"Stuck in the Middle with You": intermediate cell states are not always in transition.

The era of single-cell multiomics has led to the identification of lung epithelial cells with features of both alveolar type 1 (AT1) and alveolar type 2 (AT2) pneumocytes, leading many to infer that these cells are a distinct cell type in the process of transitioning between AT2 and AT1 cells. In this issue of the JCI, Wang and colleagues demonstrated that many so-called "transitional cells" do not actually contribute to functional repair. The findings warrant a reimagining of these cells as existing in a nondirectional, intermediate cell state, rather than moving through a transitory process from one cell type to another. We look forward to further exploration of diverse cell state expression profiles and a more refined examination of hallmark gene function beyond population labeling.

Masters of Media: A longitudinal study of parental media efficacy, media monitoring, and child problematic media use across early childhood in the United States.

The development of problematic media use in early childhood is not well understood. The current study examined long-term associations between parental media efficacy, parental media monitoring, and problematic media use across a three-year period of time during early childhood. Participants included 432 parents who reported on their own parenting and their child's use of problematic media once a year for three years (M age of child at Wave 1 = 29.68 months, SD = 3.73 months). Results revealed that early parental media efficacy predicted lower levels of child problematic media use over time. Restrictive media monitoring was also related to lower levels of child problematic media use over time. Additionally, general parental efficacy was related to parental media efficacy and lower child problematic media use, both at the cross-sectional and longitudinal levels. Discussion focuses on encouraging early parental media efficacy (and exploring other potential mechanisms) as a way to mitigate the development of problematic media use over time.

Epithelial Yap/Taz are required for functional alveolar regeneration following acute lung injury.

A hallmark of idiopathic pulmonary fibrosis (IPF) and other interstitial lung diseases is dysregulated repair of the alveolar epithelium. The Hippo pathway effector transcription factors YAP and TAZ are implicated as essential for type 1 and type 2 alveolar epithelial cell (AT1 and AT2) differentiation in the developing lung, yet aberrant activation of YAP/TAZ is a prominent feature of the dysregulated alveolar epithelium in IPF. In these studies, we sought to define the functional role of YAP/TAZ activity during alveolar regeneration. We demonstrated that Yap and Taz were normally activated in AT2 cells shortly after injury, and deletion of Yap/Taz in AT2 cells led to pathologic alveolar remodeling, failure of AT2-to-AT1 cell differentiation, increased collagen deposition, exaggerated neutrophilic inflammation, and increased mortality following injury induced by a single dose of bleomycin. Loss of Yap/Taz activity prior to an LPS injury prevented AT1 cell regeneration, led to intraalveolar collagen deposition, and resulted in persistent innate inflammation. These findings establish that AT2 cell Yap/Taz activity is essential for functional alveolar epithelial repair and prevention of fibrotic remodeling.

Can we evidence-base injury prevention and management in women's football? A scoping review.

This review aimed, as part of a larger FIFA project aiming to steer women's football research, to scope literature on any level of competitive football for women, to understand the current quantity of research on women's football injuries. The study reviewed all injury-related papers scoped by a recent scoping review mapping all published women's football research with an updated search performed on 23 February 2021. Eligibility criteria assessment followed the recent scoping review with injury-specific research focus. A total of 497 studies were scoped. Most studies contained an epidemiological (N = 226; 45%) or risk factors assessment (N = 105; 21%). Less assessed areas included financial burden (N = 1; <1%) and injury awareness (N = 5; 1%). 159 studies (32%) assessed injuries of the whole body. The most common single location assessed in the literature was the knee (N = 134, 27%), followed by head/face (N =108, 22%). These numbers were, however, substantially lowered, when subdivided by playing level and age-group. The volume of research focuses especially on descriptive research and specific body locations (head/face and knee). Although information can be taken from studies in other sports, more football-specific studies to support management and prevention of injuries are warranted.

Image-based spatial transcriptomics identifies molecular niche dysregulation associated with distal lung remodeling in pulmonary fibrosis.

