A common polymorphism in the Intelectin-1 gene influences mucus plugging in severe asthma.

By incompletely understood mechanisms, type 2 (T2) inflammation present in the airways of severe asthmatics drives the formation of pathologic mucus which leads to airway mucus plugging. Here we investigate the molecular role and clinical significance of intelectin-1 (ITLN-1) in the development of pathologic airway mucus in asthma. Through analyses of human airway epithelial cells we find that ITLN1 gene expression is highly induced by interleukin-13 (IL-13) in a subset of metaplastic MUC5AC+ mucus secretory cells, and that ITLN-1 protein is a secreted component of IL-13-induced mucus. Additionally, we find ITLN-1 protein binds the C-terminus of the MUC5AC mucin and that its deletion in airway epithelial cells partially reverses IL-13-induced mucostasis. Through analysis of nasal airway epithelial brushings, we find that ITLN1 is highly expressed in T2-high asthmatics, when compared to T2-low children. Furthermore, we demonstrate that both ITLN-1 gene expression and protein levels are significantly reduced by a common genetic variant that is associated with protection from the formation of mucus plugs in T2-high asthma. This work identifies an important biomarker and targetable pathways for the treatment of mucus obstruction in asthma.

The Puerto Rican Infant Metagenomic and Epidemiologic Study of Respiratory Outcomes (PRIMERO): Design and Baseline Characteristics for a Birth Cohort Study of Early-life Viral Respiratory Illnesses and Airway Dysfunction in Puerto Rican Children.

Epidemiologic studies demonstrate an association between early-life respiratory illnesses (RIs) and the development of childhood asthma. However, it remains uncertain whether these children are predisposed to both conditions or if early-life RIs induce alterations in airway function, immune responses, or other human biology that contribute to the development of asthma. Puerto Rican children experience a disproportionate burden of early-life RIs and asthma, making them an important population for investigating this complex interplay. PRIMERO, the Puerto Rican Infant Metagenomics and Epidemiologic Study of Respiratory Outcomes , recruited pregnant women and their newborns to investigate how the airways develop in early life among infants exposed to different viral RIs, and will thus provide a critical understanding of childhood asthma development. As the first asthma birth cohort in Puerto Rico, PRIMERO will prospectively follow 2,100 term healthy infants. Collected samples include post-term maternal peripheral blood, infant cord blood, the child's peripheral blood at the year two visit, and the child's nasal airway epithelium, collected using minimally invasive nasal swabs, at birth, during RIs over the first two years of life, and at annual healthy visits until age five. Herein, we describe the study's design, population, recruitment strategy, study visits and procedures, and primary outcomes.

Epigenomic response to albuterol treatment in asthma-relevant airway epithelial cells.

BACKGROUND: Albuterol is the first-line asthma medication used in diverse populations. Although DNA methylation (DNAm) is an epigenetic mechanism involved in asthma and bronchodilator drug response (BDR), no study has assessed whether albuterol could induce changes in the airway epithelial methylome. We aimed to characterize albuterol-induced DNAm changes in airway epithelial cells, and assess potential functional consequences and the influence of genetic variation and asthma-related clinical variables. RESULTS: We followed a discovery and validation study design to characterize albuterol-induced DNAm changes in paired airway epithelial cultures stimulated in vitro with albuterol. In the discovery phase, an epigenome-wide association study using paired nasal epithelial cultures from Puerto Rican children (n = 97) identified 22 CpGs genome-wide associated with repeated-use albuterol treatment (p < 9 × 10-8). Albuterol predominantly induced a hypomethylation effect on CpGs captured by the EPIC array across the genome (probability of hypomethylation: 76%, p value = 3.3 × 10-5). DNAm changes on the CpGs cg23032799 (CREB3L1), cg00483640 (MYLK4-LINC01600), and cg05673431 (KSR1) were validated in nasal epithelia from 10 independent donors (false discovery rate [FDR] < 0.05). The effect on the CpG cg23032799 (CREB3L1) was cross-tissue validated in bronchial epithelial cells at nominal level (p = 0.030). DNAm changes in these three CpGs were shown to be influenced by three independent genetic variants (FDR < 0.05). In silico analyses showed these polymorphisms regulated gene expression of nearby genes in lungs and/or fibroblasts including KSR1 and LINC01600 (6.30 × 10-14 ≤ p ≤ 6.60 × 10-5). Additionally, hypomethylation at the CpGs cg10290200 (FLNC) and cg05673431 (KSR1) was associated with increased gene expression of the genes where they are located (FDR < 0.05). Furthermore, while the epigenetic effect of albuterol was independent of the asthma status, severity, and use of medication, BDR was nominally associated with the effect on the CpG cg23032799 (CREB3L1) (p = 0.004). Gene-set enrichment analyses revealed that epigenomic modifications of albuterol could participate in asthma-relevant processes (e.g., IL-2, TNF-α, and NF-κB signaling pathways). Finally, nine differentially methylated regions were associated with albuterol treatment, including CREB3L1, MYLK4, and KSR1 (adjusted p value < 0.05). CONCLUSIONS: This study revealed evidence of epigenetic modifications induced by albuterol in the mucociliary airway epithelium. The epigenomic response induced by albuterol might have potential clinical implications by affecting biological pathways relevant to asthma.

