomography (EIT) can be reconstructed into various slices of functional lung images, allowing for more complete visualisation and assessment of lung physiology in health and disease. The aim of this study was to confirm the ability of 3D EIT to visualise normal lung anatomy and physiology at rest and during increased ventilation (represented by rebreathing).Approach. Two-plane EIT data, using two electrode planes 20 cm apart, were collected in 20 standing sedate horses at baseline (resting) conditions, and during rebreathing. EIT data were reconstructed into 3D EIT whereby tidal impedance variation (TIV), ventilated area, and right-left and ventral-dorsal centres of ventilation (CoVRLand CoVVD, respectively) were calculated in cranial, middle and caudal slices of lung, from data collected using the two planes of electrodes.Main results. There was a significant interaction of time and slice for TIV (p< 0.0001) with TIV increasing during rebreathing in both caudal and middle slices. The ratio of right to left ventilated area was higher in the cranial slice, in comparison to the caudal slice (p= 0.0002). There were significant effects of time and slice on CoVVDwhereby the cranial slice was more ventrally distributed than the caudal slice (p< 0.0009 for the interaction).Significance. The distribution of ventilation in the three slices corresponds with topographical anatomy of the equine lung. This study confirms that 3D EIT can accurately represent lung anatomy and changes in ventilation distribution during rebreathing in standing sedate horses.

PI3K/mTOR is a therapeutically targetable genetic dependency in diffuse intrinsic pontine glioma.

Diffuse midline glioma (DMG), including tumors diagnosed in the brainstem (diffuse intrinsic pontine glioma; DIPG), are uniformly fatal brain tumors that lack effective treatment. Analysis of CRISPR/Cas9 loss-of-function gene deletion screens identified PIK3CA and MTOR as targetable molecular dependencies across patient derived models of DIPG, highlighting the therapeutic potential of the blood-brain barrier-penetrant PI3K/Akt/mTOR inhibitor, paxalisib. At the human-equivalent maximum tolerated dose, mice treated with paxalisib experienced systemic glucose feedback and increased insulin levels commensurate with patients using PI3K inhibitors. To exploit genetic dependence and overcome resistance while maintaining compliance and therapeutic benefit, we combined paxalisib with the antihyperglycemic drug metformin. Metformin restored glucose homeostasis and decreased phosphorylation of the insulin receptor in vivo, a common mechanism of PI3K-inhibitor resistance, extending survival of orthotopic models. DIPG models treated with paxalisib increased calcium-activated PKC signaling. The brain penetrant PKC inhibitor enzastaurin, in combination with paxalisib, synergistically extended the survival of multiple orthotopic patient-derived and immunocompetent syngeneic allograft models; benefits potentiated in combination with metformin and standard-of-care radiotherapy. Therapeutic adaptation was assessed using spatial transcriptomics and ATAC-Seq, identifying changes in myelination and tumor immune microenvironment crosstalk. Collectively, this study has identified what we believe to be a clinically relevant DIPG therapeutic combinational strategy.

Ppe.RPT/SSC-1: from QTL mapping to a predictive KASP test for ripening time and soluble solids concentration in peach.

Genomic regions associated with ripening time (RPT) and soluble solids concentration (SSC) were mapped using a pedigreed population including multiple F1 and F2 families from the Clemson University peach breeding program (CUPBP). RPT and SSC QTLs were consistently identified in two seasons (2011 and 2012) and the average datasets (average of two seasons). A target region spanning 10,981,971-11,298,736 bp on chromosome 4 of peach reference genome used for haplotype analysis revealed four haplotypes with significant differences in trait values among different diplotype combinations. Favorable alleles at the target region for both RPT and SSC were determined and a DNA test for predicting RPT and SSC was developed. Two Kompetitive Allele Specific PCR (KASP) assays were validated on 84 peach cultivars and 163 seedlings from the CUPBP, with only one assay (Ppe.RPT/SSC-1) needed to predict between early and late-season ripening cultivars and low and high SSC. These results advance our understanding of the genetic basis of RPT and SSC and facilitate selection of new peach cultivars with the desired RPT and SSC.

