-
Normal and Sjogren's syndrome models of the murine lacrimal gland studied at single-cell resolution.Proceedings of the National Academy of... Oct 2023The lacrimal gland is of central interest in ophthalmology both as the source of the aqueous component of tear fluid and as the site of autoimmune pathology in the...
The lacrimal gland is of central interest in ophthalmology both as the source of the aqueous component of tear fluid and as the site of autoimmune pathology in the context of Sjogren's syndrome (SjS). To provide a foundational description of mouse lacrimal gland cell types and their patterns of gene expression, we have analyzed single-cell transcriptomes from wild-type (Balb/c) mice and from two genetically based SjS models, and (nonobese diabetic), and defined the localization of multiple cell-type-specific protein and mRNA markers. This analysis has uncovered a previously undescribed cell type, Car6+ cells, which are located at the junction of the acini and the connecting ducts. More than a dozen secreted polypeptides that are likely to be components of tear fluid are expressed by acinar cells and show pronounced sex differences in expression. Additional examples of gene expression heterogeneity within a single cell type were identified, including a gradient of Claudin4 along the length of the ductal system and cell-to-cell heterogeneity in transcription factor expression within acinar and myoepithelial cells. The patterns of expression of channels, transporters, and pumps in acinar, Car6+, and ductal cells make strong predictions regarding the mechanisms of water and electrolyte secretion. In and lacrimal glands, distinctive changes in parenchymal gene expression and in immune cell subsets reveal widespread interferon responses, a T cell-dominated infiltrate in the model, and a mixed B cell and T cell infiltrate in the model.
Topics: Female; Mice; Male; Animals; Sjogren's Syndrome; Lacrimal Apparatus; Mice, Inbred MRL lpr; Mice, Inbred NOD; Mice, Inbred BALB C; Disease Models, Animal
PubMed: 37812717
DOI: 10.1073/pnas.2311983120 -
Research Square Dec 2023Sjögren's Disease (SjD) is a systemic autoimmune disease without a clear etiology or effective therapy. Utilizing unbiased single-cell and spatial transcriptomics to...
Sjögren's Disease (SjD) is a systemic autoimmune disease without a clear etiology or effective therapy. Utilizing unbiased single-cell and spatial transcriptomics to analyze human minor salivary glands in health and disease we developed a comprehensive understanding of the cellular landscape of healthy salivary glands and how that landscape changes in SjD patients. We identified novel seromucous acinar cell types and identified a population of PRR4+CST3+WFDC2- seromucous acinar cells that are particularly targeted in SjD. Notably, GZMK+CD8 T cells, enriched in SjD, exhibited a cytotoxic phenotype and were physically associated with immune-engaged epithelial cells in disease. These findings shed light on the immune response's impact on transitioning acinar cells with high levels of secretion and explain the loss of this specific cell population in SjD. This study explores the complex interplay of varied cell types in the salivary glands and their role in the pathology of Sjögren's Disease.
PubMed: 38196575
DOI: 10.21203/rs.3.rs-3601404/v1 -
Gut Jul 2023Increasing evidence implicates mutation-induced protein misfolding and endoplasm reticulum (ER) stress in the pathophysiology of chronic pancreatitis (CP). The paucity...
OBJECTIVE
Increasing evidence implicates mutation-induced protein misfolding and endoplasm reticulum (ER) stress in the pathophysiology of chronic pancreatitis (CP). The paucity of animal models harbouring genetic risk variants has hampered our understanding of how misfolded proteins trigger CP. We previously showed that pancreatic triglyceride lipase (PNLIP) p.T221M, a variant associated with steatorrhoea and possibly CP in humans, misfolds and elicits ER stress in vitro suggesting proteotoxicity as a potential disease mechanism. Our objective was to create a mouse model to determine if PNLIP p.T221M causes CP and to define the mechanism.
DESIGN
We created a mouse model of p.T221M and characterised the structural and biochemical changes in the pancreas aged 1-12 months. We used multiple methods including histochemistry, immunostaining, transmission electron microscopy, biochemical assays, immunoblotting and qPCR.
RESULTS
We demonstrated the hallmarks of human CP in p.T221M homozygous mice including progressive pancreatic atrophy, acinar cell loss, fibrosis, fatty change, immune cell infiltration and reduced exocrine function. Heterozygotes also developed CP although at a slower rate. Immunoblot showed that pancreatic PNLIP T221M misfolded as insoluble aggregates. The level of aggregates in homozygotes declined with age and was much lower in heterozygotes at all ages. The p.T221M pancreas had increased ER stress evidenced by dilated ER, increased (BiP) mRNA abundance and a maladaptive unfolded protein response leading to upregulation of (CHOP), nuclear factor-κB and cell death.
