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Citalopram, a selective serotonin reuptake inhibitor (SSRI), is often detected in aquatic ecosystems. In this investigation, developing zebrafish were continuously exposed to one nominal concentration of either 0, 10, or 1000 µg/L citalopram for 7 days. Ribonucleic acids were then extracted from zebrafish for RNA-sequencing using the NovoSeq 6000 (Illumina). Clean reads were obtained following the removal of both the adapter and poly-N sequences. Alignment and differential gene expression analysis was conducted using programs HISAT2 and StringTie assembler. Data were converted to FPKM to quantify differentially expressed transcripts. Significant clinical subnetworks enriched following citalopram exposure included sympathetic nerve activity, blood pressure, vascular tone, and arterial pressure. Regulated transcripts were related to diseases such as mechanical hyperalgesia, pain, inflammatory pain, obstructive hypertrophic cardiomyopathy, fatigue, Diamond-Blackfan anemia, and hypertrophic cardiomyopathy. Following exposure to 10 µg/L citalopram, several transcripts were linked to brain dysfunction like prostaglandin-endoperoxide synthase 2, microtubule associated protein tau, cathepsin B, and dystrophin. Genes related to cardiac dysfunction were altered in zebrafish following exposure to 1000 µg/L citalopram. Using literature and databases that describe gene interactions, molecular networks (clinical and disease networks) were constructed to understand effects of citalopram.

Abstract Neurodegeneration is the primary driver of disease progression in multiple sclerosis (MS) resulting in permanent disability, creating an urgent need to discover its underlying mechanisms. Herein, we establish that dysfunction of the RNA binding protein heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) results in differential of binding to RNA targets causing alternative RNA splicing, which contributes to neurodegeneration in MS and its models. Using RNAseq of MS brains, we discovered differential expression and aberrant splicing of hnRNP A1 target RNAs involved in neuronal function and RNA homeostasis. We confirmed this in vivo in experimental autoimmune encephalomyelitis employing CLIPseq specific for hnRNP A1, where hnRNP A1 differentially binds and regulates RNA, including aberrantly spliced targets identified in human samples. Additionally, dysfunctional hnRNP A1 expression in neurons caused neurite loss and identical changes in splicing, corroborating hnRNP A1 dysfunction as a cause of neurodegeneration. Collectively, these data indicate hnRNP A1 dysfunction causes altered neuronal RNA splicing, resulting in neurodegeneration in MS.

Toxicity mechanisms of per- and polyfluoroalkyl substances (PFASs), a chemical class present in diverse ecosystems, as well as many of their precursors, have been increasingly characterized in aquatic species. Perfluorooctanesulfonamide (PFOSA, C8H2F17NO2S) is a common precursor of perfluorooctane sulfonic acid (PFOS), a long-chain PFAS. Here, we assessed sub-lethal endpoints related to development, oxidative stress, transcript levels, and distance moved in zebrafish embryos and larvae following continuous exposure to PFOSA beginning at 6 h post-fertilization (hpf). PFOSA decreased survival in fish treated with 1 µg/L PFOSA; however, the effect was modest relative to the controls (difference of 10%). Exposure up to 10 µg/L PFOSA did not affect hatch rate, nor did it induce ROS in 7-day-old larvae fish. The activity of larval fish treated with 100 µg/L PFOSA was reduced relative to the solvent control. Transcripts related to oxidative stress response and apoptosis were measured and BCL2-associated X, apoptosis regulator (bax), cytochrome c, somatic (cycs), catalase (cat), superoxide dismutase 2 (sod2) were induced with high concentrations of PFOSA. Genes related to neurotoxicity were also measured and transcript levels of acetylcholinesterase (ache), elav-like RNA binding protein 3 (elavl3), growth-associated protein 43 (gap43), synapsin II (syn2a), and tubulin 3 (tubb3) were all increased in larval fish with higher PFOSA exposure. These data improve our understanding of the potential sub-lethal toxicity of PFOSA in fish species.

An over-active renin-angiotensin system (RAS) is characterized by elevated angiotensin II (Ang II). While Ang II can promote metabolic and mitochondrial dysfunction in tissues, little is known about its role in the gastrointestinal system (GI). Here, we treated rat primary colonic epithelial cells with Ang II (1–5000 nM) to better define their role in the GI. We hypothesized that Ang II would negatively affect mitochondrial bioenergetics as these organelles express Ang II receptors. Ang II increased cellular ATP production but reduced the mitochondrial membrane potential (MMP) of colonocytes. However, cells maintained mitochondrial oxidative phosphorylation and glycolysis with treatment, reflecting metabolic compensation with impaired MMP. To determine whether lipid dysregulation was evident, untargeted lipidomics were conducted. A total of 1949 lipids were detected in colonocytes spanning 55 distinct (sub)classes. Ang II (1 nM) altered the abundance of some sphingosines [So(d16:1)], ceramides [Cer-AP(t18:0/24:0)], and phosphatidylcholines [OxPC(16:0_20:5(2O)], while 100 nM Ang II altered some triglycerides and phosphatidylserines [PS(19:0_22:1). Ang II did not alter the relative expression of several enzymes in lipid metabolism; however, the expression of pyruvate dehydrogenase kinase 2 (PDK2) was increased, and PDK2 can be protective against dyslipidemia. This study is the first to investigate the role of Ang II in colonic epithelial cell metabolism.

