Biology subjects

Carlotti, F.

Publications and source records attributed to Carlotti, F..

8 recordsLinked to original sources

bioRxiv2026

The Human Pancreas Cell Atlas defines a healthy reference framework for disease contextualization and translational benchmarking

A central challenge in single-cell biology is distinguishing disease-associated remodeling from normal cellular heterogeneity. Addressing this challenge requires healthy reference frameworks that capture cellular diversity across individuals, technologies, and biological contexts. Here we present the Human Pancreas Cell Atlas (HPCA), a reference atlas of the healthy human pancreas integrating 815,126 single-cell and single-nucleus transcriptomes from 109 donors across 12 studies, diverse technologies, and demographics. Using benchmarked integration and community-driven annotations, HPCA defines 94 cell types and transcriptional states spanning endocrine, exocrine, immune, and stromal compartments. The atlas identifies rare endocrine populations, including a putative, spatially supported polyhormonal alpha-beta-delta state, and provides a unified framework for interpreting pancreatic cellular variation across diverse biological and demographic covariates. Projection of disease and model-system datasets onto HPCA contextualized endocrine and epithelial remodeling relative to healthy pancreatic states. Diabetes-associated endocrine cells remained embedded within the healthy endocrine state space while exhibiting disease-specific changes, as supported by spatial and eQTL concordance analyses. Integration with a pancreatic ductal adenocarcinoma atlas resolved injury-associated and malignant epithelial ecosystem regions across donors. Finally, the HPCA enables quantitative benchmarking of murine diabetes models and stem-cell-derived islets against human pancreatic reference states. Together, the HPCA establishes a healthy transcriptional coordinate system for interpreting disease-associated pathophysiology, experimental perturbation, and regenerative fidelity, illustrating how reference atlases can function as analytical frameworks rather than static cell catalogs.

bioinformatics↗

bioRxiv2025

Beta hydroxybutyrate alters beta cell identity and function in human islets

Fasting alters insulin secretion in humans and islet cell composition in mice. Fasting triggers ketogenesis, which increases beta hydroxybutyrate (BHB) levels. BHB is a signaling molecule that alters insulin secretion in primary islets. We hypothesized that BHB alters human islet cell composition and identity. Primary human islets were cultured with R-BHB or its non-metabolizable enantiomer, S-BHB. Human islets cultured with R-BHB, but not S-BHB, resulted in an increased C-peptide-glucagon+ to C-peptide+glucagon- cell ratio, and an increased frequency of C-peptide+glucagon+ bihormonal cells and NKX6.1+glucagon+ cells. Single-cell transcriptomics revealed upregulation of alpha cell identity genes in R-BHB treated islet beta cells. Alterations in beta cell identity were accompanied by increased basal insulin secretion in response to low glucose, and a reduced insulin stimulation index in response to high glucose. The differential effects of the two BHB enantiomers on islet cell composition indicated that ketone metabolism is involved in islet cell identity change. Supporting this hypothesis, a subpopulation of beta cells in islets treated with R-BHB characterized by low ketolysis genes, OXCT1 and ACAT1, did not show altered alpha and beta cell identity genes. Additionally, primary islets from type 2 diabetes donors, which exhibited reduced OXCT1 and ACAT1 expression relative to donors without diabetes, did not display altered islet cell composition and beta cell function from R-BHB treatment. Our findings uncover a novel role of BHB in modulating beta cell identity, with potential implications for islet function in states with chronically elevated ketone concentrations.

cell biology↗

bioRxiv2025

Latent plasticity of the human pancreas across development, health, and disease.