Nasal airway transcriptome-wide association study of asthma reveals genetically driven mucus pathobiology.

To identify genetic determinants of airway dysfunction, we performed a transcriptome-wide association study for asthma by combining RNA-seq data from the nasal airway epithelium of 681 children, with UK Biobank genetic association data. Our airway analysis identified 95 asthma genes, 58 of which were not identified by transcriptome-wide association analyses using other asthma-relevant tissues. Among these genes were MUC5AC, an airway mucin, and FOXA3, a transcriptional driver of mucus metaplasia. Muco-ciliary epithelial cultures from genotyped donors revealed that the MUC5AC risk variant increases MUC5AC protein secretion and mucus secretory cell frequency. Airway transcriptome-wide association analyses for mucus production and chronic cough also identified MUC5AC. These cis-expression variants were associated with trans effects on expression; the MUC5AC variant was associated with upregulation of non-inflammatory mucus secretory network genes, while the FOXA3 variant was associated with upregulation of type-2 inflammation-induced mucus-metaplasia pathway genes. Our results reveal genetic mechanisms of airway mucus pathobiology.

Type 2 and interferon inflammation regulate SARS-CoV-2 entry factor expression in the airway epithelium.

Coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2, an emerging virus that utilizes host proteins ACE2 and TMPRSS2 as entry factors. Understanding the factors affecting the pattern and levels of expression of these genes is important for deeper understanding of SARS-CoV-2 tropism and pathogenesis. Here we explore the role of genetics and co-expression networks in regulating these genes in the airway, through the analysis of nasal airway transcriptome data from 695 children. We identify expression quantitative trait loci for both ACE2 and TMPRSS2, that vary in frequency across world populations. We find TMPRSS2 is part of a mucus secretory network, highly upregulated by type 2 (T2) inflammation through the action of interleukin-13, and that the interferon response to respiratory viruses highly upregulates ACE2 expression. IL-13 and virus infection mediated effects on ACE2 expression were also observed at the protein level in the airway epithelium. Finally, we define airway responses to common coronavirus infections in children, finding that these infections generate host responses similar to other viral species, including upregulation of IL6 and ACE2. Our results reveal possible mechanisms influencing SARS-CoV-2 infectivity and COVID-19 clinical outcomes.

Type 2 and interferon inflammation strongly regulate SARS-CoV-2 related gene expression in the airway epithelium.

Coronavirus disease 2019 (COVID-19) outcomes vary from asymptomatic infection to death. This disparity may reflect different airway levels of the SARS-CoV-2 receptor, ACE2, and the spike protein activator, TMPRSS2. Here we explore the role of genetics and co-expression networks in regulating these genes in the airway, through the analysis of nasal airway transcriptome data from 695 children. We identify expression quantitative trait loci (eQTL) for both ACE2 and TMPRSS2, that vary in frequency across world populations. Importantly, we find TMPRSS2 is part of a mucus secretory network, highly upregulated by T2 inflammation through the action of interleukin-13, and that interferon response to respiratory viruses highly upregulates ACE2 expression. Finally, we define airway responses to coronavirus infections in children, finding that these infections upregulate IL6 while also stimulating a more pronounced cytotoxic immune response relative to other respiratory viruses. Our results reveal mechanisms likely influencing SARS-CoV-2 infectivity and COVID-19 clinical outcomes.

