The intricate cellular ecosystem of human peripheral veins as revealed by single-cell transcriptomic analysis.

The venous system has been historically understudied despite its critical roles in blood distribution, heart function, and systemic immunity. This study dissects the microanatomy of upper arm veins at the single cell level, and how it relates to wall structure, remodeling processes, and inflammatory responses to injury. We applied single-cell RNA sequencing to 4 non-diseased human veins (3 basilic, 1 cephalic) obtained from organ donors, followed by bioinformatic and histological analyses. Unsupervised clustering of 20,006 cells revealed a complex ecosystem of endothelial cell (EC) types, smooth muscle cell (SMCs) and pericytes, various types of fibroblasts, and immune cell populations. The venous endothelium showed significant upregulation of cell adhesion genes, with arteriovenous zonation EC phenotypes highlighting the heterogeneity of vasa vasorum (VV) microvessels. Venous SMCs had atypical contractile phenotypes and showed widespread localization in the intima and media. MYH11+DESlo SMCs were transcriptionally associated with negative regulation of contraction and pro-inflammatory gene expression. MYH11+DEShi SMCs showed significant upregulation of extracellular matrix genes and pro-migratory mediators. Venous fibroblasts ranging from secretory to myofibroblastic phenotypes were 4X more abundant than SMCs and widely distributed throughout the wall. Fibroblast-derived angiopoietin-like factors were identified as versatile signaling hubs to regulate angiogenesis and SMC proliferation. An abundant monocyte/macrophage population was detected and confirmed by histology, including pro-inflammatory and homeostatic phenotypes, with cell counts positively correlated with age. Ligand-receptor interactome networks identified the venous endothelium in the main lumen and the VV as a niche for monocyte recruitment and infiltration. This study underscores the transcriptional uniqueness of venous cells and their relevance for vascular inflammation and remodeling processes. Findings from this study may be relevant for molecular investigations of upper arm veins used for vascular access creation, where single-cell analyses of cell composition and phenotypes are currently lacking.

Increasing Chlamydia and Gonorrhea Testing for Adolescents in the Pediatric Emergency Department.

OBJECTIVE: Adolescents who use the emergency department are more likely to engage in high-risk sexual activity and are at an increased risk of sexually transmitted infections. We aimed to increase testing for Chlamydia and gonorrhea from 12% to 50% among adolescents presenting to our pediatric emergency department with at-risk chief complaints over 12 months. METHODS: Plan-Do-Study-Act cycles were initiated in July 2020. A multidisciplinary team reviewed preexisting data and developed interventions to increase Chlamydia and gonorrhea testing in teens with at-risk complaints, including genitourinary and behavioral health complaints, and females with abdominal pain. Two categories of interventions were implemented: education and electronic medical record optimization. Process measures were the proportion of patients with a documented sexual history and the proportion of patients tested with a documented confidential phone number. Secondary outcome measures included the weekly number of positive test results and the proportion of patients testing positive who were contacted to arrange treatment. Statistical process control charts were used to examine changes in measures over time. RESULTS: Within 14 months of project initiation, the proportion of at-risk patients tested increased from 12% to 59%. Teen phone number documentation remained unchanged from 23%. Sexual history documentation remained unchanged from 46%. The number of positive test results increased from 1.8 to 3.4 per month, and the proportion of patients testing positive who were contacted to arrange treatment remained unchanged at 83%. CONCLUSIONS: We surpassed our goal and increased the proportion of at-risk patients tested for Chlamydia and gonorrhea to 59%, sustained for 4 months from the last intervention.

Eating Disorders in Youth with Chronic Health Conditions: Clinical Strategies for Early Recognition and Prevention.

Youth with chronic health conditions face an elevated risk of eating disorders and disordered eating behaviors. Contributors to this phenomenon may include the unique threats faced by this vulnerable population to their body image, their relationships with food and eating, and their mental health and self-esteem. However, youth with chronic health conditions may also experience more severe medical complications and mortality from eating disorder behaviors because of the additional risks conveyed by their underlying conditions. In this review, clinical strategies are provided to support youth with chronic health conditions through early recognition of eating disorder behaviors and prompt referral to treatment, which is important for a better prognosis. Suggestions are also given to mitigate their risk of developing eating disorders by proactively addressing risk factors and offering thoughtful anticipatory guidance that promotes a positive relationship with food and eating.

