Guidance for prevention and management of COVID-19 in children and adolescents: A consensus statement from the Pediatric Infectious Diseases Society Pediatric COVID-19 Therapies Taskforce.

BACKGROUND: Since November 2019, the SARS-CoV-2 pandemic has created challenges for preventing and managing COVID-19 in children and adolescents. Most research to develop new therapeutic interventions or to repurpose existing ones has been undertaken in adults, and although most cases of infection in pediatric populations are mild, there have been many cases of critical and fatal infection. Understanding the risk factors for severe illness and the evidence for safety, efficacy, and effectiveness of therapies for COVID-19 in children is necessary to optimize therapy. METHODS: A panel of experts in pediatric infectious diseases, pediatric infectious diseases pharmacology, and pediatric intensive care medicine from 21 geographically diverse North American institutions was re-convened. Through a series of teleconferences and web-based surveys and a systematic review with meta-analysis of data for risk factors, a guidance statement comprising a series of recommendations for risk stratification, treatment, and prevention of COVID-19 was developed and refined based on expert consensus. RESULTS: There are identifiable clinical characteristics that enable risk stratification for patients at risk for severe COVID-19. These risk factors can be used to guide the treatment of hospitalized and non-hospitalized children and adolescents with COVID-19 and to guide preventative therapy where options remain available.

Protective behavioral strategies and alcohol-related outcomes: The moderating effects of drinking refusal self-efficacy and sex.

The investigation of drinking refusal self-efficacy and alcohol protective behavioral strategies (PBSA) has revealed inconsistent results. Sex may be one factor that plays a role in these results given the demonstrable differences between the alcohol use behaviors of men and women. The current study examined the moderating effects of drinking refusal self-efficacy and sex on the relationships that PBSA subtypes have with alcohol outcomes for traditional age undergraduate students (18-25 years of age; 81% women; 60% White). Results showed negative associations between manner of drinking PBSA and alcohol consumption for individuals with high levels of drinking refusal self-efficacy but not low levels of drinking refusal self-efficacy. However, manner of drinking PBSA was positively associated with alcohol-related negative consequences for men but not for women. Results also showed negative associations between stopping and limiting drinking PBSA and alcohol related negative consequences for individuals with high levels of drinking refusal self-efficacy but not low levels of drinking refusal self-efficacy. It appears that addressing drinking refusal self-efficacy within the context of PBSA is valuable for traditional college students.

Optimizing accuracy of sampling protocols to measure nutrient content of solid manure.

Precise applications of manure to cropland can help optimize productivity and minimize environmental nutrient losses. Applying manure precisely is a challenge because the nutrient content of manures is inherently variable and the accuracy of sampling protocols are unknown. This study aimed to quantify the accuracy of sampling protocols for static solid manure piles considering both the number and depth of grab samples entering a composite sample. Over 35 grab samples were collected from each of ten static piles of dairy manure in California's Central Valley. Grab samples were individually analyzed for dry matter (DM), ash, total nitrogen, potassium, and phosphorous concentrations. Resampling simulations quantified the precision and bias of sampling protocols varying in both grab sample number and depth. Results showed that number of grab samples required for measurements to meet an accuracy standard of ±10% of the true value varied significantly by pile makeup. Over 25 grab samples were often required for multi-source manure piles, where an average of six grab samples were required from single source piles. The DM concentration of manure piles decreased at depths greater than 0.4 m, and sampling simulations showed that measurements were biased unless 70-80% of grab samples were collected from the pile interior. Both the number and location of grab samples necessary to create a representative composite require resource investments by farmers, and should be considered to manage nutrient applications cropland.

Optimizing accuracy of protocols for measuring dry matter and nutrient yield of forage crops.