CONCLUSION
Expression of PNLIP p.T221M in a preclinical mouse model results in CP caused by ER stress and proteotoxicity of misfolded mutant PNLIP.
Topics: Mice; Humans; Animals; Pancreatitis, Chronic; Pancreas; Acinar Cells; Endoplasmic Reticulum Stress; Unfolded Protein Response; Endoplasmic Reticulum Chaperone BiP
PubMed: 36631248
DOI: 10.1136/gutjnl-2022-327960 -
Cell Death and Differentiation Sep 2021Maintenance of the pancreatic acinar cell phenotype suppresses tumor formation. Hence, repetitive acute or chronic pancreatitis, stress conditions in which the acinar...
Maintenance of the pancreatic acinar cell phenotype suppresses tumor formation. Hence, repetitive acute or chronic pancreatitis, stress conditions in which the acinar cells dedifferentiate, predispose for cancer formation in the pancreas. Dedifferentiated acinar cells acquire a large panel of duct cell-specific markers. However, it remains unclear to what extent dedifferentiated acini differ from native duct cells and which genes are uniquely regulating acinar cell dedifferentiation. Moreover, most studies have been performed on mice since the availability of human cells is scarce. Here, we applied a non-genetic lineage tracing method of human pancreatic exocrine acinar and duct cells that allowed cell-type-specific gene expression profiling by RNA sequencing. Subsequent to this discovery analysis, one transcription factor that was unique for dedifferentiated acinar cells was functionally characterized. RNA sequencing analysis showed that human dedifferentiated acinar cells expressed genes in "Pathways of cancer" with a prominence of MECOM (EVI-1), a transcription factor that was not expressed by duct cells. During mouse embryonic development, pre-acinar cells also transiently expressed MECOM and in the adult mouse pancreas, MECOM was re-expressed when mice were subjected to acute and chronic pancreatitis, conditions in which acinar cells dedifferentiate. In human cells and in mice, MECOM expression correlated with and was directly regulated by SOX9. Mouse acinar cells that, by genetic manipulation, lose the ability to upregulate MECOM showed impaired cell adhesion, more prominent acinar cell death, and suppressed acinar cell dedifferentiation by limited ERK signaling. In conclusion, we transcriptionally profiled the two major human pancreatic exocrine cell types, acinar and duct cells, during experimental stress conditions. We provide insights that in dedifferentiated acinar cells, cancer pathways are upregulated in which MECOM is a critical regulator that suppresses acinar cell death by permitting cellular dedifferentiation.
Topics: Acinar Cells; Animals; Cell Death; Cell Dedifferentiation; Disease Models, Animal; Humans; MDS1 and EVI1 Complex Locus Protein; Mice; Oncogenes; Signal Transduction
PubMed: 33762742
DOI: 10.1038/s41418-021-00771-6 -
Developmental Cell Sep 2022Three-dimensional mammary epithelial acini are a model for understanding how microenvironment-driven signaling coordinates cell behavior and tissue morphogenesis. In...
Three-dimensional mammary epithelial acini are a model for understanding how microenvironment-driven signaling coordinates cell behavior and tissue morphogenesis. In this issue of Developmental Cell, Ender et al. use live-cell imaging to capture dynamic spatiotemporal patterns of ERK activity that instruct cell migration and survival fates in developing acini.
Topics: Acinar Cells; Cell Movement; Epithelial Cells; Morphogenesis; Signal Transduction
PubMed: 36167056
DOI: 10.1016/j.devcel.2022.09.001 -
Cellular and Molecular Gastroenterology... 2023Development of pancreatic ductal adenocarcinoma (PDAC) is a multistep process intensively studied; however, precocious diagnosis and effective therapy still remain...
BACKGROUND AND AIMS
Development of pancreatic ductal adenocarcinoma (PDAC) is a multistep process intensively studied; however, precocious diagnosis and effective therapy still remain unsatisfactory. The role for Notch signaling in PDAC has been discussed controversially, as both cancer-promoting and cancer-antagonizing functions have been described. Thus, an improved understanding of the underlying molecular mechanisms is necessary. Here, we focused on RBPJ, the receiving transcription factor in the Notch pathway, examined its expression pattern in PDAC, and characterized its function in mouse models of pancreatic cancer development and in the regeneration process after acute pancreatitis.
METHODS
Conditional transgenic mouse models were used for functional analysis of RBPJ in the adult pancreas, initiation of PDAC precursor lesions, and pancreatic regeneration. Pancreata and primary acinar cells were tested for acinar-to-ductal metaplasia together with immunohistology and comprehensive transcriptional profiling by RNA sequencing.