The human lung is structurally complex, with a diversity of specialized epithelial, stromal and immune cells playing specific functional roles in anatomically distinct locations, and large-scale changes in the structure and cellular makeup of this distal lung is a hallmark of pulmonary fibrosis (PF) and other progressive chronic lung diseases. Single-cell transcriptomic studies have revealed numerous disease-emergent/enriched cell types/states in PF lungs, but the spatial contexts wherein these cells contribute to disease pathogenesis has remained uncertain. Using sub-cellular resolution image-based spatial transcriptomics, we analyzed the gene expression of more than 1 million cells from 19 unique lungs. Through complementary cell-based and innovative cell-agnostic analyses, we characterized the localization of PF-emergent cell-types, established the cellular and molecular basis of classical PF histopathologic disease features, and identified a diversity of distinct molecularly-defined spatial niches in control and PF lungs. Using machine-learning and trajectory analysis methods to segment and rank airspaces on a gradient from normal to most severely remodeled, we identified a sequence of compositional and molecular changes that associate with progressive distal lung pathology, beginning with alveolar epithelial dysregulation and culminating with changes in macrophage polarization. Together, these results provide a unique, spatially-resolved characterization of the cellular and molecular programs of PF and control lungs, provide new insights into the heterogeneous pathobiology of PF, and establish analytical approaches which should be broadly applicable to other imaging-based spatial transcriptomic studies.

Multiplatform Single-Cell Analysis Identifies Immune Cell Types Enhanced in Pulmonary Fibrosis.

Immune cells have been implicated in idiopathic pulmonary fibrosis (IPF), but the phenotypes and effector mechanisms of these cells remain incompletely characterized. We performed mass cytometry to quantify immune cell subsets in lungs of 12 patients with IPF and 15 organ donors without chronic lung disease and used existing single-cell RNA-sequencing data to investigate transcriptional profiles of immune cells overrepresented in IPF. Among myeloid cells, we found increased numbers of alveolar macrophages (AMØs) and dendritic cells (DCs) in IPF, as well as a subset of monocyte-derived DCs. In contrast, monocyte-like cells and interstitial macrophages were reduced in IPF. Transcriptomic profiling identified an enrichment for IFN-γ response pathways in AMØs and DCs from IPF, as well as antigen processing in DCs and phagocytosis in AMØs. Among T cells, we identified three subsets of memory T cells that were increased in IPF, including CD4+ and CD8+ resident memory T cells (TRM) and CD8+ effector memory cells. The response to the IFN-γ pathway was enriched in CD4 TRM and CD8 TRM cells in IPF, together with T cell activation and immune response-regulating signaling pathways. Increased AMØs, DCs, and memory T cells were present in IPF lungs compared with control subjects. In IPF, these cells possess an activation profile indicating increased IFN-γ signaling and upregulation of adaptive immunity in the lungs. Together, these studies highlight critical features of the immunopathogenesis of IPF.

Awareness isn't saving lives: An experimental exploration of suicide risk reduction methods for emerging adults.

OBJECTIVE: Suicide prevention campaigns commonly employ brief informational materials aimed at emerging adults. Are such programs helpful, and do design characteristics yield differences in user outcomes? Literature is reviewed from the interpersonal theory of suicide, escape theory, and terror management theory, to inform our experimental design. METHOD: Participants (n = 977 MTurk emerging adults) reported demographics and suicide histories and were randomized to one of nine experimental cells with varying video and journaling conditions to approximate suicide prevention materials. Participants were surveyed on perceptions of the materials' risk reduction effectiveness, indicated their suicidality risk factors (e.g., hopelessness, depressiveness, purposelessness, and non-belongingness), and conducted an implicit association test of suicidality. RESULTS: Suicide risk factors did not differ between experimental and control conditions, but certain conditions were rated as more effective (i.e., essay conditions prompting reflection, and the video condition featuring a personal/affective narrative). While there was no actual comparative reduction of risk, there was a perception that certain designs were more helpful. CONCLUSION: Real-world suicide prevention campaigns often feel justified despite lacking impact. Effective suicide risk reduction requires greater time investment and deeper personal connection than brief campaigns can offer, as well as systemic changes from a public health policy perspective.

What Role Do Chronic Workloads Play in the Acute to Chronic Workload Ratio? Time to Dismiss ACWR and Its Underlying Theory.

