Enteric glia bring fresh WNT to the intestinal stem cell niche

Intestinal stem cells continuously self-renew and differentiate into a variety of specialized epithelial cells that maintain gut health. New research in this issue of Cell Stem Cell (Baghdadi et al., 2022Baghdadi M.B. Ayyaz A. Coquenlorge S. Chu B. Kumar S. Streutker C. Wrana J.L. Kim T.H. Enteric glial cell heterogeneity regulates intestinal stem cell niches.Cell Stem Cell. 2022; 29 (Published online November 1, 2021): 86-100Abstract Full Text Full Text PDF Scopus (17) Google Scholar) shows that enteric glial cells regulate the intestinal stem cell niche during regeneration and disease through the production of WNT ligands. Intestinal stem cells continuously self-renew and differentiate into a variety of specialized epithelial cells that maintain gut health. New research in this issue of Cell Stem Cell (Baghdadi et al., 2022Baghdadi M.B. Ayyaz A. Coquenlorge S. Chu B. Kumar S. Streutker C. Wrana J.L. Kim T.H. Enteric glial cell heterogeneity regulates intestinal stem cell niches.Cell Stem Cell. 2022; 29 (Published online November 1, 2021): 86-100Abstract Full Text Full Text PDF Scopus (17) Google Scholar) shows that enteric glial cells regulate the intestinal stem cell niche during regeneration and disease through the production of WNT ligands. The gastrointestinal (GI) tract is essential for nutrient and water absorption, energy balance, and host defense. It is also home to myriads of microorganisms that impact virtually all aspects of health. Central to the digestive and barrier functions of the GI tract is the intestinal epithelium, a single-cell lining that is continuously renewed and has the remarkable ability to regenerate following injury or disease. Both the turnover and regenerative capacity of the intestinal epithelium are fueled by intestinal stem cells (ISCs), which proliferate and generate progeny that differentiate into an array of functionally diverse cell types. The plasticity, self-renewal, and differentiation of this specialized stem cell population is tightly regulated by the luminal and cellular microenvironment of the intestine. New elegant research by Baghdadi et al. published in this issue of Cell Stem Cell identifies a crucial role for enteric glial cells (EGCs) in the regulation of the ISC niche (Baghdadi et al., 2022Baghdadi M.B. Ayyaz A. Coquenlorge S. Chu B. Kumar S. Streutker C. Wrana J.L. Kim T.H. Enteric glial cell heterogeneity regulates intestinal stem cell niches.Cell Stem Cell. 2022; 29 (Published online November 1, 2021): 86-100Abstract Full Text Full Text PDF Scopus (17) Google Scholar). The neural-crest-derived population of EGCs is an integral part of the enteric nervous system (ENS), an expansive neuronal network that is embedded within the gut wall and controls multiple aspects of digestive physiology and intestinal homeostasis (Seguella and Gulbransen, 2021Seguella L. Gulbransen B.D. Enteric glial biology, intercellular signalling and roles in gastrointestinal disease.Nat. Rev. Gastroenterol. Hepatol. 2021; 18: 571-587Crossref PubMed Scopus (47) Google Scholar). Although much of the research so far has focused on the “canonical” functions of EGCs, namely the support and regulation of intestinal neural circuit activity, several studies have uncovered roles of these cells in broader gut physiology and immunity. Indeed, the idea that EGCs safeguard the integrity of the intestinal epithelium has been put to the test before. Using different experimental strategies in mice, three independent groups showed over 20 years ago that ablation of EGCs leads to severe intestinal inflammation, most likely due to changes in vascular and epithelial permeability (Aikawa and Suzuki, 1985Aikawa H. Suzuki K. Enteric gliopathy in niacin-deficiency induced by CNS glio-toxin.Brain Res. 1985; 334: 354-356Crossref PubMed Scopus (15) Google Scholar) (Bush et al., 1998Bush T.G. Savidge T.C. Freeman T.C. Cox H.J. Campbell E.A. Mucke L. Johnson M.H. Sofroniew M.V. Fulminant jejuno-ileitis following ablation of enteric glia in adult transgenic mice.Cell. 1998; 93: 189-201Abstract Full Text Full Text PDF PubMed Scopus (462) Google Scholar) (Cornet et al., 2001Cornet A. Savidge T.C. Cabarrocas J. Deng W.L. Colombel J.F. Lassmann H. Desreumaux P. Liblau R.S. Enterocolitis induced by autoimmune targeting of enteric glial cells: a possible mechanism in Crohn’s disease?.Proc. Natl. Acad. Sci. USA. 2001; 98: 13306-13311Crossref PubMed Scopus (250) Google Scholar). However, more recently, Rao and colleagues reported that genetic ablation of EGCs by diphtheria toxin expressed by a proteolipid protein 1 (PLP1) transgene did not affect epithelial cell renewal or permeability and did not alter susceptibility to chemically induced colitis (Rao et al., 2017Rao M. Rastelli D. Dong L. Chiu S. Setlik W. Gershon M.D. Corfas G. Enteric Glia Regulate Gastrointestinal Motility but Are Not Required for Maintenance of the Epithelium in Mice.Gastroenterology. 