Molecular and Pathologic Characterization of YAP1-Expressing Small Cell Lung Cancer Cell Lines Leads to Reclassification as SMARCA4-Deficient Malignancies.

PURPOSE: The classification of small cell lung cancer (SCLC) into distinct molecular subtypes defined by ASCL1, NEUROD1, POU2F3, or YAP1 (SCLC-A, -N, -P, or -Y) expression, paves the way for a personalized treatment approach. However, the existence of a distinct YAP1-expressing SCLC subtype remains controversial. EXPERIMENTAL DESIGN: To better understand YAP1-expressing SCLC, the mutational landscape of human SCLC cell lines was interrogated to identify pathogenic alterations unique to SCLC-Y. Xenograft tumors, generated from cell lines representing the four SCLC molecular subtypes, were evaluated by a panel of pathologists who routinely diagnose thoracic malignancies. Diagnoses were complemented by transcriptomic analysis of primary tumors and human cell line datasets. Protein expression profiles were validated in patient tumor tissue. RESULTS: Unexpectedly, pathogenic mutations in SMARCA4 were identified in six of eight SCLC-Y cell lines and correlated with reduced SMARCA4 mRNA and protein expression. Pathologist evaluations revealed that SMARCA4-deficient SCLC-Y tumors exhibited features consistent with thoracic SMARCA4-deficient undifferentiated tumors (SMARCA4-UT). Similarly, the transcriptional profile SMARCA4-mutant SCLC-Y lines more closely resembled primary SMARCA4-UT, or SMARCA4-deficient non-small cell carcinoma, than SCLC. Furthermore, SMARCA4-UT patient samples were associated with a YAP1 transcriptional signature and exhibited strong YAP1 protein expression. Together, we found little evidence to support a diagnosis of SCLC for any of the YAP1-expressing cell lines originally used to define the SCLC-Y subtype. CONCLUSIONS: SMARCA4-mutant SCLC-Y cell lines exhibit characteristics consistent with SMARCA4-deficient malignancies rather than SCLC. Our findings suggest that, unlike ASCL1, NEUROD1, and POU2F3, YAP1 is not a subtype defining transcription factor in SCLC.

Oocyte and cumulus cell cooperativity and metabolic plasticity under the direction of oocyte paracrine factors.

Mammalian oocytes develop and mature in a mutually dependent relationship with surrounding cumulus cells. The oocyte actively regulates cumulus cell differentiation and function by secreting soluble paracrine oocyte-secreted factors (OSFs). We characterized the molecular mechanisms by which two model OSFs, cumulin and BMP15, regulate oocyte maturation and cumulus-oocyte cooperativity. Exposure to these OSFs during mouse oocyte maturation in vitro altered the proteomic and multispectral autofluorescence profiles of both the oocyte and cumulus cells. In oocytes, cumulin significantly upregulated proteins involved in nuclear function. In cumulus cells, both OSFs elicited marked upregulation of a variety of metabolic processes (mostly anabolic), including lipid, nucleotide, and carbohydrate metabolism, whereas mitochondrial metabolic processes were downregulated. The mitochondrial changes were validated by functional assays confirming altered mitochondrial morphology, respiration, and content while maintaining ATP homeostasis. Collectively, these data demonstrate that cumulin and BMP15 remodel cumulus cell metabolism, instructing them to upregulate their anabolic metabolic processes, while routine cellular functions are minimized in the oocyte during maturation, in preparation for ensuing embryonic development.NEW & NOTEWORTHY Oocyte-secreted factors (OSFs) promote oocyte and cumulus cell cooperativity by altering the molecular composition of both cell types. OSFs downregulate protein catabolic processes and upregulate processes associated with DNA binding, translation, and ribosome assembly in oocytes. In cumulus cells, OSFs alter mitochondrial number, morphology, and function, and enhance metabolic plasticity by upregulating anabolic pathways. Hence, the oocyte via OSFs, instructs cumulus cells to increase metabolic processes on its behalf, thereby subduing oocyte metabolism.

Disruption of diabetes and hypertension care during the COVID-19 pandemic and recovery approaches in the Latin America and Caribbean region: a scoping review protocol.

