Physical Activity-Related Policy and Environmental Strategies to Prevent Obesity in Rural Communities: A Systematic Review of the Literature, 2002-2013
INTRODUCTION: Health disparities exist between rural and urban residents; in particular, rural residents have higher rates of chronic diseases and obesity. Evidence supports the effectiveness of policy and environmental strategies to prevent obesity and promote health equity. In 2009, the Centers for Disease Control and Prevention recommended 24 policy and environmental strategies for use by local communities: the Common Community Measures for Obesity Prevention (COCOMO); 12 strategies focus on physical activity. This review was conducted to synthesize evidence on the implementation, relevance, and effectiveness of physical activity-related policy and environmental strategies for obesity prevention in rural communities.
METHODS: A literature search was conducted in PubMed, PsycINFO, Web of Science, CINHAL, and PAIS databases for articles published from 2002 through May 2013 that reported findings from physical activity-related policy or environmental interventions conducted in the United States or Canada. Each article was extracted independently by 2 researchers.
RESULTS: Of 2,002 articles, 30 articles representing 26 distinct studies met inclusion criteria. Schools were the most common setting (n = 18 studies). COCOMO strategies were applied in rural communities in 22 studies; the 2 most common COCOMO strategies were "enhance infrastructure supporting walking" (n = 11) and "increase opportunities for extracurricular physical activity" (n = 9). Most studies (n = 21) applied at least one of 8 non-COCOMO strategies; the most common was increasing physical activity opportunities at school outside of physical education (n = 8). Only 14 studies measured or reported physical activity outcomes (10 studies solely used self-report); 10 reported positive changes.
CONCLUSION: Seven of the 12 COCOMO physical activity-related strategies were successfully implemented in 2 or more studies, suggesting that these 7 strategies are relevant in rural communities and the other 5 might be less applicable in rural communities. Further research using robust study designs and measurement is needed to better ascertain implementation success and effectiveness of COCOMO and non-COCOMO strategies in rural communities.
Fetal Hematopoietic Stem Cell Transplantation Fails to Fully Regenerate the B-Lymphocyte Compartment
B cells are key components of cellular and humoral immunity and, like all lymphocytes, are thought to originate and renew from hematopoietic stem cells (HSCs). However, our recent single-HSC transfer studies demonstrate that adult bone marrow HSCs do not regenerate B-1a, a subset of tissue B cells required for protection against pneumonia, influenza, and other infections. Since B-1a are regenerated by transfers of fetal liver, the question arises as to whether B-1a derive from fetal, but not adult, HSCs. Here we show that, similar to adult HSCs, fetal HSCs selectively fail to regenerate B-1a. We also show that, in humanized mice, human fetal liver regenerates tissue B cells that are phenotypically similar to murine B-1a, raising the question of whether human HSC transplantation, the mainstay of such models, is sufficient to regenerate human B-1a. Thus, our studies overtly challenge the current paradigm that HSCs give rise to all components of the immune system.
Culturally adaptive storytelling method to improve hypertension control in Vietnam - "We talk about our hypertension": study protocol for a feasibility cluster-randomized controlled trial
BACKGROUND: Vietnam is experiencing an epidemiologic transition with an increased prevalence of non-communicable diseases. At present, the major risk factors for cardiovascular disease (CVD) are either on the rise or at alarming levels in Vietnam; inasmuch, the burden of CVD will continue to increase in this country unless effective prevention and control measures are put in place. A national survey in 2008 found that the prevalence of hypertension (HTN) was approximately 25 % among Vietnamese adults and it increased with advancing age. Therefore, novel, large-scale, and sustainable interventions for public health education to promote engagement in the process of detecting and treating HTN in Vietnam are urgently needed.
