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Glia-derived factors in innate lymphoid cell sensing and intestinal defence

Periodic Reporting for period 5 - GliaInnateSensing (Glia-derived factors in innate lymphoid cell sensing and intestinal defence)

Reporting period: 2020-07-01 to 2021-12-31

The interplay between intestinal microbes and immune cells ensures vital functions of the organism. However, inadequate host-microbe relationships lead to inflammatory diseases that are major public health concerns, such as inflammatory bowel disease (IBD).

Innate lymphoid cells (ILC) are an emergent family of effectors abundantly present at mucosal sites.Our study has so far revealed a novel and fully unexplored molecule that controls ILC function. If successful our work will provide better knowledge on the relationship between ILC and intestinal inflammation, which will be important for future innovative therapies in intestinal inflammatory disease.

The researchers are pursuing their work using genetically modified models with altered neurotrophic factor signals to learn more on how ILC control intestinal inflammation. Their findings will provide better knowledge on the relationships between ILC, genetic factors and intestinal bacteria in the inflamed intestine, which will help the design of future therapies and possible prevention strategies of intestinal inflammatory disease.

Group 3 innate lymphoid cells (ILC3) produce pro-inflammatory cytokines, regulate mucosal homeostasis, anti-microbial defence and adaptive immune responses. In addition to their well-established developmentally regulated program, ILC3 are also controlled by microbial and dietary signals raising the hypothesis that ILC3 possess other unexpected environmental sensing strategies.

We hypothesise that ILC3 sense their environment and exert their function as part of a novel epithelial-glial-ILC unit orchestrated by neurotrophic factors.

Our ground-breaking research will establish a novel sensing program by which ILC3 integrate environmental cues and will define a key multi-cellular unit at the core of intestinal homeostasis and defence. Finally, our work will reveal new pathways that may be targeted in inflammatory diseases that are major Public Health concerns.
We employed genetic, cellular and molecular approaches to decipher how this unconventional epithelial-glial-ILC unit is controlled and how glial-derived factors set ILC3 function and intestinal homeostasis.
We assessed ILC3-autonomous functions of neurotrophic factor receptors. ILC3-specific loss and gain of function mutant mice for neuroregulatory receptors were used to define the role of these molecules in ILC3 function, mucosal homeostasis, gut defence and microbial ecology. In addiction we deciphered the anatomical and functional basis for the enteric epithelial-glial-ILC unit. To this end we employ high-resolution imaging, genome-wide expression analysis and tissue-specific mutants for define target genes.

Group 3 innate lymphoid cells (ILC3) are major regulators of inflammation and infection at mucosal barriers. ILC3 development is thought to be programmed, but how ILC3 perceive, integrate and respond to local environmental signals remains unclear. Here we show that ILC3 in mice sense their environment and control gut defence as part of a glial–ILC3–epithelial cell unit orchestrated by neurotrophic factors. We found that enteric ILC3 express the neuroregulatory receptor RET. ILC3-autonomous Ret ablation led to decreased innate interleukin-22 (IL-22), impaired epithelial reactivity, dysbiosis and increased susceptibility to bowel inflammation and infection. Neurotrophic factors directly controlled innate Il22 downstream of the p38 MAPK/ERK-AKT cascade and STAT3 activation. Notably, ILC3 were adjacent to neurotrophic-factor-expressing glial cells that exhibited stellate-shaped projections into ILC3 aggregates. Glial cells sensed microenvironmental cues in a MYD88-dependent manner to control neurotrophic factors and innate IL-22. Accordingly, glial-intrinsic Myd88 deletion led to impaired production of ILC3-derived IL-22 and a pronounced propensity towards gut inflammation and infection. Our work sheds light on a novel multi-tissue defence unit, revealing that glial cells are central hubs of neuron and innate immune regulation by neurotrophic factor signals.
We identified a molecule, the neuroregulator RET, which controls tissue-protection and antimicrobial activity conferred by ILC3 in the intestine. Tuning RET activity in ILC3 regulate propensity to intestinal inflammation, a major hallmark and cause of IBD.

We have learned the existence of a novel glial-ILC3-epithelial cell unit orchestrated by neurotrophic factors. Notably, glial-derived neurotrophic factors operate in an ILC3-intrinsic manner by activating the tyrosine kinase RET, which directly regulates innate IL-22
downstream of p38 MAPK/ERK-AKT and STAT3 phosphorylation.

Our data reveal a novel and fully unexplored molecule that controls intestinal ILC function and gut inflammation, establishing the link between nervous system activity activity and immune responses in intestinal inflammatory disease.

Our data demonstrate that in addition to their well-established capacity to integrate dendritic cell-derived cytokines, ILC3 perceive distinct multi-tissue regulatory signals leading to STAT3 activity and IL-22 expression, notably via integration of glial cell-derived neuroregulators.

Thus, we have learned that glial cells as central hubs of neuronal and innate immune regulation. Notably, neurotrophic factors are the molecular link between glial cell sensing, innate IL-22 and intestinal epithelial defence in bowel disease. Thus, it is tempting to speculate that glial/immune cell units might be also critical to the homeostasis and defence of the intestine in IBD patients.

Recently, Henrique Veiga-Fernandes’ laboratory find that small intestine lamina propria innate lymphoid cells expresses the Neuromedin u receptor 1 (Nmur1), with a major impact in in vitro cytokine production. In order to investigate the in vivo role of Nmur1 in innate lymphoid cells function, was critical to generate an accurate genetic mouse model. With this development, we generate Nmur1 conditional knockout mice. To accomplish that, the exon 1 of Nmur1 gene are flanked with loxP sites.
In sum, these mice will provide us unique and flexible tools to investigate the role of Nmur1 in the function of innate lymphoid cell and consequently in intestinal mucosal homeostasis.