BackgroundFew studies have investigated the risk of hospitalization among patients with synucleinopathies (Parkinson disease, Dementia with Lewy Bodies, Parkinson disease dementia, Multiple System Atrophy) with associated psychosis and the impact of antipsychotic treatments on hospital admissions and duration of the stay.ObjectiveTo determine the risk of hospitalization among patients with synucleinopathies and in patients with associated psychosis. To evaluate the impact of antipsychotic treatments on hospital admission of patients with synucleinopathies and psychosis in an incident cohort study in Olmsted County, Minnesota (MN).MethodsWe used the Rochester Epidemiology Project (REP) to define an incident cohort of patients with clinically diagnosed synucleinopathies (1991–2010) in Olmsted County, MN. A movement disorder specialist reviewed all medical records to confirm the clinical diagnosis of synucleinopathies using the NINDS/NIMH unified diagnostic criteria.ResultsWe included 416 incident cases of clinically diagnosed synucleinopathies from 2,669 hospitalizations. 409 patients (98.3%) were admitted to the hospital at least once for any cause after the onset of parkinsonism. The median number of hospitalizations for a single patient was 5. In total, 195 (46.9%) patients met the criteria for psychosis: patients with psychosis had a 49% (HR = 1.49, p

Ifosfamide is a cancer-fighting chemotherapeutic that has been detected in aquatic ecosystems. Zebrafish larvae were exposed to either 0, 1 or 100 µg/L ifosfamide in the water for 7 days, and fish were subjected to total RNA extraction and RNA-seq analysis with the Illumina NovoSeq 6000 instrument. Raw sequence data were processed through fastp and clean reads obtained by removing adapter and poly-N sequences, as well as low quality reads. Differential gene expression was performed using the abundance of transcripts that mapped to the zebrafish genome. To uncover putative targets regulated by microRNAs, Pathway Studio 12.0 was used to conduct a subnetwork enrichment analysis. Expression data were used to predict which microRNAs were important for the response to ifosfamide exposure. There were 21 common microRNAs identified in both the ''IFOS1'' and ''IFOS100'' datasets. These were MIR150, MIR6515, MIR657, MIR216A, m_Mir741, MIRLET7E, miR-let-7, MIR2392, r_Mir3551, MIR181B1, MIR33A, MIR502, MIR193B, MIR146A, MIR431, MIR647, m_Mir1192, MIR297, MIR328, and MIR4717. Data can be re-used to advance adverse outcome pathways in regulatory toxicology and to refine biomarker discovery for antineoplastics in aquatic environments.

Abstract Information about regional‐level carbon (C) stocks in agroforestry systems (AFS), as well as the annual loss of agroforests and associated C stocks, is scarce, limiting our capacity for increasing C sequestration through establishing, retaining, and enhancing these systems. This study quantified regional‐level C stocks and the associated incremental economic value in the forest land‐use component of three common AFS (hedgerows, shelterbelts, and silvopastures), estimated the annual loss of hedgerow and silvopasture forests and the associated C, and assessed the potential to enhance C storage through the expansion of shelterbelts in central Alberta, Canada, using publicly available satellite imagery, previously collected field data and the Google Earth Engine platform. Results showed that forests in the three AFS stored 699.9 million tons (Mt) C across 9.5 million hectares (Mha) of land in central Alberta and were valued at $102.7 billion based on the 2021 Canadian C tax rate of $40 t−1 CO2‐equivalent. Silvopasture forests in the studied region had the highest C stocks, which were 14.2 and 67.2 times that found in hedgerow and shelterbelt forests, respectively. Between 2001 and 2020, forests in hedgerows and silvopastures declined at rates of 468.1 and 1957.1 ha year−1, respectively, leading to an 8.4 Mt decline in total C storage over the 20 years. However, there is potential to establish new shelterbelts at many road/field margins, which could increase C stocks by 2.3 times the current C stocks in shelterbelt forests. These results highlight the importance of retaining existing and establishing new AFS for increasing C sequestration, emphasizing the impact of agroforest loss on reducing C storage within agroecosystems. The development of policies that assist or reward landowners for providing the ecosystem service of C storage by retaining, establishing, and enhancing agroforests as part of existing agroecosystem management should be encouraged for mitigating climate change.