INTRODUCTION: The COVID-19 pandemic significantly disrupted primary healthcare globally, with particular impacts on diabetes and hypertension care. This review will examine the impact of pandemic disruptions of diabetes and hypertension care services and the evidence for interventions to mitigate or reverse pandemic disruptions in the Latin America and Caribbean (LAC) region. METHODS AND ANALYSES: This scoping review will examine care delivery disruption and approaches for recovery of primary healthcare in the LAC region during the COVID-19 pandemic, focusing on diabetes and hypertension awareness, detection, treatment and control. Guided by Arksey and O'Malley's scoping review methodology framework, this protocol adheres to the Joanna Briggs Institute guidelines for scoping review protocols and the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidance for protocol development and scoping reviews. We searched MEDLINE, CINAHL, Global Health, Embase, Cochrane, Scopus, Web of Science and LILACS for peer-reviewed literature published from 2020 to 12 December 2022 in English, Spanish or Portuguese. Studies will be considered eligible if reporting data on pandemic disruptions to primary care services within LAC, or interventions implemented to mitigate or reverse pandemic disruptions globally. Studies on COVID-19 or acute care will be excluded. Two reviewers will independently screen each title/abstract for eligibility, screen full texts of titles/abstracts deemed relevant and extract data from eligible full-text publications. Conflicts will be resolved through discussion and with the help of a third reviewer. Appropriate analytical techniques will be employed to synthesise the data, for example, frequency counts and descriptive statistics. Quality will be assessed using the Newcastle Ottawa Quality Assessment Scale. ETHICS AND DISSEMINATION: No ethics approval was needed as this is a scoping review of published literature. Results will be disseminated in a report to the World Bank and the Pan American Health Organization, in peer-reviewed scientific journals, and at national and international conferences.

Spatial transcriptomics and the anatomical pathologist: Molecular meets morphology.

In recent years anatomical pathology has been revolutionised by the incorporation of molecular findings into routine diagnostic practice, and in some diseases the presence of specific molecular alterations are now essential for diagnosis. Spatial transcriptomics describes a group of technologies that provide up to transcriptome-wide expression profiling while preserving the spatial origin of the data, with many of these technologies able to provide these data using a single tissue section. Spatial transcriptomics allows expression profiling of highly specific areas within a tissue section potentially to subcellular resolution, and allows correlation of expression data with morphology, tissue type and location relative to other structures. While largely still research laboratory-based, several spatial transcriptomics methods have now achieved compatibility with formalin-fixed paraffin-embedded tissue (FFPE), allowing their use in diagnostic tissue samples, and with further development potentially leading to their incorporation in routine anatomical pathology practice. This mini review provides an overview of spatial transcriptomics methods, with an emphasis on platforms compatible with FFPE tissue, approaches to assess the data and potential applications in anatomical pathology practice.

Building an Understanding of Proteostasis in Reproductive Cells: The Impact of Reactive Carbonyl Species on Protein Fate.

Significance: Stringent regulation of protein homeostasis pathways, under both physiological and pathological conditions, is necessary for the maintenance of proteome fidelity and optimal cell functioning. However, when challenged by endogenous or exogenous stressors, these proteostasis pathways can become dysregulated with detrimental consequences for protein fate, cell survival, and overall organism health. Most notably, there are numerous somatic pathologies associated with a loss of proteostatic regulation, including neurodegenerative disorders, type 2 diabetes, and some cancers. Recent Advances: Lipid oxidation-derived reactive carbonyl species (RCS), such as 4-hydroxynonenal (4HNE) and malondialdehyde, are relatively underappreciated purveyors of proteostatic dysregulation, which elicit their effects via the nonenzymatic post-translational modification of proteins. Emerging evidence suggests that a subset of germline proteins can serve as substrates for 4HNE modification. Among these, prevalent targets include succinate dehydrogenase, heat shock protein A2 and A-kinase anchor protein 4, all of which are intrinsically associated with fertility. Critical Issues: Despite growing knowledge in this field, the RCS adductomes of spermatozoa and oocytes are yet to be comprehensively investigated. Furthermore, the manner by which RCS-mediated adduction impacts protein fate and drives cellular responses, such as protein aggregation, requires further examination in the germline. Given that RCS-protein adduction has been attributed a role in infertility, there has been sparked research investment into strategies to prevent lipid peroxidation in germ cells. Future Directions: An increased depth of knowledge regarding the mechanisms and substrates of RCS-mediated protein modification in reproductive cells may reveal important targets for the development of novel therapies to improve fertility and pregnancy outcomes for future generations.

