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Group 3 innate lymphoid cells (ILC3s) are RORγT+lymphocytes that are predominately enriched in mucosal tissues and produce IL-22 and IL-17A. They are the innate counterparts of Th17 cells. While Th17 lymphocytes utilize unique metabolic pathways in their differentiation program, it is unknown whether ILC3s make similar metabolic adaptations. We employed single-cell RNA sequencing and metabolomic profiling of intestinal ILC subsets to identify an enrichment of polyamine biosynthesis in ILC3s, converging on the rate-limiting enzyme ornithine decarboxylase (ODC1). In vitro and in vivo studies demonstrated that exogenous supplementation with the polyamine putrescine or its biosynthetic substrate, ornithine, enhanced ILC3 production of IL-22. Conditional deletion of ODC1 in ILC3s impaired mouse antibacterial defense againstCitrobacter rodentiuminfection, which was associated with a decrease in anti-microbial peptide production by the intestinal epithelium. Furthermore, in a model of anti-CD40 colitis, deficiency of ODC1 in ILC3s markedly reduced the production of IL-22 and severity of inflammatory colitis. We conclude that ILC3-intrinsic polyamine biosynthesis facilitates efficient defense against enteric pathogens as well as exacerbates autoimmune colitis, thus representing an attractive target to modulate ILC3 function in intestinal disease.

The signature hallmark of adaptive immunity is the evolution of somatically rearranged antigen receptors, which confer both diversity and specificity to T and B lymphocytes. For decades, immunologists have observed cells which possess lymphoid characteristics yet lack such antigen-specific receptors. Collectively, these populations are referred to as innate lymphoid cells (ILCs) (Vivier et al. in Cell 174(5):1054-1066, 2018). Cytotoxic natural killer (NK) cells and lymphoid tissue-inducing cells (LTi), which contribute to the formation of lymphoid organs during embryogenesis, are the earliest described ILCs. Subsequently, diverse populations of ILCs have been described based on the signature cytokines they produce. Group 1 ILCs (ILC1) produce IFNγ, group 2 ILCs (ILC2) produce IL-5 and IL-13, and group 3 ILCs (ILC3) produce IL-22 and IL-17. In contrast to adaptive lymphocytes which take several days to undergo clonal expansion and acquire effector functions, ILCs secrete cytokines rapidly in response to activating signals in their tissue of residence. ILCs may also directly regulate adaptive lymphocytes and myeloid cells through co-stimulatory molecules and soluble factors. Thus, ILCs play important roles in both the initiation and amplification of the immune response. When properly regulated, ILCs maintain intestinal homeostasis and protect the host from infection by various pathogens. However, dysregulation of mucosal immunity drives intestinal inflammation and contributes to pathology, such as inflammatory bowel disease (IBD). In this review, we outline the roles that ILCs play in amplifying or regulating intestinal inflammation as well as ongoing efforts to target these disease mechanisms for IBD therapy.

Metformin is a first-line drug in the treatment of type-2 diabetes mellitus (T2DM). In addition to its antigluconeogenic and insulin-sensitizing properties, metformin has emerged as a potent inhibitor of the chronic inflammatory response of macrophages. In particular, metformin treatment has been shown to reduce expression of interleukin (IL-) 1β during long-term exposure to the pro-inflammatory stimulus lipopolysaccharide (LPS) through a reduction in reactive oxygen species (ROS), which decreases the levels of the hypoxia-inducible factor (HIF) 1-α, and through enhanced expression of IL-10. However, the effect of metformin on the acute inflammatory response, before significant levels of ROS accumulate in the cell, has not been explored. Here, we show that metformin alters the acute inflammatory response through its activation of AMP-activated protein kinase (AMPK), but independently of HIF1-α and IL-10, in primary macrophages and two macrophage-like cell lines. Thus, metformin changes the acute and the chronic inflammatory response through fundamentally distinct mechanisms. Furthermore, RNA-seq analysis reveals that metformin pretreatment affects the levels of a large yet selective subset of inflammatory genes, dampening the response to short-term LPS exposure and affecting a wide range of pathways and biological functions. Taken together, these findings reveal an unexpected complexity in the anti-inflammatory properties of this widely used drug.

