Biology subjects

McCauley, H. A.

Publications and source records attributed to McCauley, H. A..

7 recordsLinked to original sources

bioRxiv2025

Enteroendocrine Cells Regulate Intestinal Barrier Permeability

The intestinal epithelial barrier is essential for nutrient absorption and protection against ingested pathogens and foreign substances. Barrier integrity is maintained by tight junctions which are sensitive to inflammatory signals, thus creating a feed-forward loop with an increasingly permeable barrier that further drives inflammation and is the hallmark of inflammatory bowel disease. There are currently no therapeutic strategies to improve the intestinal epithelial barrier. We hypothesized that enteroendocrine cells may play an unappreciated role in maintaining barrier integrity. To test this hypothesis, we seeded human intestinal enteroids with genetic loss of enteroendocrine cells on Transwell filters and evaluated transepithelial electrical resistance, paracellular permeability, and the localization and abundance of junctional proteins. We found that enteroendocrine cells were required to maintain a healthy barrier in crypt-like "stem" and villus-like differentiated cultures. Additionally, exogenous supplementation of enteroendocrine-deficient cultures with the hormones peptide tyrosine tyrosine (PYY) and the somatostatin analog octreotide was sufficient to rescue many aspects of this barrier defect both at baseline and in the presence of the inflammatory cytokine tumor necrosis factor (TNF). Surprisingly, these improvements in barrier function occurred largely independently of changes in protein abundance of junctional proteins zona-occludens 1, occludin, and claudin-2. These findings support a novel role for enteroendocrine cells in augmenting epithelial barrier function in the presence of inflammatory stimuli and present an opportunity for developing therapies to improve the intestinal barrier. NEW & NOTEWORTHYThere are no therapies that directly improve the permeability of the intestinal epithelial barrier. This work uses a human intestinal epithelial model system to demonstrate that sensory enteroendocrine cells are necessary for healthy barrier function and that two of their secreted products, peptide YY and somatostatin, are sufficient to improve barrier function at homeostasis and in the presence of inflammatory cytokines. This could provide novel treatments for strengthening the epithelial barrier in human gastrointestinal disease.

physiology↗

bioRxiv2023

Human organoid modeling of congenital malformations caused by RFX6 mutations reveal an essential role for this transcription factor in establishing and maintaining duodenal identity upstream of PDX1

The gastrointestinal (GI) tract consists of highly specialized organs from the proximal esophagus to the distal colon, each with unique functions. Rare congenital malformations of the GI tract, including organ atresia, agenesis or mis-patterning are linked to gene mutations although the molecular basis of these malformations has been poorly studied due to lack of model systems to study human development. We identified a patient with compound heterozygous mutations in the transcription factor RFX6 with pancreatic agenesis as previously described. In addition, the patient had duodenal mal-rotation and atresia suggesting that establishment of the proximal small intestine was impaired in these patients. To identify the molecular basis of the intestinal malformation we generated induced pluripotent stem cell lines from this patient, and derived human intestinal organoid (HIOs) to identify how mutations in RFX6 impact intestinal patterning and function. We identified that the duodenal identity of HIOs and patient tissues had adopted a more distal small intestinal signature, including expression of SATB2, normally expressed in the ileum and colon. CRISPR-mediated correction of RFX6 restored duodenal identity, including expression of PDX1, which is required for duodenal development. Using transcriptomic approaches in HIOs and Xenopus embryos we identified that PDX1 is a downstream transcriptional target of RFX6 and that PDX1 expression in a RFX6 mutant background was sufficient to rescue duodenal identity. However, RFX6 had a PDX1-independent role in regulating expression of components of WNT, HH, and BMP signaling pathways that are critical for establishing early regional identity in the GI tract. In summary, we have identified that RFX6 is one of the most upstream regulators early intestinal patterning in vertebrates and that it acts by regulating key transcriptional and signaling pathways.

developmental biology↗

bioRxiv2022

Enteroendocrine cells protect the stem cell niche by regulating crypt metabolism in response to nutrients

Background and aimsThe intestinal stem cell niche is exquisitely sensitive to changes in diet, with high fat diet, caloric restriction, and fasting resulting in altered crypt metabolism and intestinal stem cell function. Unlike cells on the villus, cells in the crypt are not immediately exposed to the dynamically changing contents of the lumen. We hypothesized that enteroendocrine cells (EECs), which sense environmental cues and in response release hormones and metabolites, are essential for relaying the nutrient status of the animal to cells deep in the crypt. MethodsWe used the tamoxifen-inducible VillinCreERT2 mouse model to deplete EECs (Neurog3fl/fl) from adult intestinal epithelium and we generated human intestinal organoids from wild-type and NEUROG3-null human pluripotent stem cells. We used indirect calorimetry, 1H-NMR metabolomics, mitochondrial live imaging, and the Seahorse bioanalyzer to assess metabolism. Intestinal stem cell activity was measured by proliferation and enteroid-forming capacity. Transcriptional changes were assessed using 10X Genomics single-cell sequencing. ResultsLoss of EECs resulted in increased energy expenditure in mice, an abundance of active mitochondria, and a shift of crypt metabolism to fatty acid oxidation. Crypts from mouse and human intestinal organoids lacking EECs displayed increased intestinal stem cell activity and failed to activate phospho-S6 ribosomal protein, a marker for activity of the master metabolic regulator mammalian target of rapamycin (mTOR). These phenotypes were similar to those observed when wild-type mice were deprived of nutrients. ConclusionsDeletion of EECs recapitulated a fasting phenotype despite normal levels of ingested nutrients. These data suggest that EECs are required to relay nutritional information to the stem cell niche and are essential regulators of intestinal metabolism.