Genome-Wide Analysis Reveals Mucociliary Remodeling of the Nasal Airway Epithelium Induced by Urban PM2.5.

Air pollution particulate matter <2.5 µm (PM2.5) exposure is associated with poor respiratory outcomes. Mechanisms underlying PM2.5-induced lung pathobiology are poorly understood but likely involve cellular and molecular changes to the airway epithelium. We extracted and chemically characterized the organic and water-soluble components of air pollution PM2.5 samples, then determined the whole transcriptome response of human nasal mucociliary airway epithelial cultures to a dose series of PM2.5 extracts. We found that PM2.5 organic extract (OE), but not water-soluble extract, elicited a potent, dose-dependent transcriptomic response from the mucociliary epithelium. Exposure to a moderate OE dose modified the expression of 424 genes, including activation of aryl hydrocarbon receptor signaling and an IL-1 inflammatory program. We generated an OE-response gene network defined by eight functional enrichment groups, which exhibited high connectivity through CYP1A1, IL1A, and IL1B. This OE exposure also robustly activated a mucus secretory expression program (>100 genes), which included transcriptional drivers of mucus metaplasia (SPDEF and FOXA3). Exposure to a higher OE dose modified the expression of 1,240 genes and further exacerbated expression responses observed at the moderate dose, including the mucus secretory program. Moreover, the higher OE dose significantly increased the MUC5AC/MUC5B gel-forming mucin expression ratio and strongly downregulated ciliated cell expression programs, including key ciliating cell transcription factors (e.g., FOXJ1 and MCIDAS). Chronic OE stimulation induced mucus metaplasia-like remodeling characterized by increases in MUC5AC+ secretory cells and MUC5AC mucus secretions. This epithelial remodeling may underlie poor respiratory outcomes associated with high PM2.5 exposure.

Identification of CFTR variants in Latino patients with cystic fibrosis from the Dominican Republic and Puerto Rico.

BACKGROUND: In cystic fibrosis (CF), the spectrum and frequency of CFTR variants differ by geography and race/ethnicity. CFTR variants in White patients are well-described compared with Latino patients. No studies of CFTR variants have been done in patients with CF in the Dominican Republic or Puerto Rico. METHODS: CFTR was sequenced in 61 Dominican Republican patients and 21 Puerto Rican patients with CF and greater than ​​​​60 mmol/L sweat chloride. The spectrum of CFTR variants was identified and the proportion of patients with 0, 1, or 2 CFTR variants identified was determined. The functional effects of identified CFTR variants were investigated using clinical annotation databases and computational prediction tools. RESULTS: Our study found 10% of Dominican patients had two CFTR variants identified compared with 81% of Puerto Rican patients. No CFTR variants were identified in 69% of Dominican patients and 10% of Puerto Rican patients. In Dominican patients, there were 19 identified CFTR variants, accounting for 25 out of 122 disease alleles (20%). In Puerto Rican patients, there were 16 identified CFTR variants, accounting for 36 out of 42 disease alleles (86%) in Puerto Rican patients. Thirty CFTR variants were identified overall. The most frequent variants for Dominican patients were p.Phe508del and p.Ala559Thr and for Puerto Rican patients were p.Phe508del, p.Arg1066Cys, p.Arg334Trp, and p.I507del. CONCLUSIONS: In this first description of the CFTR variants in patients with CF from the Dominican Republic and Puerto Rico, there was a low detection rate of two CFTR variants after full sequencing with the majority of patients from the Dominican Republic without identified variants.

Crosstalk of LKB1- and PTEN-regulated signals in liver morphogenesis and tumor development.

