Analysis of donor pancreata defines the transcriptomic signature and microenvironment of early neoplastic lesions.
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ABSTRACT: The adult healthy human pancreas has been poorly studied given lack of indication to obtain tissue from the pancreas in the absence of disease and rapid postmortem degradation. We obtained pancreata from brain dead donors thus avoiding any warm ischemia time. We found that neoplastic lesions occur frequently in healthy organs, in donors as young as in their 3rd decade of life. Using the 10X Genomics Platform, we performed single cell RNA sequencing on 6 donor pancreata, 5 of which we separately sequenced tissue from the pancreas head and tail, for a total of 11 sequencing runs. This is a robust single-cell dataset of healthy, nonpathologic pancreas tissue and includes acinar, ductal, and non-epithelial cells (myeloid cells, fibroblasts, endothelial cells, lymphocytes). We compared this dataset of normal pancreata to single cell sequencing of tumor samples previously published in Steele, et al, Nature Cancer 2020, (raw fastq in dbGap phs002071) realigned to GRCh38 reference genome (CellRanger 6.0). This GEO series contains the raw and filtered feature matrices for the donor pancreata as well as the realigned tumor samples. Supplementary files contain the aggregate feature matrices and the metadata of the donor pancreata samples and the tumor samples.
Project description:The adult healthy human pancreas has been poorly studied given lack of indication to obtain tissue from the pancreas in the absence of disease and rapid postmortem degradation. We obtained pancreata from brain dead donors thus avoiding any warm ischemia time. The 30 donors were diverse in age and race and had no known pancreas disease. Histopathological analysis of the samples revealed PanIN lesions in most individuals irrespective of age. Using a combination of multiplex immunohistochemistry, single cell RNA sequencing, and spatial transcriptomics, we provide the first ever characterization of the unique microenvironment of the adult human pancreas and of sporadic PanIN lesions. We compared healthy pancreata to pancreatic cancer and peritumoral tissue and observed distinct transcriptomic signatures in fibroblasts, and, to a lesser extent, macrophages. PanIN epithelial cells from healthy pancreata were remarkably transcriptionally similar to cancer cells, suggesting that neoplastic pathways are initiated early in tumorigenesis.
Project description:The decreasing numbers of Donation after Brain Death (DBD) donors necessitates the comprehensive evaluation of Donation after Cardiac Death Donors (DCD) as a source of pancreata. The aim of this study was to characterize pancreata and islets from DCD and DBD donors with respect to markers of cellular stress that may indicate compromised islet quality. Immunohistochemical staining of pre-isolation pancreas biopsies found increased numbers of caspase 3 positive islets in DBD, while markers of oxidative stress (nitrotyrosine, CML, and HNE) were elevated in DCD. Assessment of islet quality by standard (yield, morphology, fluorescence microscopy, and glucose stimulated insulin secretion) and novel methods (flow cytometry, HPLC quantification of ATP) did not reveal significant differences. However, the post culture loss of DCD islets was increased compared to DBD, and DCD islets showed delayed functional potency when transplanted into diabetic NOD.scid mice. Microarray analysis of cultured islets showed increased expression of multiple stress pathway related genes in DCD compared to DBD. Together these data indicate that the current standard donor management, pancreas recovery and preservation practices are insufficient to quench the oxidative stress injury suffered by DCD islets which leads to loss in culture and may complicate their use in clinical transplant. Keywords: cell type comparison
Project description:We investigated the distinct state of pancreas resident memory T cells (TRM) by whole transcriptome profiling of sorted CD8+TRM cells from donor-matched pancreata, jejunum, and PLN compared to circulating blood CD8+TEM cells from living healthy donors. We identified 2029 genes differentially expressed in pancreas TRM with comparison to both PLN and jejunum TRM. Strong correlation was observed in the differentially upregulated and downregulated genes between pancreas vs. jejunum TRM and pancreas vs. PLN TRM which together define a pancreas-specific gene signature.