Genetic studies have highlighted microglia as pivotal in orchestrating Alzheimer's disease (AD). Microglia that adhere to Aβ plaques acquire a transcriptional signature, "disease-associated microglia" (DAM), which largely emanates from the TREM2-DAP12 receptor complex that transmits intracellular signals through the protein tyrosine kinase SYK. The human TREM2

In a recent study, Masuda and colleagues (Nature 2019;566:388–392) used single-cell RNA-sequencing (scRNA-seq) to profile microglia across different anatomical compartments, developmental stages, and types of brain pathology in mice. Moreover, the authors performed a novel transcriptomic characterization of microglia from multiple sclerosis patients and identified phenotypically conserved microglial subsets between species. These findings, together with seminal prior results from various groups, provide valuable insights into the spatiotemporal heterogeneity of microglia during brain development and disease.

Innate lymphoid cells (ILCs) are tissue-resident lymphocytes categorized on the basis of their core regulatory programs and the expression of signature cytokines. Human ILC3s that produce the cytokine interleukin-22 convert into ILC1-like cells that produce interferon-γ in vitro, but whether this conversion occurs in vivo remains unclear. In the present study we found that ILC3s and ILC1s in human tonsils represented the ends of a spectrum that included additional discrete subsets. RNA velocity analysis identified an intermediate ILC3-ILC1 cluster, which had strong directionality toward ILC1s. In humanized mice, the acquisition of ILC1 features by ILC3s showed tissue dependency. Chromatin studies indicated that the transcription factors Aiolos and T-bet cooperated to repress regulatory elements active in ILC3s. A transitional ILC3-ILC1 population was also detected in the human intestine. We conclude that ILC3s undergo conversion into ILC1-like cells in human tissues in vivo, and that tissue factors and Aiolos were required for this process.

Qian et al. shows that ILC2s can be generated from not only thymic multipotent progenitors but also committed T cell precursors. These processes are greatly suppressed by E protein transcription factors. Thymic ILC2s show functional differences from those made elsewhere., Current models propose that group 2 innate lymphoid cells (ILC2s) are generated in the bone marrow. Here, we demonstrate that subsets of these cells can differentiate from multipotent progenitors and committed T cell precursors in the thymus, both in vivo and in vitro. These thymic ILC2s exit the thymus, circulate in the blood, and home to peripheral tissues. Ablation of E protein transcription factors greatly promotes the ILC fate while impairing B and T cell development. Consistently, a transcriptional network centered on the ZBTB16 transcription factor and IL-4 signaling pathway is highly up-regulated due to E protein deficiency. Our results show that ILC2 can still arise from what are normally considered to be committed T cell precursors, and that this alternative cell fate is restrained by high levels of E protein activity in these cells. Thymus-derived lung ILC2s of E protein–deficient mice show different transcriptomes, proliferative properties, and cytokine responses from wild-type counterparts, suggesting potentially distinct functions., Graphical Abstract

Lymphoid tissue inducer (LTi)-like cells are tissue resident innate lymphocytes that rapidly secrete cytokines that promote gut epithelial integrity and protect against extracellular bacterial infections.Here, we report that the retention of LTi-like cells in conventional solitary intestinal lymphoid tissue (SILT) is essential for controlling LTi-like cell function and is maintained by expression of the chemokine receptor CXCR5. Deletion of Cxcr5 functionally unleashed LTi-like cells in a cell intrinsic manner, leading to uncontrolled IL-17 and IL-22 production. The elevated production of IL-22 in Cxcr5-deficient mice improved gut barrier integrity and protected mice during infection with the opportunistic pathogen Clostridium difficile. Interestingly, Cxcr5(−/−) mice developed LTi-like cell aggregates that were displaced from their typical niche at the intestinal crypt, and LTi-like cell hyperresponsiveness was associated with the local formation of this unconventional SILT. Thus, LTi-like cell positioning within mucosa controls their activity via niche-specific signals that temper cytokine production during homeostasis.