AIM: The aim of this study was to examine the associations between the injury risk and the acute (AL) to chronic (CL) workload ratio (ACWR) by substituting the original CL with contrived values to assess the role of CL (i.e., the presence and implications of statistical artefacts). METHODS: Using previously published data, we generated a contrived ACWR by dividing the AL by fixed and randomly generated CLs, and we compared these results to real data. We also reproduced previously reported subgroup analyses, including dichotomising players' data above and below the median CL. Our analyses follow the same, previously published modelling approach. RESULTS: The analyses with original data showed effects compatible with higher injury risk for ACWR only (odd ratios, OR: 2.45, 95% CI 1.28-4.71). However, we observed similar effects by dividing AL by the "contrived" fixed and randomly generated CLs: OR 1.95 (1.18-3.52) dividing by 1510 (average CL); and OR ranging from 1.16 to 2.07, using random CL 1.53 (mean). Random ACWRs reduced the variance relative to the original AL and further inflated the ORs (mean OR 1.89, from 1.42 to 2.70). ACWR causes artificial reclassification of players compared to AL alone. Finally, neither ACWR nor AL alone confer a meaningful predictive advantage to an intercept-only model, even within the training sample (c-statistic 0.574/0.544 vs. 0.5 in both ACWR/AL and intercept-only models, respectively). DISCUSSION: ACWR is a rescaling of the explanatory variable (AL, numerator), in turn magnifying its effect estimates and decreasing its variance despite conferring no predictive advantage. Other ratio-related transformations (e.g., reducing the variance of the explanatory variable and unjustified reclassifications) further inflate the OR of AL alone with injury risk. These results also disprove the etiological theory behind this ratio and its components. We suggest ACWR be dismissed as a framework and model, and in line with this, injury frameworks, recommendations, and consensus be updated to reflect the lack of predictive value of and statistical artefacts inherent in ACWR models.

Parent-child joint media engagement in infancy.

Early conversations are an important source in shaping children's cognitive and emotional development, and it is vital to understand how parents use media as a platform to engage in conversations with their young children and what might predict the quality of these interactions. Thus, in the current study we explored the nature of parent-child discourse while engaging in media (i.e., joint media engagement) with infants, and how parent (empathic concern and responsiveness) and child (negative emotionality and regulatory capacity) variables might be associated with the quality of engagement. The current study consisted of 269 infants (50% female, Mage  = 17.09 months, SD = 3.93; 59% White) and their primary caregiver (98% mothers) who engaged in a variety of in-home tasks and parental questionnaires. Results established three meaningful codes for both parent and child that assessed positive and negative joint media engagement. Further, results suggested that parental empathic concern was associated with positive parent and child media engagement, while child negative emotionality was associated with lower levels of distraction. Discussion focuses on the importance of studying parent-child discourse in the context of joint media engagement and recommends limiting media exposure before 18 months of age.

Inhibitory Anti-Peroxidasin Antibodies in Pulmonary-Renal Syndromes.

BACKGROUND: Goodpasture syndrome (GP) is a pulmonary-renal syndrome characterized by autoantibodies directed against the NC1 domains of collagen IV in the glomerular and alveolar basement membranes. Exposure of the cryptic epitope is thought to occur via disruption of sulfilimine crosslinks in the NC1 domain that are formed by peroxidasin-dependent production of hypobromous acid. Peroxidasin, a heme peroxidase, has significant structural overlap with myeloperoxidase (MPO), and MPO-ANCA is present both before and at GP diagnosis in some patients. We determined whether autoantibodies directed against peroxidasin are also detected in GP. METHODS: We used ELISA and competitive binding assays to assess the presence and specificity of autoantibodies in serum from patients with GP and healthy controls. Peroxidasin activity was fluorometrically measured in the presence of partially purified IgG from patients or controls. Clinical disease severity was gauged by Birmingham Vasculitis Activity Score. RESULTS: We detected anti-peroxidasin autoantibodies in the serum of patients with GP before and at clinical presentation. Enriched anti-peroxidasin antibodies inhibited peroxidasin-mediated hypobromous acid production in vitro. The anti-peroxidasin antibodies recognized peroxidasin but not soluble MPO. However, these antibodies did crossreact with MPO coated on the polystyrene plates used for ELISAs. Finally, peroxidasin-specific antibodies were also found in serum from patients with anti-MPO vasculitis and were associated with significantly more active clinical disease. CONCLUSIONS: Anti-peroxidasin antibodies, which would previously have been mischaracterized, are associated with pulmonary-renal syndromes, both before and during active disease, and may be involved in disease activity and pathogenesis in some patients.