Introduction: Severe obesity among children and adolescents has emerged as a public health concern in multiple places around the world. Methods: We searched the Medline database for articles on severe obesity rates in children published between January 1960 and January 2020. For studies with available prevalence rates for an early and a more recent time period, the relative increase in prevalence was imputed. Results: In total, 874 publications were identified, of which 38 contained relevant epidemiological data. Rates of severe obesity varied significantly according to age, gender, geographic area, and the definition of severe obesity. The highest rates of class II and III obesity in the USA according to the Centers of Disease Control cut-off were 9.5% and 4.5%, respectively. Seventeen studies reported prevalence rates in at least two time periods. Data for 9,190,718 individuals showed a 1.71 (95% CI, 1.53–1.90) greater odds for severe obesity in 2006–2017 (N = 5,029,584) versus 1967–2007 (N = 4,161,134). In an analysis limited to studies from 1980s with a minimum follow-up of 20 years, a 9.16 (95% CI, 7.76–10.80) greater odds for severe obesity in recent versus earlier time was found. An analysis limited to studies from 2000, with a follow-up of 5–15 years, a 1.09 (95% CI, 0.99–1.20) greater odds was noted when comparing (2011–2017; N = 4,991,831) versus (2000–2011; N = 4,134,340). Conclusion: Severe pediatric obesity is escalating with a marked increase from the 1980s and a slower rate from 2000.

Mortalin (GRP75, HSPA9A), a heat shock protein (HSP), regulates a wide range of cellular processes, including cell survival, growth, and metabolism. The regulatory functions of mortalin are mediated through a diverse set of protein partners associated with different cellular compartments, which allows mortalin to perform critical functions under physiological conditions, including mitochondrial protein quality control. However, alteration of mortalin’s activities, its abnormal subcellular compartmentalization, and its protein partners turn mortalin into a disease-driving protein in different pathological conditions, including cancers. Here, mortalin’s contributions to tumorigenic pathways are explained. Pathology information based on mortalin’s RNA expression extracted from The Cancer Genome Atlas (TCGA) transcriptomic database indicates that mortalin has an independent prognostic value in common tumors, including lung, breast, and colorectal cancer (CRC). Subsequently, the binding partners of mortalin reported in different cellular models, from yeast to mammalian cells, and its regulation by post-translational modifications are discussed. Finally, we focus on colorectal cancer and discuss how mortalin and its tumorigenic downstream protein targets are regulated by a ubiquitin-like protein through the 26S proteasomal degradation machinery. A broader understanding of the function of mortalin and its positive and negative regulation in the formation and progression of human diseases, particularly cancer, is essential for developing new strategies to treat a diverse set of human diseases critically associated with dysregulated mortalin.

Neurodegeneration, the progressive loss or damage to neurons and axons, underlies permanent disability in multiple sclerosis (MS); yet its mechanisms are incompletely understood. Recent data indicates autoimmunity to several intraneuronal antigens, including the RNA binding protein (RBP) heterogenous nuclear ribonucleoprotein A1 (hnRNP A1), as contributors to neurodegeneration. We previously showed that addition of anti-hnRNP A1 antibodies, which target the same immunodominant domain of MS IgG, to mice with experimental autoimmune encephalomyelitis (EAE) worsened disease and resulted in an exacerbation of hnRNP A1 dysfunction including cytoplasmic mislocalization of hnRNP A1, stress granule (SG) formation and neurodegeneration in the chronic stages of disease. Because this previous study focused on a singular timepoint during EAE, it is unclear whether anti-hnRNP A1 antibody induced hnRNP A1 dysfunction caused neurodegeneration or was result of it. In the present study, we analyzed in vivo and in vitro models of anti-hnRNP A1 antibody-mediated autoimmunity for markers of hnRNP A1 dysfunction and neurodegeneration over a time course to gain a better understanding of the connection between hnRNP A1 dysfunction and neurodegeneration. Anti-hnRNP A1 antibody treatment resulted in increased neuronal hnRNP A1 mislocalization and nuclear depletion temporally followed by altered RNA expression and SG formation, and lastly an increase in necroptotic signalling and neuronal cell death. Treatment with necrostatin-1s inhibited necroptosis and partially rescued anti-hnRNP A1 antibody-mediated neurodegeneration while clathrin knockdown specifically inhibited anti-hnRNP A1 antibody uptake into neurons. This data identifies a novel antibody-mediated mechanism of neurodegeneration, which may be targeted to inhibit neurodegeneration and prevent permanent neurological decline in persons living with MS.

