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In this chapter, we show how megachurches satisfy attendees’ desire to belong in a community of like-minded others. We describe the process by which attendees go from being spectators to being active members, and the key role that small groups play in this process. Small groups connect attendees to others, allow them to feel accepted, make a large church feel small, and provide them with emotional energy in between weekend services. Megachurches offer a wide variety of small group options in an effort to meet their attendees’ every need, which include small groups focusing on marriage that become a critical factor in creating what we call megachurch marriage culture.

In this chapter, we show how megachurches meet attendees’ desire for a reliable leader. In megachurches, for those who attend, their desire and the force of feeling, the zenith of emotional energy, is squarely centered on the megachurch pastor. We identify how the charismatic bond between the attendees and their senior pastor is solidified through the demonstration of his perceived extraordinary and ordinary qualities. Allowing followers to see their human side makes charismatic leaders more relatable, authentic, and trustworthy. The pastor is both the central figure around which most activities, and in some sense the lives of individuals, revolve, and the mouthpiece for the explicit articulation of the values, beliefs, morals, and symbols that will define the group. The pastor is the “mutual focus of attention” that contributes to binding the group together and amplifying the emotional energy experienced by attendees.

In this chapter, we show how megachurches meet their attendees’ desire for acceptance and belonging. It all begins with co-presence—being together at the same place and time—in an atmosphere of excitement and acceptance as one is welcomed and invited into the worship service. Megachurches have crafted particular strategies to make attendees feel welcome, which include having few barriers to entry. The entire experience is designed to require no prior knowledge and to make individuals feel comfortable. This is accomplished in part by incorporating aspects of secular culture. In this way, megachurches both embody the world (i.e., secularity), and introduce and bring out the sacred at the heart of the secular.

Megachurches are growing in size and number in the United States with no indication of slowing down. We argue that their success is due to motivating their congregations with emotional energy that stimulates intense loyalty and a desire to come back repeatedly to get recharged. Megachurches are like drug dealers offering members and nonmembers alike their next hit of emotional energy. Ritual life is critical for the generation of emotional energy, but so are the minimally counterintuitive ideas that capture attention, channel the emotional energy, and rally loyalty and motivation to keep coming back for more. However, these ideas are not sufficient on their own but need to be charged with emotional energy from rituals to inspire loyalty from participants.

We describe the paradigm of micro-sociology using Randall Collins’s work on interaction ritual chains to understand how emotional energy is produced in megachurches. We argue that individuals are motivated to participate in megachurches through a process of interaction ritual chains that produce and evoke deep desires satisfied through emotional energy, which attracts and keeps so many coming to megachurches. We describe Collins’s ingredients for successful rituals (co-presence, a shared mood, a mutual focus of attention, and barriers to outsiders) and how megachurches meet or transform these ingredients for their purposes. As we narrate Collins’s interactive ritual structure, we briefly outline the ways in which the six desires, described in Chapter 3, are met in a cyclical manner within megachurches. We show how they are evoked and addressed in overlapping and synchronic ways, which reinforces the power of the collective effervescence of these churches.

We examine Fourth Presbyterian Church in Chicago as a rare example of a liberal Protestant megachurch that provides a case study of broader changes in American religion. We argue that there are four important lessons that can be learned from American religious history: (1) culture controls churches—churches are shaped by the cultural climates of their time, (2) emotion always trumps the mind—the emotional capacity of churches wins over cognitive claims, (3) leadership counts—charismatic leaders are vital for the success of churches, and (4) congregations tell the real story of what is going on in American religious culture.

In this chapter, we identify how megachurches meet attendees’ desire for deliverance. The preaching and music produce high levels of emotional energy that need to be released. In particular, altar calls and healing services are the emotional peaks of the church service, following the worship singing and sermons. These moments allow attendees to release the emotional energy that has built up from the service and in their lives. These are collective experiences in which individuals transform and watch others do the same. In response to the experience of deliverance, individuals are asked to serve their church and communities as an expression of their faith.

This chapter examines the dark side of megachurches by quantifying and theorizing megachurch scandals. We collected data on megachurch scandals and identified fifty-six such scandals discussed in forty-eight online newspaper articles for the years 2006 to 2017. Most of the scandals that we studied came from the malfeasance of megachurch senior pastors, and the vast majority were sexual in nature. The charismatic bond that megachurch pastors form with their attendees and the power that stems from it can be used for their own personal gain. We theorize a form of soft patriarchalism as a way to name this power differential. We contend that the model of soft patriarchalism gives power to men and creates opportunities for them to manipulate and ruin the lives of women under their care. These scandals often lead to the implosion of megachurches, where the charismatic bond between the pastor and congregation is irrevocably broken.

We provide and support our definition of religion. Religion is (1) a social enactment of a desire for the ultimate. It is (2) embodied in ritual practices; (3) described by systems of symbols and beliefs; (4) developed in communal settings, and often institutionally legitimated. (5) Religion interacts and negotiates with powers and forces that are experienced as within and beyond the self and group. (6) This power or force is most often referred to as god/spirit or gods/spirits. (7) The affective experience of ritual, and the symbolic and social boundaries constructed in rites, mobilize group identity and bind the group into a moral community. Last, (8) these moral communities produce networks of solidarity, and carry the potential for tension and, more rarely, conflict and violence within and between groups.

In this chapter, we demonstrate how megachurches provide individuals with purpose through service. The genius in megachurch socialization is that it not only encourages service to others, but also conceptually roots this service as an expression of one’s “gifts.” It makes individuals feel as though their service—their gifts—are specifically and uniquely needed. Encouraging their attendees to engage in service results in a community overflowing with ministries for members and the community. Megachurches are like small towns—communities that welcome all and where every conceivable form of care is given, which includes hospital clinics for “every sinner and sickness.”

