Isthmic progenitor cells, tissue-specific stem cells in the stomach corpus, maintain mucosal homeostasis by balancing between proliferation and differentiation to gastric epithelial cell lineages. The progenitor cells rapidly adopt an active state in response to mucosal injury. However, it remains unclear how the isthmic progenitor cell niche is controlled during the regeneration of damaged mucosa.

We recapitulated tissue recovery process after acute mucosal injury in the mouse stomach with a parietal cell toxic drug. BrdU incorporation was utilized to trace newly generated cells during the injury and recovery phases. To define epithelial lineage commitment process during recovery, we performed single cell RNA-sequencing (scRNA-seq) on epithelial cells from the mouse stomachs. We validated the effects of amphiregulin (AREG) on the mucosal recovery, utilizing recombinant AREG treatment or AREG deficient mice.

In homeostatic conditions of the stomach corpus, most of the BrdU+ cells were located in the isthmus area without expression of differentiated gastric cell lineage markers, or in the surface area with expression of IL33, indicating that isthmic progenitor cells are normally keeping their own population or committed to foveolar cells. After injury and during early recovery phase (1 day of recovery), BrdU+ cells were still restricted in the isthmus and surface area, and BrdU-incorporated cells then appeared in the oxyntic gland area below isthmus with expression of H⁺K⁺ATPase, a marker of parietal cells, after 2 days of recovery. Based on the identification of lineage-committed subpopulations in the corpus epithelium through scRNA-seq, we showed that isthmic progenitor cells give rise to epithelial cell lineages through an ordered paradigm during mucosal regeneration. Consistent with increased expression of EGFR phosphorylated at tyrosine 1068 in the isthmus area after damage, a predictive model based on the scRNA-seq data demonstrated that interaction between EGFR and EGFR ligands can actively occur within isthmic progenitor, pre-surface and pre-parietal cells from damaged epithelium. AREG, an EGFR ligand, treatment promoted repopulation with parietal cells, while suppressing surface cell commitment of progenitors. In contrast, TGF-alpha, another ligand for EGFR, did not alter parietal cell regeneration but did induce expansion of surface cell populations. In the same context, AREG deficiency impaired parietal cell regeneration, but increased surface cell commitment.

Therefore, these data demonstrate that different EGF receptor ligands can distinctly regulate isthmic progenitor-driven lineage commitment and mucosal regeneration.

David Gibbs

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