Summary: Background: School-based programmes, including hearing screening, provide essential preventive services for rural children. However, minimal evidence on screening methodologies, loss to follow-up, and scarcity of specialists for subsequent care compound rural health disparities. We hypothesised telemedicine specialty referral would improve time to follow-up for school hearing screening compared with standard primary care referral. Methods: In this cluster-randomised controlled trial conducted in 15 rural Alaskan communities, USA, we randomised communities to telemedicine specialty referral (intervention) or standard primary care referral (control) for school hearing screening. All children (K–12; aged 4–21 years) enrolled in Bering Straight School District were eligible. Community randomisation occurred within four strata using location and school size. Participants were masked to group allocation until screening day, and assessors were masked throughout data collection. Screening occurred annually, and children who screened positive for possible hearing loss or ear disease were monitored for 9 months from the screening date for follow-up. Primary outcome was the time to follow-up after a positive hearing screen; analysis was by intention to treat. The trial was registered with ClinicalTrials.gov, NCT03309553. Findings: We recruited participants between Oct 10, 2017, and March 28, 2019. 15 communities were randomised: eight (750 children) to telemedicine referral and seven (731 children) to primary care referral. 790 (53·3%) of 1481 children screened positive in at least one study year: 391 (52∤1%) in the telemedicine referral communities and 399 (50∤4%) in the primary care referral communities. Of children referred, 268 (68·5%) in the telemedicine referral communities and 128 (32·1%) in primary care referral communities received follow-up within 9 months. Among children who received follow-up, mean time to follow-up was 41·5 days (SD 55·7) in the telemedicine referral communities and 92·0 days (75·8) in the primary care referral communities (adjusted event-time ratio 17·6 [95% CI 6·8–45·3] for all referred children). There were no adverse events. Interpretation: Telemedicine specialty referral significantly improved the time to follow-up after hearing screening in Alaska. Telemedicine might apply to other preventive school-based services to improve access to specialty care for rural children. Funding: Patient-Centered Outcomes Research Institute.

Plain English summary Community involvement is important in good research practice. We led a community-based study to improve early detection and treatment of childhood hearing loss in rural Alaska. This study evaluated a cell phone-based hearing screening process and compared a new telemedicine specialty referral pathway to the standard primary care referral pathway. The study included community involvement, engagement, and participation from the very beginning to inform how to best design the trial. We obtained insight and feedback from community members through involvement of a core stakeholder team and through community engagement and participation in focus groups and community events. Feedback received through community involvement and participation influenced the design of the trial at key decision points. Community member guidance shaped the research question, the outcomes to be measured, and the procedures for completing the project, such as participant recruitment. This study offers an example of community involvement, engagement and participation that could be mirrored in future research to maintain the interests of participating communities. Abstract Background Effective systems for early identification and treatment of childhood hearing loss are essential in rural Alaska, where data indicate a high prevalence of childhood ear infections and hearing loss. However, loss to follow-up from school hearing screening programs is pervasive. The Hearing Norton Sound study was a mixed methods community randomized controlled trial that was developed to address this gap. The study engaged community members and participants in the design of the trial, including involvement of stakeholders as collaborators. Methods Community engagement and participation in research design occurred through focus groups and through the integration of stakeholders into the study team. Representation was cross-sectoral, involving individuals from multiple levels of the school and health system, as well as community members from each of the 15 communities. Feedback obtained between April 2017 and August 2017 informed the final design of the randomized trial, which began enrollment of children in October 2017 and concluded in March 2019. Results Stakeholder involvement and community participation shaped the design of specific trial elements (research question; comparators; outcomes and measures; telemedicine protocols; and recruitment and retention). Community involvement was strengthened by the use of multiple modalities of involvement and by the positionality of lead stakeholders on the study team. Conclusions This study highlights the effectiveness of multifaceted stakeholder involvement and participation in the design of health research conducted within Alaska Native communities. It offers an example of involvement and reporting that could be mirrored in future research in order to protect and further the interests of the participating community. Trial registration ClinicalTrials.gov, NCT03309553 , First registered 10/9/2017

Abstract Severe obesity among children and adolescents is a significant global public health concern. The prevalence has markedly increased over the last decades, becoming common in many countries. Overwhelming rates of obesity among youth have prompted efforts to identify an evidence-based immediate- and long-term cardiometabolic risk factor profile in childhood-onset severe obesity, and to highlight gaps that require further investigation. The PubMed database was systematically searched in accordance with PRISMA guidelines. The search yielded 831 results, of which 60 fulfilled stringent criteria and were summarized in this review. The definition of severe obesity was variable, with only one half the publications using the definition BMI > 120% of the 95th percentile. Point estimates of the prevalence of at least one cardiometabolic risk factor in children with severe obesity reportedly range from 67 to 86%. Cross-sectional studies indicate that children and adolescents with severe obesity are at greater risk than those with mild obesity for type 2 diabetes, hypertension, fatty liver disease and dyslipidemia, already at childhood and adolescence. Robust epidemiological data on the long-term risk and actual point estimates in adulthood are lacking for these diseases as well as for other diseases (coronary heart disease, stroke, chronic kidney disease and cancer). Recent longitudinal studies indicate an increased risk for cardiomyopathy, heart failure, cardiovascular mortality and all-cause mortality in adulthood for adolescents with severe obesity compared to those with mild obesity. Given the alarming increase in the prevalence of severe obesity, the persistence of adiposity from childhood to adulthood and the precarious course of young adults with chronic comorbidities, the economic and clinical services burden on the healthcare system is expected to rise.