Repetitive schistosoma exposure causes perivascular lung fibrosis and persistent pulmonary hypertension.

BACKGROUND: Pulmonary hypertension (PH) can occur as a complication of schistosomiasis. In humans, schistosomiasis-PH persists despite antihelminthic therapy and parasite eradication. We hypothesized that persistent disease arises as a consequence of exposure repetition. METHODS: Following intraperitoneal sensitization, mice were experimentally exposed to Schistosoma eggs by intravenous injection, either once or three times repeatedly. The phenotype was characterized by right heart catheterization and tissue analysis. RESULTS: Following intraperitoneal sensitization, a single intravenous Schistosoma egg exposure resulted in a PH phenotype that peaked at 7-14 days, followed by spontaneous resolution. Three sequential exposures resulted in a persistent PH phenotype. Inflammatory cytokines were not significantly different between mice exposed to one or three egg doses, but there was an increase in perivascular fibrosis in those who received three egg doses. Significant perivascular fibrosis was also observed in autopsy specimens from patients who died of this condition. CONCLUSIONS: Repeatedly exposing mice to schistosomiasis causes a persistent PH phenotype, accompanied by perivascular fibrosis. Perivascular fibrosis may contribute to the persistent schistosomiasis-PH observed in humans with this disease.

Malnutrition from anorexia nervosa triggers severe complications of hereditary spherocytosis in an adolescent girl: A case report.

Hemolytic crises and aplastic crises in hereditary spherocytosis (HS) are most commonly triggered by viral infections. We present the case of an adolescent girl with HS who developed unexpected and life-threatening complications of her inherited hemolytic anemia as a consequence of anorexia nervosa and severe malnutrition.

Maternal immune response and placental antibody transfer after COVID-19 vaccination across trimester and platforms.

The availability of three COVID-19 vaccines in the United States provides an unprecedented opportunity to examine how vaccine platforms and timing of vaccination in pregnancy impact maternal and neonatal immunity. Here, we characterize the antibody profile after Ad26.COV2.S, mRNA-1273 or BNT162b2 vaccination in 158 pregnant individuals and evaluate transplacental antibody transfer by profiling maternal and umbilical cord blood in 175 maternal-neonatal dyads. These analyses reveal lower vaccine-induced functions and Fc receptor-binding after Ad26.COV2.S compared to mRNA vaccination and subtle advantages in titer and function with mRNA-1273 versus BN162b2. mRNA vaccines have higher titers and functions against SARS-CoV-2 variants of concern. First and third trimester vaccination results in enhanced maternal antibody-dependent NK-cell activation, cellular and neutrophil phagocytosis, and complement deposition relative to second trimester. Higher transplacental transfer ratios following first and second trimester vaccination may reflect placental compensation for waning maternal titers. These results provide novel insight into the impact of platform and trimester of vaccination on maternal humoral immune response and transplacental antibody transfer.

Chronic Daily House Dust Mite Exposure in Mice is an Effective Model to Quantify the Effect of Pharmacologic Agents on Discrete Stages of Artery Remodeling in Pulmonary Hypertension.