Naloxone use by Aotearoa New Zealand emergency medical services, 2017-2021.

OBJECTIVE: Emergency medical services (EMS) use of naloxone in the prehospital setting is indicated in patients who have significantly impaired breathing or level of consciousness when opioid intoxication is suspected. The present study characterised naloxone use in a nationwide sample of Aotearoa New Zealand road EMS patients to establish a baseline for surveillance of any changes in the future. METHODS: A retrospective analysis of rates of patients with naloxone administrations was conducted using Hato Hone St John (2017-2021) and Wellington Free Ambulance (2018-2021) electronic patient report form datasets. Patient demographics, presenting complaints, naloxone dosing, and initial and last vital sign clinical observations were described. RESULTS: There were 2018 patients with an equal proportion of males and females, and patient median age was 47 years. There were between 8.0 (in 2018) and 9.0 (in 2020) naloxone administrations per 100 000 population-years, or approximately one administration per day for the whole country of 5 million people. Poisoning by unknown agent(s) was the most common presenting complaint (61%). The median dose of naloxone per patient was 0.4 mg; 85% was administered intravenously. The median observed change in Glasgow Coma Scale score was +1, and respiratory rate increased by +2 breaths/min. CONCLUSIONS: A national rate of EMS naloxone patients was established; measured clinical effects of naloxone were modest, suggesting many patients had reasons other than opioid toxicity contributing to their symptoms. Naloxone administration rates provide indirect surveillance information about suspected harmful opioid exposures but need to be interpreted with care.

Survival rates and factors associated with survival and laminitis of horses with acute diarrhoea admitted to referral institutions.

BACKGROUND: Clinicopathological findings and their association with the outcome and development of laminitis in horses with acute diarrhoea has not been investigated in a multicentre study across different geographic regions. OBJECTIVES: Describe and compare clinicopathologic findings of diarrhoeic horses between different geographic regions, survival rates and factors associated with non-survival and laminitis. STUDY DESIGN: Multicentre retrospective case series. METHODS: Information from horses with acute diarrhoea presenting to participating institutions between 2016 and 2020 was collected, and clinicopathological data were compared between surviving and non-surviving horses and horses that did and did not develop laminitis. Survival rates and seasonal and geographic differences were also investigated. RESULTS: One thousand four hundred thirty-eight horses from 26 participating institutions from 4 continents were included; 76% survived to discharge with no differences identified between geographic regions. The survival proportion of horses with SIRS and creatinine concentrations > 159 µmol/L was 55% (154/279) compared with 81% (358/437) for those with SIRS and creatinine concentrations < 159 µmol/L (p < 0.001). The survival proportion of horses with SIRS that had an L-lactate concentration > 2.8 mmol/L was 59% (175/298) compared with 81% (240/296) in horses with SIRS and L-lactate concentration < 2.8 mmol/L (p < 0.001). The proportion of horses that developed laminitis was lower in Europe (4%, 19/479) compared with North America (8%, 52/619), Australia (8%, 12/138) and Latin America (11%, 16/146) (p < 0.05). More horses developed laminitis in the summer (46%, 39/85) compared with winter (18%, 15/85), spring (18%, 15/85) and fall (19%, 16/85) (p < 0.01). Horses with laminitis had greater odds of non-survival than those without laminitis (OR: 3.73, 95% CI: 2.47-5.65). MAIN LIMITATIONS: Not all variables were available for all horses due to the retrospective nature. CONCLUSIONS: Clinicopathological findings in horses with acute diarrhoea and their association with survival are similar across geographic regions. However, developing laminitis secondary to diarrhoea is less common in Europe. In addition, factors associated with non-survival were indicative of disease severity and subsequent cardiovascular compromise.