Farmers around the world must precisely manage nutrients applied to and removed from crop fields to maintain production and without causing nutrient pollution. This study is the first to quantify the baseline accuracy of current industry measurement protocols and achievable accuracy from intensifying protocols for measuring dry matter (DM), nitrogen (N), potassium (K), and phosphorus (P) yields from forage crops harvested for silage. The 'true' DM and nutrient yields of three fields each of corn, sorghum, and small grain were intensively measured by weighing and sampling every truckload of harvested forage. Simulations quantified the accuracy of practical sampling protocols by repeatedly subsampling the complete dataset for each field to measure average truckload weight and average DM and nutrient concentrations. Then uncertainty was propagated to DM, N, P, and K yield calculations using standard error equations. Yields measured using current industry protocols diverged from the true yields of some fields by more than ±40%, emphasizing the need for improved protocols. This study shows that improving average DM and nutrient concentration measurements is unlikely to improve accuracy of yield measurements if average load weight is not precisely measured. Accuracy did not come within 27% of true yields without weighing all truckloads on some fields even when DM and nutrient concentration measurements were perfectly accurate. Once all truckloads were weighed, the timing of forage sample collection to measure average DM concentration had the greatest impact on accuracy; precision improved by an average of 6.2% when >3 samples were evenly spaced throughout the harvest compared to the same number of consecutive samples. All crop fields are affected by within field variation in growing conditions that results in heterogeneity in DM and nutrient yield. Globally, this study provides foundational methodology to quantitatively evaluate and improve yield measurement protocols that ultimately support sustainable crop production.

Mass balance analyses of nutrients on California dairies to evaluate data quality for regulatory review.

Effective regulations may help reduce nitrate contamination of groundwater from agriculture. Dairy farmers in California must maintain a ratio below 1.4 of total nitrogen (N) applied to total N-removed (N-Ratio) on cropland receiving manure application. In annual reports to the regulatory agency, farmers detail nutrients applied to cropland, removed in harvests, and exported off farm. Data were extracted from all available annual reports for 62 dairies from 2011, 2012, and 2013. Excretions of N, phosphorus (P), and potassium (K) were calculated using reported herd demographics and standard excretion equations from the American Society of Agricultural and Biological Engineers. Calculated nutrient excretion values were compared to the reported values of manure nutrients applied to cropland and exported off farm. Reported N-Ratios were compared to mass balance simulations exploring variable crop yields and alfalfa management. In the nutrient excretion balance, the distribution of the percent of N and P recovered in manures applied or exported peaked at 24% (median=31%) and 26% (median=53%) of excreted, respectively. The distribution of recovered K was fairly uniform from 0% to 300% (median=146%) of excreted K. In N-ratio simulations, 62% and 66% of all reported N-ratios were lower than their respective simulated N-ratio, assuming alfalfa crops received no N fertilization and minimal fertilization (26% of N-removed in harvest) respectively. When simulated crop yields were normally (sd=0.25) or Student's t distributed (df=154) around expected crop yields, 28% and 57% of all reported ratios fell within the 95% confidence interval of the simulations, respectively. Low and erratic recovery rates of excreted P and K existed. Additionally, reported N-Ratios were generally lower and more varied than necessary for farmers to maintain crop yields while complying with regulations. Greater understanding of low recovery rates is needed before data are used to assess the impact of regulations.

EGLN1 Inhibition and Rerouting of α-Ketoglutarate Suffice for Remote Ischemic Protection.

Ischemic preconditioning is the phenomenon whereby brief periods of sublethal ischemia protect against a subsequent, more prolonged, ischemic insult. In remote ischemic preconditioning (RIPC), ischemia to one organ protects others organs at a distance. We created mouse models to ask if inhibition of the alpha-ketoglutarate (αKG)-dependent dioxygenase Egln1, which senses oxygen and regulates the hypoxia-inducible factor (HIF) transcription factor, could suffice to mediate local and remote ischemic preconditioning. Using somatic gene deletion and a pharmacological inhibitor, we found that inhibiting Egln1 systemically or in skeletal muscles protects mice against myocardial ischemia-reperfusion (I/R) injury. Parabiosis experiments confirmed that RIPC in this latter model was mediated by a secreted factor. Egln1 loss causes accumulation of circulating αKG, which drives hepatic production and secretion of kynurenic acid (KYNA) that is necessary and sufficient to mediate cardiac ischemic protection in this setting.