Pulmonary hypertension (PH) is a heterogenous and incurable disease marked by varying degrees of pulmonary vascular remodeling. This vascular remodeling, which includes thickening of the smooth muscle layer (an early finding) and formation of occlusive neointimal lesions (a late finding) in the pulmonary arteries, is a major driver of morbidity and mortality in PH. Available PH therapies consist of vasodilators that do not specifically target lesion formation or expansion and neither prevent progression nor reverse disease. This paucity of curative treatments highlights the need for new drug discovery targeting crucial steps of artery remodeling in PH. The cell dynamics and molecular signals driving neointimal lesion formation have been difficult to elucidate as classic mouse models of PH do not develop neointima. Here, we detail the methods to generate a robust and non-genetic mouse model of PH with medial thickening and neointimal lesion formation in the pulmonary arteries, through chronic exposure to an inflammatory stimulus-house dust mite (HDM). This model rapidly generates human-like pulmonary arterial lesions following a reproducible time course, allowing scrutiny of the cellular and molecular mechanisms controlling each stage of artery remodeling. Further, we outline optimal tissue handling, sectioning, and staining methodologies for detailed quantitative analysis of artery medial thickening and neointimal lesion formation and expansion. Finally, we present a method for staged pharmacologic intervention to identify molecules and pathways required at each step of the pulmonary arterial remodeling process. The advantages of this mouse model of PH over currently available animal models are five-fold. (i) It allows the use of the full range of genetic and single cell tools available in mice to manipulate and study the process of vascular remodeling seen in human disease, including the formation of neointimal lesions in a controlled and cell specific manner. (ii) The vascular lesions develop in a stereotyped manner with predictable timing, allowing for pharmacologic manipulation at discrete stages of vessel remodeling. (iii) It is rapid, with development of PH and vascular remodeling in a timeframe of two to eight weeks. (iv) It uses simple techniques and requires neither surgery, unusual equipment, or extensive personnel training. (v) The staining and quantitation methodologies we present are a significant improvement over those currently in use in the field. We hope that dissemination of this model and the associated detailed methods will speed up the development of novel and more effective PH therapeutics. Graphic abstract: Chronic perivascular inflammation induces medial thickening and neointima formation in pulmonary arteries, following a stereotyped time course, and allowing staged pharmacologic intervention during specific remodeling events, as well as quantitative assessment of vascular changes.

A Notch3-Marked Subpopulation of Vascular Smooth Muscle Cells Is the Cell of Origin for Occlusive Pulmonary Vascular Lesions.

BACKGROUND: Pulmonary arterial hypertension (PAH) is a fatal disease characterized by profound vascular remodeling in which pulmonary arteries narrow because of medial thickening and occlusion by neointimal lesions, resulting in elevated pulmonary vascular resistance and right heart failure. Therapies targeting the neointima would represent a significant advance in PAH treatment; however, our understanding of the cellular events driving neointima formation, and the molecular pathways that control them, remains limited. METHODS: We comprehensively map the stepwise remodeling of pulmonary arteries in a robust, chronic inflammatory mouse model of pulmonary hypertension. This model demonstrates pathological features of the human disease, including increased right ventricular pressures, medial thickening, neointimal lesion formation, elastin breakdown, increased anastomosis within the bronchial circulation, and perivascular inflammation. Using genetic lineage tracing, clonal analysis, multiplexed in situ hybridization, immunostaining, deep confocal imaging, and staged pharmacological inhibition, we define the cell behaviors underlying each stage of vascular remodeling and identify a pathway required for neointima formation. RESULTS: Neointima arises from smooth muscle cells (SMCs) and not endothelium. Medial SMCs proliferate broadly to thicken the media, after which a small number of SMCs are selected to establish the neointima. These neointimal founder cells subsequently undergoing massive clonal expansion to form occlusive neointimal lesions. The normal pulmonary artery SMC population is heterogeneous, and we identify a Notch3-marked minority subset of SMCs as the major neointimal cell of origin. Notch signaling is specifically required for the selection of neointimal founder cells, and Notch inhibition significantly improves pulmonary artery pressure in animals with pulmonary hypertension. CONCLUSIONS: This work describes the first nongenetically driven murine model of pulmonary hypertension (PH) that generates robust and diffuse occlusive neointimal lesions across the pulmonary vascular bed and does so in a stereotyped timeframe. We uncover distinct cellular and molecular mechanisms underlying medial thickening and neointima formation and highlight novel transcriptional, behavioral, and pathogenic heterogeneity within pulmonary artery SMCs. In this model, inflammation is sufficient to generate characteristic vascular pathologies and physiological measures of human PAH. We hope that identifying the molecular cues regulating each stage of vascular remodeling will open new avenues for therapeutic advancements in the treatment of PAH.

A Brief Provider Training Video Improves Comfort With Recommending the Human Papillomavirus Vaccine.