CONTEXTO: Achados clínico-patológicos e suas associações com o sobrevivência e desenvolvimento de laminite em cavalos com diarreia aguda não foram investigados em um estudo multicêntrico envolvendo diferentes regiões geográficas. OBJETIVOS: Descrever e comparar achados clínico-patológicos de cavalos com diarreia em diferentes regiões geográficas, taxa de sobrevivência e fatores associados com mortalidade e laminite. DELINEAMENTO DO ESTUDO: Estudo multicêntrico retrospectivo de série de casos. METODOLOGIA: Informação sobre equinos com diarreia aguda apresentados às instituições participantes entre 2016 e 2020 foram coletados, e dados clínico-patológicos foram comparados entre sobreviventes e não-sobreviventes, e cavalos que desenvolveram ou não laminite. Taxa de sobrevivência, e diferenças sazonais e geográficas também foram investigadas. RESULTADOS: 1438 cavalos de 26 instituições participantes de 4 continentes foram incluídos; 76% sobreviveram e receberam alta e nenhuma diferença foi observada entre as diferentes regiões geográficas. A proporção de cavalos que sobreviveram com SIRS e concentração de creatinina > 1.8 mg/dL foi 55% (154/279) comparado com 81% (358/437) dos cavalos com SIRS e concentração de creatinina < 1.8 mg/dL (p < 0.001). A proporção de cavalos com SIRS que tinham concentração de L-lactato > 2.8 mmol que sobreviveram foi 59% (175/298) comparado com 81% (240/296) dos cavalos com SIRS e concentração de L-lactato < 2.8 mmol/L (p < 0.001). A proporção de cavalos que desenvolveram laminite foi menor na Europa (4%, 19/479) comparado com a América do Norte (8%, 52/619), Austrália (8%, 12/138) e América Latina (11% 16/146) (p < 0.05). Mais cavalos desenvolveram laminite no verão (46%, 39/8) comparado com inverno (18%, 15/85), primavera (18%, 15/85) e outono (19%, 16/85) (p < 0.01). Cavalos com laminite tiveram chances maior de não sobreviver do que aqueles que não desenvolveram laminite (OR: 3.73, 95% CI: 2.47 a 5.65). PRINCIPAIS LIMITAÇÕES: Algumas variáveis não estavam disponíveis para alguns cavalos devido à natureza retrospectiva deste estudo. CONCLUSÕES: Achados clínico-patológicos em equinos com diarreia aguda e sua associação com sobrevivência é similar entre as diferentes regiões geográficas. Contudo, o desenvolvimento de laminite secundário à diarreia é menos comum na Europa. Além disso, fatores associados com não-sobrevivência foram indicativos de severidade da doença e subsequente comprometimento cardiovascular.

Diagnostic approaches, aetiological agents and their associations with short-term survival and laminitis in horses with acute diarrhoea admitted to referral institutions.

BACKGROUND: An international description of the diagnostic approaches used in different institutions to diagnose acute equine diarrhoea and the pathogens detected is lacking. OBJECTIVES: To describe the diagnostic approach, aetiological agents, outcome, and development of laminitis for diarrhoeic horses worldwide. STUDY DESIGN: Multicentre retrospective case series. METHODS: Information from horses with acute diarrhoea presenting to participating institutions between 2016 and 2020, including diagnostic approaches, pathogens detected and their associations with outcomes, were compared between institutions or geographic regions. RESULTS: One thousand four hundred and thirty-eight horses from 26 participating institutions from 4 continents were included. Overall, aetiological testing was limited (44% for Salmonella spp., 42% for Neorickettsia risticii [only North America], 40% for Clostridiodes difficile, and 29% for ECoV); however, 13% (81/633) of horses tested positive for Salmonella, 13% (35/262) for N. risticii, 9% (37/422) for ECoV, and 5% (27/578) for C. difficile. C. difficile positive cases had greater odds of non-survival than horses negative for C. difficile (OR: 2.69, 95%CI: 1.23-5.91). In addition, horses that were positive for N. risticii had greater odds of developing laminitis than negative horses (OR: 2.76, 95%CI: 1.12-6.81; p = 0.029). MAIN LIMITATIONS: Due to the study's retrospective nature, there are missing data. CONCLUSIONS: This study highlighted limited diagnostic investigations in cases of acute equine diarrhoea. Detection rates of pathogens are similar to previous reports. Non-survival and development of laminitis are related to certain detected pathogens.