2017; 153: 1068-1081.e7Abstract Full Text Full Text PDF PubMed Scopus (84) Google Scholar). The reason for this discrepancy remained unclear, but the possibility that the Plp1 transgene targeted only a subset of EGCs could not be discounted. Indeed, distinct glial subpopulations with unique transcriptional profiles have been described in the ENS of adult mice and humans (Progatzky et al., 2021Progatzky F. Shapiro M. Chng S.H. Garcia-Cassani B. Classon C.H. Sevgi S. Laddach A. Bon-Frauches A.C. Lasrado R. Rahim M. et al.Regulation of intestinal immunity and tissue repair by enteric glia.Nature. 2021; 599: 125-130Crossref PubMed Scopus (28) Google Scholar; Seguella and Gulbransen, 2021Seguella L. Gulbransen B.D. Enteric glial biology, intercellular signalling and roles in gastrointestinal disease.Nat. Rev. Gastroenterol. Hepatol. 2021; 18: 571-587Crossref PubMed Scopus (47) Google Scholar) but the functional significance of EGC heterogeneity remained unclear. In their current study, Baghdadi and colleagues provide convincing evidence that adult EGCs are functionally heterogeneous, suggest a complex equilibrium between enteric glia subpopulations, and demonstrate unequivocally the key role of enteric glia in maintaining the integrity of the intestinal epithelium. In addition to PLP1, the intermediate filament glial fibrillary acidic protein (GFAP) has also been used as a marker of enteric glia but, in contrast to central nervous system astrocytes, which universally express this protein, only a relatively small percentage of EGCs are positive for GFAP. However, it is well established that in response to infectious challenge, enteric glia change their morphology and upregulate GFAP (Seguella and Gulbransen, 2021Seguella L. Gulbransen B.D. Enteric glial biology, intercellular signalling and roles in gastrointestinal disease.Nat. Rev. Gastroenterol. Hepatol. 2021; 18: 571-587Crossref PubMed Scopus (47) Google Scholar). To examine the functional roles of these two major EGC populations in vivo, Baghdadi and colleagues took advantage of transgenic tools that allowed them to specifically ablate the PLP1- or the GFAP-expressing subsets of enteric glia in mice. Using in vivo characterization of transgenic mice and analysis of organoid cultures, they were able to demonstrate that ablation of the PLP1+ subset of enteric glia had no measurable effects on the intestinal epithelium, but in contrast, selective elimination of the GFAP-expressing EGCs was sufficient to disrupt normal ISC dynamics. However, even in this case, the effect was only transient, pointing to redundant roles of the two glia subpopulations and suggesting that PLP1+ EGCs can functionally compensate for the ablation of their GFAP+ counterparts. These predictions were confirmed by the simultaneous ablation of both EGC subpopulations, which led to the rapid breakdown of intestinal epithelial barrier function and lethality. Although the phenotype of the compound transgenics may not have been unexpected, the severity and the speed at which it appeared must have surprised the authors. It appears therefore that the “activated” GFAP+ EGCs function as an effector subpopulation of enteric glia that is actively involved in the maintenance of the intestinal epithelium and that the population of PLP1+ EGCs is capable of replenishing the GFAP+ pool in case their number is reduced or is in higher demand due to barrier-damaging infectious challenge (Figure 1). Beyond the phenotypic characterization of EGC-deficient transgenic mice, Baghdadi et al. also provide mechanistic insight into the regulatory effects of enteric glia on the intestinal epithelium. Specifically, they show that GFAP+ EGCs are a source of WNT ligands, which drive the regeneration of the intestinal epithelium by activating LGR5+ ISCs (Figure 1). The role of WNT signaling in the regulation of ISC dynamics is well established, but the specific requirement of glia-produced WNT ligands is another unexpected finding of the study, as several other cell types in the intestinal mucosa are known to produce these signaling molecules. What is then so special about EGC-derived WNTs