The American Academy of Pediatrics California Chapter 3 created a 20-minute training video targeting barriers to strong provider recommendation of the human papillomavirus (HPV) vaccine. The video included clinical vignettes featuring pediatricians modeling counseling techniques with vaccine-hesitant families. Ninety-six multidisciplinary providers (including pediatric residents) at 6 sites viewed the video and completed baseline and posttest questionnaires assessing their vaccine knowledge, attitudes toward vaccination, and comfort with skills needed to facilitate vaccination. Following the intervention, providers had substantial and statistically significant ( P < .05) improvements in multiple areas assessed, particularly knowledge of the burden of HPV-related disease in males and changes in vaccine response with age; likelihood of "strongly agreeing" that vaccination should not be delayed beyond preadolescence and that HPV vaccine is safe; and feeling "very comfortable" counseling vaccine-hesitant parents and facilitating vaccine completion. This cost-effective and easily disseminated training modality shows promise in increasing provider comfort with HPV vaccine counseling.

Severe iodine deficiency from dietary restriction and subsequent iodine excess from seaweed snack overuse in an adolescent with disordered eating.

Individuals with disordered eating commonly exclude salt and animal products from their diets, which may predispose them to iodine deficiency even without significant weight loss. Overconsumption of dietary supplements and "natural" foods are also commonly observed among eating disorder patients. This report describes an adolescent female with disordered eating presenting with growth and pubertal delay, found to have severe iodine deficiency (urine iodine of 18 mcg/L) and abnormal thyroid function resulting from strict avoidance of salt and animal products despite adequate caloric intake. To avoid having to start eating animal products, she began consuming excessive quantities of seaweed supplements to increase her iodine intake resulting in excessive iodine levels (urine iodine of >1,500 mcg/L) and worsening of thyroid function. When her parents began supervising her nutritional intake, her abnormal iodine levels, abnormal thyroid function tests, and hypogonadotropic hypogonadism all resolved. This is the first report of both iodine deficiency and iodine excess developing from disordered eating behavior.

Inattentive Attention-Deficit/Hyperactivity Disorder, Stimulant Medication, and Weight Loss in a 15-Year-Old Girl: Are We Enabling the Development of an Eating Disorder?

CASE: Nicole is a 15-year-old girl presenting to the Developmental Behavioral Pediatrics Clinic with symptoms of the inattentive type of Attention-Deficit/Hyperactivity Disorder (ADHD) and declining school performance over the last year. She expressed frustration over her inability to concentrate on schoolwork. Assuming that her poor grades were secondary to lack of effort, her parents withdrew privileges. Nicole became increasingly depressed. She stopped participating in activities, she previously enjoyed, and her parents reported that she stopped singing in the shower. After talking to a cousin with ADHD, Nicole concluded that she had ADHD as well. She asked her parents to arrange for an evaluation.Nicole met DSM-5 criteria for the diagnosis of inattentive ADHD and was started on a stimulant medication (mixed amphetamine salts). She had symptoms of a coexisting depression, although she did not meet criteria for diagnosis of a depressive disorder. At a 3-week follow-up visit, she showed improvement in targeted ADHD symptoms; homework was now easier and her grades improved. At a 2-month follow-up, Nicole's weight dropped from 53 kg (47th percentile) prestimulant treatment to 49 kg (31st percentile). She reported appetite suppression after taking the stimulant but did not feel that her eating habits had changed significantly. Her father reported that she had a preference for junk food and snacks. Nicole did not enjoy exercising and did not participate in extracurricular sports.She weighed herself several times a day, as she was worried about losing too much weight. Nicole's mood continued to be low, despite the fact that her grades improved, and her parents were more understanding of her challenges. She was otherwise healthy and reported regular menstrual cycles. Nicole requested an increase in the dose of stimulant medication for greater improvement in concentration during homework and in school.Her pediatric clinician was concerned about the possibility of an eating disorder in addition to depression. She asked herself, "Are we treating inattentive ADHD effectively or are we enabling an eating disorder?"

Outcomes of an inpatient refeeding protocol in youth with anorexia nervosa: Rady Children's Hospital San Diego/University of California, San Diego.