Persistence of UBTF tandem duplications in remission in acute myeloid leukaemia.

UBTF tandem duplications are recurrent in adult and paediatric acute myeloid leukaemia and have been reported to be associated with a poor prognosis. Co-mutations in WT1 and FLT3 are common while morphological dysplasia is frequent. The role of UBTF-TDs in leukemogenesis is yet to be elucidated; however they have been proposed as early initiating events, making them attractive for assessment of MRD and a potential therapeutic target. We present two cases where the UBTF-TD was observed in remission and discuss the implications of these findings in the clinicobiological understanding of this emerging entity.

Right dorsal colitis in horses: A multicenter retrospective study of 35 cases.

BACKGROUND: Right dorsal colitis (RDC) is a nonsteroidal anti-inflammatory drug (NSAID) induced, protein losing enteropathy in horses associated with a high case fatality rate. OBJECTIVES: To describe signalment, NSAID usage, clinical presentations, clinical pathology, ultrasonographic findings, treatments, outcomes, and factors associated with survival in horses diagnosed with RDC. ANIMALS: Thirty-five horses from 7 Australian equine hospitals diagnosed with RDC. METHODS: Retrospective case series. Clinical records of cases were accepted if definitively or presumptively diagnosed by an internist with RDC and had ≥3 of: hypoproteinemia or hypoalbuminemia; diarrhea with negative test results for infectious diseases; colic for which other diseases were excluded or right dorsal colon thickening on ultrasound. Descriptive data analysis was performed for categorical and continuous variables. Univariate binominal logistic regressions were used to assess factors associated with survival. RESULTS: An overdose of NSAIDs occurred in 84% (21/25) cases where dose was known. Common clinical presentations included diarrhea (69%; 22/32), colic (61%; 20/33), and tachycardia (53%, 17/32). Common clinicopathological findings included hypoalbuminemia (83%; 26/31), hypocalcaemia (79%, 23/29), and hyperlactatemia (77%, 14/18). The right dorsal colon wall appeared subjectively thickened in 77% (24/31) cases using ultrasonography. Case fatality rate was 43% (15/35). Odds of survival significantly decreased with increasing heart rate (odds 0.84, 95% CI = 0.71-0.92, P = .01), packed cell volume (odds 0.91, 95% CI 0.82-0.98, P = .05) and abnormal appearance of mucous membranes (odds 0.05, 95% CI 0.005-0.28, P = .001) on hospital presentation. CONCLUSIONS AND CLINICAL IMPORTANCE: An overdose of NSAIDs is common in horses diagnosed with RDC. Serum albumin concentrations should be monitored in horses receiving a prolonged course of NSAIDs. Overall prognosis for RDC remains fair.

Proteomic and phosphoproteomic characterisation of primary mouse embryonic fibroblasts.

Fibroblasts are the most common cell type in stroma and function in the support and repair of most tissues. Mouse embryonic fibroblasts (MEFs) are amenable to isolation and rapid growth in culture. MEFs are therefore widely used as a standard model for functional characterisation of gene knockouts, and can also be used in co-cultures, commonly to support embryonic stem cell cultures. To facilitate their use as a research tool, we have performed a comprehensive proteomic and phosphoproteomic characterisation of wild-type primary MEFs from C57BL/6 mice. EIF2/4 and MTOR signalling pathways were abundant in both the proteome and phosphoproteome, along with extracellular matrix (ECM) and cytoskeleton associated pathways. Consistent with this, kinase enrichment analysis identified activation of P38A, P90RSK, P70S6K, and MTOR. Cell surface markers and matrisome proteins were also annotated. Data are available via ProteomeXchange with identifier PXD043244. This provides a comprehensive catalogue of the wild-type MEF proteome and phosphoproteome which can be utilised by the field to guide future work.