RESULTS
We identified reduced RBPJ expression in a subset of human PDAC specimens. Ptf1α-Cre-driven depletion of RBPJ in transgenic mice revealed that its function is dispensable for the homeostasis and maintenance of adult acinar cells. However, primary RBPJ-deficient acinar cells underwent acinar-to-ductal differentiation in ex vivo. Importantly, oncogenic KRAS expression in the context of RBPJ deficiency facilitated the development of pancreatic intraepithelial neoplasia lesions with massive fibrotic stroma formation. Interestingly, RNA-sequencing data revealed a transcriptional profile associated with the cytokine/chemokine and extracellular matrix changes. In addition, lack of RBPJ delays the course of acute pancreatitis and critically impairs it in the context of KRAS expression.
CONCLUSIONS
Our findings imply that downregulation of RBPJ in PDAC patients derepresses Notch targets and promotes KRAS-mediated pancreatic acinar cells transformation and desmoplasia development.
Topics: Animals; Humans; Mice; Acinar Cells; Acute Disease; Carcinoma in Situ; Carcinoma, Pancreatic Ductal; Immunoglobulin J Recombination Signal Sequence-Binding Protein; Mice, Transgenic; Pancreatic Neoplasms; Pancreatitis; Proto-Oncogene Proteins p21(ras)
PubMed: 37543088
DOI: 10.1016/j.jcmgh.2023.07.013 -
Cell Death & Disease Oct 2022Noninflammatory clearance of dying cells by professional phagocytes, termed efferocytosis, is fundamental in both homeostasis and inflammatory fibrosis disease but has...
Noninflammatory clearance of dying cells by professional phagocytes, termed efferocytosis, is fundamental in both homeostasis and inflammatory fibrosis disease but has not been confirmed to occur in chronic pancreatitis (CP). Here, we investigated whether efferocytosis constitutes a novel regulatory target in CP and its mechanisms. PRSS1 transgenic (PRSS1) mice were treated with caerulein to mimic CP development. Phospholipid metabolite profiling and epigenetic assays were performed with PRSS1 CP models. The potential functions of Atp8b1 in CP model were clarified using Atp8b1-overexpressing adeno-associated virus, immunofluorescence, enzyme-linked immunosorbent assay(ELISA), and lipid metabolomic approaches. ATAC-seq combined with RNA-seq was then used to identify transcription factors binding to the Atp8b1 promoter, and ChIP-qPCR and luciferase assays were used to confirm that the identified transcription factor bound to the Atp8b1 promoter, and to identify the specific binding site. Flow cytometry was performed to analyze the proportion of pancreatic macrophages. Decreased efferocytosis with aggravated inflammation was identified in CP. The lysophosphatidylcholine (LPC) pathway was the most obviously dysregulated phospholipid pathway, and LPC and Atp8b1 expression gradually decreased during CP development. H3K27me3 ChIP-seq showed that increased Atp8b1 promoter methylation led to transcriptional inhibition. Atp8b1 complementation substantially increased the LPC concentration and improved CP outcomes. Bhlha15 was identified as a transcription factor that binds to the Atp8b1 promoter and regulates phospholipid metabolism. Our study indicates that the acinar Atp8b1/LPC pathway acts as an important "find-me" signal for macrophages and plays a protective role in CP, with Atp8b1 transcription promoted by the acinar cell-specific transcription factor Bhlha15. Bhlha15, Atp8b1, and LPC could be clinically translated into valuable therapeutic targets to overcome the limitations of current CP therapies.