BACKGROUND: Current guidelines for nutritional rehabilitation in hospitalized restrictive eating disorder patients recommend a cautious approach to refeeding. Several studies suggest that higher calorie diets may be safe and effective, but have traditionally excluded severely malnourished patients. The goal of this study was to evaluate the safety of a higher calorie nutritional rehabilitation protocol (NRP) in a broad sample of inpatients with restrictive eating disorders, including those who were severely malnourished. METHODS: A retrospective chart review was conducted among eating disorder inpatients between January 2015 and March 2016. Patients were started on a lower calorie diet (≤1500 kcals/day) or higher calorie diet (≥1500 kcals/day). Calorie prescription on admission was based on physician clinical judgement. The sample included patients aged 8-20 years with any DSM-5 restrictive eating disorder. Those who were severely malnourished (<75% expected body weight [EBW]) or required tube feeding during admission were included. Multivariable regression models were used to determine whether level of nutritional rehabilitation was associated with hypophosphatemia, hypomagnesemia, or hypokalemia. RESULTS: The sample included 87 patients; mean age was 14.4 years (S.D. 32.7); 29% were <75% EBW. The majority (75.8%) was started on higher calorie diets (mean 1781 kcal/day). Controlling for rate of calorie change, initial %EBW, age, race/ethnicity, insurance, diagnosis, and NG/NJ tube placement, higher calorie diets were not associated with hypophosphatemia, hypomagnesemia, or hypokalemia on admission or within the first 72 h. Increased risk of hypophosphatemia on admission was associated with lower baseline %EBW. CONCLUSION: A higher calorie NRP was tolerated in this broad population of inpatients with restrictive eating disorders. Lower %EBW on admission was a more important predictor of hypophosphatemia than initial calorie level. Larger studies are required to demonstrate the safety of higher calorie diets in severely malnourished patients.

Chronic Headaches After a Concussion in an Obese 16-Year-Old Girl.

CASE: Jennifer is a 16-year-old Latina girl who is new to your practice. During her first well visit, she mentions that she has had daily headaches for 2 years. They began after sustaining a concussion in a car accident. Typically, her headaches are bilateral and "squeezing"; they occur in the afternoons and last for a few hours. Her concussion also resulted in depressed mood, which has improved over time.When you ask if her headaches have changed recently, she says that they have been worse for the last few days. The quality and severity are unchanged; however, they now occur first thing in the morning, are worse when supine, and no longer remit. In the last 2 days, she has developed new-onset blurry vision, nausea, dizziness, photophobia, and sonophobia. Although she previously experienced sadness with her concussion, she now feels irritable. She has never used tobacco, alcohol, or drugs, and she takes no medications.On examination, her body mass index is above the 99th percentile. You note mild papilledema bilaterally. She has no focal neurological deficits. The remainder of her examination is normal.You send her to an emergency department. Her head computed tomography is normal. A lumbar puncture demonstrates an opening pressure of 32 cm H2O; she feels relief after the procedure. She is admitted with a diagnosis of benign intracranial hypertension and is started on acetazolamide. What is the differential diagnosis of chronic headaches in an obese adolescent? How should a busy community pediatrician manage Jennifer acutely? What follow-up care should Jennifer receive?

Transcription factor TBX4 regulates myofibroblast accumulation and lung fibrosis.

Progressive tissue fibrosis is a major cause of the morbidity and mortality associated with repeated epithelial injuries and accumulation of myofibroblasts. Successful treatment options are limited by an incomplete understanding of the molecular mechanisms that regulate myofibroblast accumulation. Here, we employed in vivo lineage tracing and real-time gene expression transgenic reporting methods to analyze the early embryonic transcription factor T-box gene 4 (TBX4), and determined that TBX4-lineage mesenchymal progenitors are the predominant source of myofibroblasts in injured adult lung. In a murine model, ablation of TBX4-expressing cells or disruption of TBX4 signaling attenuated lung fibrosis after bleomycin-induced injury. Furthermore, TBX4 regulated hyaluronan synthase 2 production to enable fibroblast invasion of matrix both in murine models and in fibroblasts from patients with severe pulmonary fibrosis. These data identify TBX4 as a mesenchymal transcription factor that drives accumulation of myofibroblasts and the development of lung fibrosis. Targeting TBX4 and downstream factors that regulate fibroblast invasiveness could lead to therapeutic approaches in lung fibrosis.