Immortalized mouse caput epididymal epithelial (mECap18) cell line recapitulates the in-vivo environment.

Residing between the testes and the vas deferens, the epididymis is a highly convoluted tubule whose unique luminal microenvironment is crucial for the functional maturation of spermatozoa. This microenvironment is created by the combined secretory and resorptive activity of the lining epididymal epithelium, including the release of extracellular vesicles (epididymosomes), which encapsulate fertility modulating proteins and a myriad of small non-coding RNAs (sncRNAs) that are destined for delivery to recipient sperm cells. To enable investigation of this intercellular communication nexus, we have previously developed an immortalized mouse caput epididymal epithelial cell line (mECap18). Here, we describe the application of label-free mass spectrometry to characterize the mECap18 cell proteome and compare this to the proteome of native mouse caput epididymal epithelial cells. We report the identification of 5,313 mECap18 proteins, as many as 75.8% of which were also identified in caput epithelial cells wherein they mapped to broadly similar protein classification groupings. Furthermore, key pathways associated with protein synthesis (e.g., EIF2 signaling) and cellular protection in the male reproductive tract (e.g., sirtuin signaling) were enriched in both proteomes. This comparison supports the utility of the mECap18 cell line as a tractable in-vitro model for studying caput epididymal epithelial cell function.

Observational and model evidence together support wide-spread exposure to noncompensable heat under continued global warming.

As our planet warms, a critical research question is when and where temperatures will exceed the limits of what the human body can tolerate. Past modeling efforts have investigated the 35°C wet-bulb threshold, proposed as a theoretical upper limit to survivability taking into account physiological and behavioral adaptation. Here, we conduct an extreme value theory analysis of weather station observations and climate model projections to investigate the emergence of an empirically supported heat compensability limit. We show that the hottest parts of the world already experience these heat extremes on a limited basis and that under moderate continued warming parts of every continent, except Antarctica, will see a rapid increase in their extent and frequency. To conclude, we discuss the consequences of the emergence of this noncompensable heat and the need for incorporating different critical thermal limits into heat adaptation planning.

Maternal breastfeeding is associated with offspring microbiome diversity; a secondary analysis of the MicrobeMom randomized control trial.

Background: Microbial dysbiosis in infancy can influence long-term health outcomes such as childhood obesity. The aim of this study is to explore relationships among maternal well-being during pregnancy, breastfeeding, and the infant gut microbiome. Methods: This is a secondary analysis of healthy pregnant women from the MicrobeMom study, a double-blind randomized control trial of maternal probiotic supplementation (Bifidobacterium breve 702258) versus placebo antenatally and up to 3 months postpartum. Maternal well-being was assessed using the WHO-5 well-being index at 16 weeks' and 34 weeks' gestation. Breastfeeding practices were recorded at discharge from hospital and at 1 month postpartum. Infant stool samples were obtained at 1 month of age. Next generation shotgun sequencing determined infant microbial diversity. Independent sample t-tests and Mann-Whitney U tests informed adjusted regression analysis, which was adjusted for delivery mode, antibiotics during delivery, maternal age and body mass index (BMI), and probiotic vs. control study group. Results: Women (n = 118) with at least one measure of well-being were on average 33 years (SD 3.93) of age and 25.09 kg/m2 (SD 3.28) BMI. Exclusive breastfeeding was initiated by 65% (n = 74). Any breastfeeding was continued by 69% (n = 81) after 1 month. In early and late pregnancy, 87% (n = 97/111) and 94% (n = 107/114) had high well-being scores. Well-being was not associated with infant microbial diversity at 1 month. In adjusted analysis, exclusive breastfeeding at discharge from hospital was associated with infant microbial beta diversity (PC2; 0.254, 95% CI 0.006, 0.038). At 1 month postpartum, any breastfeeding was associated with infant microbial alpha diversity (Shannon index; -0.241, 95% CI -0.498, -0.060) and observed species; (-0.325, 95% CI -0.307, -0.060), and infant microbial beta diversity (PC2; 0.319, 95% CI 0.013, 0.045). Exclusive breastfeeding at 1 month postpartum was associated with infant alpha diversity (Shannon index -0.364, 95% CI -0.573, -0.194; Simpson index 0.339, 95% CI 0.027, 0.091), and infant's number of observed microbial species (-0.271, 95% CI -0.172, -0.037). Conclusion: Breastfeeding practices at 1 month postpartum were associated with lower microbial diversity and observed species in infants at 1 month postpartum, which is potentially beneficial to allow greater abundance of Bifidobacterium. Clinical trial registration: ISRCTN53023014.