A rational strategy to achieve optimal vaccine responses is to potentiate Tfh cells and the germinal center response. This work shows the adjuvant GLA-SE enhances circulating Tfh cells and enduring antibody responses to a malaria vaccine in Tanzanian adults., The generation of protective humoral immunity after vaccination relies on the productive interaction between antigen-specific B cells and T follicular helper (Tfh) cells. Despite the central role of Tfh cells in vaccine responses, there is currently no validated way to enhance their differentiation in humans. From paired human lymph node and blood samples, we identify a population of circulating Tfh cells that are transcriptionally and clonally similar to germinal center Tfh cells. In a clinical trial of vaccine formulations, circulating Tfh cells were expanded in Tanzanian volunteers when an experimental malaria vaccine was adjuvanted in GLA-SE but not when formulated in Alum. The GLA-SE–formulated peptide was associated with an increase in the extrafollicular antibody response, long-lived antibody production, and the emergence of public TCRβ clonotypes in circulating Tfh cells. We demonstrate that altering vaccine adjuvants is a rational approach for enhancing Tfh cells in humans, thereby supporting the long-lived humoral immunity that is required for effective vaccines., Graphical Abstract

BACKGROUND A lung transplant patient with invasive aspergillosis (IA) manifested symptoms of voriconazole-induced transaminitis with systemic voriconazole and progression of IA after switching to oral posaconazole. With limited options for standard triazole therapy, aerosolized delivery with one of the second-generation triazoles was considered. METHODS Feasibility for aerosolized delivery was evaluated using cascade impactor and analysis of physicochemical characteristics of voriconazole (10 mg/mL) and posaconazole (6, 12 mg/mL) solutions. RESULTS Both triazoles showed favorable characteristics for aerosol delivery with mass median aerodynamic diameter, geometric standard deviation, respirable fraction (

We describe the 'Crescendo Mouse', a human V

Although much has been done to understand how rearrangement of the Igκ locus is regulated during B-cell development, little is known about the way the variable (V) segments themselves are selected. Here we show, using B6/Cast hybrid pre-B-cell clones, that a limited number of V segments on each allele is stochastically activated as characterized by the appearance of non-coding RNA and histone modifications. The activation states are clonally distinct, stable across cell division and developmentally important in directing the Ig repertoire upon differentiation. Using a new approach of allelic ATAC-seq, we demonstrate that the Igκ V alleles have differential chromatin accessibility, which may serve as the underlying basis of clonal maintenance at this locus, as well as other instances of monoallelic expression throughout the genome. These findings highlight a new level of immune system regulation that optimizes gene diversity., B cell development involves sequential rearrangement of the immunoglobulin chains, but fine control over the selection process remains a mystery. Here the authors show that individual alleles in pre-B cells are clonally unique and result from stochastic activation of V gene segments to induce optimal generation of a diverse repertoire.

Hypothesis Surfactant-driven Marangoni flow on liquid films is predicted to depend on subphase depth and initial surface tension difference between the subphase and deposited surfactant solution drop. Changes in flow behavior will impact transport of soluble species entrained in the Marangoni flow along the surface. In extreme cases, the subphase film may rupture, limiting transport. Understanding this behavior is important for applications in drug delivery, coatings, and oil spill remediation. Experiments A trans-illumination optical technique measured the subphase height profiles and drop content transport after drop deposition when varying initial subphase depth, surfactant concentration, and subphase viscosity. Findings Three distinct flow regimes were identified depending on the subphase depth and surfactant concentration and mapped onto an operating diagram. These are characterized as a “central depression” bounded by an outwardly traveling ridge, an “annular depression” bounded by a central dome and the traveling ridge, and an “annular dewetting” when the subphase ruptures. Well above the critical micelle concentration, transitions between regimes occur at characteristic ratios of gravitational and initial surface tension gradient stresses; transitions shift when surfactant dilution during spreading weakens the stress before the completion of the spreading event. Drop contents travel with the ridge, but dewetting hinders transport.

Cystic fibrosis (CF) disease is caused by mutations affecting the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR), an anion channel expressed in the mucosal side of epithelial tissue. In the airway, dysfunctional CFTR results in a transepithelial osmotic imbalance leading to hyperabsorption of airway surface liquid mucostasis, chronic inflammation, and eventual respiratory failure. Human nasal epithelial cell cultures from healthy and CF donors were used to perform studies of liquid and solute transport dynamics at an air/liquid interface in order to emulate the in vivo airway. Then, these results were used to inform a quantitative systems pharmacology model of airway epithelium describing electrically and chemically driven transcellular ionic transport, contributions of both convective and diffusive paracellular solute transport, and osmotically driven transepithelial water dynamics. Model predictions showed CF cultures, relative to non-CF ones, have increased apical and basolateral water permeabilities, and increase paracellular permeability and transepithelial chemical driving force for a radiolabeled tracer used to track small molecule absorption. These results provide a computational platform to better understand and probe the mechanisms behind the liquid hyperabsorption and small molecule retention profiles observed in the CF airway.

Pediatric patients are very dependent on inhaled aerosol medications. There are significant differences in how these aerosols deposit in the lungs of children vs. adults that may affect the efficacy of the therapies. Inefficient aerosol delivery to children, caused by factors such as high mouth and throat deposition during oral inhalation, significant losses within adjunct devices such as masks, and high rates of nasal deposition during cannula delivery, can lead to dosing that is difficult to control. Here we discuss the methods, such as deposition scintigraphy, that are used to assess inhaled dose in vivo and review previous studies where these techniques have been applied to measure dosing in children. This includes studies of nebulizers and metered dose inhalers and delivery through adjuncts such as facemasks and nasal cannulas. We discuss the factors that can lead to inefficient inhaled drug delivery and high levels of mouth and throat deposition in children. Finally, we propose areas of innovation to improve inhaled drug delivery to this population. There is a need for child-specific technologies for inhaled drug delivery. This includes the use of smart devices that can guide pediatric breathing patterns and better engage children during treatments, the use of smaller aerosols which are less likely to deposit in the upper airways after inhalation, and the design of better nasal cannula interfaces for aerosol delivery to infants.

