Cellular and Molecular Gastroenterology and Hepatology最新文献

Background & aims: Transmembrane protein 41B (TMEM41B) and vacuolar membrane protein 1 (VMP1) are endoplasmic reticulum (ER) scramblases whose roles in hepatic lipoprotein secretion and autophagy in metabolic-associated steatotic liver disease (MASLD) remain unclear.

Methods: We undertook lipidomic and functional studies in liver- and hepatocyte-specific Tmem41b knockout (KO) mice; Tmem41b knock-in (KI) mice, Tmem41b/Vmp1 double KO (DKO); Tmem41b KO/Vmp1 KI, and Vmp1 KO/Tmem41b KI mice.

Results: TMEM41B protein levels decreased in the livers of human subjects with MASLD. Loss of hepatic Tmem41b impaired very low-density lipoprotein (VLDL) secretion, with steatosis, inflammation and fibrosis, while hepatic TMEM41B overexpression mitigated these effects. Tmem41b/Vmp1 DKO mice showed further impairment in VLDL secretion compared to single Tmem41b KO. Lipidomic analysis revealed decreased phosphatidylcholine and phosphatidylethanolamine, with increased neutral lipids in Tmem41b KO mice. VMP1 and TMEM41B localize at the mitochondrial-associated membrane (MAM) with reduced mitochondria-ER contact in VMP1 and TMEM41B KO mice. Loss of hepatic VMP1 or TMEM41B increased levels of LC3B-II and p62/SQSTM1, which were not further changed in DKO mice. Restoring VMP1 in Tmem41b KO mice and TMEM41B in Vmp1 KO mice partially corrected defective VLDL secretion and hepatic steatosis in these single KO mice, respectively. Restoring VMP1 at a low but not a high dose corrected defective autophagy in Tmem41b KO mice, whereas overexpression of TMEM41B dose-dependently improved defective autophagy in Vmp1 KO mice.

Conclusions: Loss of hepatic VMP1 or TMEM41B reduces VLDL secretion and promotes MASLD via overlapping but distinct mechanisms that regulate lipoprotein secretion and autophagy.

Background & aims: Intraductal papillary mucinous neoplasms (IPMNs) are pancreatic cysts that represent one of the few radiologically identifiable precursors to pancreatic ductal adenocarcinoma (PDAC). Though the IPMN-bearing patient population represents a unique opportunity for early detection and interception, current guidelines provide insufficient accuracy in determining which patients should undergo resection versus surveillance, resulting in a sizable fraction of resected IPMNs only harboring low-grade dysplasia, suggesting that there may be overtreatment of this clinical entity.

Methods: To investigate the transcriptional changes that occur during IPMN progression, we performed spatial transcriptomics using the Nanostring GeoMx on patient samples containing the entire spectrum of IPMN disease including low-grade dysplasia, high-grade dysplasia, and IPMN-derived carcinoma. Single cell RNA sequencing was performed on side branch and main duct IPMN biospecimens.

Results: We identified a subpopulation of histologically low-grade IPMN epithelial cells that express malignant transcriptional features including KRT17, S100A10 and CEACAM5, markers that are enriched in PDAC. We validated this high-risk gene signature in both single-cell RNA sequenced samples and an external spatial transcriptomic dataset containing a larger number of IPMN samples with non-tumor bearing IPMN (i.e. low-grade IPMN in isolation). Immunofluorescence staining of a large cohort of patient tissues confirmed the presence of KRT17-positive cells, which were found to comprise a small subset of epithelial cells within histologically low-grade IPMN in a patchy distribution.

Conclusions: Our study demonstrates that KRT17 marks a distinct transcriptional signature in a subpopulation of epithelial cells within histologically low-grade IPMN. This population of cells likely represents a transitional state of histologically low-grade epithelial cells undergoing progression to a higher grade of dysplasia and thus may represent a higher risk of progression to carcinoma.

Background & aims: Tuft cells, a type of epithelial cell in the gut, play a pivotal role in regulating type 2 immunity and maintaining the gut barrier. However, their role in cognitive impairments remains unclear.