METHODS: A feasibility randomized trial will be conducted in Hung Yen province, Vietnam to evaluate the feasibility and acceptability of a novel community-based intervention using the "storytelling" method to enhance the control of HTN in adults residing in four rural communities. The intervention will center on stories about living with HTN, with patients speaking in their own words. The stories will be obtained from particularly eloquent patients, or "video stars," identified during Story Development Groups. The study will involve two phases: (i) developing a HTN intervention using the storytelling method, which is designed to empower patients to facilitate changes in their lifestyle practices, and (ii) conducting a feasibility cluster-randomized trial to investigate the feasibility, acceptability, and potential efficacy of the intervention compared with usual care in HTN control among rural residents. The trial will be conducted at four communes, and within each commune, 25 individuals 50 years or older with HTN will be enrolled in the trial resulting in a total sample size of 100 patients.
DISCUSSION: This feasibility trial will provide the necessary groundwork for a subsequent large-scale, fully powered, cluster-randomized controlled trial to test the efficacy of our novel community-based intervention. Results from the full-scale trial will provide health policy makers with practical evidence on how to combat a key risk factor for CVD using a feasible, sustainable, and cost-effective intervention that could be used as a national program for controlling HTN in Vietnam and other developing countries.
TRIAL REGISTRATION: ClinicalTrials.gov.
REGISTRATION NUMBER: https://clinicaltrials.gov/ct2/show/NCT02483780 (registration date June 22, 2015).
Multipolar mitosis and aneuploidy after chrysotile treatment: a consequence of abscission failure and cytokinesis regression
Chrysotile, like other types of asbestos, has been associated with mesothelioma, lung cancer and asbestosis. However, the cellular abnormalities induced by these fibers involved in cancer development have not been elucidated yet. Previous works show that chrysotile fibers induce features of cancer cells, such as aneuploidy, multinucleation and multipolar mitosis. In the present study, normal and cancer derived human cell lines were treated with chrysotile and the cellular and molecular mechanisms related to generation of aneuploid cells was elucidated. The first alteration observed was cytokinesis regression, the main cause of multinucleated cells formation and centrosome amplification. The multinucleated cells formed after cytokinesis regression were able to progress through cell cycle and generated aneuploid cells after abnormal mitosis. To understand the process of cytokinesis regression, localization of cytokinetic proteins was investigated. It was observed mislocalization of Anillin, Aurora B, Septin 9 and Alix in the intercellular bridge, and no determination of secondary constriction and abscission sites. Fiber treatment also led to overexpression of genes related to cancer, cytokinesis and cell cycle. The results show that chrysotile fibers induce cellular and molecular alterations in normal and tumor cells that have been related to cancer initiation and progression, and that tetraploidization and aneuploid cell formation are striking events after fiber internalization, which could generate a favorable context to cancer development.
Apremilast, an oral phosphodiesterase 4 inhibitor, in patients with psoriatic arthritis and current skin involvement: a phase III, randomised, controlled trial (PALACE 3)
OBJECTIVE: To evaluate apremilast treatment in patients with active psoriatic arthritis, including current skin involvement, despite prior therapy with conventional disease-modifying antirheumatic drugs and/or biologic agents.
METHODS: Patients (N=505) were randomised (1:1:1) to placebo, apremilast 20 mg twice daily, or apremilast 30 mg twice daily. Rescue therapy with apremilast was designated at week 16 for placebo patients not achieving 20% improvement in swollen and tender joint counts. At week 24, the remaining placebo patients were then randomised to apremilast 20 mg twice daily or 30 mg twice daily. The efficacy and safety of apremilast were assessed over 52 weeks.
RESULTS: At week 16, significantly more patients receiving apremilast 20 mg twice daily (28%) and 30 mg twice daily (41%) achieved 20% improvement in American College of Rheumatology response criteria versus placebo (18%; p=0.0295 and p < 0.0001, respectively), and mean decrease in the Health Assessment Questionnaire-Disability Index score was significantly greater with apremilast 30 mg twice daily (-0.20) versus placebo (-0.07; p=0.0073). In patients with baseline psoriasis body surface area involvement > /=3%, significantly more apremilast 30 mg twice daily patients achieved 50% reduction from baseline Psoriasis Area and Severity Index score (41%) versus placebo (24%; p=0.0098) at week 16. At week 52, observed improvements in these measures demonstrated sustained response with continued apremilast treatment. Most adverse events were mild to moderate in severity; the most common were diarrhoea, nausea, headache and upper respiratory tract infection.