Getting around the blood–brain barrier The meninges comprise three membranes that surround and protect the central nervous system (CNS). Recent studies have noted the existence of myeloid cells resident there, but little is known about their ontogeny and function, and whether other meningeal immune cell populations have important roles remains unclear (see the Perspective by Nguyen and Kubes). Cugurra et al. found in mice that a large proportion of continuously replenished myeloid cells in the dura mater are not blood derived, but rather transit from cranial bone marrow through specialized channels. In models of CNS injury and neuroinflammation, the authors demonstrated that these myeloid cells have an immunoregulatory phenotype compared with their more inflammatory blood-derived counterparts. Similarly, Brioschi et al. show that the meninges host B cells that are also derived from skull bone marrow, mature locally, and likely acquire a tolerogenic phenotype. They further found that the brains of aging mice are infiltrated by a second population of age-associated B cells, which come from the periphery and may differentiate into autoantibody-secreting plasma cells after encountering CNS antigens. Together, these two studies may inform future treatment of neurological diseases. Science , abf7844, abf9277, this issue p. eabf7844 , p. eabf9277 ; see also abj8183, p. 396

Natural killer (NK) cells and type 1 innate lymphoid cells (ILC1s) are heterogenous innate lymphocytes broadly defined in mice as Lin(−)NK1.1(+)NKp46(+) cells that express the transcription factor T-BET and produce interferon-γ. The ILC1 definition primarily stems from studies on liver and small intestinal populations. However, NK1.1(+)NKp46(+) cells in the salivary glands, uterus, adipose, and other tissues exhibit nonuniform programs that differ from those of liver or intestinal ILC1s or NK cells. Here, we performed single-cell RNA sequencing on murine NK1.1(+)NKp46(+) cells from blood, spleen, various tissues, and solid tumors. We identified gene expression programs of tissue-specific ILC1s, tissue-specific NK cells, and non-tissue-specific populations in blood, spleen, and other tissues largely corresponding to circulating cells. Moreover, we found that circulating NK cell programs were reshaped in tumor-bearing mice. Core programs of circulating and tumor NK cells paralleled conserved human NK cells signatures, advancing our understanding of the human NK-ILC1 spectrum.

The basic helix-loop-helix transcription factors collectively called E proteins powerfully suppress the differentiation of group2 innate lymphoid cells from bone marrow and thymic progenitors. Here we investigated the underlying molecular mechanisms using inducible gain and loss of function approaches in ILC2s and their precursors, respectively. Cross-examination of RNA sequencing and ATAC sequencing data obtained at different time points reveals a set of genes which are likely direct targets of E proteins. Consequently, a widespread down-regulation of chromatin accessibility occurs at a later time point, possibly due to the activation of transcriptional repressor genes such as Cbfa2t3 and Jdp2. The large number of genes repressed by gain of E protein function leads to the down-regulation of a transcriptional network important for ILC2 differentiation.SummaryDifferentiation of group 2 innate lymphoid cells is forcefully repressed by E protein transcription factors. This report elucidates how E proteins repress a transcriptional network important for ILC2 differentiation by up-regulating the expression of transcriptional repressors.

The diversity of natural killer cells between mouse and human is poorly understood. In this issue of Immunity, Crinier et al. (2018) utilize single-cell RNA-seq to profile splenic and blood NK cells from both organisms, uncovering both tissue- and species-specific transcriptomic signatures.