Efforts to address the coronavirus disease 2019 (COVID-19) pandemic have encountered skepticism among the public, but COVID-19 is not the only medical or scientific issue that receives such skepticism. How does COVID-19 skepticism relate to other forms of science skepticism? Using new data from a nationally representative survey of U.S. adults, this study reveals that skepticism toward COVID-19 is similar to patterns of skepticism toward vaccines in general and, more interestingly, skepticism toward climate change. Patterns of skepticism toward evolution and genetically modified foods are more distinct from COVID-19 skepticism. Notably, even after accounting for other forms of science skepticism, political conservatism is significantly associated with greater skepticism toward COVID-19. Finally, contrary to some media narratives, the analysis reveals few racial or ethnic differences in skepticism toward COVID-19, and the differences that do exist indicate less skepticism among Black and Asian individuals relative to White individuals.

Summary Variable (V), diversity (D), and joining (J) (V(D)J) recombination is the first determinant of antigen receptor diversity. Understanding how recombination is regulated requires a comprehensive, unbiased readout of V gene usage. We have developed VDJ sequencing (VDJ-seq), a DNA-based next-generation-sequencing technique that quantitatively profiles recombination products. We reveal a 200-fold range of recombination efficiency among recombining V genes in the primary mouse Igh repertoire. We used machine learning to integrate these data with local chromatin profiles to identify combinatorial patterns of epigenetic features that associate with active VH gene recombination. These features localize downstream of VH genes and are excised by recombination, revealing a class of cis-regulatory element that governs recombination, distinct from expression. We detect two mutually exclusive chromatin signatures at these elements, characterized by CTCF/RAD21 and PAX5/IRF4, which segregate with the evolutionary history of associated VH genes. Thus, local chromatin signatures downstream of VH genes provide an essential layer of regulation that determines recombination efficiency., Graphical Abstract, Highlights • VDJ-seq enables precise quantification of antibody V(D)J recombination products • Two distinct cis-regulatory designs characterize actively recombining V genes • Putative recombination regulatory elements map downstream of mouse Igh V genes • Recombination regulatory architecture reflects the V genes’ evolutionary history, Bolland et al. develop a technique to quantitatively profile antigen receptor diversity. Using VDJ-seq in the mouse Igh locus, they uncover the regulatory logic underlying the highly varying recombination rates of V gene segments, with implications for immune disorders and aberrant recombination in cancer.

Reducing the risk of rearrangement As lymphocytes develop, they rearrange their antigen receptor genes and proliferate extensively, potentially putting their genomes at risk. Galloway et al. found that two RNA-binding proteins, ZFP36L1 and ZFP36L2, ensure careful entry and exit into the cell cycle. This helps developing B lymphocytes maintain their genomic integrity. Mice deficient in ZFP36L1 and ZFP36L2 exhibited a profound block in B cell development. ZFP36L1 and ZFP36L2 suppress mRNAs that help B cells progress through the cell cycle, ensuring that cells can enter quiescence and keep their genomes safe when they undergo the risky process of rearranging their antigen receptors. Science , this issue p. 453

How do insurgents engaged in high-risk collective action maintain solidarity when faced with increasing costs and dangers? Based on a combination of process tracing through qualitative evidence and an event-history analysis of a unique data set assembled from naval archives concerning a mass mutiny in the Royal Navy in 1797, this article explains why insurgent solidarity varied among the ships participating in the mutiny. Maintaining solidarity was the key problem that the organizers of the mutiny faced in confronting government repression and inducements for ships’ companies to defect. Solidarity, proxied here as the duration of a ship's company's adherence to the mutiny, relied on techniques used by the mutiny leadership that increased dependence and imposed control over rank-and-file seamen. In particular, mutiny leaders monitored and sanctioned compliance and exploited informational asymmetries to persuade seamen to stand by the insurgency, even as prospects for its success faded.

Background: The cryopyrin-associated periodic syndromes (CAPS) are a group of inherited disorders associated with systemic auto-inflammation. CAPS result from gain-of-function mutations in NLRP3, which result in formation of an intracellular protein complex known as the NLRP3 inflammasome. This leads to overproduction of IL-1β and other pro-inflammatory signals, resulting in inflammatory symptoms. Treatments for NLRP3-related diseases are biologic agents that directly target IL-1β. We sought to determine if the orally available small molecule NLRP3 inhibitor MCC950 could inhibit IL-1β ex vivo in a cohort of patients with autoinflammatory disease. Methods: Patients were recruited to donate blood, from which PBMCs were isolated and assayed in the presence of MCC950 to determine inhibitory efficacy. Results: We found that apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) and mature IL-1β was higher in ex vivo PBMCs from CAPS patients than healthy donors. MCC950 inhibited production of mature IL-1β in PBMC from CAPS patients with a range of mutations and blocked NLRP3 activity in an in vitro mutation reconstitution assay. Similar results were observed with PBMC from two patients with Schnitzler’s Syndrome, another auto-inflammatory disease. Conclusions: The NLRP3 inflammasome inhibitor MCC950 blocked constitutive activation of NLRP3 observed in the PBMCs of CAPS patients. This study highlights the potential utility of NLRP3 inhibition by a small molecule for rare autoinflammatory diseases that are driven by NLRP3.

Background The ability to restore cystic fibrosis transmembrane regulator (CFTR) function with effective small molecule modulators in patients with cystic fibrosis provides an opportunity to study relationships between CFTR ion channel function, organ level physiology, and clinical outcomes. Methods We performed a multisite, prospective, observational study of ivacaftor, prescribed in patients with the G551D-CFTR mutation. Measurements of lung mucociliary clearance (MCC) were performed before and after treatment initiation (1 and 3 months), in parallel with clinical outcome measures. Results Marked acceleration in whole lung, central lung, and peripheral lung MCC was observed 1 month after beginning ivacaftor and was sustained at 3 months. Improvements in MCC correlated with improvements in forced expiratory volume in the first second (FEV1) but not sweat chloride or symptom scores. Conclusions Restoration of CFTR activity with ivacaftor led to significant improvements in MCC. This physiologic assessment provides a means to characterize future CFTR modulator therapies and may help to predict improvements in lung function. Trial registration ClinicialTrials.gov, NCT01521338. Funding CFF Therapeutics (GOAL11K1).