CONCLUSIONS: Apremilast demonstrated clinically meaningful improvements in psoriatic arthritis and psoriasis at week 16; sustained improvements were seen with continued treatment through 52 weeks. Apremilast was generally well tolerated and demonstrated an acceptable safety profile.
TRIAL REGISTRATION NUMBER: NCT01212770.
Aberrant Activation of p38 MAP Kinase-Dependent Innate Immune Responses Is Toxic to Caenorhabditis elegans
Inappropriate activation of innate immune responses in intestinal epithelial cells underlies the pathophysiology of inflammatory disorders of the intestine. Here we examine the physiological effects of immune hyperactivation in the intestine of the nematode Caenorhabditis elegans. We previously identified an immunostimulatory xenobiotic that protects C. elegans from bacterial infection by inducing immune effector expression via the conserved p38 MAP kinase pathway, but was toxic to nematodes developing in the absence of pathogen. To investigate a possible connection between the toxicity and immunostimulatory properties of this xenobiotic, we conducted a forward genetic screen for C. elegans mutants that are resistant to the deleterious effects of the compound, and identified five toxicity suppressors. These strains contained hypomorphic mutations in each of the known components of the p38 MAP kinase cassette (tir-1, nsy-1, sek-1, and pmk-1), demonstrating that hyperstimulation of the p38 MAPK pathway is toxic to animals. To explore mechanisms of immune pathway regulation in C. elegans, we conducted another genetic screen for dominant activators of the p38 MAPK pathway, and identified a single allele that had a gain-of-function (gf) mutation in nsy-1, the MAP kinase kinase kinase that acts upstream of p38 MAPK pmk-1. The nsy-1(gf) allele caused hyperinduction of p38 MAPK PMK-1-dependent immune effectors, had greater levels of phosphorylated p38 MAPK, and was more resistant to killing by the bacterial pathogen Pseudomonas aeruginosa compared to wild-type controls. In addition, the nsy-1(gf) mutation was toxic to developing animals. Together, these data suggest that the activity of the MAPKKK NSY-1 is tightly regulated as part of a physiological mechanism to control p38 MAPK-mediated innate immune hyperactivation, and ensure cellular homeostasis in C. elegans.
Patients are increasingly using online social networks (ie, social media) to connect with other patients and health care professionals--a trend called peer-to-peer health care. Because online social networks provide a means for health care professionals to communicate with patients, and for patients to communicate with each other, an opportunity exists to use social media as a modality to deliver behavioral interventions. Social media-delivered behavioral interventions have the potential to reduce the expense of behavioral interventions by eliminating visits, as well as increase our access to patients by becoming embedded in their social media feeds. Trials of online social network-delivered behavioral interventions have shown promise, but much is unknown about intervention development and methodology. In this paper, we discuss the process by which investigators can translate behavioral interventions for social media delivery. We present a model that describes the steps and decision points in this process, including the necessary training and reporting requirements. We also discuss issues pertinent to social media-delivered interventions, including cost, scalability, and privacy. Finally, we identify areas of research that are needed to optimize this emerging behavioral intervention modality.
Incidence of Dengue Virus Infection in Adults and Children in a Prospective Longitudinal Cohort in the Philippines
BACKGROUND: The mean age of dengue has been increasing in some but not all countries. We sought to determine the incidence of dengue virus (DENV) infection in adults and children in a prospective cohort study in the Philippines where dengue is hyperendemic.
METHODOLOGY/PRINCIPAL FINDINGS: A prospective cohort of subjects > /=6 months old in Cebu City, Philippines, underwent active community-based surveillance for acute febrile illnesses by weekly contact. Fever history within the prior seven days was evaluated with an acute illness visit followed by 2, 5, and 8-day, and 3-week convalescent visits. Blood was collected at the acute and 3-week visits. Scheduled visits took place at enrolment and 12 months that included blood collections. Acute samples were tested by DENV PCR and acute/convalescent samples by DENV IgM/IgG ELISA to identify symptomatic infections. Enrolment and 12-month samples were tested by DENV hemagglutination inhibition (HAI) assay to identify subclinical infections. Of 1,008 enrolled subjects, 854 completed all study activities at 12 months per-protocol undergoing 868 person-years of surveillance. The incidence of symptomatic and subclinical infections was 1.62 and 7.03 per 100 person-years, respectively. However, in subjects > 15 years old, only one symptomatic infection occurred whereas 27 subclinical infections were identified. DENV HAI seroprevalence increased sharply with age with baseline multitypic HAIs associated with fewer symptomatic infections. Using a catalytic model, the historical infection rate among dengue naive individuals was estimated to be high at 11-22%/year.