cell biology↗

bioRxiv2022

High fat diet initiates rapid adaptation of the intestine

While the systemic impacts of overnutrition are well-known to cause obesity and metabolic syndrome over the course of months, the immediate adaptive response of the intestinal epithelium to dietary changes remains poorly understood. Here we used physiological metrics and single cell analyses to interrogate the adaptive response of intestinal epithelial cells when moved to a high fat diet (HFD). Within 1 day of HFD exposure, mice exhibited altered feeding behavior, an increase in energy expenditure and an increase in intestinal epithelial proliferation. Single cell transcriptional analysis demonstrated several acute cellular changes on day 1, including a cell-stress response in intestinal crypts, de-granularization of Paneth cells, and a shift towards fatty acid cellular metabolism. By 3 days of HFD, there was an emergence of uncommitted progenitors with a transcriptional profile indicative of a shift from secretory populations and towards an absorptive fate. In enterocytes, genes regulating lipid transport and absorption increased over the first 3 days which paralleled a functional increase in lipid absorption in vivo over the course of 7 days on HFD. These findings demonstrate that intestinal epithelial cell populations respond rapidly to changes in diet through initial changes in cellular function followed by a shift in cellular composition.

cell biology↗

bioRxiv2021

Using Human Induced Pluripotent Stem Cell Derived Organoids to Identify New Pathologies in Patients with PDX1 Mutations

Two patients with mutations in PDX1 presented with pancreatic agenesis, chronic diarrhea and poor weight gain, the causes of which were not identified through routine clinical testing. We generated patient derived organoids as a novel diagnostic strategy and observed that PDX1188delC/188delC antral organoids convert to an intestinal phenotype, while intestinal organoids undergo gastric metaplasia with significant reduction in enteroendocrine cells. This prompted a re-examination of gastric and intestinal biopsies from both PDX1188delC/188delC patients, which recapitulated organoid phenotypes. Antral biopsies had increased parietal cells and lacked G-cells suggesting loss of antral identity. These patients will now be monitored for the progression of metaplasia. This study demonstrates the utility of organoids for patient diagnoses and treatment.

developmental biology↗

bioRxiv2020

Ontogeny and function of the circadian clock in intestinal organoids

Circadian rhythms regulate diverse aspects of gastrointestinal physiology ranging from the composition of microbiota to motility. However, development of the intestinal circadian clock and detailed molecular mechanisms regulating circadian physiology of the intestine remain largely unknown. The lack of appropriate human model systems that enable organ- and/or diseasespecific interrogation of clock functions is a major obstacle hindering advancements of translational applications using chronotherapy. In this report, we show that both pluripotent stem cell-derived human intestinal organoids engrafted into mice and patient-derived human intestinal enteroids (HIEs) possess robust circadian rhythms, and demonstrate circadian phase-dependent necrotic cell death responses to Clostridium difficile toxin B (TcdB). Intriguingly, mouse and human enteroids demonstrate anti-phasic necrotic cell death responses. RNA-Seq data show ~4% of genes are rhythmically expressed in HIEs. Remarkably, we observe anti-phasic gene expression of Rac1, a small GTPase directly inactivated by TcdB, between mouse and human enteroids. Importantly, the observed circadian time-dependent necrotic cell death response is abolished in both mouse enteroids and human intestinal organoids (HIOs) lacking robust circadian rhythms. Our findings uncover robust functions of circadian rhythms regulating critical clock-controlled genes (CCGs) in human enteroids governing organism-specific, circadian phasedependent necrotic cell death responses. Our data highlight unique differences between mouse and human enteroids, and lay a foundation for human organ- and disease-specific investigation of clock functions using human organoids for translational applications.

molecular biology↗

bioRxiv2020

Enteroendocrine cells couple nutrient sensing to nutrient absorption by regulating ion transport

The ability to absorb ingested nutrients is an essential function of all metazoans and utilizes a wide array of nutrient transporters found on the absorptive enterocytes of the small intestine. A unique population of patients has previously been identified with severe congenital malabsorptive diarrhea upon ingestion of any enteral nutrition. The intestines of these patients are macroscopically normal, but lack enteroendocrine cells (EECs), suggesting an essential role for this rare population of nutrient-sensing cells in regulating macronutrient absorption. We used human and mouse models of EEC deficiency to identify a new role for the EEC hormone peptide YY in regulating ion-coupled absorption of glucose and dipeptides the small intestine. We found that peptide YY is required in to maintain normal electrophysiology in the presence of vasoactive intestinal polypeptide, a potent stimulator of ion secretion produced by enteric neurons. Administration of peptide YY to EEC-deficient mice restored normal electrophysiology, improved glucose and peptide absorption, diminished diarrhea and rescued postnatal survival. These data suggest that peptide YY is a key regulator of macronutrient absorption in the small intestine and may be a viable therapeutic option to treat patients with malabsorption.

physiology↗