whose activity cannot be compensated for by other cellular sources? Imaging of cleared intestinal preparations has previously demonstrated that enteric glia form basket-like cellular networks that surround, and perhaps at the same time partially sequester, the bases of the crypts, while high resolution confocal microscopy has revealed that EGCs often make direct contacts with intestinal epithelial cells. It is possible therefore that EGCs have a privileged position within the ISC niche that allows glia-derived cues (including WNTs) to have direct and easy access to epithelial cells, whether by diffusion, tethering to extracellular matrix bridges, or even exosomes. Dissecting the complexity of multiple WNT sources of the ISC niche during homeostasis and repair and understanding the cellular and molecular basis of the “special relationship” between GFAP+ enteric glia and intestinal epithelial cells is a fascinating area for future investigation. The WNT-mediated communication between EGCs and ISCs in the mucosa is the latest example of an anatomical and functional neuro-immune cell unit (NICU) between enteric glia and non-neuroectodermal cell types in the gut wall. Previous work from Veiga-Fernandes and colleagues has shown that glia-derived GDNF is a key regulator of ILC3 cells and mucosal immunity (Ibiza et al., 2016Ibiza S. García-Cassani B. Ribeiro H. Carvalho T. Almeida L. Marques R. Misic A.M. Bartow-McKenney C. Larson D.M. Pavan W.J. et al.Glial-cell-derived neuroregulators control type 3 innate lymphoid cells and gut defence.Nature. 2016; 535: 440-443Crossref PubMed Scopus (205) Google Scholar), while more recent reports have provided evidence for specialized NICUs between EGCs and several cell types in tunica muscularis (including innate and adaptive immune cells, mesothelial cells, and fibroblasts) that are critical regulators of immune responses and immune homeostasis (Grubišić et al., 2020Grubišić V. McClain J.L. Fried D.E. Grants I. Rajasekhar P. Csizmadia E. Ajijola O.A. Watson R.E. Poole D.P. Robson S.C. et al.Enteric Glia Modulate Macrophage Phenotype and Visceral Sensitivity following Inflammation.Cell Rep. 2020; 32: 108100Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar; Progatzky et al., 2021Progatzky F. Shapiro M. Chng S.H. Garcia-Cassani B. Classon C.H. Sevgi S. Laddach A. Bon-Frauches A.C. Lasrado R. Rahim M. et al.Regulation of intestinal immunity and tissue repair by enteric glia.Nature. 2021; 599: 125-130Crossref PubMed Scopus (28) Google Scholar). These gut-focused studies, together with the demonstration that nerve-associated glial cells in the skin are important for wound repair (Parfejevs et al., 2018Parfejevs V. Debbache J. Shakhova O. Schaefer S.M. Glausch M. Wegner M. Suter U. Riekstina U. Werner S. Sommer L. Injury-activated glial cells promote wound healing of the adult skin in mice.Nat. Commun. 2018; 9: 236Crossref PubMed Scopus (86) Google Scholar), suggest that peripheral glia represent important regulatory nodes for maintaining and restoring the function of all barrier organs. Although the current report focuses on loss-of-function studies, it raises the possibility that an overdrive of the EGC-ISC functional unit could lead to abnormal epithelial cell growth and tumor formation. Indeed, the majority of colorectal cancers show hyperactivation of the WNT pathway, which is believed to be critical for tumor initiation and growth. By demonstrating key roles of the EGC(GFAP+)-WNT-ISC signaling axis, Baghdadi and colleagues advance our fundamental understanding of intestinal physiology and open new and clear directions for the development of novel therapies for the treatment of common GI disorders, such as inflammatory bowel disease and colon cancer. Work in the Pachnis lab is funded by the Francis Crick Institute, which receives core funding from Cancer Research UK (FC001128, FC001159), the UK Medical Research Council (FC001128, FC001159) and the Wellcome Trust (FC001128, FC001159). We also acknowledge additional funding from a Wellcome Trust Investigator Award (212300/Z/18/Z). Enteric glial cell heterogeneity regulates intestinal stem cell nichesBaghdadi et al.Cell Stem CellNovember 1, 2021In BriefBaghdadi et al. provide evidence that enteric glial cells are important for intestinal function and control stem cell activity in homeostasis and regeneration. Single-cell analysis and mouse genetics studies show that a specific EGC population becomes activated and promotes stem cell response in the early stages of repair and chronic inflammation. Full-Text PDF