Methods: We compared behavioral performance between male tuft cell-absent mice (Pou2f3^-/-) and their wild-type littermates (WT). We analyzed gut microbiota using fecal 16S rRNA, measured gut permeability via FITC-dextran assay, and detected CD4⁺-T cells and type 2 innate lymphoid cells by flow cytometry in both genotypes. Co-housing and fecal microbiota transplantation (FMT) experiments were conducted to explore the role of gut microbiota in cognitive diseases. Single-cell RNA sequencing and fluorescence imaging were used to examine tuft cell changes in the colon of WT and Alzheimer's disease (AD) model mice. Colonic organoids were used to assess the effect of β-amyloid on tuft cell differentiation. Succinic acid, a promoter of tuft cells, was administered, and tuft cell-deficient AD mice were generated to evaluate its impact on behavior and gut homeostasis.

Results: Increased gut permeability, immune imbalance, neuroinflammation, and cognitive dysfunction occurred in 10-month-old mice lacking tuft cells. These alterations were mediated by gut microbiota, evidenced by shifts in microbiota composition and abundance, and supported by co-housing and FMT experiments. AD model mice had fewer tuft cells and impaired type 2 immunity in the gut, potentially because of β-amyloid inhibiting tuft cell differentiation. Succinic acid, a tuft cell activator, restored cognitive function and gut homeostasis in AD mice.

Conclusion: Tuft cells may be necessary for maintaining gut homeostasis in cognitive disorders.

Background & aims: Tuft cells play protective roles in infection, inflammation, and tumorigenesis through the secretion of cytokines and eicosanoids. Tuft cells are known for their tall, blunt microvilli, thought to be analogous to mechanosensory hair cell stereocilia; however, a functional role for the microvillar apparatus has not been identified. POU2F3 is the master regulator transcription factor for tuft cells, yet how POU2F3 drives formation of this unique structure is unknown. Here, we aimed to identify POU2F3 target genes and commonalities between tuft and hair cells to better understand this unique structure.

Methods: POU2F3 ChIP-seq was performed on tuft cells and compared to the hair cell transcriptome. Tuft cell RNA-seq datasets were interrogated for hair cell structural and mechanosensory genes; expression was validated. Intestinal and gallbladder tuft cells were examined using multiple light and electron microscopy (EM) modalities. PCDH20 was knocked down in mouse models and ultrastructural analyses were performed. The tuft cell cytoskeleton was modeled using AlphaFold3 prediction.

Results: Genes encoding structural and mechanosensory proteins common to both tuft and hair cells, including Pcdh20, were identified. Imaging localized PCDH20 to tuft cell microvilli and hair cell stereocilia. Genetic ablation of Pcdh20 in mice resulted in structural defects in tuft cell microvilli, including loss of rigidity and organization. Molecular modeling suggests PCDH20 homodimers link adjacent microvilli.

Conclusions: Pcdh20 is a POU2F3 target gene in tuft cells, critical to maintain the rigid microvillar apparatus. These findings, together with the shared expression of mechanosensory components like TMC1, support the hypothesis that tuft cells could have mechanosensory capabilities analogous to cochlear hair cells.

Background & aims: Dietary fat increases the risk of intestinal cancer, but the effect of the fatty acid composition on tumorigenesis is unclear. The aim of this study is to investigate the impact of diets with different fatty acids on carcinogenesis in the intestine.

Methods: Mice were fed a linoleic acid (LA)-rich or stearic acid (SA)-rich high-fat diet (HFD) from the age of 4 weeks. The Apc^Min/+ mice and an azoxymethane- and a dextran sulfate sodium-induced colorectal cancer (CRC) mouse model were used to examine the effects of different dietary fatty acids on CRC development. fatty acid-binding protein 5 (FABP5) knockout mice and SBFI-26, an inhibitor of FABP5, were used to assess its contribution.

Results: We found that an SA-rich HFD more strongly accelerated tumorigenesis in murine CRC models than an LA-rich HFD, with fewer obesity phenotypes compared with LA-rich HFD-fed mice. Dietary SA more strongly promoted epithelial cell proliferation and Paneth cell differentiation than LA, whereas no differences in the numbers of leucine-rich repeat-containing G protein-coupled receptor 5⁺ and B lymphoma Mo-MLV insertion region 1 homolog⁺ intestinal stem cells were detected between the groups. In murine and human intestinal organoids, SA promoted crypt formation. We found that FABP5 was expressed in a small population of Ki67⁺ proliferative cells in crypts, and the number of Ki67⁺ FABP5⁺ cells was increased by SA-rich HFD feeding. FABP5 inhibition suppressed SA-induced epithelial cell proliferation, Paneth cell differentiation, and tumorigenesis.

