Enteric Glial Functions in Gastrointestinal Homeostasis

Abstract

Enteric glia are highly abundant, heterogeneous cells found throughout the entire length of the digestive system. In recent decades, the cells have been shown to regulate neuronal function, modulate the intestinal epithelium, and respond to inflammatory challenges and infections. Despite these advancements, questions remain regarding the extent of the cells’ involvement in these processes. Here, we investigated two enteric glial roles: their purported regulation of the intestinal epithelium and their phagocytic function. A primary aim of our study was to determine whether the cells perform these roles in the intestine. Additionally, we sought to understand the mechanisms underlying these glial functions and their physiological significance within the GI tract. To better define the roles of enteric glia in a steady state, we first focused on investigating their putative role in regulating the intestinal epithelium. To this end, we characterized glia in the intestine, focusing on ones that closely appose intestinal epithelial cells. We identified PLP1 as a specific and widely expressed marker of these cells in mice and humans. Having established that, we used a genetic mouse model of PLP1+ cell depletion and transcriptional profiling to assess the effects of glial loss on the GI tract in a steady state. We discovered that glial loss had a minimal impact on the transcriptional programs in the intestine. However, the minor changes we did detect varied along the longitudinal axis of the gastrointestinal tract. In the ileum, where enteric glia had been considered most essential for epithelial integrity, glial depletion similarly failed to significantly alter epithelial gene expression. It did, however, cause a modest enrichment in signatures associated with Paneth cells, a secretory cell type critical for innate immunity. In the absence of PLP1+ glia, Paneth cell number was unchanged, but a subset of these cells appeared abnormal with irregular and heterogenous cytoplasmic granules, suggestive of a secretory defect. Consistent with this possibility, secretions from ileal explants of glial-depleted mice contained significantly reduced levels of functional lysozyme compared to the littermate control explants. In line with these results, glial deficiency resulted in shifts in gut microbial composition. Taken together, these observations suggest that enteric glia are important for regulating Paneth cell function and gut microbial ecology. Glial cells throughout the nervous system act as phagocytes, fostering the proper development and functioning of neuronal circuits and facilitating appropriate responses to injury. In the second part of the project described in this thesis, we sought to test whether enteric glia similarly function as phagocytes in the intestine. To that end, we first investigated their expression of engulfment machinery and found that the cells express several known phagocytic receptors, including MEGF10 and AXL. We next demonstrated that enteric glia are phagocytic cells as they efficiently engulf material from apoptotic cells in an AXL- and MEGF10-dependent manner in vitro. To determine the physiologic relevance of enteric glial phagocytosis, we utilized MEGF10-deficient mice and assessed their steady-state ENS function and architecture, as well as the response to acute inflammatory challenges to the GI tract. Although we observed no effects on the progression and outcomes of GI pathology between MEGF10-deficient and MEGF10-sufficient animals, we discovered that loss of MEGF10 caused age-dependent dysmotility and failure to thrive. These collective findings suggest that enteric glial are phagocytes whose engulfment capacity may influence the regulation of gut homeostasis. Together, the results of the two parts of my thesis project provide evidence for two novel functions of enteric glia – regulation of Paneth cell function and phagocytosis, which might play a role in the modulation of steady-state functions of the GI tract.