Background: No studies have reported the rate of motor complications (MC) and response to medical and surgical treatment in a population-based cohort of young-onset Parkinson's Disease (YOPD) patients and a cohort of sex-matched late-onset Parkinson's Disease (LOPD).Objective: To assess the outcomes of dopaminergic treatment in YOPD and LOPD, explore treatment-induced MC, medical adjustment, and rate of deep brain stimulation (DBS).Methods: We used the expanded Rochester Epidemiology Project (eREP) to investigate a population-based cohort of YOPD between 2010 and 2015 in 7 counties in Minnesota. Cases with onset ≤55 years of age were included as YOPD. An additional sex-matched cohort of LOPD (onset at ≥56 years of age) was included for comparison. All medical records were reviewed to confirm the diagnoses.Results: In the seven counties 2010–15, there were 28 YOPD patients, which were matched with a LOPD cohort. Sixteen (57%) YOPD had MC, as compared to 9 (32%) LOPD. In YOPD, 9 had motor fluctuations (MF) and Levodopa-induced dyskinesia (LID) together, whereas 3 had LID only and 4 MF only. In LOPD, 3 had MF and LID, 3 MF only, and 3 LID only. Following medical treatment for MC, 6/16 YOPD (38%) and 3/9 (33%) LOPD had symptoms resolution. In YOPD, 11/16 (69%) were considered for DBS implantation, in LOPD they were 2/9 (22%), but only 7 (6 YOPD and 1 LOPD) underwent the procedure. YOPD had significantly higher rates in both DBS candidacy and DBS surgery (respectively, p = 0.03 and p = 0.04). Among DBS-YOPD, 5/6 (83%) had positive motor response to the surgery; the LOPD case had a poor response. We report the population-based incidence of both YOPD with motor complications and YOPD undergoing DBS, which were 1.17 and 0.44 cases per 100,000 person-years, respectively.Conclusion: Fifty-seven percent of our YOPD patients and 32% of the LOPD had motor complications. Roughly half of both YOPD and LOPD were treatment resistant. YOPD had higher rates of DBS candidacy and surgery. Six YOPD and 1 LOPD underwent DBS implantation and most of them had a positive motor response after the surgery.

Background: Cognitive function among apparently healthy adolescents has been associated with cardiovascular morbidity and mortality. We examined the relationship between global and subdomain cognitive scores in adolescence and early-onset type 2 diabetes (T2D) in men and women. Methods: A nationwide, population-based study of 971,677 Israeli born adolescents (56% men; mean age 17.4 years) who were medically examined and their cognitive performance was assessed before compulsory military service during 1992–2010. Data included global and subdomain cognitive Z-scores (problem-solving, verbal abstraction and categorization, verbal comprehension, and mathematical abilities). Data were linked to the Israeli National Diabetes Registry. The relations between global and subdomain scores and incident T2D was determined using Cox proportional hazard models and logistic regression models. Analyses were conducted separately for men and women. Findings: During 16,095,122 person-years, 3,570 individuals developed T2D. After adjustment, those in the low compared to the high quintile of global cognitive Z-score had the highest risk for T2D; HR 2.46, (95% CI 2.10–2.88) for men and 2.33 (95% CI 1.88–2.89) for women. A one-unit lower global cognitive Z-score was associated with 1.41 (95% CI 1.34–1.48) and 1.46 (95% CI 1.36–1.56) increased risks for men and women, respectively. The relationship was noted for the cognitive subdomains scores as well as for the global cognitive score, with no heterogeneity across cognitive subdomains. Interpretation: This large nationally representative cohort suggests relationship between global, as well as subdomain cognitive scores in late adolescence, and incident early onset T2D in both sexes, which was independent of socioeconomic status.

Greenhouse gas (GHG) emissions from crop residue management have been studied extensively, yet the effects of harvesting more than one crop residue in a rotation have not been reported. Here, we measured the short-term changes in methane (CH4), nitrous oxide (N2O), and carbon dioxide (CO2) emissions in response to residue removal from continuous corn (Zea mays L.) (CC) and corn–wheat (Triticum aestivum L.)–soybean (Glycine max L. Merr.) (CWS) rotations in the Mid-Atlantic USA. A first experiment retained five corn stover rates (0, 3.33, 6.66, 10, and 20 Mg ha−1) in a continuous corn (CC) in Blacksburg, VA, in 2016 and 2017. Two other experiments, initiated during the wheat and corn phases of the CWS rotation in New Kent, VA, utilized a factorial combination of retained corn (0, 3.33, 6.66, and 10.0 Mg ha−1) and wheat residue (0, 1, 2, and 3 Mg ha−1). Soybean residue was not varied. Different crop retention rates did not affect CO2 fluxes in any of the field studies. In Blacksburg, retaining 5 Mg ha−1 stover or more increased CH4 and N2O emissions by ~25%. Maximum CH4 and N2O fluxes (4.16 and 5.94 mg m−2 day−1) occurred with 200% (20 Mg ha−1) retention. Two cycles of stover management in Blacksburg, and one cycle of corn or wheat residue management in New Kent did not affect GHG fluxes. This study is the first to investigate the effects of crop residue on GHG emissions in a multi-crop system in humid temperate zones. Longer-term studies are warranted to understand crop residue management effects on GHG emissions in these systems.