CONCLUSIONS/SIGNIFICANCE: In this hyperendemic area with high seroprevalence of multitypic DENV HAIs in adults, symptomatic dengue rarely occurred in individuals older than 15 years. Our findings demonstrate that dengue is primarily a pediatric disease in areas with high force of infection. However, the average age of dengue could increase if force of infection decreases over time, as is occurring in some hyperendemic countries such as Thailand.
Heterotaxy, a birth defect involving left-right patterning defects, and primary ciliary dyskinesia (PCD), a sinopulmonary disease with dyskinetic/immotile cilia in the airway are seemingly disparate diseases. However, they have an overlapping genetic etiology involving mutations in cilia genes, a reflection of the common requirement for motile cilia in left-right patterning and airway clearance. While PCD is a monogenic recessive disorder, heterotaxy has a more complex, largely non-monogenic etiology. In this study, we show mutations in the novel dynein gene DNAH6 can cause heterotaxy and ciliary dysfunction similar to PCD. We provide the first evidence that trans-heterozygous interactions between DNAH6 and other PCD genes potentially can cause heterotaxy. DNAH6 was initially identified as a candidate heterotaxy/PCD gene by filtering exome-sequencing data from 25 heterotaxy patients stratified by whether they have airway motile cilia defects. dnah6 morpholino knockdown in zebrafish disrupted motile cilia in Kupffer's vesicle required for left-right patterning and caused heterotaxy with abnormal cardiac/gut looping. Similarly DNAH6 shRNA knockdown disrupted motile cilia in human and mouse respiratory epithelia. Notably a heterotaxy patient harboring heterozygous DNAH6 mutation was identified to also carry a rare heterozygous PCD-causing DNAI1 mutation, suggesting a DNAH6/DNAI1 trans-heterozygous interaction. Furthermore, sequencing of 149 additional heterotaxy patients showed 5 of 6 patients with heterozygous DNAH6 mutations also had heterozygous mutations in DNAH5 or other PCD genes. We functionally assayed for DNAH6/DNAH5 and DNAH6/DNAI1 trans-heterozygous interactions using subthreshold double-morpholino knockdown in zebrafish and showed this caused heterotaxy. Similarly, subthreshold siRNA knockdown of Dnah6 in heterozygous Dnah5 or Dnai1 mutant mouse respiratory epithelia disrupted motile cilia function. Together, these findings support an oligogenic disease model with broad relevance for further interrogating the genetic etiology of human ciliopathies.
Palliative cytoreductive surgery and hyperthermic intraperitoneal chemoperfusion: current clinical practice or misnomer
Cytoreductive surgery and hyperthermic intraperitoneal chemoperfusion (CRS/HIPEC) is being used more and more frequently for the management of peritoneal carcinomatosis. Despite significant improvements in oncologic outcomes and the risk of complications and mortality, CRS/HIPEC remains one of the most morbid treatments offered for advanced cancers. Consequently CRS/HIPEC is still considered controversial by many, even in the setting of cancers that are potentially curable. However, as high volume surgical oncologists become more experienced with CRS/HIPEC, the potential role of "palliative CRS/HIPEC" in the management of peritoneal carcinomatosis is being raised. Given the often limited survival benefit expected after CRS/HIPEC, understanding the impact of the treatment on quality of life (QOL) needs to be an essential part of the decision to proceed and is critical to optimizing recovery afterwards. This article reviews the potential definitions of "palliative CRS/HIPEC" in various clinical contexts and describes the current state of the QOL experience after CRS/HIPEC.
