Iron capture through CD71 drives perinatal and tumor-associated Treg expansion.

Besides suppressing immune responses, regulatory T cells (Tregs) maintain tissue homeostasis and control systemic metabolism. Whether iron is involved in Treg-mediated tolerance is completely unknown. Here, we showed that the transferrin receptor CD71 was upregulated on activated Tregs infiltrating human liver cancer. Mice with a Treg-restricted CD71 deficiency spontaneously developed a scurfy-like disease, caused by impaired perinatal Treg expansion. CD71-null Tregs displayed decreased proliferation and tissue-Treg signature loss. In perinatal life, CD71 deficiency in Tregs triggered hepatic iron overload response, characterized by increased hepcidin transcription and iron accumulation in macrophages. Lower bacterial diversity, and reduction of beneficial species, were detected in the fecal microbiota of CD71 conditional knockout neonates. Our findings indicate that CD71-mediated iron absorption is required for Treg perinatal expansion and is related to systemic iron homeostasis and bacterial gut colonization. Therefore, we hypothesize that Tregs establish nutritional tolerance through competition for iron during bacterial colonization after birth.

Divergent roles for STAT4 in shaping differentiation of cytotoxic ILC1 and NK cells during gut inflammation.

Natural killer (NK) cells and type 1 innate lymphoid cells (ILC1) require signal transducer and activator of transcription 4 (STAT4) to elicit rapid effector responses and protect against pathogens. By combining genetic and transcriptomic approaches, we uncovered divergent roles for STAT4 in regulating effector differentiation of these functionally related cell types. Stat4 deletion in Ncr1-expressing cells led to impaired NK cell terminal differentiation as well as to an unexpected increased generation of cytotoxic ILC1 during intestinal inflammation. Mechanistically, Stat4-deficient ILC1 exhibited upregulation of gene modules regulated by STAT5 in vivo and an aberrant effector differentiation upon in vitro stimulation with IL-2, used as a prototypical STAT5 activator. Moreover, STAT4 expression in NCR+ innate lymphocytes restrained gut inflammation in the dextran sulfate sodium-induced colitis model limiting pathogenic production of IL-13 from adaptive CD4+ T cells in the large intestine. Collectively, our data shed light on shared and distinctive mechanisms of STAT4-regulated transcriptional control in NK cells and ILC1 required for intestinal inflammatory responses.

SCF and IL-33 regulate mouse mast cell phenotypic and functional plasticity supporting a pro-inflammatory microenvironment.

Mast cells (MCs) are multifaceted innate immune cells often present in the tumor microenvironment (TME). Several recent findings support their contribution to the transition from chronic inflammation to cancer. However, MC-derived mediators can either favor tumor progression, inducing the spread of the tumor, or exert anti-tumorigenic functions, limiting tumor growth. This apparent controversial role likely depends on the plastic nature of MCs that under different microenvironmental stimuli can rapidly change their phenotype and functions. Thus, the exact effect of unique MC subset(s) during tumor progression is far from being understood. Using a murine model of colitis-associated colorectal cancer, we initially characterized the MC population within the TME and in non-lesional colonic areas, by multicolor flow cytometry and confocal microscopy. Our results demonstrated that tumor-associated MCs harbor a main connective tissue phenotype and release high amounts of Interleukin (IL)-6 and Tumor Necrosis Factor (TNF)-α. This MC phenotype correlates with the presence of high levels of Stem Cell Factor (SCF) and IL-33 inside the tumor. Thus, we investigated the effect of SCF and IL-33 on primary MC cultures and underscored their ability to shape MC phenotype eliciting the production of pro-inflammatory cytokines. Our findings support the conclusion that during colonic transformation a sustained stimulation by SCF and IL-33 promotes the accumulation of a prevalent connective tissue-like MC subset that through the secretion of IL-6 and TNF-α maintains a pro-inflammatory microenvironment.

The Regulatory Activity of Noncoding RNAs in ILCs.

Innate lymphoid cells (ILCs) are innate lymphocytes playing essential functions in protection against microbial infections and participate in both homeostatic and pathological contexts, including tissue remodeling, cancer, and inflammatory disorders. A number of lineage-defining transcription factors concurs to establish transcriptional networks which determine the identity and the activity of the distinct ILC subsets. However, the contribution of other regulatory molecules in controlling ILC development and function is also recently emerging. In this regard, noncoding RNAs (ncRNAs) represent key elements of the complex regulatory network of ILC biology and host protection. ncRNAs mostly lack protein-coding potential, but they are endowed with a relevant regulatory activity in immune and nonimmune cells because of their ability to control chromatin structure, RNA stability, and/or protein synthesis. Herein, we summarize recent studies describing how distinct types of ncRNAs, mainly microRNAs, long ncRNAs, and circular RNAs, act in the context of ILC biology. In particular, we comment on how ncRNAs can exert key effects in ILCs by controlling gene expression in a cell- or state-specific manner and how this tunes distinct functional outputs in ILCs.

NK cell and ILC heterogeneity in colorectal cancer. New perspectives from high dimensional data.

Innate lymphoid cells (ILCs) and tissue-resident natural killer (NK) cells ensure immunity at environmental interfaces and help maintain barrier integrity of the intestinal tract. This wide range of innate lymphocytes is able to provide fast and potent inflammatory responses that, when deregulated, have been associated with pathogenesis of inflammatory bowel disease (IBD) and colorectal cancer (CRC). While the presence of tumor-infiltrating NK cells is generally associated with a favorable outcome in CRC patients, emerging evidence reveals distinct roles for ILCs in regulating CRC pathogenesis and progression. Advances in next generation sequencing technology, and in particular of single-cell RNA-seq approaches, along with multidimensional flow cytometry analysis, have helped to deconvolute the complexity and heterogeneity of the ILC system both in homeostatic and pathological contexts. In this review, we discuss the protective and detrimental roles of NK cells and ILCs in the pathogenesis of CRC, focusing on the phenotypic and transcriptional modifications these cells undergo during CRC development and progression.

