Biology subjects

Liddle, R. A.

Publications and source records attributed to Liddle, R. A..

6 recordsLinked to original sources

bioRxiv2025

Intestinal enteroendocrine cell subtype differentiation and hormone production in zebrafish

Enteroendocrine cells (EECs) are rare sensory cells in the intestinal epithelium that coordinate digestive physiology by secreting a diverse repertoire of peptide hormones. These hormones are the main effectors of EEC function, and their characterization requires direct observation by mass spectrometry due to the specialized protein cleavage and posttranslational modifications that yield their mature forms. Based on the distinct subset of hormones they predominantly secrete, EECs can be categorized into subtypes. How each EEC subtype is specified, however, remains poorly understood. Here we describe EEC subtype differentiation and hormone production in the zebrafish. Using single-cell RNA sequencing data, we identified EEC progenitors and six EEC subtypes in zebrafish and revealed that their expression profiles are consistent across larval and adult stages. Mass spectrometry analysis of isolated zebrafish EECs identified highly processed peptides derived from 18 of 21 hormone coding genes expressed by EECs, yielding a catalog of >400 unique EEC hormone peptides. We assembled reporters for zebrafish EEC subtypes to test the lineage relationships between EEC subtypes and the EEC progenitor population, which expresses neurogenin3. Despite its essential role in mammalian EEC differentiation, we found that selective cytotoxic ablation of neurogenin3+ cells in zebrafish only reduced a subset of EEC subtypes. Finally, we discovered that selective ablation of ghrelin+ EECs reduced a different subset of EEC subtypes, together suggesting that neurogenin3+ and ghrelin+ cells serve as distinct precursors for separate EEC subtypes. We anticipate these observations and resources will facilitate future studies in the zebrafish to discern the developmental biology, physiology, and endocrinology of EEC subtypes.

developmental biology↗

bioRxiv2023

Piezo1-induced durotaxis of pancreatic stellate cells depends on TRPC1 and TRPV4 channels

Pancreatic stellate cells (PSCs) are primarily responsible for producing the stiff tumor tissue in pancreatic ductal adenocarcinoma (PDAC). Thereby, PSCs generate a stiffness gradient between the healthy pancreas and the tumor. This gradient induces durotaxis, a form of directional cell migration driven by differential stiffness. The molecular sensors behind durotaxis are still unclear. To investigate the role of mechanosensitive ion channels in PSC durotaxis, we established a two-dimensional stiffness gradient mimicking PDAC. Using pharmacological and genetic methods, we investigated the role of the ion channels Piezo1, TRPC1, and TRPV4 in PSC durotaxis. We found that PSC migration towards a stiffer substrate is diminished by altering Piezo1 activity. Moreover, disrupting TRPC1 along with TRPV4 abolishes PSC durotaxis even when Piezo1 is functional. Hence, PSC durotaxis is optimal with an intermediary level of mechanosensitive channel activity, which we simulated using a numerically discretized mathematical model. Our findings suggest that mechanosensitive ion channels, particularly Piezo1, detect the mechanical microenvironment to guide PSC migration. SummaryCells move towards regions with higher stiffness in a process called durotaxis. This study shows that ion channels Piezo1, TRPV4, and TRPC1 are crucial sensors in pancreatic stellate cells. They act together to orchestrate durotaxis of pancreatic stellate cells which may be relevant in pancreatic cancer.

physiology↗

bioRxiv2023

Gut mucosal cells transfer α-synuclein to the vagus nerve

Epidemiological and histopathological findings have raised the possibility that misfolded -synuclein protein might spread from the gut to the brain and increase the risk of Parkinsons disease (PD). While past experimental studies in mouse models have relied on gut injections of exogenous recombinant -synuclein fibrils to study gut to brain -synuclein transfer, the possible origins of misfolded -synuclein within the gut have remained elusive. We recently demonstrated that sensory cells of the gut mucosa express -synuclein. In this study, we employed mouse intestinal organoids expressing human -synuclein to observe the transfer of -synuclein protein from gut epithelial cells in organoids co-cultured with vagal nodose neurons that are otherwise devoid of -synuclein expression. In intact mice that express pathological human -synuclein, but no mouse -synuclein, -synuclein fibril templating activity emerges in -synuclein seeded fibril aggregation assays in tissues from the gut, vagus nerve, and dorsal motor nucleus. In newly engineered transgenic mice that restrict pathological human -synuclein expression to intestinal epithelial cells, -synuclein fibril-templating activity transfers to the vagus nerve and to the dorsal motor nucleus. Subdiaphragmatic vagotomy prior to the induction of -synuclein expression in the gut epithelial cells effectively protects the hindbrain from the emergence of -synuclein fibril templating activity. Overall, these findings highlight a novel potential non-neuronal source of fibrillar -synuclein protein that might arise in gut mucosal cells.