Liver kinase B 1 (LKB1 or STK11) and PTEN (phosphatase and tensin homologue deleted on chromosome 10) are two tumor suppressors that regulate the mTOR signaling pathway. Deletion studies show that loss of either Lkb1 (Lkb+/- ) or Pten (PtenloxP/loxP; Alb-Cre+ ) leads to liver injury and development of hepatocarcinoma. In this study, we investigated the crosstalk of LKB1 and PTEN loss during tumorigenesis and liver development. We show here that haplo-insufficiency of Lkb1 in the liver leads to advanced tumor development in the Pten null mice (PtenloxP/loxP; LkbloxP/+; Alb-Cre+ ). Our analysis shows that LKB1 and PTEN interacted with each other in their regulation of fatty acid synthase as well as p21 expression. The combined loss of LKB1 and PTEN (PtenloxP/loxP; LkbloxP/loxP; Alb-Cre+ ) also led to the inability to form zonal structures in the liver. The lack of metabolic zonal structures is consistent with the inability of the livers to store glycogen as well as elevated plasma bilirubin and alanine aminotransferase (ALT), indicative of liver dysfunction. These structural and functional defects are associated with cytoplasm distribution of a canalicular membrane protein MRP2 (multidrug resistant protein 2) which is responsible for clearing bilirubin. This observed regulation of MRP2 by LKB1 likely contributed to the lack of cellular polarity and the early lethality phenotype associated with homozygous loss of Lkb1 alone or in combination with Pten. Finally, Pten deletion does not rescue the precocious ductal plate formation reported for Lkb1 deleted livers. CONCLUSION: Our study dissected the functional and molecular crosstalk of PTEN and LKB1 and elucidate key molecular targets for such interaction.

Telemonitoring Adherence to Medications in Heart Failure Patients (TEAM-HF): A Pilot Randomized Clinical Trial.

BACKGROUND: Medication nonadherence contributes to hospitalizations in recently discharged patients with heart failure (HF). We aimed to test the feasibility of telemonitoring medication adherence in patients with HF. METHODS AND RESULTS: We randomized 40 patients (1:1) hospitalized for HF to 30 days of loop diuretic adherence monitoring with telephonic support or to passive adherence monitoring alone. Eighty-three percent of eligible patients agreed to participate. The median age of patients was 64 years, 25% were female, and 45% were Hispanic. Overall, 67% of patients were nonadherent (percentage of days that the correct number of doses were taken <88%). There were no differences between intervention and passive monitoring group patients, respectively, in adherence (median correct dosing adherence 82% vs 73%; P = .41) or in the proportion readmitted within 30 days (30% vs 20%; P = .72). Eighty-eight percent of patients rated the wireless electronic adherence device as somewhat or very easy to use, and 88% agreed to use it again. CONCLUSIONS: Adherence telemonitoring was acceptable to most patients with HF. Diuretic nonadherence was common even when patients knew they were being monitored. Future studies should assess whether adherence telemonitoring can improve adherence and reduce readmissions among patients with HF.

Activation of hepatic stellate cell in Pten null liver injury model.

BACKGROUND: Hepatic fibrosis is a prominent pathological feature associated with chronic liver disease including non-alcoholic hepatosteatosis (NASH), and a precursor for liver cancer development. We previously reported that PTEN loss in the liver, which leads to hyperactivated liver insulin signaling results in NASH development. Here we used the same mouse model to study the progression from steatosis to fibrosis. RESULTS: The Pten null livers develop progressive liver fibrosis as indicated by Sirius Red staining and increased expression of collagen I, Timp 1, SMAα, and p75NTR. Consistently, hepatic stellate cells (HSCs) isolated from Pten null livers are readily activated when compared with that from mice with intact PTEN. Deletion of AKT2, the downstream target of PTEN signal, blocked NASH development, and alleviated fibrosis. HSCs from the Pten/Akt2 double null mice are quiescent like those isolated from the control livers. Our analysis shows that the activation of HSCs does not depend on the intrinsic signals regulated by PI3K/AKT, the target of PTEN, but does depend on steatosis and injury to the liver. During the progression of liver fibrosis in the Pten null model, Wnt ligands and signaling receptor are induced, concurrent with the reduction of sFRP5, a Wnt antagonist. We showed that treatment of HSCs with Wnt receptor antagonist blocks the observed morphological changes when HSCs undergo activation in culture. This signal appears to be mediated by ß-catenin, as manipulating ß-catenin signaling alters marker gene expressions of HSC activation. CONCLUSIONS: Wnt/ß-catenin activation serves as an important mediator for fibrosis development resulting from NASH using a mouse model where NASH is mimicked by PTEN loss.

Adult-onset deletion of Pten increases islet mass and beta cell proliferation in mice.