Aim Nasal cannulas are used to provide oxygen support for infants and have been considered as a means for delivering aerosols to the lungs. To measure mucociliary clearance in the lungs of infants with congenital heart defects, we delivered radiopharmaceutical aerosols via a nasal cannula. Here we report on the pulmonary and nasal deposition of these aerosols. Method A total of 18 infants (median age = 26 days; quartiles = 11-74 days) performed clearance measurements soon before or after corrective cardiac surgery. The regional aerosol deposition was assessed using gamma camera imaging. Results Cannula flow rate significantly affected pulmonary dosing. Flow rates useful for oxygen support were associated with low pulmonary deposition (2 L/min; mean, 4.5% of deposited dose; range, 2%-9%; n = 7) and high nasal deposition. Much lower cannula flow rates increased the pulmonary deposition (0.2 L/min; mean, 33.5% of deposited dose; range, 15%-51%; n = 5; P = 0.005 vs 2 L/min). The ratio of nose/lung dosing was approximately 26:1 at 2 L/min and 2:1 at 0.2 L/min. Bench studies demonstrated cannula output rates of 10.2 ± 1.7% (2 L/min) and 3.3 ± 0.4% (0.2 L/min) of the loaded nebulizer dose during a 2-minute delivery. Combining in vitro and in vivo results, we estimate that 0.46% of the loaded nebulizer dose reaches the lungs at 2 L/min vs 1.10% at 0.2 L/min during a 2-minute delivery. Conclusion With the delivery system used here, pulmonary aerosol delivery via nasal cannula was very inefficient at the flow rates required to provide oxygen support. Even at low flows, nasal deposition was substantial and local toxicity must be considered.

Background Aging is characterized by loss of function of the adaptive immune system, but the underlying causes are poorly understood. To assess the molecular effects of aging on B cell development, we profiled gene expression and chromatin features genome-wide, including histone modifications and chromosome conformation, in bone marrow pro-B and pre-B cells from young and aged mice. Results Our analysis reveals that the expression levels of most genes are generally preserved in B cell precursors isolated from aged compared with young mice. Nonetheless, age-specific expression changes are observed at numerous genes, including microRNA encoding genes. Importantly, these changes are underpinned by multi-layered alterations in chromatin structure, including chromatin accessibility, histone modifications, long-range promoter interactions, and nuclear compartmentalization. Previous work has shown that differentiation is linked to changes in promoter-regulatory element interactions. We find that aging in B cell precursors is accompanied by rewiring of such interactions. We identify transcriptional downregulation of components of the insulin-like growth factor signaling pathway, in particular downregulation of Irs1 and upregulation of Let-7 microRNA expression, as a signature of the aged phenotype. These changes in expression are associated with specific alterations in H3K27me3 occupancy, suggesting that Polycomb-mediated repression plays a role in precursor B cell aging. Conclusions Changes in chromatin and 3D genome organization play an important role in shaping the altered gene expression profile of aged precursor B cells. Components of the insulin-like growth factor signaling pathways are key targets of epigenetic regulation in aging in bone marrow B cell precursors. Electronic supplementary material The online version of this article (10.1186/s13059-018-1489-y) contains supplementary material, which is available to authorized users.

VDJ-seq is a DNA-based, high-throughput sequencing method that uses biotinylated J gene primers and UMIs to quantify immunoglobulin diversity in an unbiased manner on a per-cell basis. For high-throughput sequencing and quantification of immunoglobulin repertoires, most methodologies use RNA. However, output varies enormously between recombined genes due to different promoter strengths and differential activation of lymphocyte subsets, precluding quantitation of recombinants on a per-cell basis. To date, DNA-based approaches have used V gene primer cocktails, with substantial inherent biases. Here, we describe VDJ sequencing (VDJ-seq), which accurately quantitates immunoglobulin diversity at the DNA level in an unbiased manner. This is accomplished with a single primer-extension step using biotinylated J gene primers. By addition of unique molecular identifiers (UMIs) before primer extension, we reliably remove duplicate sequences and correct for sequencing and PCR errors. Furthermore, VDJ-seq captures productive and nonproductive VDJ and DJ recombination events on a per-cell basis. Library preparation takes 3 d, with 2 d of sequencing and 1 d of data processing and analysis.

Understanding the fundamentals of surface transport on thin viscous films has important application in pulmonary drug delivery. The human lung contains a large-area interface between its complex fluid lining and inhaled air. Marangoni flows driven by surface tension gradients along this interface would promote enhanced distribution of inhaled therapeutics by carrying them from where they are deposited in the upper airways, along the fluid interface to deeper regions of the lung. Motivated by the potential to improve therapies for acute and chronic lung diseases, we review recent progress in modeling and experimental studies of Marangoni transport induced by the deposition of surfactant-containing microliter drops and liquid aerosols (picoliter drops) onto a fluid interface. The roles of key system variables are identified, including surfactant solubility, drop miscibility with the subphase, and the thickness, composition and surface properties of the subphase liquid. Of particular interest is the unanticipated but crucial role of aerosol processing to achieve Marangoni transport via phospholipid vesicle dispersions, which are likely candidates for a biocompatible delivery system. Progress in this field has the potential to not only improve outcomes in patients with chronic and acute lung diseases, but also to further our understanding of surface transport in complex systems.