Safety of anticoagulation with uninterrupted warfarin vs. interrupted dabigatran in patients requiring an implantable cardiac device
BACKGROUND: The optimal strategy of peri-procedural anticoagulation in patients undergoing permanent cardiac device implantation is controversial. Our objective was to compare the major bleeding and thromboembolic complications in patients managed with uninterrupted warfarin (UW) vs. interrupted dabigatran (ID) during permanent pacemaker (PPM) or implantable cardioverter defibrillators (ICD) implantation.
METHODS: A retrospective cohort study of all eligible patients from July 2011 through January 2012 was performed. UW was defined as patients who had maintained a therapeutic international normalized ratio (INR) on the day of the procedure. ID was defined as stopping dabigatran > /=12 hours prior to the procedure and then resuming after implantation. Major bleeding events included hemothorax, hemopericardium, intracranial hemorrhage, gastrointestinal bleed, epistaxis, or pocket hematoma requiring surgical intervention. Thromboembolic complications included stroke, transient ischemic attack, deep venous thrombosis, pulmonary embolism, or arterial embolism.
RESULTS: Of the 133 patients (73.4+/-11.0 years; 91 males) in the study, 86 received UW and 47 received ID. One (1.2%) patient in the UW group sustained hemopericardium perioperatively and died. In comparison, the ID patients had no complications. As compared to the ID group, the UW group had a higher median CHADS2 score (2 vs. 3, P=0.04) and incidence of Grade 1 pocket hematoma (0% vs. 7%, P=0.09). Neither group developed any thromboembolic complications.
CONCLUSIONS: Major bleeding rates were similar among UW and ID groups. Perioperative ID appears to be a safe anticoagulation strategy for patients undergoing PPM or ICD implantation.
Novel oral transforming growth factor-beta signaling inhibitor EW-7197 eradicates CML-initiating cells
Recent strategies for treating CML patients have focused on investigating new combinations of tyrosine kinase inhibitors (TKIs) as well as identifying novel translational research agents that can eradicate CML leukemia-initiating cells (CML-LICs). However, little is known about the therapeutic benefits such CML-LIC targeting therapies might bring to CML patients. In this study, we investigated the therapeutic potential of EW-7197, an orally bioavailable transforming growth factor-beta signaling inhibitor which has recently been approved as an Investigational New Drug (NIH, USA), to suppress CML-LICs in vivo. Compared to TKI treatment alone, administration of TKI plus EW-7197 to CML-affected mice significantly delayed disease relapse and prolonged survival. Notably, combined treatment with EW-7197 plus TKI was effective in eliminating CML-LICs even if they expressed the TKI-resistant T315I mutant BCR-ABL1 oncogene. Collectively, these results indicate that EW-7197 may be a promising candidate for a new therapeutic that can greatly benefit CML patients by working in combination with TKIs to eradicate CML-LICs.
The eastern North American monarch butterfly, Danaus plexippus, is an emerging model system to study the neural, molecular, and genetic basis of animal long-distance migration and animal clockwork mechanisms. While genomic studies have provided new insight into migration-associated and circadian clock genes, the general lack of simple and versatile reverse-genetic methods has limited in vivo functional analysis of candidate genes in this species. Here, we report the establishment of highly efficient and heritable gene mutagenesis methods in the monarch butterfly using transcriptional activator-like effector nucleases (TALENs) and CRISPR-associated RNA-guided nuclease Cas9 (CRISPR/Cas9). Using two clock gene loci, cryptochrome 2 and clock (clk), as candidates, we show that both TALENs and CRISPR/Cas9 generate high-frequency nonhomologous end-joining (NHEJ)-mediated mutations at targeted sites (up to 100%), and that injecting fewer than 100 eggs is sufficient to recover mutant progeny and generate monarch knockout lines in about 3 months. Our study also genetically defines monarch CLK as an essential component of the transcriptional activation complex of the circadian clock. The methods presented should not only greatly accelerate functional analyses of many aspects of monarch biology, but are also anticipated to facilitate the development of these tools in other nontraditional insect species as well as the development of homology-directed knock-ins.
