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Altered enteric microorganisms in concert with host genetics shape inflammatory bowel disease (IBD) phenotypes. However, insight is limited to bacteria and fungi. We found that eukaryotic viruses and bacteriophages (collectively, the virome), enriched from non-IBD, noninflamed human colon resections, actively elicited atypical anti-inflammatory innate immune programs. Conversely, ulcerative colitis or Crohn’s disease colon resection viromes provoked inflammation, which was successfully dampened by non-IBD viromes. The IBD colon tissue virome was perturbed, including an increase in the enterovirus B species of eukaryotic picornaviruses, not previously detected in fecal virome studies. Mice humanized with non-IBD colon tissue viromes were protected from intestinal inflammation, whereas IBD virome mice exhibited exacerbated inflammation in a nucleic acid sensing–dependent fashion. Furthermore, there were detrimental consequences for IBD patient–derived intestinal epithelial cells bearing loss-of-function mutations within virus sensor MDA5 when exposed to viromes. Our results demonstrate that innate recognition of IBD or non-IBD human viromes autonomously influences intestinal homeostasis and disease phenotypes. Thus, perturbations in the intestinal virome, or an altered ability to sense the virome due to genetic variation, contribute to the induction of IBD. Harnessing the virome may offer therapeutic and biomarker potential.

SummaryAltered enteric microorganisms in concert with host genetics shape inflammatory bowel disease (IBD) phenotypes. However, insight is limited to bacteria and fungi. We found virus like particles (VLPs) enriched from normal human colon resections, containing eukaryotic viruses and bacteriophages (collectively, the virome), actively elicited atypical anti-inflammatory innate immune programs. Conversely, IBD patient VLPs provoked inflammation, which was successfully dampened by healthy VLPs. The IBD colon tissue virome was perturbed, including enriched PicornovirusEnterovirus B,not previously observed in fecal virome studies. Mice with humanized healthy colon tissue viromes had attenuated intestinal inflammation while those with IBD-derived viromes exhibited exacerbated inflammation in a nucleic acid sensing-dependent fashion. Furthermore, there were detrimental consequences for IBD-associated MDA5 loss-of-function on patient intestinal epithelial cells exposed to healthy or IBD viromes. Our results demonstrate that innate recognition of either healthy or IBD human viromes autonomously influences disease phenotypes in IBD. Harnessing the virome may offer therapeutic and biomarker potential.One Sentence SummaryHuman viromes divergently shape host immunity and disease

The generation of radiation chimeras allows researchers to substitute the hematopoietic system of a mouse with that of one or more donors. A suspension of hematopoietic stem cells (HSCs) is prepared from the bone marrow (BM) or the fetal liver (FL) of a donor mouse and adoptively transferred into an irradiated recipient. Within days, the donor's HSCs will engraft, and their progeny will quickly replace the blood cells of the recipient. This simple tool, together with the large availability of genetically modified mouse lines, can be harnessed to manipulate and study various aspects of blood cell biology in vivo. We present here protocols to generate three types of radiation chimera: (1) BM chimeras, which can assist in determining whether the origin of a genetically based phenotype is the hematopoietic or radio-resistant compartment and which are also conducive for studying the ecology of blood cells and for manipulating the environment hematopoietic cells live; (2) FL chimeras, which allow the study of hematopoietic systems from animals that carry genetic modifications incompatible with postnatal life; and (3) mixed BM chimeras, in which the hematopoietic system comprises blood cells of two different genotypes. Mixed BM chimeras can be used to identify genes that affect hematopoietic cell fitness and to establish whether secreted factors mediate a phenotype of interest. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Generation of bone marrow chimera Basic Protocol 2: Generation of fetal liver chimera Basic Protocol 3: Generation of mixed bone marrow chimera Support Protocol 1: Isolation of bone marrow cells Support Protocol 2: Cell counting by flow cytometry Support Protocol 3: Assessment of chimerism.

Summary: Stimulator of Interferon Genes (STING) is a critical component of host innate immune defense but can contribute to chronic autoimmune or autoinflammatory disease. Once activated, the cyclic guanosine monophosphate (GMP)-adenosine monophosphate (AMP) (cGAMP) synthase (cGAS)-STING pathway induces both type I interferon (IFN) expression and nuclear factor-κB (NF-κB)-mediated cytokine production. Currently, these two signaling arms are thought to be mediated by a single upstream kinase, TANK-binding kinase 1 (TBK1). Here, using genetic and pharmacological approaches, we show that TBK1 alone is dispensable for STING-induced NF-κB responses in human and mouse immune cells, as well as in vivo. We further demonstrate that TBK1 acts redundantly with IκB kinase ε (IKKε) to drive NF-κB upon STING activation. Interestingly, we show that activation of IFN regulatory factor 3 (IRF3) is highly dependent on TBK1 kinase activity, whereas NF-κB is significantly less sensitive to TBK1/IKKε kinase inhibition. Our work redefines signaling events downstream of cGAS-STING. Our findings further suggest that cGAS-STING will need to be targeted directly to effectively ameliorate the inflammation underpinning disorders associated with STING hyperactivity. : Activation of NF-κB via STING is considered to be exclusively dependent on TBK1. Balka et al. now show that, although TBK1 and its kinase activity are critical for IRF3 activation and type I IFNs, it is dispensable for NF-κB. Instead, TBK1 and IKKε act redundantly to mediate STING-induced NF-κB responses. Keywords: STING, cGAS, innate immunity, NF-κB, TBK1, IKKε, signal transduction, type I interferons, cytokines, protein kinases