Aerosol drug delivery is a viable option for treating diseased airways, but airway obstructions associated with diseases such as cystic fibrosis cause non-uniform drug distribution and limit efficacy. Marangoni stresses produced by surfactant addition to aerosol formulations may enhance delivery uniformity by post-deposition spreading of medications over the airway surface, improving access to poorly ventilated regions. We examine the roles of different variables affecting the maximum post-deposition spreading of a dye (drug mimic).Entangled aqueous solutions of either poly(acrylamide) (PA) or porcine gastric mucin (PGM) serve as airway surface liquid (ASL) mimicking subphases for in vitro models of aerosol deposition. Measured aerosol deposition fluxes indicate that the experimental delivery conditions are representative of aerosol delivery to the conducting airways. Post-deposition spreading beyond the locale of direct aerosol deposition is tracked by fluorescence microscopy. Aqueous aerosols formulated with either nonionic surfactant (tyloxapol) or fluorosurfactant (FS-3100) are compared with surfactant-free control aerosols.Significant enhancement of post-deposition spreading is observed with surfactant solutions relative to surfactant-free control solutions, provided the surfactant solution surface tension is less than that of the subphase. Amongst the variables considered--surfactant concentration, aerosol flow-rate, total deposited volume, time of delivery, and total deposited surfactant mass--surfactant mass is the primary predictor of maximum spread distance. This dependence is also observed for solutions deposited as a single, microliter-scale drop with a volume comparable to the total volume of deposited aerosol.Marangoni stress-assisted spreading after surfactant-laden aerosol deposition at high fluxes on a complex fluid subphase is capable of driving aerosol contents over significantly greater distances compared to surfactant-free controls. Total delivered surfactant mass is the primary determinant of the extent of spreading, suggesting a great potential to extend the reach of aerosolized medication in partially obstructed airways via a purely physical mechanism.

Cystic fibrosis (CF) is associated with the accumulation of dehydrated mucus in the pulmonary airways. This alters ventilation and aerosol deposition patterns in ways that limit drug delivery to peripheral lung regions. We investigated the use of surfactant-based, self-dispersing aerosol carriers that produce surface tension gradients to drive two-dimensional transport of aerosolized medications via Marangoni flows after deposition on the airway surface liquid (ASL). We considered the post-deposition spreading of individual aerosol droplets and two-dimensional expansion of a field of aerosol droplets, when deposited at low fluxes that are representative of aerosol deposition in the small airways.We used physically entangled aqueous solutions of poly(acrylamide) or porcine gastric mucin as simple ASL mimics that adequately capture the full miscibility but slow penetration of entangled macromolecular chains of the ASL into the deposited drop. Surfactant formulations were prepared with aqueous solutions of nonionic tyloxapol or FS-3100 fluorosurfactant. Fluorescein dye served as a model "drug" tracer and to visualize the extent of post-deposition spreading.The surfactants not only enhanced post-deposition spreading of individual aerosol droplets due to localized Marangoni stresses, as previously observed with macroscopic drops, but they also produced large-scale Marangoni stresses that caused the deposited aerosol fields to expand into initially unexposed regions of the subphase. We show that the latter is the main mechanism for spreading drug over large distances when aerosol is deposited at low fluxes representative of the small airways. The large scale convective expansion of the aerosol field drives the tracer (drug mimic) over areas that would cover an entire airway generation or more, in peripheral airways, where sub-monolayer droplet deposition is expected during aerosol inhalation.The results suggest that aerosolized surfactant formulations may provide the means to maximize deposited drug uniformity in and access to small airways.

Serotonin (5-HT)-synthesizing neurons of the medullary raphe are putative central chemoreceptors, proposed to be one of potentially multiple brain stem chemosensitive cell types and loci interacting to produce the respiratory chemoreflex. Hypocretin-synthesizing neurons of the lateral hypothalamus are important contributors to arousal state, thermoregulation, and feeding behavior and are also reportedly involved in the hypercapnic ventilatory response. Recently, a functional interaction was found between the hypocretin system and 5-HT neurons of the dorsal raphe. The validity and potential significance of hypocretin modulation of medullary raphe 5-HT neurons, however, is unknown. As such, the purpose of this study was to explore functional interactions between the hypocretin system and 5-HT system of the medullary raphe on baseline respiratory output and central chemosensitivity. To explore such interactions, we used the neonatal in vitro medullary slice preparation derived from wild-type (WT) mice (normal 5-HT function) and a knockout strain lacking all central 5-HT neurons ( Lmx1b f/f/p mice). We examined effects of acidosis, hypocretin-1, a hypocretin receptor antagonist (SB-408124), and the effect of the antagonist on the response to acidosis. We confirmed the critical role of 5-HT neurons in central chemosensitivity given that the increased hypoglossal burst frequency with acidosis, characteristic of WT mice, was absent in preparations derived from Lmx1b f/f/p mice. We also found that hypocretin facilitated baseline neural ventilatory output in part through 5-HT neurons. Although the impact of hypocretin on 5-HT neuronal sensitivity to acidosis is still unclear, hypocretins did appear to mediate the burst duration response to acidosis via serotonergic mechanisms.

Prior studies on perceptions of structural disadvantage and injustice, efficacy, and collective action have suffered from two major limitations: (1) they have used singlecountry samples, usually of economically advanced countries, and (2) generally theorized and investigated perceptions of structural injustice and efficacy separately. Drawing on value-expectancy theory, we provide an integrated theory to predict direct and conditional effects of efficacy and perceptions of structural disadvantage and injustice on collective action within countries. To address the limitations of previous research, we use cross-national data of 29 countries, including economically advanced and less advanced nations, to test how well these hypotheses explain within-country variation in collective action. We find that internal efficacy is significantly and positively associated with lowand moderate-cost collective action, whereas organizational embeddedness, a proxy for political efficacy, is significantly and positively associated with low-, moderate-, and high-cost collective action. Perceptions of legitimate and unjust structural disadvantage are also positively associated with all types of collective action. Importantly, the positive effects of both types of efficacy on high-cost collective action are conditional on perceptions of structural injustice. That is, participation in high-cost collective action is more likely for those who are both efficacious and perceive structural disadvantage as unjust.