AIMS/HYPOTHESIS: Adult beta cells have a diminished ability to proliferate. Phosphatase and tensin homologue (PTEN) is a lipid phosphatase that antagonises the function of the mitogenic phosphatidylinositol 3-kinase (PI3K) pathway. The objective of this study was to understand the role of PTEN and PI3K signalling in the maintenance of beta cells postnatally. METHODS: We developed a Pten (lox/lox); Rosa26 (lacZ); RIP-CreER (+) model that permitted us to induce Pten deletion by treatment with tamoxifen in mature animals. We evaluated islet mass and function as well as beta cell proliferation in 3- and 12-month-old mice treated with tamoxifen (Pten deleted) vs mice treated with vehicle (Pten control). RESULTS: Deletion of Pten in juvenile (3-month-old) beta cells significantly induced their proliferation and increased islet mass. The expansion of islet mass occurred concomitantly with the enhanced ability of the Pten-deleted mice to maintain euglycaemia in response to streptozotocin treatment. In older mice (>12 months of age), deletion of Pten similarly increased islet mass and beta cell proliferation. This novel finding suggests that PTEN-regulated mechanisms may override the age-onset diminished ability of beta cells to respond to mitogenic stimulation. We also found that proteins regulating G1/S cell-cycle transition, such as cyclin D1, cyclin D2, p27 and p16, were altered when PTEN was lost, suggesting that they may play a role in PTEN/PI3K-regulated beta cell proliferation in adult tissue. CONCLUSIONS/INTERPRETATION: The signals regulated by the PTEN/PI3K pathway are important for postnatal maintenance of beta cells and regulation of their proliferation in adult tissues.

PTEN controls ß-cell regeneration in aged mice by regulating cell cycle inhibitor p16ink4a.

Tissue regeneration diminishes with age, concurrent with declining hormone levels including growth factors such as insulin-like growth factor-1 (IGF-1). We investigated the molecular basis for such decline in pancreatic ß-cells where loss of proliferation occurs early in age and is proposed to contribute to the pathogenesis of diabetes. We studied the regeneration capacity of ß-cells in mouse model where PI3K/AKT pathway downstream of insulin/IGF-1 signaling is upregulated by genetic deletion of Pten (phosphatase and tensin homologue deleted on chromosome 10) specifically in insulin-producing cells. In this model, PTEN loss prevents the decline in proliferation capacity in aged ß-cells and restores the ability of aged ß-cells to respond to injury-induced regeneration. Using several animal and cell models where we can manipulate PTEN expression, we found that PTEN blocks cell cycle re-entry through a novel pathway leading to an increase in p16(ink4a), a cell cycle inhibitor characterized for its role in cellular senescence/aging. A downregulation in p16(ink4a) occurs when PTEN is lost as a result of cyclin D1 induction and the activation of E2F transcription factors. The activation of E2F transcriptional factors leads to methylation of p16(ink4a) promoter, an event that is mediated by the upregulation of polycomb protein, Ezh2. These analyses establish a novel PTEN/cyclin D1/E2F/Ezh2/p16(ink4a) signaling network responsible for the aging process and provide specific evidence for a molecular paradigm that explain how decline in growth factor signals such as IGF-1 (through PTEN/PI3K signaling) may control regeneration and the lack thereof in aging cells.

Centralized, stepped, patient preference-based treatment for patients with post-acute coronary syndrome depression: CODIACS vanguard randomized controlled trial.

IMPORTANCE: Controversy remains about whether depression can be successfully managed after acute coronary syndrome (ACS) and the costs and benefits of doing so. OBJECTIVE: To determine the effects of providing post-ACS depression care on depressive symptoms and health care costs. DESIGN: Multicenter randomized controlled trial. SETTING: Patients were recruited from 2 private and 5 academic ambulatory centers across the United States. PARTICIPANTS: A total of 150 patients with elevated depressive symptoms (Beck Depression Inventory [BDI] score ≥10) 2 to 6 months after an ACS, recruited between March 18, 2010, and January 9, 2012. INTERVENTIONS: Patients were randomized to 6 months of centralized depression care (patient preference for problem-solving treatment given via telephone or the Internet, pharmacotherapy, both, or neither), stepped every 6 to 8 weeks (active treatment group; n = 73), or to locally determined depression care after physician notification about the patient's depressive symptoms (usual care group; n = 77). MAIN OUTCOME MEASURES: Change in depressive symptoms during 6 months and total health care costs. RESULTS: Depressive symptoms decreased significantly more in the active treatment group than in the usual care group (differential change between groups, -3.5 BDI points; 95% CI, -6.1 to -0.7; P = .01). Although mental health care estimated costs were higher for active treatment than for usual care, overall health care estimated costs were not significantly different (difference adjusting for confounding, -$325; 95% CI, -$2639 to $1989; P = .78). CONCLUSIONS: For patients with post-ACS depression, active treatment had a substantial beneficial effect on depressive symptoms. This kind of depression care is feasible, effective, and may be cost-neutral within 6 months; therefore, it should be tested in a large phase 3 pragmatic trial. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01032018.

