Human Mucin-2–Producing Colonic Goblet-Like Cells Secrete the Chemokine CXCL8 by Activating Multiple Proinflammatory Pathways in Response to Entamoeba histolytica

Abstract

The mucus layer produced by highly stressed goblet cells forms a protective shield in the gut to protect the underlying mucosal epithelial cells from external threats. Hypersecretion and depletion of mucin-2 (MUC2) mucin from goblet cells is characteristic of symptomatic Entamoeba histolytica infections. It was hypothesized that MUC2 depleted goblet cells could mount a second line of innate host defense by producing proinflammatory cytokines. To investigate this, whether E. histolytica could stimulate proinflammatory responses in wild-type (WT) high MUC2 mucin-producing goblet-like cells and in clustered regularly interspaced palindromic repeats and CRISPR-associated protein 9 (CRISPR-Cas9) gene-edited MUC2KO cells was investigated. In response to live E. histolytica and soluble E. histolytica proteins, WT, and to a lesser extent, MUC2KO cells produced high levels of CXCL8. Entamoeba histolytica temporally induced greater levels of CXCL8 mRNA expression and protein secretion in WT versus MUC2KO cells, which was abrogated with alleviation of endoplasmic reticulum stress with the NADPH-oxidase inhibitor diphenyleneiodonium chloride. WT cells produced elevated reactive oxygen species that induced longer half-lives of CXCL8 transcripts, which was abrogated with diphenyleneiodonium chloride. Western blot and proteomic analyses revealed that WT cells, but not MUC2KO cells, were basally primed to respond to external stressors and responded to E. histolytica through rapid activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase, mitogen-activated protein kinase/p38, and phosphatidylinositol 3-kinase/Akt pathways, to induce CXCL8. These results suggest that colonic goblet-like cells defend against E. histolytica infections by hypersecreting mucus and produce the chemokine, CXCL8, to recruit neutrophils.