The pancreas plays a central role in major human diseases, yet our understanding of its cellular diversity and plasticity remains incomplete. Here, we present a single-cell multiomics atlas of the human pancreas, profiling over four million cells and nuclei from 57 donors across fetal development, adult homeostasis, and type 2 diabetes (T2D). Integrating sc/snRNA-seq, snATAC-seq, VASA-seq, spatial transcriptomics (Xenium), and multiplexed proteomics (CODEX), we resolve gene expression, chromatin accessibility, and spatial organization at high resolution. We identify transcriptionally plastic centroacinar-like cells (pCACs) in adults with fetal-like features, delineate endocrine and exocrine lineage trajectories during development, and uncover HNF1A-defined beta cell epigenetic states. In T2D, we observe shifts in beta cell subtypes and altered regulatory programs. Glucose perturbation of healthy islets reveals cell-type-specific adaptation and stress responses. This atlas provides a foundational framework to understand pancreas biology and the role of cellular plasticity in regeneration and disease.

genomics↗

bioRxiv2025

Cell type-specific eQTL detection from single-cell RNA-seq reveals post-transcriptional regulatory mechanisms in human islets

Gene regulatory networks (GRNs) must be robust to maintain cellular identity across individuals yet flexible to accommodate population genetic variants. Comparing expression quantitative trait loci (eQTLs) in health and disease can therefore reveal hidden players in gene regulation. Here, we developed a computational pipeline to infer post-transcriptional regulatory events and their functional consequences from eQTL signals located in the 3' untranslated region of genes using single-cell RNA sequencing. As a case study, we repurposed datasets from human islets of donors with and without type 2 diabetes (T2D). We identified eQTL landscapes differing by cell type and diabetes status, with cell-type specificity associated with gene differential expression and RNA-binding proteins. Integrating eQTLs with GWAS and miRNA hits linked variants in G6PC2, QDPR, and RELL1 to insulin secretion, endoplasmic reticulum stress, and oxidative phosphorylation. Our pipeline provides a framework to unravel post-transcriptional regulatory mechanisms in health and disease at cell type resolution.

genetics↗

bioRxiv2024

ER stress relief drives beta cell proliferation

Regenerating endogenous pancreatic {beta}-cells is a potentially curative yet currently elusive strategy for diabetes therapy. Mimicking the microenvironment of the developing pancreas and leveraging vascular signals that support pancreatic endocrinogenesis may promote {beta}-cell regeneration. We aimed to investigate whether recovery from experimental hypovascularization of the endocrine pancreas, achieved by modulating the transgenic production of a VEGF-A blocker in {beta}-cells, could trigger mouse {beta}-cell proliferation. Serendipitously, we found that transgene overexpression in {beta}-cells induces endoplasmic reticulum (ER) stress and that subsequent relief from this stress stimulates {beta}-cell proliferation independent of vessel recovery. Transient GFP overexpression in vivo and chemical induction of ER stress in vitro replicated this {beta}-cell cycling response. Our findings highlight the potential side effects of ER stress due to transgene overexpression in {beta}-cells and assert that ER stress relief serves as a potent regenerative stimulus.

molecular biology↗

bioRxiv2023

Compromised beta-cell identity in type 2 diabetes

Compromised beta-cell identity is emerging as an important contributor of beta-cell dysfunction in type 2 diabetes (T2D). Several studies suggest that hyperglycemia induces the inactivation of transcription factors involved in mature beta-cell identity. More specifically, chronic hyperglycemia leads to the downregulation of PDX1 and MAFA, two important beta-cell identity transcription factors and regulators of insulin promoter activity. Regulation of these transcription factors depends on interactions between multiple signaling cascades and processes driven by complex non-linear dynamics and taking place in different cellular compartments. To better understand these non-linear dynamics, we developed an integrated mathematical model of the underlying mechanisms regulating these key transcription factors. Our model was able to reproduce experimentally measured variations in the levels of PDX1, MAFA and insulin mRNA under different glucose concentrations. We used this model to simulate scenarios that could allow to restore PDX1 and MAFA levels and therefore insulin gene expression. From these simulations, FOXO1 emerged as an important target for the restoration of beta-cell identity. Author summaryGlucose regulation depends on the secretion of insulin by beta-cells and uptake of glucose by the peripheral cells mediated by the action of insulin. In type 2 diabetes both aspects can be compromised. Defective insulin secretion results from compromised beta-cell function. One of the reasons behind compromised beta-cell function is that beta-cells fail to express one or more of the genes involved in insulin production and secretion and thus maintenance of beta-cell identity. The processes involved in the regulation of insulin production and secretion are complex. In this work, we are particularly interested in the role and downregulation of transcription factors, PDX1 and MAFA, which are critical regulators of insulin production, in relation with compromised beta-cell identity and function in the presence of chronic hyperglycemia. To understand better these complex processes, we use mathematical modelling which enables to generate hypotheses and simulate scenarios to extend our understanding of the mechanisms leading to compromised beta-cell function in the presence of chronic hyperglycemia. Our model and similar models can serve to identify therapeutical targets in beta-cells in order to restore their function.