Ticks transmit more pathogens to humans and animals than any other arthropod. We describe the 2.1 Gbp nuclear genome of the tick, Ixodes scapularis (Say), which vectors pathogens that cause Lyme disease, human granulocytic anaplasmosis, babesiosis and other diseases. The large genome reflects accumulation of repetitive DNA, new lineages of retro-transposons, and gene architecture patterns resembling ancient metazoans rather than pancrustaceans. Annotation of scaffolds representing approximately 57% of the genome, reveals 20,486 protein-coding genes and expansions of gene families associated with tick-host interactions. We report insights from genome analyses into parasitic processes unique to ticks, including host 'questing', prolonged feeding, cuticle synthesis, blood meal concentration, novel methods of haemoglobin digestion, haem detoxification, vitellogenesis and prolonged off-host survival. We identify proteins associated with the agent of human granulocytic anaplasmosis, an emerging disease, and the encephalitis-causing Langat virus, and a population structure correlated to life-history traits and transmission of the Lyme disease agent.
Improved cell metabolism prolongs photoreceptor survival upon retinal-pigmented epithelium loss in the sodium iodate induced model of geographic atrophy
Age-related macular degeneration (AMD) is characterized by malfunction and loss of retinal-pigmented epithelium (RPE) cells. Because the RPE transfers nutrients from the choriocapillaris to photoreceptor (PR), PRs are affected as well. Geographic atrophy (GA) is an advanced form of AMD characterized by severe vision impairment due to RPE loss over large areas. Currently there is no treatment to delay the degeneration of nutrient deprived PRs once RPE cells die. Here we show that cell-autonomous activation of the key regulator of cell metabolism, the kinase mammalian target of rapamycin complex 1 (mTORC1), delays PR death in the sodium iodate induced model of RPE atrophy. Consistent with this finding loss of mTORC1 in cones accelerates cone death as cones fail to balance demand with supply. Interestingly, promoting rod survival does not promote cone survival in this model of RPE atrophy as both, rods and cones suffer from a sick and dying RPE. The findings suggest that activation of metabolic genes downstream of mTORC1 can serve as a strategy to prolong PR survival when RPE cells malfunction or die.
Reconstruction of 60 Years of Chikungunya Epidemiology in the Philippines Demonstrates Episodic and Focal Transmission
Proper understanding of the long-term epidemiology of chikungunya has been hampered by poor surveillance. Outbreak years are unpredictable and cases often misdiagnosed. Here we analyzed age-specific data from 2 serological studies (from 1973 and 2012) in Cebu, Philippines, to reconstruct both the annual probability of infection and population-level immunity over a 60-year period (1952-2012). We also explored whether seroconversions during 2012-2013 were spatially clustered. Our models identified 4 discrete outbreaks separated by an average delay of 17 years. On average, 23% (95% confidence interval [CI], 16%-37%) of the susceptible population was infected per outbreak, with > 50% of the entire population remaining susceptible at any point. Participants who seroconverted during 2012-2013 were clustered at distances of < 230 m, suggesting focal transmission. Large-scale outbreaks of chikungunya did not result in sustained multiyear transmission. Nevertheless, we estimate that > 350,000 infections were missed by surveillance systems. Serological studies could supplement surveillance to provide important insights on pathogen circulation.
Transcriptional activation of follistatin by Nrf2 protects pulmonary epithelial cells against silica nanoparticle-induced oxidative stress
Silica nanoparticles (SiO2 NPs) cause oxidative stress in respiratory system. Meanwhile, human cells launch adaptive responses to overcome SiO2 NP toxicity. However, besides a few examples, the regulation of SiO2 NP-responsive proteins and their functions in SiO2 NP response remain largely unknown. In this study, we demonstrated that SiO2 NP induced the expression of follistatin (FST), a stress responsive gene, in mouse lung tissue as well as in human lung epithelial cells (A549). The levels of Ac-H3(K9/18) and H3K4me2, two active gene markers, at FST promoter region were significantly increased during SiO2 NP treatment. The induction of FST transcription was mediated by the nuclear factor erythroid 2-related factor 2 (Nrf2), as evidenced by the decreased FST expression in Nrf2-deficient cells and the direct binding of Nrf2 to FST promoter region. Down-regulation of FST promoted SiO2 NP-induced apoptosis both in cultured cells and in mouse lung tissue. Furthermore, knockdown of FST increased while overexpression of FST decreased the expression level of NADPH oxidase 1 (NOX1) and NOX5 as well as the production of cellular reactive oxygen species (ROS). Taken together, these findings demonstrated a protective role of FST in SiO2 NP-induced oxidative stress and shed light on the interaction between SiO2 NPs and biological systems.
