Molecular analysis of single cells lining the digestive tract found high CFTR activity — the gene associated with cystic fibrosis (CF) — in some cells in the duodenum, or first part, of the small intestine.
A comparison of cells lining the stomach of mice and humans also showed differences that may affect how well mouse models capture human disease, and serve as models for therapy development.
The study, “Human gastrointestinal epithelia of the esophagus, stomach, and duodenum resolved at single-cell resolution,” was published in the journal Cell Reports.
Mutations in the CFTR gene cause CF, which mainly manifests in the lungs. But patients also experience problems associated with a blockage in the digestive (gastrointestinal GI) tract due to the buildup of sticky mucus.
The CFTR protein’s role as a membrane channel is to regulate the transport of chloride ions into and out of cells, controlling water movement in tissues that supports thin and freely flowing mucus.
The upper GI tract’s primary responsibilities are digestion, nutrient uptake, defense against microbes, and hormone production.
Layers of epithelial cells line the esophagus that leads to the stomach, protect that tube from abrasion by incoming raw food. In contrast, the stomach, small intestine, and colon have a single layer of epithelial cells that secrete acid and digestive enzymes, besides absorbing nutrients. These processes mainly involve channels and transporter proteins located across the cell membrane.
Most cells lining the GI tract on the large intestine have been identified, particularly in various disease stages. But an examination of cells lining the upper GI tract — the esophagus, stomach, and duodenum — has been lacking, the researchers wrote.
Using a technique called single-cell RNA sequencing (scRNA-seq), scientists in the Netherlands identified various types of epithelial cell that line the upper GI tract using healthy human tissue.
Different cell types are characterized by the presence of specific proteins. As the information for proteins is carried in genes (DNA) and then transcribed into RNA for protein production, scRNA-seq can be used to identify individual cell types based on gene activity.
Here, the team analyzed healthy human tissue taken in biopsies from the esophagus, stomach, and duodenum.
In relation to CF, a small cluster of cells — comprising 1.3% of duodenum epithelial cells — showed high gene activity (expression) of BEST4, GUCA2A, GUCA2B, as well as the CF-related gene, CFTR. Collectively, these cells are called BCHE and likely mediate high-volume fluid secretion, the scientists said. BEST4 encodes for bestrophin 4, a chloride channel like CFTR.
Among all duodenal cells, BCHE cells expressed the highest level of CFTR, which was found only in the duodenum but not in BEST4-positive cells in the colon. This “strongly suggests that the colonic [BEST4-postive cells] and duodenal BCHE cells may have distinct functions,” the scientists wrote.
Duodenal cells known as enterochromaffin cells (EC cells) secrete the hormone serotonin, involved in modulating mood, cognition, learning, and memory. In contrast, EC-like cells in the stomach secrete histamine as part of local immune responses.
The analysis unexpectedly found that EC-like cells in the stomach also expressed genes for the luteinizing hormone (LH) — known for its role in ovulation in females and in regulating testosterone in males. “Future investigations will be required to elucidate the role of locally produced LH in the stomach,” the researchers wrote.
Cells from the multilayered esophagus showed a continuum of cell types based on the marker proteins KRT4, KRT13, and KRT14. Marker KRT5 exhibited a broad expression pattern.
Two RNA markers — KRT15 and COL17A1 — were highly expressed in dormant esophageal basal cells, “which suggests that these cells may function as stem/progenitor cells in the human esophagus,” the team wrote. Notably, mutations in the COL17A1 gene are associated with junctional epidermolysis bullosa, leading to the formation of blisters.
“The results provide insight into the molecular characteristics of individual cell types and how they may function in the healthy epithelium,” Georg Busslinger, PhD, the study’s first author, said in a press release. “Moreover, these data serve as a resource for scientists everywhere.”
The team also investigated the difference in stomach cells between people and mice, which are known to be anatomically comparable. However, the expression of several genes were detected in some areas of the human stomach but not in mice, while others were expressed in mice but not humans. In particular, cells expressing LH in the stomach were only found in human tissue.
“The observed differences between human and mouse are more extensive than expected and may complicate the extrapolation of gene expression and [disease characteristics] obtained in the mouse model organism to the respective human gastrointestinal organs,” the scientists wrote.
To avoid differences between mouse models and humans, and support the translation from preclinical to clinical research, the team also established organoid cultures from healthy esophageal biopsies. The cultures mimic the structure of the epithelium and can be maintained for up to two months.
“We could, for example, use organoids to follow-up on our current findings,” added Busslinger.
“We have now performed an in-depth analysis of the healthy human epithelia of the esophagus, [stomach], and the duodenum, which provided interesting insight into cellular and molecular functions of the distinct cell types of these organs,” the investigators wrote.
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