Conclusions: Dietary SA can promote CRC via FABP5 without promoting obesity.

Background & aims: Mast cells (MCs) play a critical role in the pathogenesis of inflammatory bowel diseases (IBD), encompassing ulcerative colitis and Crohn's disease. Nevertheless, their regulatory impact on intestine stem cells (ISCs) compartment remains poorly characterized. We aim to study the effect of MCs on ISCs-mediated epithelial regeneration in IBD.

Methods: The bulk RNA-seq data of intestine tissues from IBD patients was collected from the Mount Sinai Crohn's and Colitis registry (MSCCR) to explore the direct and indirect correlation of MCs with ISCs, stemness, and related pathways. Subsequently, the results were verified by experiments such as Dextran sulfate sodium (DSS)-induced colitis in MCs-deficient rats and C57BL/6 mice, and co-culture of bone marrow-derived mast cells (BMMCs) and small intestinal organoids.

Results: Bulk RNA-seq data analysis demonstrated significant MCs activation in inflamed mucosa of IBD patients, showing negative correlations with transcriptional signatures of ISC, stemness markers, and Wnt pathway activity. Genetic ablation of MCs in Kit^W-sh/W-sh rats conferred protection against DSS-induced epithelial damage, exhibiting enhanced Lgr5 expression and Wnt/lrp6/β-catenin signaling activation compared to wild-type Kit^+/+ rats. Pharmacological stabilization of MCs with cromolyn sodium (CS) or intervention with a carboxypeptidase A (CPA) inhibitor during the recovery phase of DSS-induced colitis promoted epithelial restoration in mice, evidenced by improved crypt architecture and upregulation of ISC-associated genes and proteins. In vitro co-culture experiments demonstrated MCs-mediated suppression of intestinal organoid growth and Wnt/lrp6/β-catenin signaling pathway, reversible through Lrp6 activation and carboxypeptidase A3 (CPA3) inhibition. Mediation analysis coupled with neutrophils detection revealed an additional indirect regulatory axis involving MC-driven neutrophil recruitment to inhibit ISC-mediated epithelial repair.

Conclusions: Our findings establish MCs play a pivotal role in inhibiting ISCs-mediated epithelial regeneration by suppressing Wnt/lrp6/β-catenin pathway in IBD, directly through CPA3 secretion and indirectly through neutrophils recruitment.

Background & aims: The exocrine pancreas has a limited regenerative capacity, but to what extent all acinar cells are involved in this process is unclear. Nevertheless, the heterogenous nature of acinar cells suggests that cells exhibiting higher plasticity might play a more prominent role in acinar regeneration. In that regard, Stmn1-expressing acinar cells have been identified as potential facultative progenitor-like cells in the adult pancreas. Here, we studied Stmn1-progeny under physiological conditions, during regeneration, and in the context of Kras^G12D expression.

Methods: We followed the fate of Stmn1-progenies both under baseline conditions, following caerulein-induced acute or chronic pancreatitis, pancreatic duct ligation, and in the context of Kras^G12D expression.

Results: The Stmn1-lineage contributes to baseline acinar cell turnover under physiological conditions. Furthermore, these cells rapidly proliferate and repopulate the acinar compartment in response to acute injury in an ADM-independent manner. Moreover, acinar regeneration during chronic pancreatitis progression is in conjunction with a decline in the proliferative capacity of the Stmn1-lineage. Interestingly, newly generated acinar cells display increased susceptibility to additional injury during recurrent acute pancreatitis (RAP). Finally, given their inability to form ADMs, the Stmn1-lineage fails to form PanINs upon oncogenic Kras expression.

Conclusions: Our findings establish the Stmn1-lineage as a pivotal subpopulation for acinar regeneration. The ability of these cells to restore acinar tissue in an ADM-independent manner distinguishes them as a critical regenerative population. This study presents a new paradigm for acinar regeneration and repair in the context of pancreatitis and neoplasia.