Extracellular chloride signals collagen IV network assembly during basement membrane formation.

Basement membranes are defining features of the cellular microenvironment; however, little is known regarding their assembly outside cells. We report that extracellular Cl(-) ions signal the assembly of collagen IV networks outside cells by triggering a conformational switch within collagen IV noncollagenous 1 (NC1) domains. Depletion of Cl(-) in cell culture perturbed collagen IV networks, disrupted matrix architecture, and repositioned basement membrane proteins. Phylogenetic evidence indicates this conformational switch is a fundamental mechanism of collagen IV network assembly throughout Metazoa. Using recombinant triple helical protomers, we prove that NC1 domains direct both protomer and network assembly and show in Drosophila that NC1 architecture is critical for incorporation into basement membranes. These discoveries provide an atomic-level understanding of the dynamic interactions between extracellular Cl(-) and collagen IV assembly outside cells, a critical step in the assembly and organization of basement membranes that enable tissue architecture and function. Moreover, this provides a mechanistic framework for understanding the molecular pathobiology of NC1 domains.

Stem Cell-Soluble Signals Enhance Multilumen Formation in SMG Cell Clusters.

Saliva plays a major role in maintaining oral health. Patients with salivary hypofunction exhibit difficulty in chewing and swallowing foods, tooth decay, periodontal disease, and microbial infections. At this time, treatments for hyposalivation are limited to medications (e.g., muscarinic receptor agonists: pilocarpine and cevimeline) that induce saliva secretion from residual acinar cells as well as artificial salivary substitutes. Therefore, advancement of restorative treatments is necessary to improve the quality of life in these patients. Our previous studies indicated that salivary cells are able to form polarized 3-dimensional structures when grown on growth factor-reduced Matrigel. This basement membrane is rich in laminin-III (L1), which plays a critical role in salivary gland formation. Mitotically inactive feeder layers have been used previously to support the growth of many different cell types, as they provide factors necessary for cell growth and organization. The goal of this study was to improve salivary gland cell differentiation in primary cultures by using a combination of L1 and a feeder layer of human hair follicle-derived mesenchymal stem cells (hHF-MSCs). Our results indicated that the direct contact of mouse submandibular (mSMG) cell clusters and hHF-MSCs was not required for mSMG cells to form acinar and ductal structures. However, the hHF-MSC conditioned medium enhanced cell organization and multilumen formation, indicating that soluble signals secreted by hHF-MSCs play a role in promoting these features.

External Responsiveness of the Yo-Yo IR Test Level 1 in High-level Male Soccer Players.

The aim of this study was to assess the external responsiveness, construct validity and internal responsiveness of the Yo-Yo Intermittent Recovery test level 1 and its sub-maximal version in semi-professional players. Tests and friendly matches were performed during the preseason and regular season. The distance covered above 15 km·h(-1) was considered as an indicator of the physical match performance. Construct validity and external responsiveness were examined by correlations between test and physical match performance (preseason and regular season) and training-induced changes. Internal responsiveness was determined as Cohen's effect size, standardized response mean and signal-to-noise ratio. The physical match performance increased after training (34.8%). The Yo-Yo Intermittent Recovery test level 1 improved after training (40.2%), showed longitudinal (r=0.69) and construct validity (r=0.73 and 0.59, preseason and regular season) and had higher internal responsiveness compared to its sub-maximal version. The heart rate at the 6(th) minute in the sub-maximal version did not show longitudinal (r=-0.38) and construct validity (r=0.01 and -0.06, preseason and regular season) and did not significantly change after training (-0.3%). The rate of perceived exertion decreased in the sub-maximal version (- 29.8%). In conclusion, the Yo-Yo Intermittent Recovery test level 1 is valid and responsive, while the validity of its sub-maximal version is questionable.

Bromine is an essential trace element for assembly of collagen IV scaffolds in tissue development and architecture.