Background & Aims: Fatigue is frequent and disabling in patients with inflammatory bowel diseases (IBD) but its mechanisms are poorly understood. We investigated alterations in fecal microbiomes and serum metabolomes and proteomes in patients with quiescent IBD, with vs without fatigue. Methods: We performed a prospective observational study of patients (44% women; mean age, 39.8 y) with clinically and endoscopically quiescent Crohn's disease (n = 106) or ulcerative colitis (n = 60) at a tertiary hospital, from March 2016 through December 2018. Fatigue was assessed using the functional assessment of chronic illness therapy-fatigue scoring system and defined as a score of 43 or less. We performed metabolomic analysis of serum samples using liquid chromatography-mass spectrometry methods and proteomic analysis using multiplex proximity extension assay (PEA) technology. Stool samples were obtained from 50 patients and analyzed by shotgun metagenomic sequencing on Illumina HiSeq platform. Results: Of the 166 study participants, 91 (55%) were fatigued. Serum samples from patients with fatigue (n = 59) did not have significant increases in levels of inflammatory cytokines compared with serum samples from nonfatigued patients (n = 72). We found a statistically significant difference in a cluster of 18 serum metabolites between patients with fatigue (n = 84) vs without fatigue (n = 72) (P = .033); serum samples from patients with fatigue had significant reductions in levels of methionine (P = .020), tryptophan (P = .042), proline (P = .017), and sarcosine (P = .047). Fecal samples from patients with fatigue had a less diverse gut microbiome, with significant reductions in butyrate-producing bacteria, including Faecalibacterium prausnitzii (P = .0002, q =.007) and Roseburia hominis (P = .0079, q = 0.105). This fatigue-like microbiome was associated with fatigue scales and correlated with progressive depletion of metabolites from serum samples. Conclusions: In an analysis of fecal and serum samples from 166 patients with IBD, we found alterations in serum metabolites and fecal microbes that were associated with fatigue.

Innate lymphoid cells (ILCs) are tissue-resident lymphocytes devoid of antigen-specific receptors that provide an immediate source of cytokines in early immune responses and are enriched in mucosal tissues. Human ILC3s that produce the cytokine interleukin-22 and express the transcription factor RORγt have been shown to be able to convert into ILC1-like cells that produce mainly IFN-γ and express T-bet, in vitro. Whether this conversion also occurs in vivo remains unclear. In the present study we found that ILC3s and ILC1s in human tonsils represented the ends of a spectrum that included additional discrete subsets with intermediate transcriptional signatures. This finding strongly suggests that the ILC3 to ILC1-like conversion also occurs in vivo in humans. Gene expression analysis showed that the transcription factors Aiolos and T-bet were induced in the subsets closer to the canonical ILC1 subset. Chromatin studies indicated that Aiolos and T-bet cooperate to repress regulatory elements active in ILC3s. Ectopic expression of Aiolos and T-bet in the ILC3-like cell line MNK-3 repressed the production of IL-22 and induced IFN-γ. We conclude that Aiolos and T-bet cooperate to repress the ILC3 transcriptional program therefore contributing to the ILC3 to ILC1-like conversion.

Under conventional conditions, mice deficient in core 1-derived O-glycans (TM-IEC C1galt1(-/-)), which have a defective mucus layer, experienced spontaneous inflammation of the colon. Analysis of fecal bacterial populations by pyrosequencing of 16S rRNA gene showed that disease in conventional TM-IEC C1galt1(-/-) was associated with shifts in the microbiota manifested by increases in Lactobacillus and Clostridium species, and decreases in unclassified Ruminococcaceae and Lachnospiraceae. Under germ-free (GF) conditions, TM-IEC C1galt1(-/-) presented decreased goblet cells, but did not develop inflammation. Monoassociation of GF TM-IEC C1galt1(-/-) revealed that bacterial species differ significantly in their ability to induce inflammatory changes. Bacteroides thetaiotaomicron caused inflammation, while Lactobacillus johnsonii (enriched during colitis) did not. These observations demonstrate that not all microbiota shifts that correlate with disease contribute to pathogenesis.

Measuring investment risk precisely is critical to investment strategies. In common practice, the most popular measure of risk is standard deviation. However, standard deviation makes no distinction between positive and negative deviations from the mean. Such risk measurement could lead to biased decision making, given that asset returns are generally not symmetrically distributed.In this paper, risk is confined to adverse outcomes that are measured by the negative semi-deviation. By examining Australian Listed Property Trusts (LPTs) for the period of 1985- 2004, this paper illustrates how the differentiation between risk (semi-deviation) and uncertainty (standard deviation) make significant differences in performance measurement and optimal portfolio construction. This paper demonstrates that the concept and application of downside risk is a valuable construct to LPT investment strategies.