Osteogenesis imperfecta (OI) is characterized by short stature, skeletal deformities, low bone mass, and motor deficits. A subset of OI patients also present with joint hypermobility; however, the role of tendon dysfunction in OI pathogenesis is largely unknown. Using the Crtap-/- mouse model of severe, recessive OI, we found that mutant Achilles and patellar tendons were thinner and weaker with increased collagen cross-links and reduced collagen fibril size at 1- and 4-months compared to wildtype. Patellar tendons from Crtap-/- mice also had altered numbers of CD146+CD200+ and CD146-CD200+ progenitor-like cells at skeletal maturity. RNA-seq analysis of Achilles and patellar tendons from 1-month Crtap-/- mice revealed dysregulation in matrix and tendon marker gene expression concomitant with predicted alterations in TGF-β, inflammatory, and metabolic signaling. At 4-months, Crtap-/- mice showed increased αSMA, MMP2, and phospho-NFκB staining in the patellar tendon consistent with excess matrix remodeling and tissue inflammation. Finally, a series of behavioral tests showed severe motor impairments and reduced grip strength in 4-month Crtap-/- mice – a phenotype that correlates with the tendon pathology.

That conditional, if-then reasoning does not emerge until 4–5 years has long been accepted. Here we show that children barely 3 years old can do conditional reasoning. All that was needed was a superficial change to the stimuli: When color was a property of the shapes (line drawings of a star and truck) rather than of the background (as in all past conditional discrimination [CD] testing), 3-year-olds could succeed. Three-year-olds do not seem to use color to inform them which shape is correct unless color is a property of the shapes themselves. While CD requires integrating color and shape information, the dimensional change card sort (DCCS) task requires keeping those dimension cognitively separate – inhibiting attention to one (e.g., shape) when sorting by the other (e.g., color). For DCCS, a superficial change to the stimuli that is the inverse of what helps on CD enables 3-year-olds to succeed when normally they do not until ∼4⁤12 years. As we and others have previously shown, 3-year-olds can succeed at DCCS when color is a property of the background (e.g., a white truck on a red background), instead of a property of the stimulus (e.g., a red truck on a white background, as in standard DCCS). Our findings on CD and DCCS suggest that scaffolding preschoolers’ emerging conceptual skills by changing the way stimuli look (perceptual bootstrapping) enables 3-year-olds to demonstrate reasoning abilities long thought beyond their grasp. Evidently, children of 3 years have difficulty mentally separating dimensions (e.g., color and shape) of the same object and difficulty mentally integrating dimensions not part of the same object. Our present CD findings plus our earlier DCCS findings provide strong evidence against prominent cognitive complexity, conditional reasoning, and graded memory theories for why 3-year-olds fail these two tasks. The ways we have traditionally queried children may have obscured the budding reasoning competencies present at 3 years of age.

Efforts to reduce greenhouse gas (GHG) emissions from cattle production have led to modifications of livestock diet composition aimed at reducing CH4 emissions from enteric fermentation. These diet modifications can result in varied manure types that may differentially affect GHG emissions when applied to soil. The purpose of this experiment was to examine the effect of different manure types on GHG emissions. We conducted an incubation experiment, comparing the manure from livestock fed a corn-based diet (CM) to that from livestock fed a traditional barley-based diet (BM). The manures were applied to three soil types (with varied soil fertility and pH) and compared to a control (without manure application). Carbon dioxide (CO2) emissions were greater from CM than from BM across all soil types (29.1 and 14.7 mg CO2-C kg−1, respectively). However, CM resulted in lower N2O emissions relative to BM in the low fertility soil (4.21 and 72.67 μg N2O-N kg−1, respectively) and in lower CH4 emissions relative to BM in the two acidic soils (0.5 and 2.5 μg CH4-C kg−1, respectively). Total GHG emissions (sum of CO2, N2O, and CH4) were similar between CM and BM across all soil types, but CM (unlike BM) had 52–66% lower emissions in the low fertility soil relative to both CM and BM in the high fertility soil. Our study shows that manure and soil type interact to affect GHG emissions and that CM may mitigate N2O emissions relative to BM when applied to low fertility soils.

Cattle production is a large source of greenhouse gas (GHG) emissions from the Canadian livestock sector. Efforts to reduce CH4 emissions from enteric fermentation have led to modifications of diet composition for livestock, resulting in a corresponding change in manure properties. We studied the effect of applying manure from cattle fed a barley-based diet with and without the methane inhibitor supplement, 3-nitrooxypropanol (3-NOP), on soil GHG emissions. Three soils common to Alberta, Canada, were used: a Black Chernozem, a Dark Brown Chernozem, and a Gray Luvisol. We compared the supplemented (3-NOPM) and non-supplemented manure (BM) amendments to a composted 3-NOPM (3-NOPC) amendment and a control with no manure amendment (CK). In an 84-day laboratory incubation experiment, 3-NOPM had significantly lower cumulative CO2 emissions compared to BM in both the Black Chernozem and Gray Luvisol. The cumulative N2O emissions were lowest for 3-NOPC and CK and highest for 3-NOPM across all soil types. Cumulative CH4 emissions were only affected by soil type, with a net positive flux from the fine-textured Gray Luvisol and Dark Brown Chernozem and a net negative flux from the coarse-textured Black Chernozem. Cumulative anthropogenic GHG emissions (CO2-equivalent) from soil amended with 3-NOPM were significantly higher than those for both BM and CK amendments in the Black Chernozem, while the cumulative anthropogenic GHG emissions from the 3-NOPC treatment were similar to or significantly lower than those for the BM and CK treatments across all soil types. We conclude that soil GHG emissions resulting from the 3-NOPM amendment are dependent on soil type and 3-NOPM could potentially increase soil GHG emissions compared to BM or CK. Although we show that the composting of 3-NOPM prior to soil application can reduce soil GHG emissions, the composting process also releases GHGs, which should also be considered in assessing the life-cycle of manure application. Our results provide a first look at the potential effect of the next stage in the life cycle of 3-NOP on GHG emissions. Further research related to the effect of soil properties, particularly in field studies, is needed to assess the best management practices related to the use of manure from cattle-fed diets supplemented with 3-NOP as a soil amendment.