Skin damage is a clinical manifestation of both urinary incontinence (UI) and/or fecal incontinence (FI). Incontinence associated dermatitis (IAD) is characterized by irritation and inflammation of the skin from prolonged exposure to urine or stool. IAD can affect many different groups of patients but is most prevalent in the elderly and those with medical comorbidities. Prevention should be targeted to this “at risk” population, however it is known that the prevalence and incidence rates for IAD are significant across all care settings. Complications include infections, pain, skin damage, and pressure ulcers. Management goals should address skin maintenance, protection from fecal and urinary irritants, pain management, and treatment of secondary infections (www.wocn.org, Doughty et al., J Wound Ostomy Continence Nurs 39(3):303–315, 2012).

V(D)J recombination is essential for the generation of diverse antigen receptor (AgR) repertoires. In B cells, immunoglobulin kappa (Igκ) light chain recombination follows immunoglobulin heavy chain (Igh) recombination. We recently developed the DNA-based VDJ-seq assay for the unbiased quantitation of Igh VH and DH repertoires. Integration of VDJ-seq data with genome-wide datasets revealed that two chromatin states at the recombination signal sequence (RSS) of VH genes are highly predictive of recombination in mouse pro-B cells. It is unknown whether local chromatin states contribute to Vκ gene choice during Igκ recombination. Here we adapt VDJ-seq to profile the Igκ VκJκ repertoire and present a comprehensive readout in mouse pre-B cells, revealing highly variable Vκ gene usage. Integration with genome-wide datasets for histone modifications, DNase hypersensitivity, transcription factor binding and germline transcription identified PU.1 binding at the RSS, which was unimportant for Igh, as highly predictive of whether a Vκ gene will recombine or not, suggesting that it plays a binary, all-or-nothing role, priming genes for recombination. Thereafter, the frequency with which these genes recombine was shaped both by the presence and level of enrichment of several other chromatin features, including H3K4 methylation and IKAROS binding. Moreover, in contrast to the Igh locus, the chromatin landscape of the promoter, as well as of the RSS, contributes to Vκ gene recombination. Thus, multiple facets of local chromatin features explain much of the variation in Vκ gene usage. Together, these findings reveal shared and divergent roles for epigenetic features and transcription factors in AgR V(D)J recombination and provide avenues for further investigation of chromatin signatures that may underpin V(D)J-mediated chromosomal translocations.

Aim: Inhaled hypertonic saline increases mucociliary clearance, improves pulmonary function, and decreases exacerbations in cystic fibrosis (CF) but contributes to the already significant treatment burden of CF. Overnight delivery of inhaled medications via a specially designed nasal cannula-aerosol device (Trans-nasal Pulmonary Aerosol Delivery [tPAD]) is an alternative approach. Here, we test whether overnight inhalation of hypertonic saline via tPAD improves mucociliary clearance and assess the tolerability of the device. Method: In this study, 12 CF subjects inhaled 7% hypertonic saline (HS) for 8 h overnight using the tPAD system. Safety and tolerability were assessed and measurements of mucociliary and absorptive clearance (MCC/ABS) were performed after the treatment. Comparisons were made versus sham treatment where the same subjects wore the nasal cannula overnight but did not receive aerosol. Results: Both the HS and sham treatments were well-tolerated. Only one subject did not complete the overnight HS treatment. There were no significant differences in MCC associated with HS inhalation at any time point (90 min, 3 h, 6 h) in any lung zone. Changes in FEV1 on both study days were similar. There were no differences in quality of sleep between HS and sham nights as assessed with the modified Leeds Sleep Evaluation Questionnaire (mLSEQ). Sino-Nasal Outcome Test (SNOT-14) questionnaires demonstrated significant increases (worsening) in 2/14 symptom categories with HS. Conclusions: The most likely cause for the failure to accelerate MCC was under-dosing of HS relative to the active transport of salt from the airways.

Human pluripotent stem cells (PSCs) exist in naive and primed states and provide important models to investigate the earliest stages of human development. Naive cells can be obtained through primed-to-naive resetting, but there are no reliable methods to prospectively isolate unmodified naive cells during this process. Here we report comprehensive profiling of cell surface proteins by flow cytometry in naive and primed human PSCs. Several naive-specific, but not primed-specific, proteins were also expressed by pluripotent cells in the human preimplantation embryo. The upregulation of naive-specific cell surface proteins during primed-to-naive resetting enabled the isolation and characterization of live naive cells and intermediate cell populations. This analysis revealed distinct transcriptional and X chromosome inactivation changes associated with the early and late stages of naive cell formation. Thus, identification of state-specific proteins provides a robust set of molecular markers to define the human PSC state and allows new insights into the molecular events leading to naive cell resetting.

Summary Serotonergic neurons modulate behavioral and physiological responses from aggression and anxiety to breathing and thermoregulation. Disorders involving serotonin (5HT) dysregulation are commensurately heterogeneous and numerous. We hypothesized that this breadth in functionality derives in part from a developmentally determined substructure of distinct subtypes of 5HT neurons each specialized to modulate specific behaviors. By manipulating developmentally defined subgroups one by one chemogenetically, we find that the Egr2-Pet1 subgroup is specialized to drive increased ventilation in response to carbon dioxide elevation and acidosis. Furthermore, this subtype exhibits intrinsic chemosensitivity and modality-specific projections—increasing firing during hypercapnic acidosis and selectively projecting to respiratory chemosensory but not motor centers, respectively. These findings show that serotonergic regulation of the respiratory chemoreflex is mediated by a specialized molecular subtype of 5HT neuron harboring unique physiological, biophysical, and hodological properties specified developmentally and demonstrate that the serotonergic system contains specialized modules contributing to its collective functional breadth.