Bromine is ubiquitously present in animals as ionic bromide (Br(-)) yet has no known essential function. Herein, we demonstrate that Br(-) is a required cofactor for peroxidasin-catalyzed formation of sulfilimine crosslinks, a posttranslational modification essential for tissue development and architecture found within the collagen IV scaffold of basement membranes (BMs). Bromide, converted to hypobromous acid, forms a bromosulfonium-ion intermediate that energetically selects for sulfilimine formation. Dietary Br deficiency is lethal in Drosophila, whereas Br replenishment restores viability, demonstrating its physiologic requirement. Importantly, Br-deficient flies phenocopy the developmental and BM defects observed in peroxidasin mutants and indicate a functional connection between Br(-), collagen IV, and peroxidasin. We establish that Br(-) is required for sulfilimine formation within collagen IV, an event critical for BM assembly and tissue development. Thus, bromine is an essential trace element for all animals, and its deficiency may be relevant to BM alterations observed in nutritional and smoking-related disease. PAPERFLICK:

Treating type 1 diabetes: from strategies for insulin delivery to dual hormonal control.

Type 1 diabetes is a disorder where slow destruction of pancreatic ß-cells occurs through autoimmune mechanisms. The result is a progressive and ultimately complete lack of endogenous insulin. Due to ß-cell lack, secondary abnormalities in glucagon and likely in incretins occur. These multiple hormonal abnormalities cause metabolic instability and extreme glycemic variability, which is the primary phenotype. As the disease progresses patients often develop hypoglycemia unawareness and defects in their counterregulatory defenses. Intensive insulin therapy may thus lead to 3-fold excess of severe hypoglycemia and severely hinder the effective and safe control of hyperglycemia. The main goal of the therapy for type 1 diabetes has long been physiological mimicry of normal insulin secretion based on monitoring which requires considerable effort and understanding of the underlying physiology. Attainment of this goal is challenged by the nature of the disease and our current lack of means to fully repair the abnormal endocrine pancreas interactive functions. As a result, various insulin preparations have been developed to partially compensate for the inability to deliver timely exogenous insulin directly to the portal/intrapancreatic circulation. It remains an ongoing task to identify the ideal routes and regimens of their delivery and potentially that of other hormones to restore the deficient and disordered hormonal environment of the pancreas to achieve a near normal metabolic state. Several recent technological advances help addressing these goals, including the rapid progress in insulin pumps, continuous glucose sensors, and ultimately the artificial pancreas closed-loop technology and the recent start of dual-hormone therapies.

Recovery after an intermittent test.

The purpose of this study was to analyse the impact of an intermittent test reproducing the soccer running activity profile on physical performance, subjective ratings and biochemical parameters throughout 72 h recovery. 8 professional soccer players performed the intermittent test on a non-motorised treadmill and data was collected before, immediately after, 24, 48 and 72 h after the test. Squat jump (SJ), countermovement jump (CMJ), peak isometric force (IFpeak), 6-s sprint, repeated sprints test (RS), perceptual ratings (fatigue, muscle soreness, stress), creatine kinase ([CK]) and uric acid ([UA]) were analyzed. After the test, a mean reduction in countermovement jump performance of -8.2% (CI: -12.9 to -3.4, p<0.01) was observed, while perceived fatigue (+2.1±1.7 a.u.; p<0.05), perceived muscle soreness (+1.8±1.5 a.u.; p<0.05), perceived stress (+1.6±1.5 a.u.; p<0.05), creatine kinase (+171±77 IU x l(-1); p<0.01) and uric acid (+168±89 Umol x l(-1); p<0.01) concentrations were significantly increased relative to baseline. No significant effect was found for SJ, IFpeak, 6-s sprint, RS immediately after and throughout the 72 h following the test. In conclusion, soccer running performance does not appear to be the main cause of post soccer match-induced fatigue. Physical data provided by video match analysis systems is insufficient to accurately estimate the level of match fatigue.

Optimizing reduction in basal hyperglucagonaemia to repair defective glucagon counterregulation in insulin deficiency.

