Enteric glial reactivity in intestinal inflammation and colorectal cancer

Abstract

Enteric glial cells (EGCs) are important regulators of gastrointestinal functions in health and disease. Over the last few years, their role in intestinal inflammation and their interplay with the local tissue microenvironment has raised increasing attention. As EGCs can react to various environmental stimuli, such as cytokines and chemokines, and contribute to the inflammatory milieu, it is critical to understand the signaling pathways inducing their reactive phenotype and the resulting actions on their tissue microenvironment. Using primary EGC cultures and glial-specific genetic modifications, we revealed specialized extracellular matrix compositions, the sympathetic nervous system, and interleukin (IL)-1 as important regulators of EGCs' immune functions. Applying brightfield and immunofluorescent imaging and RNA-sequencing, we demonstrated distinct effects of extracellular matrix substrates on EGC primary cultures, pinpointing Matrigel and laminin as superior coating substrates in EGC purity, network formation, and reactivity. Postoperative RNA sequencing results expanded our knowledge about EGC reactivity by identifying the sympathetic nervous system as an early EGC activator during acute intestinal inflammation. Using EGC cultures, we discovered norepinephrine signaling via β-1/2-adrenergic receptors as a responsible inducer of immediate postoperative glial reactivity. Moreover, we used a targeted approach to analyze the effect of the well-known and strong immune activator IL-1 on glial-specific reactivity in acute postoperative intestinal inflammation. Strikingly, glial activation via IL-1 led to increased expression of genes related to ‘enteric gliosis' both in vitro and in intestinal inflammatory models in vivo. Furthermore, glial reactivity correlated with macrophage activation and migration, highlighting EGCs' contribution to the inflammatory microenvironment. Finally, we transferred the insights gained from the acute postoperative intestinal inflammation studies into the field of tumor immunology, representing a chronic inflammatory condition as it occurs in colorectal cancer (CRC). Applying three murine CRC models, including EGC-specific molecular reporters and gene deficiencies, we identified a bidirectional signaling cascade between EGCs and tumor-promoting macrophages in CRC, leading to cancer progression. Unraveling the molecular pathways involved, monocyte-derived IL-1 was identified as an EGC activator via glial IL-1 receptor 1 signaling, triggering the secretion of IL-6 into the tumor microenvironment, which promotes the differentiation of infiltrating monocytes into tumor-promoting SPP1+ macrophages. Together, we highlighted the significance of EGCs in intestinal inflammatory diseases. Identifying the extracellular matrix, the sympathetic nervous system, and IL-1 as crucial regulators of glial biology and immune reactivity, we substantiated our understanding of the molecular pathways involved in EGCs' reactive phenotype. Importantly, IL-1-mediated glial activation was proven essential for glia-macrophage interactions in intestinal inflammation and CRC, underlining EGCs' vital role as immune regulators actively modifying disease outcomes in inflammation-driven intestinal diseases and disorders.