Previous studies have reported subsets of medullary raphé neurons that are either stimulated or inhibited by CO2/pH in vitro, in situ, and in vivo. We tested the hypothesis that medullary raphé CO2-inhibited neurons are GABAergic. Extracellular recordings in unanesthetized juvenile in situ rat preparations showed reversible hypercapnia-induced suppression of 19% (63/323) of medullary raphé neurons, and this suppression persisted after antagonism of NMDA, AMPA/kainate, and GABAA receptors. We stained a subset of CO2-inhibited cells and found that most (11/12) had glutamic acid decarboxylase 67 immunoreactivity (GAD67-ir). These data indicate that the majority of acidosis-inhibited medullary raphé neurons are GABAergic, and that their chemosensitivity is independent of major fast synaptic inputs. Thus, CO2-sensitive GABAergic neurons may play a role in central CO2/pH chemoreception.

Megachurches have been criticized as superficial sources of entertainment that do not produce significant feelings of belonging, moral responsibility, or spirituality. This article challenges popular criticisms of megachurches and, drawing on interaction ritual theory, proposes that megachurches are successful interaction ritual venues and powerful purveyors of emotional religious experience. We predict that these interaction rituals produce positive emotional energy, membership symbols that are charged with emotional significance, feelings of morality, and a heightened sense of spirituality. From a census of 1,250 known megachurches in America, 12 were selected that closely represent the national megachurch profile. At each church, focus groups were conducted and attendees participated in a survey. We combine these data sources to provide a more comprehensive picture of the megachurch interaction ritual. The combined qualitative and quantitative results provide strong support for our predictions.

Inhaled cyclosporine (CsA) is being investigated as a prophylaxis for lung transplant rejection. Lung deposition and systemic exposure of nebulized CsA in lung transplant patients was evaluated as part of the Phase 3 cyclosporine inhalation solution (CIS) trial (CYCLIST).Ten patients received 300 mg of CIS (62.5 mg/mL CsA in propylene glycol) admixed with 148 MBq of Tc-DTPA (technetium-99m bound to diethylenetriaminepentaacetic acid) administered using a Sidestream(®) disposable jet nebulizer. Deposition was assessed using a dual-headed gamma camera. Blood samples were collected over a 24-hr time period after aerosol dosing and analyzed for CsA levels. A pharmacokinetic analysis of the resulting blood concentration versus time profiles was performed.The average total deposited dose was 53.7 ± 12.7 mg. Average pulmonary dose was 31.8 ± 16.3 mg, and stomach dose averaged 15.5 ± 11.1 mg. Device performance was consistent, with breathing maneuvers influencing dose variation. Predose coaching with five of 10 patients reduced stomach deposition (22.6 ± 11.2 vs. 8.3 ± 5.2 mg; p=0.03). Blood concentrations declined quickly from a maximum of 372 ± 140 ng/mL to 15.3 ± 9.7 ng/mL at 24 hr post dose. Levels of AUC(0-24) [area under the concentration vs. time curve from 0 to 24 hr] averaged 1,493 ± 746 ng hr/mL. On a three times per week dose regimen, this represents5% of the weekly systemic exposure of twice per day oral administration.Substantial doses of CsA can be delivered to the lungs of lung transplant patients by inhaled aerosol. Systemic levels are small relative to typical oral CsA administration.

New measures are needed to rapidly assess emerging treatments for cystic fibrosis (CF) lung disease. Using an imaging approach, we evaluated the absorptive clearance of the radiolabeled small molecule probe diethylene triamine penta-acetic acid (DTPA) as an in vivo indicator of changes in airway liquid absorption. DTPA absorption and mucociliary clearance rates were measured in 21 patients with CF (12 adults and nine children) and nine adult controls using nuclear imaging. The effect of hypertonic saline on DTPA absorption was also studied. In addition, in vitro studies were conducted to identify the determinants of transepithelial DTPA absorption. CF patients had significantly increased rates of DTPA absorption compared with control subjects but had similar mucociliary clearance rates. Treatment with hypertonic saline resulted in a decrease in DTPA absorption and an increase in mucociliary clearance in 11 out of 11 adult CF patients compared with treatment with isotonic saline. In vitro studies revealed that ∼ 50% of DTPA absorption can be attributed to transepithelial fluid transport. Apically applied mucus impedes liquid and DTPA absorption. However, mucus effects become negligible in the presence of an osmotic stimulus. Functional imaging of DTPA absorption provides a quantifiable marker of immediate response to treatments that promote airway surface liquid hydration.

Dehydration of mucus in the lungs of cystic fibrosis (CF) patients can inhibit mucociliary clearance (MCC), resulting in mucus build-up and blockages. Prior mathematical models have shown differences in MCC from the large airways and airway surface liquid (ASL) absorption dynamics between CF and non-CF patients. The model developed here captures MCC and ASL absorption dynamics over 250 minutes with higher accuracy than previous models.

Inhibitory 5-HT1areceptors are located on serotonin (5-HT) neurons (autoreceptors) as well as neurons of the respiratory network (heteroreceptors). Thus, effects on breathing of 5-HT1aagonists, such as (R)-(+)-8-hydroxy-2-(di-N-propylamino) tetralin (8-OH-DPAT), could either be due to decreased firing of 5-HT neurons or direct effects on the respiratory network. Mice in which the transcription factor LMX1B is genetically deleted selectively inPet1-1-expressing cells (Lmx1bf/f/p) essentially have complete absence of central 5-HT neurons, providing a unique opportunity to separate the effect of activation of downstream 5-HT1aheteroreceptors from that of autoreceptors. We used rhythmically active medullary slices from wild-type (WT) andLmx1bf/f/pneonatal mice to differentiate autoreceptor versus heteroreceptor effects of 8-OH-DPAT on hypoglossal nerve respiratory output. 8-OH-DPAT transiently increased respiratory burst frequency inLmx1bf/f/ppreparations, but not in WT slices. This excitation was abolished when synaptic inhibition was blocked by GABAergic/glycinergic receptor antagonists. Conversely, after 10 min of application, frequency inLmx1bf/f/pslices was not different from baseline, whereas it was significantly depressed in WT slices. In WT micein vivo, subcutaneous injection of 8-OH-DPAT produced similar biphasic respiratory effects as inLmx1bf/f/pmice. We conclude that 5-HT1areceptor agonists have two competing effects: rapid stimulation of breathing due to excitation of the respiratory network, and delayed inhibition of breathing due to autoreceptor inhibition of 5-HT neurons. The former effect is presumably due to inhibition of inhibitory interneurons embedded in the respiratory network.