This study focuses on the efficient market hypothesis (EMH) and inspects the existence of calendar anomalies in the Australian capital markets. It is the first study that provides a comprehensive examination of calendar anomalies in the Australian capital markets, particularly in the Australian Listed Property Trust (LPT) market. The study reveals that calendar anomalies exist in both the Australian LPT market and the broader equity market, providing evidence of market inefficiency. It is also found that some of these return irregularities are diminishing and dissipating over time, suggesting the move towards a higher level of efficiency.The results from this study provide support to active investing. Moreover, this study quantifies the potential for profiting on the calendar anomalies by active investing in a practical sense and provides significant practical implications for LPT portfolio managers and investors.

This study evaluates the performance of Australian Listed Property Trusts (LPTs) in the context of selectivity and timing over June 1998 to May 2003, and provides significant practical implications for investment strategies for LPT managers. The importance of benchmark indices and model specifications to performance evaluation is tested and highlighted in this paper. After specifying the appropriate model, the author has defined and constructed the appropriate benchmark indices for evaluating the performance of Australian LPTs. The study results provide evidence of superior strategic skills of Australian LPT managers over the study period based on study results.This paper further illustrates the importance of longer-term market forecasts, especially in the circumstance where a lengthy time is required to apply timing skills. It suggests that managers who are superior in longer-term macro forecasts will have the edge to outperform the market portfolio and competitors, providing significant implicat...

This study focuses on weak form EMH and investigates whether the Australian Listed Property Trust (LPT) market and broader equity market (All Ordinaries) are efficient in the weak form. It examines the daily closing price indices of LPTs and All Ordinaries for the period of 1 June 1992 to 30 May 2003. The results suggest that Australian LPTs and the broader equity markets are efficient in the weak form, and securities prices are not predictable based on historical price movements.The evidence of weak form efficiency has significant implications for investment strategies. It questions the relevance of technical analysis in the process of formulating investment/trading strategies. In the wake of questionable significance of technical analysis, this study provides active managers with suggestions for formulating investment strategies and investment decision making.

T cell lymphoblastic leukemia (T-ALL) is known to be associated with chromosomal abnormalities that lead to aberrant expression of a number of transcription factors such as TAL1, which dimerizes with basic helix-loop-helix (bHLH) E proteins and inhibits their function. Activated Notch receptors also efficiently induce T cell leukemogenesis in mouse models. Interestingly, gain-of-function mutations or cryptic transcription initiation of the Notch1 gene have been frequently found in both human and mouse T-ALL. However, the correlations between these alterations and overall Notch activities or leukemogenesis have not been thoroughly evaluated. Therefore, we made use of our collection of T cell lymphomas developed in transgenic mice expressing Id1, which like TAL1, inhibits E protein function. By comparing expression levels of Notch target genes in Id1-expressing tumors to those in tumors induced by a constitutively active form of Notch1, N1C, we were able to assess the overall activities of Notch pathways and conclude that the majority of Id1-expressing tumors had elevated Notch function to a varying degree. However, 26% of the Id1-expressing tumors had no evidence of enhanced Notch activation, but that did not delay the onset of tumorigenesis. Furthermore, we examined the genetic or epigenetic alterations thought to contribute to ligand-independent activation or protein stabilization of Notch1 and found that some of the Id1-expressing tumors acquired these changes, but they are not uniformly associated with elevated Notch activities in Id1 tumor samples. In contrast, N1C-expressing tumors do not harbor any PEST domain mutations nor exhibit intragenic transcription initiation. Taken together, it appears that Notch activation provides Id1-expressing tumor cells with selective advantages in growth and survival. However, this may not be absolutely essential for lymphomagenesis in Id1 transgenic mice and additional factors could also cooperate with Id1 to induce T cell lymphoma. Therefore, a broad approach is necessary in designing T-ALL therapy.

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