QTL discovery for resistance to black spot and cercospora leaf spot, and defoliation in two interconnected F1 bi-parental tetraploid garden rose populations.

Garden roses are an economically important horticultural crop worldwide, and two major fungal pathogens, black spot (Diplocarpon rosae F.A. Wolf) and cercospora leaf spot of rose (Rosisphaerella rosicola Pass.), affect both the health and ornamental value of the plant. Most studies on black spot disease resistance have focused on diploid germplasm, and little work has been performed on cercospora leaf spot resistance. With the use of newly developed software tools for autopolyploid genetics, two interconnected tetraploid garden rose F1 populations (phenotyped over the course of 3 years) were used for quantitative trait locus (QTL) analysis of black spot and cercospora leaf spot resistance as well as plant defoliation. QTLs for black spot resistance were mapped to linkage groups (LGs) 1-6. QTLs for cercospora resistance and susceptibility were found in LGs 1, 4, and 5 and for defoliation in LGs 1, 3, and 5. The major locus on LG 5 for black spot resistance coincides with the previously discovered Rdr4 locus inherited from Rosa L. 'Radbrite' (Brite Eyes™), the common parent used in these mapping populations. This work is the first report of any QTL for cercospora resistance/susceptibility in tetraploid rose germplasm and the first report of defoliation QTL in roses. A major QTL for cercospora susceptibility coincides with the black spot resistance QTL on LG 5 (Rdr4). A major cercospora resistance QTL was found on LG 1. These populations provide a genetic resource that will further the knowledge base of rose genetics as more traits are studied. Studying more traits from these populations will allow for the stacking of various QTLs for desirable traits.

Pedigree-based QTL analysis of flower size traits in two multi-parental diploid rose populations.

Rose (Rosa spp.) is one of the most economically important ornamental species worldwide. Flower diameter, flower weight, and the number of petals and petaloids are key flower-size parameters and attractive targets for DNA-informed breeding. Pedigree-based analysis (PBA) using FlexQTL software was conducted using two sets of multi-parental diploid rose populations. Phenotypic data for flower diameter (Diam), flower weight (fresh (FWT)/dry (DWT)), number of petals (NP), and number of petaloids (PD) were collected over six environments (seasons) at two locations in Texas. The objectives of this study were to 1) identify new and/or validate previously reported QTL(s); 2) identify SNP haplotypes associated with QTL alleles (Q-/q-) of a trait and their sources; and 3) determine QTL genotypes for important rose breeding parents. Several new and previously reported QTLs for NP and Diam traits were identified. In addition, QTLs associated with flower weight and PD were identified for the first time. Two major QTLs with large effects were mapped for all traits. The first QTL was at the distal end of LG1 (60.44-60.95 Mbp) and was associated with Diam and DWT in the TX2WOB populations. The second QTL was consistently mapped in the middle region on LG3 (30.15-39.34 Mbp) and associated with NP, PD, and flower weight across two multi-parent populations (TX2WOB and TX2WSE). Haplotype results revealed a series of QTL alleles with differing effects at important loci for most traits. This work is distinct from previous studies by conducting co-factor analysis to account for the DOUBLE FLOWER locus while mapping QTL for NP. Sources of high-value (Q) alleles were identified, namely, 'Old Blush' and Rosa wichuraiana from J14-3 for Diam, while 'Violette' and PP-J14-3 were sources for other traits. In addition, the source of the low-value (q) alleles for Diam was 'Little Chief', and Rosa wichuraiana through J14-3 was the source for the remaining traits. Hence, our results can potentially inform parental/seedling selections as means to improve ornamental quality in roses and a step towards implementing DNA-informed techniques for use in rose breeding programs.