Magnesium Absorption in Intestinal Cells: Evidence of Cross-Talk between EGF and TRPM6 and Novel Implications for Cetuximab Therapy.

Hypomagnesemia is very commonly observed in cancer patients, most frequently in association with therapy with cetuximab (CTX), a monoclonal antibody targeting the epithelial growth factor receptor (EGFR). CTX-induced hypomagnesemia has been ascribed to renal magnesium (Mg) wasting. Here, we sought to clarify whether CTX may also influence intestinal Mg absorption and if Mg supplementation may interfere with CTX activity. We used human colon carcinoma CaCo-2 cells as an in vitro model to study the mechanisms underlying Mg transport and CTX activity. Our findings demonstrate that TRPM6 is the key channel that mediates Mg influx in intestinal cells and that EGF stimulates such influx; consequently, CTX downregulates TRPM6-mediated Mg influx by interfering with EGF signaling. Moreover, we show that Mg supplementation does not modify either the CTX IC50 or CTX-dependent inhibition of ERK1/2 phosphorylation. Our results suggest that reduced Mg absorption in the intestine may contribute to the severe hypomagnesemia that occurs in CTX-treated patients, and Mg supplementation may represent a safe and effective nutritional intervention to restore Mg status without impairing the CTX efficacy.

JAK Inhibition Differentially Affects NK Cell and ILC1 Homeostasis.

Janus kinase (JAK) inhibitors are widely used in the treatment of multiple autoimmune and inflammatory diseases. Immunologic and transcriptomic profiling have revealed major alterations on natural killer (NK) cell homeostasis associated with JAK inhibitions, while information on other innate lymphoid cells (ILCs) is still lacking. Herein, we observed that, in mice, the homeostatic pool of liver ILC1 was less affected by JAK inhibitors compared to the pool of NK cells present in the liver, spleen and bone marrow. JAK inhibition had overlapping effects on the transcriptome of both subsets, mainly affecting genes regulating cell cycle and apoptosis. However, the differential impact of JAK inhibition was linked to the high levels of the antiapoptotic gene Bcl2 expressed by ILC1. Our findings provide mechanistic explanations for the effects of JAK inhibitors on NK cells and ILC1 which could be of major clinically relevance.

TRPM6 is Essential for Magnesium Uptake and Epithelial Cell Function in the Colon.

Intestinal magnesium (Mg) uptake is essential for systemic Mg homeostasis. Colon cells express the two highly homologous transient receptor potential melastatin type (TRPM) 6 and 7 Mg2+ channels, but their precise function and the consequences of their mutual interaction are not clear. To explore the functional role of TRPM6 and TRPM7 in the colon, we used human colon cell lines that innately express both channels and analyzed the functional consequences of genetic knocking-down, by RNA interference, or pharmacological inhibition, by NS8593, of either channel. TRPM7 silencing caused an increase in Mg2+ influx, and correspondingly enhanced cell proliferation and migration, while downregulation of TRPM6 did not affect significantly either Mg2+ influx or cell proliferation. Exposure to the specific TRPM6/7 inhibitor NS8593 reduced Mg2+ influx, and consequently cell proliferation and migration, but Mg supplementation rescued the inhibition. We propose a model whereby in colon cells the functional Mg2+ channel at the plasma membrane may consist of both TRPM7 homomers and TRPM6/7 heteromers. A different expression ratio between the two proteins may result in different functional properties. Altogether, our findings confirm that TRPM6 cannot be replaced by TRPM7, and that TRPM6/7 complexes and TRPM6/7-mediated Mg2+ influx are indispensable in human epithelial colon cells.

Dietary Magnesium Alleviates Experimental Murine Colitis Through Upregulation of the Transient Receptor Potential Melastatin 6 Channel.

Background: Magnesium (Mg) is essential for human health and is absorbed mainly in the intestine. In view of the likely occurrence of an Mg deficit in inflammatory bowel disease (IBD) and the documented role of Mg in modulating inflammation, the present study addresses whether Mg availability can affect the onset and progression of intestinal inflammation. Methods: To study the correlation between Mg status and disease activity, we measured magnesemia by atomic absorption spectroscopy in a cohort of IBD patients. The effects of dietary Mg modulation were assessed in a murine model of dextran sodium sulfate (DSS)-induced colitis by monitoring magnesemia, weight, fecal occult blood, diarrhea, colon length, and histology. Expression of the transient receptor potential melastatin (TRPM) 6 channel was assessed by real-time reverse transcription polymerase chain reaction and immunohistochemistry in murine colon tissues. The effect of Mg on epithelial barrier formation/repair was evaluated in human colon cell lines. Results: Inflammatory bowel disease patients presented with a substantial Mg deficit, and serum Mg levels were inversely correlated with disease activity. In mice, an Mg-deficient diet caused hypomagnesemia and aggravated DSS-induced colitis. Colitis severely compromised intestinal Mg2+ absorption due to mucosal damage and reduction in TRPM6 expression, but Mg supplementation resulted in better restoration of mucosal integrity and channel expression. Conclusions: Our results highlight the importance of evaluating and correcting magnesemia in IBD patients. The murine model suggests that Mg supplementation may represent a safe and cost-effective strategy to reduce inflammation and restore normal mucosal function.