Picocyanobacteria are the numerically dominant photoautotrophs of the oligotrophic regions of Earth's oceans. These organisms are characterized by their small size and highly reduced genomes. Strains partition to different light intensity and nutrient level niches, with differing photosynthetic apparatus stoichiometry, light harvesting machinery and susceptibility to photoinactivation. In this study, we grew three strains of picocyanobacteria: the low light, high nutrient strain Prochlorococcus marinus MIT 9313; the high light, low nutrient Prochlorococcus marinus MED 4; and the high light, high nutrient marine Synechococcus strain WH 8102; under low and high growth light levels. We then performed matched photophysiology, protein and transcript analyses. The strains differ significantly in their rates of Photosystem II repair under high light and in their capacity to remove the PsbA protein as the first step in the Photosystem II repair process. Notably, all strains remove the PsbD subunit at the same rate that they remove PsbA. When grown under low light, MIT 9313 loses active Photosystem II quickly when shifted to high light, but has no measurable capacity to remove PsbA. MED 4 and WH 8102 show less rapid loss of Photosystem II and considerable capacity to remove PsbA. MIT 9313 has less of the FtsH protease thought to be responsible for the removal of PsbA in other cyanobacteria. Furthermore, by transcript analysis the predominant FtsH isoform expressed in MIT 9313 is homologous to the FtsH 4 isoform characterized in the model strain Synechocystis PCC 6803, rather than the FtsH 2 and 3 isoforms thought to be responsible for PsbA degradation. MED 4 on the other hand shows high light inducible expression of the isoforms homologous to FtsH 2 and 3, consistent with its faster rate of PsbA removal. MIT 9313 has adapted to its low light environment by diverting resources away from Photosystem II content and repair.

Marine Synechococcus and Prochlorococcus are picocyanobacteria predominating in subtropical, oligotrophic marine environments, a niche predicted to expand with climate change. When grown under common low light conditions Synechococcus WH 8102 and Prochlorococcus MED 4 show similar Cytochrome b6f and Photosystem I contents normalized to Photosystem II content, while Prochlorococcus MIT 9313 has twice the Cytochrome b6f content and four times the Photosystem I content of the other strains. Interestingly, the Prochlorococcus strains contain only one third to one half of the RUBISCO catalytic subunits compared to the marine Synechococcus strain. The maximum Photosystem II electron transport rates were similar for the two Prochlorococcus strains but higher for the marine Synechococcus strain. Photosystem II electron transport capacity is highly correlated to the molar ratio of RUBISCO active sites to Photosystem II but not to the ratio of cytochrome b6f to Photosystem II, nor to the ratio of Photosystem I: Photosystem II. Thus, the catalytic capacity for the rate-limiting step of carbon fixation, the ultimate electron sink, appears to limit electron transport rates. The high abundance of Cytochrome b6f and Photosystem I in MIT 9313, combined with the slower flow of electrons away from Photosystem II and the relatively low level of RUBISCO, are consistent with cyclic electron flow around Photosystem I in this strain.

Research on classroom motivational climates and microclimates--students' shared and idiosyncratic perceptions of motivational classroom features--demonstrates their importance for fostering adaptive motivational and achievement-related outcomes. However, a lack of coherent theoretical guidance about the nature of students' classroom climate perceptions has yielded numerous conceptualizations and measurement approaches for these processes. Further, although existing theories and conceptualizations vary in the specific motivational climate features they propose, considerable conceptual overlap exists among them. Working toward conceptual clarity, theoretical integration, and guidance for measurement, we performed a systematic review to identify prominent measurement trends in motivational climate research. Results revealed teacher autonomy support and classroom goal structures as the most frequently measured classroom climate qualities. We observed a wide variety of validity evidence for the measures; in particular, a low incidence of studies assessed the factor structure and considered the multilevel nature of climate data, with most treating climate perceptions as student-level phenomena. In addition to providing a much-needed guide of existing measurement practices, this systematic review lays a foundation for the continued theoretical advancement of motivational climate. We call for more rigorous reporting of validity evidence, rationales for measure selection, and the underlying assumptions guiding measure selection when conducting classroom climate research, as well as focused development of instruments targeting less-frequently measured climate constructs.

