ArticleBMC genomics2024
NKX2-2 based nuclei sorting on frozen human archival pancreas enables the enrichment of islet endocrine populations for single-nucleus RNA sequencing.
Article in BMC genomics, 2024. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
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4 citing papers in PubMed.
- Decoding adult murine pancreatic islet cell diversity through cell type-resolved proteomics and phosphoproteomics.Communications biology · 2025Article
- Evaluating cell-specific gene expression using single-cell and single-nuclei RNA-sequencing data from human pancreatic islets of the same donors.Scientific reports · 2025Article
- Molecular Mechanisms of Human Pancreatic Islet Dysfunction Under Overnutrition Metabolic Stress.Diabetes · 2025Article
- β-cell heterogeneity and molecular plasticity in type 2 diabetes: multi-omics perspectives and the role of gut microbiota.Frontiers in cell and developmental biology · 2025Review
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9 authors.
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Abstract
backgroundCurrent approaches to profile the single-cell transcriptomics of human pancreatic endocrine cells almost exclusively rely on freshly isolated islets. However, human islets are limited in availability. Furthermore, the extensive processing steps during islet isolation and subsequent single cell dissolution might alter gene expressions. In this work, we report the development of a single-nucleus RNA sequencing (snRNA-seq) approach with targeted islet cell enrichment for endocrine-population focused transcriptomic profiling using frozen archival pancreatic tissues without islet isolation.
resultsWe cross-compared five nuclei isolation protocols and selected the citric acid method as the best strategy to isolate nuclei with high RNA integrity and low cytoplasmic contamination from frozen archival human pancreata. We innovated fluorescence-activated nuclei sorting based on the positive signal of NKX2-2 antibody to enrich nuclei of the endocrine population from the entire nuclei pool of the pancreas. Our sample preparation procedure generated high-quality single-nucleus gene-expression libraries while preserving the endocrine population diversity. In comparison with single-cell RNA sequencing (scRNA-seq) library generated with live cells from freshly isolated human islets, the snRNA-seq library displayed comparable endocrine cellular composition and cell type signature gene expression. However, between these two types of libraries, differential enrichments of transcripts belonging to different functional classes could be observed.
conclusionsOur work fills a technological gap and helps to unleash frozen archival pancreatic tissues for molecular profiling targeting the endocrine population. This study opens doors to retrospective mappings of endocrine cell dynamics in pancreatic tissues of complex histopathology. We expect that our protocol is applicable to enrich nuclei for transcriptomics studies from various populations in different types of frozen archival tissues.
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