In health, the pancreatic islet cells work as a network with highly co-ordinated signals over time to balance glycaemia within a narrow range. In type 1 diabetes (T1DM), with autoimmune destruction of the ß-cells, lack of insulin is considered the primary abnormality and is the primary therapy target. However, replacing insulin alone does not achieve adequate glucose control and recent studies have focused on controlling the endogenous glucagon release as well. In T1DM, glucagon secretion is disordered but not absolutely deficient; it may be excessive postprandially yet it is characteristically insufficient and delayed in response to hypoglycaemia. We review our system-level analysis of the pancreatic endocrine network mechanisms of glucagon counterregulation (GCR) and their dysregulation in T1DM and focus on possible use of α-cell inhibitors (ACIs) to manipulate the glucagon axis to repair the defective GCR. Our results indicate that the GCR abnormalities are of 'network origin'. The lack of ß-cell signalling is the primary deficiency that contributes to two separate network abnormalities: (i) absence of a ß-cell switch-off trigger and (ii) increased intraislet basal glucagon. A strategy to repair these abnormalities with ACI is proposed, which could achieve better control of glycaemia with reduced hypoglycaemia risk.

Bis-N-heterocyclic carbene palladium(IV) tetrachloride complexes: synthesis, reactivity, and mechanisms of direct chlorinations and oxidations of organic substrates.

This Article describes the preparation and isolation of novel octahedral CH(2)-bridged bis-(N-heterocyclic carbene)palladium(IV) tetrachlorides of the general formula LPd(IV)Cl(4) [L = (NHC)CH(2)(NHC)] from LPd(II)Cl(2) and Cl(2). In intermolecular, nonchelation-controlled transformations LPd(IV)Cl(4) reacted with alkenes and alkynes to 1,2-dichlorination adducts. Aromatic, benzylic, and aliphatic C-H bonds were converted into C-Cl bonds. Detailed mechanistic investigations in the dichlorinations of alkenes were conducted on the 18VE Pd(IV) complex. Positive solvent effects as well as kinetic measurements probing the impact of cyclohexene and chloride concentrations on the rate of alkene chlorination support a Pd(IV)-Cl ionization in the first step. Product stereochemistry and product distributions from various alkenes also support Cl(+)-transfer from the pentacoordinated Pd(IV)-intermediate LPd(IV)Cl(3)(+) to olefins. 1-Hexene/3-hexene competition experiments rule out both the formation of π-complexes along the reaction coordinate as well as in situ generated Cl(2) from a reductive elimination process. Instead, a ligand-mediated direct Cl(+)-transfer from LPd(IV)Cl(3)(+) to the π-system is likely to occur. Similarly, C-H bond chlorinations proceed via an electrophilic process with in situ formed LPd(IV)Cl(3)(+). The presence of a large excess of added Cl(-) slows cyclohexene chlorination while the presence of stoichiometric amounts of chloride accelerates both Pd(IV)-Cl ionization and Cl(+)-transfer from LPd(IV)Cl(3)(+). (1)H NMR titrations, T1 relaxation time measurements, binding isotherms, and Job plot analysis point to the formation of a trifurcated Cl(-)···H-C bond in the NHC-ligand periphery as a supramolecular cause for the accelerated chemical events involving the metal center.

Avian coronavirus infectious bronchitis virus susceptibility to botanical oleoresins and essential oils in vitro and in vivo.

Anti-coronaviral activity of a mixture of oleoresins and essential oils from botanicals, designated QR448(a), was examined in vitro and in vivo. Treatment of avian infectious bronchitis virus (IBV) with QR448(a) reduced the virus titer as measured in two laboratory host systems, Vero E6 cells and embryonating eggs. The effect of QR448(a) on IBV in chickens was also investigated. Administering QR448(a) to chickens at a 1:20 dilution by spray, 2h before challenge with IBV was determined to be the most effective treatment. Treatment decreased the severity of clinical signs and lesions in the birds, and lowered the amount of viral RNA in the trachea. Treatment with QR448(a) protected chickens for up to 4 days post-treatment from clinical signs of disease (but not from infection) and decreased transmission of IBV over a 14-day period. Anti-IBV activity of QR448(a) was greater prior to virus attachment and entry indicating that the effect is virucidal. In addition, QR448(a) had activity against both Massachusetts and Arkansas type IB viruses, indicating that it can be expected to be effective against IBV regardless of serotype. To our knowledge, this is the first report on the in vivo use of a virucidal mixture of compounds effective against the coronavirus IBV.