systems biology↗

bioRxiv2021

PCSK9 affects expression of key surface proteins in human pancreatic beta cells through intra- and extracellular regulatory circuits

Aims/hypothesisProprotein convertase subtilisin/kexin 9 (PCSK9) is involved in the degradation of LDLR. However, PCSK9 can target other proteins in a cell-type specific manner. While PCSK9 has been detected in pancreatic islets, its expression in insulin-producing pancreatic beta cells is debated. Herein, we studied PCSK9 expression, regulation and function in the human pancreatic beta cell line EndoC-{beta}H1. MethodsWe assessed PCSK9 expression in mouse and human pancreatic islets, and in the pancreatic beta cell line EndoC-{beta}H1. We also studied PCSK9 regulation by cholesterol, lipoproteins, Mevastatin, and by SREBPs transcription factors. To evaluate PCSK9 function in pancreatic beta cells, we performed PCSK9 gain-and loss-of-function experiments in EndoC-{beta}H1 using siPCSK9 or recombinant PCSK9 treatments, respectively. ResultsWe demonstrate that PCSK9 is expressed and secreted by pancreatic beta cells. In EndoC-{beta}H1 cells, PCSK9 expression is regulated by cholesterol and by SREBPs transcription factors. Importantly, PCSK9 knockdown results in multiple transcriptome, proteome and secretome deregulations and impaired insulin secretion. By gain- and loss-of-function experiments, we observed that PCSK9 regulates the expression levels of LDLR and VLDLR through an extracellular mechanism while CD36, PD-L1 and HLA-ABC are regulated through an intracellular mechanism. Conclusions/interpretationCollectively, these results highlight PCSK9 as an important regulator of CD36, PD-L1 and HLA-ABC cell surface expression in pancreatic beta cells. Data availabilityRNA-seq data have been deposited to GEO database with accession number GSE182016. Mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the following identifiers: PXD027921, PXD027911 and PXD027913.

cell biology↗

bioRxiv2021

Organization and dynamics of the cortical complexes controlling insulin secretion in β-cells.

Insulin secretion in pancreatic {beta}-cells is regulated by cortical complexes that are enriched at the sites of adhesion to extracellular matrix facing the vasculature. Many components of these complexes, including Bassoon, RIM, ELKS and liprins, are shared with neuronal synapses. Here, we show that insulin secretion sites also contain non-neuronal proteins LL5{beta} and KANK1, which in migrating cells organize exocytotic machinery in the vicinity of integrin-based adhesions. Depletion of LL5{beta} or focal adhesion disassembly triggered by myosin II inhibition perturbed the clustering of secretory complexes and attenuated the first wave of insulin release. While previous analyses in vitro and in neurons suggested that secretory machinery might assemble through liquid-liquid phase separation, analysis of endogenously labeled ELKS in pancreatic islets indicated that its dynamics is inconsistent with such a scenario. Instead, fluorescence recovery after photobleaching and single molecule imaging showed that ELKS turnover is driven by binding and unbinding to low-mobility scaffolds. Both the scaffold movements and ELKS exchange were stimulated by glucose treatment. Our findings help to explain how integrin-based adhesions control spatial organization of glucose-stimulated insulin release.

cell biology↗