Antigen- and cytokine-driven accumulation of regulatory T cells in visceral adipose tissue of lean mice.

A unique population of Foxp3(+)CD4(+) regulatory T (Treg) cells, with a distinct transcriptome and antigen-receptor repertoire, resides in visceral adipose tissue (VAT) of lean individuals. These cells regulate local inflammation and both local and systemic metabolic indices. Here we focus on expansion of the VAT Treg compartment in aging lean mice-assessing these cells' phenotypic conversion from conventional CD4(+) T cells, influx from lymphoid organs, and local population dynamics. Our findings establish that the VAT Treg compartment is seeded from thymocytes generated during the first weeks of life and expands beyond 10 weeks of age due to indolent proliferation, of certain clones in particular, coupled with enhanced survival. Accumulation of VAT Tregs depends on the antigen(s) presented by MHC class-II molecules and soluble mediators, notably interleukin(IL)-33. Addressing such factors therapeutically promises novel approaches for harnessing Tregs to stem the growing epidemic of obesity and consequent metabolic abnormalities.

Young, proliferative thymic epithelial cells engraft and function in aging thymuses.

The thymus reaches its maximum size early in life and then begins to shrink, producing fewer T cells with increasing age. This thymic decline is thought to contribute to age-related T cell lymphopenias and hinder T cell recovery after bone marrow transplantation. Although several cellular and molecular processes have been implicated in age-related thymic involution, their relative contributions are not known. Using heterochronic parabiosis, we observe that young circulating factors are not sufficient to drive regeneration of the aged thymus. In contrast, we find that resupplying young, engraftable thymic epithelial cells (TECs) to a middle-aged or defective thymus leads to thymic growth and increased T cell production. Intrathymic transplantation and in vitro colony-forming assays reveal that the engraftment and proliferative capacities of TECs diminish early in life, whereas the receptivity of the thymus to TEC engraftment remains relatively constant with age. These results support a model in which thymic growth and subsequent involution are driven by cell-intrinsic changes in the proliferative capacity of TECs, and further show that young TECs can engraft and directly drive the growth of involuted thymuses.

Diminished Schwann cell repair responses underlie age-associated impaired axonal regeneration.

The regenerative capacity of the peripheral nervous system declines with age. Why this occurs, however, is unknown. We demonstrate that 24-month-old mice exhibit an impairment of functional recovery after nerve injury compared to 2-month-old animals. We find no difference in the intrinsic growth capacity between aged and young sensory neurons in vitro or in their ability to activate growth-associated transcriptional programs after injury. Instead, using age-mismatched nerve transplants in vivo, we show that the extent of functional recovery depends on the age of the nerve graft, and not the age of the host. Molecular interrogation of the sciatic nerve reveals that aged Schwann cells (SCs) fail to rapidly activate a transcriptional repair program after injury. Functionally, aged SCs exhibit impaired dedifferentiation, myelin clearance, and macrophage recruitment. These results suggest that the age-associated decline in axonal regeneration results from diminished Schwann cell plasticity, leading to slower myelin clearance.

Vascular and neurogenic rejuvenation of the aging mouse brain by young systemic factors.

In the adult central nervous system, the vasculature of the neurogenic niche regulates neural stem cell behavior by providing circulating and secreted factors. Age-related decline of neurogenesis and cognitive function is associated with reduced blood flow and decreased numbers of neural stem cells. Therefore, restoring the functionality of the niche should counteract some of the negative effects of aging. We show that factors found in young blood induce vascular remodeling, culminating in increased neurogenesis and improved olfactory discrimination in aging mice. Further, we show that GDF11 alone can improve the cerebral vasculature and enhance neurogenesis. The identification of factors that slow the age-dependent deterioration of the neurogenic niche in mice may constitute the basis for new methods of treating age-related neurodegenerative and neurovascular diseases.

Growth differentiation factor 11 is a circulating factor that reverses age-related cardiac hypertrophy.