Topics: Animals; Mice; Acinar Cells; Adenosine Triphosphatases; Ceruletide; Histones; Inflammation; Lysophosphatidylcholines; Macrophages; Pancreatitis, Chronic; Phospholipid Transfer Proteins; Transcription Factors
PubMed: 36273194
DOI: 10.1038/s41419-022-05322-6 -
Japanese Journal of Clinical Oncology Mar 2024Pancreatic acinar cell carcinoma is a rare form (0.2-4.3%) of pancreatic neoplasm with unique clinical and molecular characteristics, which largely differ from... (Review)
Review
Pancreatic acinar cell carcinoma is a rare form (0.2-4.3%) of pancreatic neoplasm with unique clinical and molecular characteristics, which largely differ from pancreatic ductal adenocarcinoma. Pancreatic acinar cell carcinoma occurs more frequently in males and can occur in children. Serum lipase is elevated in 24-58% of patients with pancreatic acinar cell carcinoma. Pancreatic acinar cell carcinomas tend to be large at diagnosis (median tumour size: ~5 cm) and are frequently located in the pancreas head. Radiologically, pancreatic acinar cell carcinoma generally exhibits a solid appearance; however, necrosis, cystic changes and intratumoral haemorrhage can occur in larger lesions. Immunostaining is essential for the definitive diagnosis of pancreatic acinar cell carcinoma. Compared with pancreatic ductal adenocarcinoma, pancreatic acinar cell carcinoma has a more favourable prognosis. Although radical surgery is recommended for patients with pancreatic acinar cell carcinoma who do not have distant metastases, the recurrence rate is high. The effectiveness of adjuvant therapy for pancreatic acinar cell carcinoma is unclear. The response to FOLFIRINOX is generally favourable, and some patients achieve a complete response. Pancreatic acinar cell carcinoma has a different genomic profile compared with pancreatic ductal adenocarcinoma. Although genomic analyses have shown that pancreatic acinar cell carcinoma rarely has KRAS, TP53 and CDKN2A mutations, it has a higher prevalence of homologous recombination-related genes, including BRCA1/2 and ATM, than pancreatic ductal adenocarcinoma, suggesting high sensitivity to platinum-containing regimens and PARP inhibitors. Targeted therapies for genomic alternations are beneficial. Therefore, genetic testing is important for patients with pancreatic acinar cell carcinoma to choose the optimal therapeutic strategy.
Topics: Male; Child; Humans; Pancreatic Neoplasms; Carcinoma, Acinar Cell; Antineoplastic Combined Chemotherapy Protocols; BRCA1 Protein; BRCA2 Protein; Carcinoma, Pancreatic Ductal
PubMed: 38109477
DOI: 10.1093/jjco/hyad176 -
Nature Communications Oct 2023Exocrine acinar cells in salivary glands (SG) are critical for oral health and loss of functional acinar cells is a major clinical challenge. Fibroblast growth factor...
Exocrine acinar cells in salivary glands (SG) are critical for oral health and loss of functional acinar cells is a major clinical challenge. Fibroblast growth factor receptors (FGFR) are essential for early development of multiple organs, including SG. However, the role of FGFR signaling in specific populations later in development and during acinar differentiation are unknown. Here, we use scRNAseq and conditional deletion of murine FGFRs in vivo to identify essential roles for FGFRs in craniofacial, early SG development and progenitor function during duct homeostasis. Importantly, we also discover that FGFR2 via MAPK signaling is critical for seromucous acinar differentiation and secretory gene expression, while FGFR1 is dispensable. We show that FGF7, expressed by myoepithelial cells (MEC), activates the FGFR2-dependent seromucous transcriptional program. Here, we propose a model where MEC-derived FGF7 drives seromucous acinar differentiation, providing a rationale for targeting FGFR2 signaling in regenerative therapies to restore acinar function.
Topics: Animals; Mice; Cell Differentiation; Homeostasis; Orosomucoid; Salivary Glands; Signal Transduction
PubMed: 37838739
DOI: 10.1038/s41467-023-42243-0 -
Diabetology International Oct 2020Distinct features of the pancreas of fulminant type 1 diabetes (FT1DM) include (1) enterovirus infection of the islets and exocrine acinar tissue. (2) Activated innate... (Review)
Review
Distinct features of the pancreas of fulminant type 1 diabetes (FT1DM) include (1) enterovirus infection of the islets and exocrine acinar tissue. (2) Activated innate immune responses: MDA5 and RIG-I expression and TLR4 and TLR9 expression in the islets of FT1DM. (3) Combined activation of the STAT/JNK and NFkB pathways, resulting in Type I interferon (IFN) and proinflammatory cytokine (i.e., IFNγ) expression in islet beta cells and MHC class I hyper-expression. (4) Activation of dendritic cells followed by effector cell infiltration of CD8 T cells and CD68 macrophages, resulting in apoptosis and neurosis of islet cells and exocrine acinar cells. (5) Many chemo-attractants (i.e., CXCL10) and chemotactic activators (i.e., l-plastin) were induced by a viral infection. (6) Mutual stimulating effect of cytokines expressed in beta cells in autocrine and paracrine mechanisms may enhance beta-cell destruction through the STA1-caspase pathway. (7) Proteomics analysis using laser capture microdissection followed by mass spectrometry found 38 molecules in inflamed islets of FT1DM, which were not highlighted before. Our pathologically verified model of beta-cell destruction in FT1DM will contribute to anti-virus therapy of type 1 diabetes in the near future.
PubMed: 33088638
DOI: 10.1007/s13340-020-00462-6