Design and baseline data from the vanguard of the Comparison of Depression Interventions after Acute Coronary Syndrome (CODIACS) randomized controlled trial.

This paper describes the rationale and design of the vanguard for the Comparison of Depression Interventions after Acute Coronary Syndrome (CODIACS), a multicenter, randomized, controlled trial of a patient preference-based, stepped care protocol for persistent depressive symptoms after acute coronary syndrome (ACS). The overall aim of the vanguard phase was to determine whether the patient-preference, stepped care protocol, which is based on the intervention used in the recent Coronary Psychosocial Evaluation Studies (COPES) trial, was feasible in patients with recent ACS who were recruited from 5 geographically diverse sites. Innovative design features of this trial include randomization to either initial patient-preference of treatment or to a referred care arm in which the primary care provider decided upon care. Additionally, delivery of psychotherapy was accomplished by telephone, or webcam, depending upon patient preference. The vanguard phase provides estimates of eligibility and screening/enrollment ratios, patient acceptance of screening, and retention. In this report, we describe the innovative features and the baseline results of the vanguard phase of CODIACS. The data from this vanguard study will be used to finalize planning for a large, phase III clinical trial designed to evaluate the effect of treatment on depressive symptoms, coronary events, and death.

Enhanced depression care for patients with acute coronary syndrome and persistent depressive symptoms: coronary psychosocial evaluation studies randomized controlled trial.

BACKGROUND: Depressive symptoms are an established predictor of mortality and major adverse cardiac events (defined as nonfatal myocardial infarction or hospitalization for unstable angina or urgent/emergency revascularizations) in patients with acute coronary syndrome (ACS). This study was conducted to determine the acceptability and efficacy of enhanced depression treatment in patients with ACS. METHODS: A 3-month observation period to identify patients with ACS and persistent depressive symptoms was followed by a 6-month randomized controlled trial. From January 1, 2005, through February 29, 2008, 237 patients with ACS from 5 hospitals were enrolled, including 157 persistently depressed patients randomized to intervention (initial patient preference for problem-solving therapy and/or pharmacotherapy, then a stepped-care approach; 80 patients) or usual care (77 patients) and 80 nondepressed patients who underwent observational evaluation. The primary outcome was patient satisfaction with depression care. Secondary outcomes were depressive symptom changes (assessed with the Beck Depression Inventory), major adverse cardiac events, and death. RESULTS: At the end of the trial, the proportion of patients who were satisfied with their depression care was higher in the intervention group (54% of 80) than in the usual care group (19% of 77) (odds ratio, 5.4; 95% confidence interval [CI], 2.2-12.9 [P < .001]). The Beck Depression Inventory score decreased significantly more (t(155) = 2.85 [P = .005]) for intervention patients (change, -5.7; 95% CI, -7.6 to -3.8; df = 155) than for usual care patients (change, -1.9; 95% CI, -3.8 to -0.1; df = 155); the depression effect size was 0.59 of the standard deviation. At the end of the trial, 3 intervention patients and 10 usual care patients had experienced major adverse cardiac events (4% and 13%, respectively; log-rank test, chi(2)(1) = 3.93 [P = .047]), as well as 5 nondepressed patients (6%) (for the intervention vs nondepressed cohort, chi(2)(1) = 0.48 [P = .49]). CONCLUSION: Enhanced depression care for patients with ACS was associated with greater satisfaction, a greater reduction in depressive symptoms, and a promising improvement in prognosis. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00158054.