All social exchanges involve some degree of uncertainty, although the amount depends on the conditions of the exchange. Uncertainty may arise due to the unverifiable quality of the good exchanged or the indeterminable quality of the exchange partner (i.e., whether she is trustworthy). Social exchange theory offers several mechanisms by which uncertainty may be reduced or mitigated such as through trust, repeated exchanges, reputation, information regarding the exchanges of others, and institutions. This paper applies principles from social exchange theory to religious behavior and argues that the same mechanisms that reduce uncertainty in social exchanges also reduce uncertainty in ‘religious’ exchanges, resulting in higher levels of religious commitment. In particular, this paper proposes that having experiences believed to be supernatural, having close ties to others within one’s religious congregation, and being affiliated with a higher tension church should increase religious commitment through decreasing uncertainty. Analysis of the 1988 General Social Survey and the 2007 Baylor Religion Survey supports these hypotheses.

Recently, there has been a call for social scientific studies of religion to start taking “place” seriously. This paper adds to this growing literature by embedding religious commitment within place. We propose that the tension religious groups experience with their surroundings partly depends on local contexts and that certain types of local contexts may generate more tension. These contexts include regions in which the nonreligiously affiliated constitute the majority of the population, such as the American Pacific Northwest (PNW) region. Analysis of qualitative data on evangelical and liberal Protestants from western Oregon and western Washington suggests that they are aware of and experience tension with the regional PNW culture, which shapes their religious experience.

Motivated by the possibility of enhancing aerosol drug delivery to mucus-obstructed lungs, the spreading of a drop of aqueous surfactant solution on a physically entangled aqueous poly(acrylamide) solution subphase that mimics lung airway surface liquid was investigated. Sodium dodecyl sulfate was used as the surfactant. To visualize spreading of the drop and mimic the inclusion of a drug substance, fluorescein, a hydrophilic and non-surface active dye, was added to the surfactant solution. The spreading progresses through a series of events. Marangoni stresses initiate the convective spreading of the drop. Simultaneously, surfactant escapes across the drop’s contact line within a second of deposition and causes a change in subphase surface tension outside the drop on the order of 1 mN/m. Convective spreading of the drop ends within 2–3 seconds of drop deposition, when a new interfacial tension balance is achieved. Surfactant escape depletes the drop of surfactant and the residual drop takes the form of a static lens of non-zero contact angle. On longer time scales, the surfactant dissolves into the subphase. The lens formed by the water in the deposited drop persists for as long as 3 minutes after the convective spreading process ends due to the long diffusional timescales associated with the underlying entangled polymer solution. The persistence of the lens suggests that the drop phase behaves as if it were immiscible with the subphase during this time period. Whereas surfactant escapes the spreading drop and advances on the subphase/vapor interface, hydrophilic dye molecules in the drop do not escape, but remain with the drop throughout the convective spreading. The quasi-immiscible nature of the spreading event suggests that the chemical properties of the surfactant and subphase are much less important than their physical properties, consistent with prior qualitative studies of spreading of different types of surfactants on entangled polymer subphases: the selection of surfactant for pulmonary delivery applications may be limited only by physical and toxicological considerations. Further, the escape of surfactant from individual drops may provide an additional spreading mechanism in the lung as hydrodynamic and/or surface pressure repulsions may drive individual droplets apart after deposition.

The sudden infant death syndrome is associated with a reduction in brainstem serotonin 5-hydroxytryptamine (5-HT) and 5-HT(1A) receptor binding, yet it is unknown if and how these findings are linked. In this study, we used quantitative tissue autoradiography to determine if post-natal development of brainstem 5-HT(1A) receptors is altered in two mouse models where the development of 5-HT neurons is defective, the Lmx1b(f/f/p) , and the Pet-1⁻/⁻ mouse. 5-HT(1A) receptor agonist-binding sites were examined in both 5-HT-source nuclei (autoreceptors) and in sites that receive 5-HT innervation (heteroreceptors). In control mice between post-natal day (P) 3 and 10, 5-HT(1A) receptor binding increased in several brainstem sites; by P25, there were region-specific increases and decreases, refining the overall binding pattern. In the Lmx1b(f/f/p) and Pet-1⁻/⁻ mice, 5-HT(1A)-autoreceptor binding was significantly lower than in control mice at P3, and remained low at P10 and P25. In contrast, 5-HT(1A) heteroreceptor levels were comparable between control and 5-HT-deficient mice. These data define the post-natal development of 5-HT(1A)-receptor binding in the mouse brainstem. Furthermore, the data suggest that 5-HT(1A)-heteroreceptor deficits detected in sudden infant death syndrome are not a direct consequence of a 5-HT neuron dysfunction nor reduced brain 5-HT levels. To elucidate the developmental relationship between serotonin (5-HT) levels and 5-HT(1A) receptors in the brainstem, we examined 5-HT(1A) binding in two 5-HT-deficient mouse models. In nuclei containing 5-HT neurons, 5-HT(1A) binding was decreased (autoreceptors), while binding was maintained in projection sites (heteroreceptors). Thus, brainstem 5-HT(1A)-heteroreceptor-binding sites do not appear developmentally sensitive to reduced brain 5-HT levels.