Manipulation of IL-1beta and IL-18 production by Yersinia pestis effectors YopJ and YopM and redundant impact on virulence
Innate immunity plays a central role in resolving infections by pathogens. Host survival during plague, caused by the Gram-negative bacterium Yersinia pestis, is favored by a robust early innate immune response initiated by Interleukin-1beta (IL-1beta) and IL-18. These cytokines are produced by a two-step mechanism involving NF-kB mediated pro-cytokine production and inflammasome-driven maturation into bioactive inflammatory mediators. Because of the anti-microbial effects induced by IL-1beta/IL-18, it may be desirable for pathogens to manipulate their production. Y. pestis type III secretion system effectors YopJ and YopM can interfere with different parts of this process. Both effectors have been reported to influence inflammasome caspase-1 activity; YopJ promotes caspase-8 dependent cell death and caspase-1 cleavage, while YopM inhibits caspase-1 activity via an incompletely understood mechanism. Yet neither effector appears essential for full virulence in vivo. Here we report that the sum of influences by YopJ and YopM on IL-1beta/IL-18 release is suppressive. In the absence of YopM, YopJ minimally affects caspase-1 cleavage, but suppresses IL-1beta, IL-18 and other cytokines and chemokines. Importantly, we find that Y. pestis containing combined deletions of YopJ and YopM induces elevated levels of IL-1beta/IL-18 in vitro and in vivo, and is significantly attenuated in a mouse model of bubonic plague. The reduced virulence of the YopJ-YopM mutant is dependent on the presence of IL-1beta, IL-18 and caspase-1. Thus, we conclude that Y. pestis YopJ and YopM can both exert a tight control of host IL-1beta/IL-18 production to benefit the bacteria, resulting in a redundant impact on virulence.
Humans that are heterozygous for the common S180L polymorphism in the Toll-like receptor (TLR) adaptor Mal (encoded by TIRAP) are protected from a number of infectious diseases, including tuberculosis (TB), whereas those homozygous for the allele are at increased risk. The reason for this difference in susceptibility is not clear. We report that Mal has a TLR-independent role in interferon-gamma (IFN-gamma) receptor signaling. Mal-dependent IFN-gamma receptor (IFNGR) signaling led to mitogen-activated protein kinase (MAPK) p38 phosphorylation and autophagy. IFN-gamma signaling via Mal was required for phagosome maturation and killing of intracellular Mycobacterium tuberculosis (Mtb). The S180L polymorphism, and its murine equivalent S200L, reduced the affinity of Mal for the IFNGR, thereby compromising IFNGR signaling in macrophages and impairing responses to TB. Our findings highlight a role for Mal outside the TLR system and imply that genetic variation in TIRAP may be linked to other IFN-gamma-related diseases including autoimmunity and cancer.
Traditional cloning methods have limitations on the number of DNA fragments that can be simultaneously manipulated, which dramatically slows the pace of molecular assembly. Here we describe GMAP, a Gibson assembly-based modular assembly platform consisting of a collection of promoters and genes, which allows for one-step production of DNA constructs. GMAP facilitates rapid assembly of expression and viral constructs using modular genetic components, as well as increasingly complicated genetic tools using contextually relevant genomic elements. Our data demonstrate the applicability of GMAP toward the validation of synthetic promoters, identification of potent RNAi constructs, establishment of inducible lentiviral systems, tumor initiation in genetically engineered mouse models, and gene-targeting for the generation of knock-in mice. GMAP represents a recombinant DNA technology designed for widespread circulation and easy adaptation for other uses, such as synthetic biology, genetic screens, and CRISPR-Cas9.