The marine picocyanobacteria Synechococcus and Prochlorococcus numerically dominate open ocean phytoplankton. Although evolutionarily related they are ecologically distinct, with different strategies to harvest, manage and exploit light. We grew representative strains of Synechococcus and Prochlorococcus and tracked their susceptibility to photoinactivation of Photosystem II under a range of light levels. As expected blue light provoked more rapid photoinactivation than did an equivalent level of red light. The previous growth light level altered the susceptibility of Synechococcus, but not Prochlorococcus, to this photoinactivation. We resolved a simple linear pattern when we expressed the yield of photoinactivation on the basis of photons delivered to Photosystem II photochemistry, plotted versus excitation pressure upon Photosystem II, the balance between excitation and downstream metabolism. A high excitation pressure increases the generation of reactive oxygen species, and thus increases the yield of photoinactivation of Photosystem II. Blue photons, however, retained a higher baseline photoinactivation across a wide range of excitation pressures. Our experiments thus uncovered the relative influences of the direct photoinactivation of Photosystem II by blue photons which dominates under low to moderate blue light, and photoinactivation as a side effect of reactive oxygen species which dominates under higher excitation pressure. Synechococcus enjoyed a positive metabolic return upon the repair or the synthesis of a Photosystem II, across the range of light levels we tested. In contrast Prochlorococcus only enjoyed a positive return upon synthesis of a Photosystem II up to 400 μmol photons m-2 s-1. These differential cost-benefits probably underlie the distinct photoacclimation strategies of the species.

Most of our terrestrial carbon (C) storage occurs in soils as organic C derived from living organisms. Therefore, the fate of soil organic C (SOC) in response to changes in climate, land use, and management is of great concern. Here we provide a unified conceptual model for SOC cycling by gathering the available information on SOC sources, dissolved organic C (DOC) dynamics, and soil biogeochemical processes. The evidence suggests that belowground C inputs (from roots and microorganisms) are the dominant source of both SOC and DOC in most ecosystems. Considering our emerging understanding of SOC protection mechanisms and long-term storage, we highlight the present need to sample (often ignored) deeper soil layers. Contrary to long-held biases, deep SOC—which contains most of the global amount and is often hundreds to thousands of years old—is susceptible to decomposition on decadal timescales when the environmental conditions under which it accumulated change. Finally, we discuss the vulnerability of SOC in different soil types and ecosystems globally, as well as identify the need for methodological standardization of SOC quality and quantity analyses. Further study of SOC protection mechanisms and the deep soil biogeochemical environment will provide valuable information about controls on SOC cycling, which in turn may help prioritize C sequestration initiatives and provide key insights into climate-carbon feedbacks.

Shorter lectures punctuated with activities to engage students in the learning process can increase student understanding, critical thinking, and overall learning. However, some students have negative responses to active teaching strategies. Here we explore the topic of student resistance to active learning, including reasons for this opposition and strategies to prevent or respond to it. Recognizing factors that lead to students' resistance to active learning is important to mitigating these barriers to learning. Equally critical to mitigating student resistance is the promotion of student resilience. Structuring classrooms to promote resilience includes community building, structured activities, and policies that recognize student diversity, and the complexity of learning processes.

ContextThe azide anion (N3-) is highly toxic. It exists most commonly as sodium azide, which is used widely and is readily available, raising the potential for occupational incidents and use as a weapon of mass destruction. Azide-poisoned patients present with vomiting, seizures, hypotension, metabolic acidosis, and coma; death can occur. No specific azide antidote exists, with treatment being solely supportive. Azide inhibits mitochondrial cytochrome c oxidase and is likely oxidized to nitric oxide in vivo. Cytochrome c oxidase inhibition depletes intracellular adenosine triphosphate and increases oxidative stress, while increased nitric oxide causes hypotension and exacerbates oxidative damage. Here, we tested whether the cobalamin (vitamin B12) analog cobinamide, a strong and versatile antioxidant that also neutralizes nitric oxide, can reverse azide toxicity in mammalian cells, Drosophila melanogaster, and mice.ResultsWe found cobinamide bound azide with a moderate affinity (Ka 2.87 × 105 M-1). Yet, cobinamide improved growth, increased intracellular adenosine triphosphate, and reduced apoptosis and malondialdehyde, a marker of oxidative stress, in azide-exposed cells. Cobinamide rescued Drosophila melanogaster and mice from lethal exposure to azide and was more effective than hydroxocobalamin. Azide likely generated nitric oxide in the mice, as evidenced by increased serum nitrite and nitrate, and reduced blood pressure and peripheral body temperature in the animals; the reduced temperature was likely due to reflex vasoconstriction in response to the hypotension. Cobinamide improved recovery of both blood pressure and body temperature.ConclusionWe conclude cobinamide likely acted by neutralizing both oxidative stress and nitric oxide, and that it should be given further consideration as an azide antidote.