Implementation of evidence-based practices can improve efficiency and effectiveness of public health efforts. Few studies have explored the political contextual factors that impact implementation of evidence-based non-communicable disease prevention (EBNCDP). This study aimed to do so in Australia, Brazil, China and the United States. Investigators conducted 10-13 qualitative, semi-structured interviews of public health practitioners working in functionally similar public health organizations in each country (total N = 50). Study participants were identified through purposive sampling and interviews were structured around an interview guide covering six domains related to EBNCDP. Interviewees from all four countries identified funding as the primary politically-influenced barrier to implementing EBNCDP. Similarly widespread barriers included government funding priorities that shift based on who is in power and the difficulty of convincing policy-makers and funders that non-communicable disease prevention is a wise investment of political capital. Policymakers who are not evidence-driven was another common barrier even in the United States and Australia, where EBNCDP is more established. Findings suggest that political contextual factors influence EBNCDP and vary to an extent by country, though certain factors seem to be universal. This can aid public health practitioners, political leaders, and policymakers in advocating for conditions and policies that encourage evidence-based practice. Refereed/Peer-reviewed

Background Little is known about the contextual factors affecting the uptake of evidence-based chronic disease interventions in the United States and in other countries. This study sought to better understand the contextual similarities and differences influencing the dissemination and implementation of evidence-based chronic disease prevention (EBCDP) in Australia, Brazil, China, and the United States. Methods Between February and July 2015, investigators in each country conducted qualitative, semi-structured interviews (total N = 50) with chronic disease prevention practitioners, using interview guides that covered multiple domains (e.g., use of and access to EBCDP interventions, barriers and facilitators to the implementation of EBCDP interventions). Results Practitioners across the four countries reported only a few programmatic areas in which repositories of EBCDP interventions were used within their workplace. Across countries, academic journals were the most frequently cited channels for accessing EBCDP interventions, though peers were commonly cited as the most useful. Lack of time and heavy workload were salient personal barriers among practitioners in Australia and the United States, while lack of expertise in developing and implementing EBCDP interventions was more pertinent among practitioners from Brazil and China. Practitioners in all four countries described an organizational culture that was unsupportive of EBCDP. Practitioners in Brazil, China and the United States cited an inadequate number of staff support to implement EBCDP interventions. A few practitioners in Australia and China cited lack of access to evidence. Partnerships were emphasized as key facilitators to implementing EBCDP interventions across all countries. Conclusions This study is novel in its cross-country qualitative exploration of multilevel constructs of EBCDP dissemination and implementation. The interviews produced rich findings about many contextual similarities and differences with EBCDP that can inform both cross-country and country-specific research and practice to address barriers and improve EBCDP implementation among the four countries long-term.

Epigenetic "readers" that recognize defined posttranslational modifications on histones have become desirable therapeutic targets for cancer and inflammation. SP140 is one such bromodomain- and plant homeodomain (PHD)-containing reader with immune-restricted expression, and single-nucleotide polymorphisms (SNPs) within SP140 associate with Crohn's disease (CD). However, the function of SP140 and the consequences of disease-associated SP140 SNPs have remained unclear. We show that SP140 is critical for transcriptional programs that uphold the macrophage state. SP140 preferentially occupies promoters of silenced, lineage-inappropriate genes bearing the histone modification H3K27me3, such as the HOXA cluster in human macrophages, and ensures their repression. Depletion of SP140 in mouse or human macrophages resulted in severely compromised microbe-induced activation. We reveal that peripheral blood mononuclear cells (PBMCs) or B cells from individuals carrying CD-associated SNPs within SP140 have defective SP140 messenger RNA splicing and diminished SP140 protein levels. Moreover, CD patients carrying SP140 SNPs displayed suppressed innate immune gene signatures in a mixed population of PBMCs that stratified them from other CD patients. Hematopoietic-specific knockdown of Sp140 in mice resulted in exacerbated dextran sulfate sodium (DSS)-induced colitis, and low SP140 levels in human CD intestinal biopsies correlated with relatively lower intestinal innate cytokine levels and improved response to anti-tumor necrosis factor (TNF) therapy. Thus, the epigenetic reader SP140 is a key regulator of macrophage transcriptional programs for cellular state, and a loss of SP140 due to genetic variation contributes to a molecularly defined subset of CD characterized by ineffective innate immunity, normally critical for intestinal homeostasis.

Pre-eclampsia (PE) is a leading cause of maternal and fetal death, characterised by an imbalance of placental growth factors and hypertension at >20 weeks gestation. Impaired maternal systemic vascular adaptations and fetal growth restriction are features of both PE and pregnant relaxin-deficient (Rln–/–) mice. The aim of the present study was to investigate whether these phenotypes in Rln–/– mice are associated with abnormal placental growth factor expression, increased soluble fms-like tyrosine kinase-1 (sFlt-1), proteinuria and/or hypertension during pregnancy. In addition, we examined relaxin and relaxin receptor (relaxin/insulin like family peptide receptor 1 (RXFP1)) mRNA expression in placentas of women with PE. There was no significant difference in placental vascular endothelial growth factor A (VegfA) and placenta growth factor (Plgf) gene expression between Rln–/– and wild-type mice. Circulating plasma sFlt-1 concentrations in pregnant mice of both genotypes and ages were increased compared with non-pregnant mice but were lower in younger pregnant Rln–/– mice compared with aged-matched Rln+/+ mice. Aged pregnant Rln–/– mice had higher urinary albumin : creatinine ratios compared with age-matched Rln+/+ mice, indicative of proteinuria. Systolic and diastolic blood pressures did not differ between genotypes. In addition, PE in women was not associated with altered placental mRNA expression of RLN2 or RXFP1 at term. Overall, the data demonstrate that pregnant Rln–/– mice do not have the typical characteristics of PE. However, these mice show evidence of proteinuria, but we suggest that this results from systemic renal vascular dysfunction before pregnancy.