The most common form of heart failure occurs with normal systolic function and often involves cardiac hypertrophy in the elderly. To clarify the biological mechanisms that drive cardiac hypertrophy in aging, we tested the influence of circulating factors using heterochronic parabiosis, a surgical technique in which joining of animals of different ages leads to a shared circulation. After 4 weeks of exposure to the circulation of young mice, cardiac hypertrophy in old mice dramatically regressed, accompanied by reduced cardiomyocyte size and molecular remodeling. Reversal of age-related hypertrophy was not attributable to hemodynamic or behavioral effects of parabiosis, implicating a blood-borne factor. Using modified aptamer-based proteomics, we identified the TGF-ß superfamily member GDF11 as a circulating factor in young mice that declines with age. Treatment of old mice to restore GDF11 to youthful levels recapitulated the effects of parabiosis and reversed age-related hypertrophy, revealing a therapeutic opportunity for cardiac aging.

Sarcomas induced in discrete subsets of prospectively isolated skeletal muscle cells.

Soft-tissue sarcomas are heterogeneous cancers that can present with tissue-specific differentiation markers. To examine the cellular basis for this histopathological variation and to identify sarcoma-relevant molecular pathways, we generated a chimeric mouse model in which sarcoma-associated genetic lesions can be introduced into discrete, muscle-resident myogenic and mesenchymal cell lineages. Expression of Kirsten rat sarcoma viral oncogene [Kras(G12V)] and disruption of cyclin-dependent kinase inhibitor 2A (CDKN2A; p16p19) in prospectively isolated satellite cells gave rise to pleomorphic rhabdomyosarcomas (MyoD-, Myogenin- and Desmin-positive), whereas introduction of the same oncogenetic hits in nonmyogenic progenitors induced pleomorphic sarcomas lacking myogenic features. Transcriptional profiling demonstrated that myogenic and nonmyogenic Kras; p16p19(null) sarcomas recapitulate gene-expression signatures of human rhabdomyosarcomas and identified a cluster of genes that is concordantly up-regulated in both mouse and human sarcomas. This cluster includes genes associated with Ras and mechanistic target of rapamycin (mTOR) signaling, a finding consistent with activation of the Ras and mTOR pathways both in Kras; p16p19(null) sarcomas and in 26-50% of human rhabdomyosarcomas surveyed. Moreover, chemical inhibition of Ras or mTOR signaling arrested the growth of mouse Kras; p16p19(null) sarcomas and of human rhabdomyosarcoma cells in vitro and in vivo. Taken together, these data demonstrate the critical importance of lineage commitment within the tumor cell-of-origin in determining sarcoma histotype and introduce an experimental platform for rapid dissection of sarcoma-relevant cellular and molecular events.

Plexiform-like lesions and increased tissue factor expression in a rat model of severe pulmonary arterial hypertension.

Severe pulmonary arterial hypertension (PAH) occurs in idiopathic form and in association with diverse diseases. The pathological hallmarks are distal smooth muscle hypertrophy, obliteration of small pulmonary arteriole lumens, and disorganized cellular proliferation in plexiform lesions. In situ thrombosis is also observed. A detailed understanding of the disease progression has been hampered by the absence of an animal model bearing all the pathological features of human disease. To create a model with these characteristics, we gave young (200-g) rats monocrotaline 1 wk following left pneumonectomy; controls with vehicle treatment or sham operation were also studied. In experimental rats, pulmonary arteries had distal smooth muscle hypertrophy and proliferative perivascular lesions. The lesions had a plexiform appearance, occurred early in disease development, and were composed of cells expressing endothelial antigens. Three-dimensional microangiography revealed severe vascular pruning and disorganized vascular networks. We found that expression of tissue factor (TF), the membrane glycoprotein that initiates coagulation, facilitates angiogenesis, and mediates arterial injury in the systemic circulation, was increased in the pulmonary arterioles and plexiform-like lesions of the rats. TF was also heavily expressed in the vessels and plexiform lesions of humans with pulmonary arterial hypertension. We conclude that plexiform-like lesions can be reproduced in rats, and this model will facilitate experiments to address controversies about the role of these lesions in PAH. Increased TF expression may contribute to the prothrombotic diathesis and vascular cell proliferation typical of human disease.