neuroscience↗

bioRxiv2020

Shear stress-induced pathological changes in endothelial cells occur through Piezo1 activation of TRPV4

Although the ion channels Piezo1 and TRPV4 have been implicated in high venous pressure- and fluid shear stress-induced vascular hyperpermeability, they have been described as working independently. Moreover, the mechanism by which Piezo1 and TRPV4 channels in endothelial cells execute the same function is poorly understood. Here we demonstrate that Piezo1 regulates TRPV4 channel activation in endothelial cells and that Piezo1-mediated TRPV4 channel opening is a function of the strength and duration of fluid shear stress. Application of the Piezo1 antagonist, GsMTx4, completely blocked the elevation in intracellular calcium ([Ca2+]i) induced by both fluid shear stress and the Piezo1 agonist, Yoda1. High and prolonged shear stress caused sustained [Ca2+]i elevation which required TRPV4 opening and was responsible for fluid shear stress- and Piezo1-mediated disruption of adherens junctions and actin remodeling. We found that Piezo1’s effects were mediated by phospholipase A2 activation. Blockade of TRPV4 channels with the selective TRPV4 blocker, HC067047, prevented the loss of endothelial cell integrity and actin disruption induced by Yoda1 or shear stress and prevented Piezo1-induced monocyte adhesion to endothelial cell monolayers. These findings demonstrate that Piezo1 activation by fluid shear stress initiates a calcium signal that causes TRPV4 opening which in turn is responsible for the sustained phase calcium elevation that triggers pathological events in endothelial cells.The abbreviations used areHUVEChuman umbilical vein endothelial cellTRPV4transient receptor potential vanilloid subfamily 4PLA2phospholipase A2AJsadherens junctions5′,6′-EET5′,6′-epoxyeicosatrienoic acidHC067HC067047GSK101GSK1016790APVPpulmonary venous pressureView Full Text

cell biology↗

bioRxiv2020

Enteroendocrine cells sense bacterial tryptophan catabolites to activate enteric and vagal neuronal pathways

The intestinal epithelium senses nutritional and microbial stimuli using epithelial sensory enteroendocrine cells (EECs). EECs can communicate nutritional information to the nervous system, but similar mechanisms for microbial information are unknown. Using in vivo real-time measurements of EEC and nervous system activity in zebrafish, we discovered that the bacteria Edwardsiella tarda specifically activates EECs through the receptor transient receptor potential ankyrin A1 (Trpa1) and increases intestinal motility in an EEC-dependent manner. Microbial, pharmacological, or optogenetic activation of Trpa1+EECs directly stimulates vagal sensory ganglia and activates cholinergic enteric neurons through 5-HT. We identified a subset of indole derivatives of tryptophan catabolism produced by E. tarda and other gut microbes that potently activates zebrafish EEC Trpa1 signaling and also directly stimulates human and mouse Trpa1 and intestinal 5-HT secretion. These results establish a molecular pathway by which EECs regulate enteric and vagal neuronal pathways in response to specific microbial signals.

physiology↗

bioRxiv2019

High fat diet induces microbiota-dependent silencing of enteroendocrine cells

Enteroendocrine cells (EECs) are specialized sensory cells in the intestinal epithelium that sense and transduce nutrient information. Consumption of dietary fat contributes to metabolic disorders, but EEC adaptations to high fat feeding were unknown. Here, we established a new experimental system to directly investigate EEC activity in vivo using a zebrafish reporter of EEC calcium signaling. Our results reveal that high fat feeding alters EEC morphology and converts them into a nutrient insensitive state that is coupled to endoplasmic reticulum (ER) stress. We called this novel adaptation \"EEC silencing\". Gnotobiotic studies revealed that germ-free zebrafish are resistant to high fat diet induced EEC silencing. High fat feeding altered gut microbiota composition including enrichment of Acinetobacter species, and we identified an Acinetobacter strain sufficient to induce EEC silencing. These results establish a new mechanism by which dietary fat and gut microbiota modulate EEC nutrient sensing and signaling.

developmental biology↗