Project description:Gene expression profiling of monocyte-derived Dendritic cells manufactured several times from one healthy donor and one single time from 8 different healthy donors to test the sources of variability Sources tested: Assay-related variability (8 samples), Manufacture-related variability (5 samples), Intra-donor-related variability (5 samples), inter-donor-related variability (9 samples)
Project description:Experiment designed to study the effect of deregulated myc on pancreatic cancer progression and regression. In the KRasG12D/RosaMycER mouse model KRas and MycER are expressed exclusively in the pancreas, but MycER activity depends on Tamoxifen presence. RNAseq was preformed on pancreata from mice treated with Tamoxifen as follows: 1- Normal food. 2- Tamoxifen food 2 weeks. 3- Tamoxifen food 2 weeks, followed by normal food 1 day. 4- Tamoxifen food 2 weeks, followed by normal food 3 days.
Project description:LFQ analysis of matrisome fractions enriched from 12 murine PDA or PanIN samples (samples 1 - 12) and a normal pancreas pool (samples 13) pooled from 10 pancreata of C57-Bl6 animals. Three independent enrichments have been conducted for each sample with each having had four injections.
Project description:Purpose: Single cell sequencing has advanced our understanding of kidney biology. The goals of this study are to compare single nucleus RNA (snRNA-seq) and single nucleus assay for transposase accessible chromatin sequencing (snATAC-seq) from healthy control donors and donors with type 2 diabetic kidney disease to identify altered signaling pathways and chromatin accessibility patterns observed in diabetic kidney disease. Methods: Nuclear dissociation of human kidney cortex samples was performed on samples obtained from patients undergoing tumor nephrectromy or deceased organ donors. snRNA-seq and snATAC-seq libraries were prepared using 10X Genomics kits according to manufacturers instructions. For this study, two healthy control libraries from one donor were prepared with the following chemistries (1 - single cell 3-prime v3, 1 - single cell ATAC v1) and nine DKD libraries from seven donors were prepared with the following chemistries (2 - single cell 5-prime v2 paired-end, 7 - single cell ATAC v1). Raw data for three healthy control and three DKD snRNA-seq libraries sequenced with single nucleus 5-prime R2 only can be found in GSE13188213. Raw data for two healthy control snRNA-seq sequenced with single nucleus 5-prime paired-end and five snATAC-seq single nucleus ATAC v1 libraries can be found in GSE151302. snRNA-seq and snATAC-seq libraries from these earlier studies were recounted with cellranger (v4.0) or cellranger-atac (v2.0) to provide consistency across processed data files.
Project description:How human islet antigen reactive CD4+ memory T cells (IAR T cells) from peripheral blood affect Type 1 Diabetes (T1D) progression in the pancreas is poorly understood. We identified paired alpha/beta (TRA/TRB) T cell receptors (TCRs) in IAR T cells from the blood of healthy, at-risk, new onset, and established T1D donors, and measured sequence overlap with TCRs in pancreata from organ donors. We detected extensive TRA junction sharing between IAR T cells and pancreatic infiltrating T cells (PIT), with perfect or single mismatched TRA junction amino acid sequences comprising ~34% of unique IAR TRA junctions. PIT-matched TRA junctions were largely public, and showed significant nucleotide sequence convergence, increased use of germline-encoded residues in epitope engagement, and a potential for cross-reactivity. The link with T cells in the pancreas implicates autoreactive IAR T cells with shared TRA junctions in the prediabetic and new onset phases of T1D progression.
Project description:How human islet antigen reactive CD4+ memory T cells (IAR T cells) from peripheral blood affect Type 1 Diabetes (T1D) progression in the pancreas is poorly understood. We identified paired alpha/beta (TRA/TRB) T cell receptors (TCRs) in IAR T cells from the blood of healthy, at-risk, new onset, and established T1D donors, and measured sequence overlap with TCRs in pancreata from organ donors. We detected extensive TRA junction sharing between IAR T cells and pancreatic infiltrating T cells (PIT), with perfect or single mismatched TRA junction amino acid sequences comprising ~34% of unique IAR TRA junctions. PIT-matched TRA junctions were largely public, and showed significant nucleotide sequence convergence, increased use of germline-encoded residues in epitope engagement, and a potential for cross-reactivity. The link with T cells in the pancreas implicates autoreactive IAR T cells with shared TRA junctions in the prediabetic and new onset phases of T1D progression.