To defend the gut from microbes and pathogens, Paneth cells rely on TMEM16A, a calcium-activated chloride channel, and TMEM16F, a phospholipid scramblase, according to a new study published in Gastro Hep Advances.
The Paneth cells in mice missing TMEM16A or TMEM16F showed defects in signaling and release of secretary factors, researchers reported.
Dr. Rainer Schreiber
Inhibiting or activating TMEM16A and TMEM16F is likely to affect microbial content and immune functions in the small intestine, concluded Rainer Schreiber, Dr. rer. nat., of the Institute of Physiology at Universität Regensburg, Germany, and colleagues.
“Many small molecules and numerous natural or herbal compounds have been identified that either inhibit or activate TMEM16A or TMEM16F,” they wrote. “Some of these compounds may turn out to be useful therapeutics in inflammatory bowel disease, intestinal allergies, or abnormal colonization of the gut.”
Paneth cells play a central role in intestinal innate immune response, the authors wrote. Located at the base of small intestinal crypts and occasionally found in the proximal colon, these cells have defensive functions, such as protecting stem cells in response to invading microbes and eradicating ingested pathogens from intestinal crypts. Through secretion, they also regulate the composition and number of commensal intestinal bacteria. In inflammatory bowel disease, the Paneth cell zone expands due to an increase in cell size and cell number.
In previous studies, cholinergic stimulation provided enhanced protection in animals orally infected with virulent Salmonella enterica. However, the mechanisms of luminal stimulation of Paneth cell secretion in response to bacteria or lipopolysaccharide are unclear. Recent reports show that TMEM16A (also known as anoctamin 1, or ANO1) and TMEM16F (anoctamin 6, or ANO6) control intracellular calcium (Ca2+) signaling and that high local Ca2+ levels support exocytosis in intestinal cells.
The researchers analyzed the roles of the two molecules in Paneth cell secretion using mice with intestinal epithelial-specific knockout of TMEM16A or TMEM16F. They examined tissue structures and Paneth cells in the mice, as well as Paneth cell exocytosis in small intestinal organoids in vitro. They also compared Ca2+ signals between wild-type and knockout mice and analyzed bacterial colonization and intestinal apoptosis.
In wild-type mice, TMEM16A was detected at the apical pole of crypt epithelial cells, while TMEM16F was located predominantly at the basolateral side. Notably, TMEM16A was also located in intestinal smooth muscle cells.
Compared with wild-type mice, the TMEM16 knockout mice had pronounced accumulation of lysozyme in jejunal Paneth cells. This suggests a defect in Paneth cell secretion in the absence of TMEM16A and TMEM16F, the authors wrote.
Previous studies had found an accumulation of mucus in intestinal goblet cells in mice with tissue-specific knockout of TMEM16A and TMEM16F. In this study, a more detailed analysis of mucus using periodic acid-Schiff staining of duodenum, jejunum, and ileum confirmed those results and demonstrated enhanced mucus in the small intestine of knockout mice. This suggests that a lack of TMEM16A or TMEM16F causes a broad secretion defect in secretory cells, including Paneth cells, the authors wrote.
Because granules of Paneth cells contain antimicrobial peptides, cytokines, and other factors that control proliferation or epithelial cell death, the researchers analyzed the presence of Gram-positive and Gram-negative bacteria in the jejunum and ileum. Compared to wild-type mice, the number of bacteria was higher in the ileum of both TMEM16A and TMEM16F knockout mice and in the jejunum of TMEM16F knockout mice, suggesting reduced antimicrobial activity in the absence of TMEM16 proteins.
The researchers also compared regulated cell death of intestinal epithelial cells in jejuna of wild-type and knockout mice. They found largely reduced cell death in both TMEM16A and TMEM16F knockout mice.
“Intestinal inflammatory diseases such as Crohn’s disease, necrotizing enterocolitis, and intestinal microbiota dysbiosis have been related to abnormal Paneth cell physiology,” the authors wrote. “The present findings may therefore provide the basis for a novel anti-inflammatory therapy for intestinal diseases and may improve our understanding of the molecular mechanism of some of the currently available drugs.”
The study was supported by the Deutsche Forschungsgemeinschaft funding program. The authors disclosed no conflicts of interest.
