Cross-species single-cell analysis of pancreatic ductal adenocarcinoma reveals cancer-associated fibroblasts expressing MHC class II
Ontology highlight
ABSTRACT: This study used 10X Genomics, single-cell RNA-sequencing to examine the cell types present in the KrasLSL-G12D; Trp53LSL-R172H; Pdx1-Cre (KPC) mouse model for pancreatic ductal adenocarcinoma. The study analyzed tumors from 4 different mice. For each tumor, we performed flow sorting to isolate all viable cells, and to isolate a fibroblast-enriched population of cells for single-cell RNA-seq to determine the transcriptomes of individual cells in KPC pancreatic ductal adenocarcinoma tumors.
Project description:This study used 10X Genomics, single-cell RNA-sequencing to examine the differentiation states of cancer cells present in tumors derived from the KrasLSL-G12D; Trp53LSL-R172H; Pdx1-Cre (KPC) mouse model of pancreatic ductal adenocarcinoma. The study analyzed tumors from 8 different mice.
Project description:This study used Illumina strand-specific, paired-end RNA-sequencing to examine gene expression differences between matched murine tumor- and metastasis-derived mouse pancreatic ductal adenocarcinoma (PDAC) cells grown as three-dimensional, organoid cultures. The study analyzed 16 organoid lines derived from matched primary PDAC tumors and PDAC metastases from 6 KPC (KrasLSL-G12D; Trp53LSL-R172H; Pdx1-Cre) mice.
Project description:We harvested and sequenced the spontaneous pancreatic tumor generated by a 6-month-old KPC mouse (KrasLSL-G12D; Trp53LSL-R172H; Ptf1a-Cre).
Project description:Constitutive Kras and NF-kB activation is identified as signature alterations in human pancreatic ductal adenocarcinoma (PDAC). Here, we report that pancreas-targeted IKK2/beta inactivation inhibited NF-kB activation and completely suppressed PDAC development. Our findings demonstrated that NF-kB is required for development of pancreatic ductal adenocarcinoma that was initiated by Kras activation. Pancreatic tissue from 4 groups of mice were used in this project: (1) the pancreas normal appearance of Pdx1-cre;KrasLSL-G12D;IKK2/beta mice, (2) the normal pancreas of Pdx1-cre;KrasLSL-G12D mice, (3) the pancreatic lesion of pancreatic intraepithelial neoplasia (PanIN) of Pdx1-cre;KrasLSL-G12D mice, and (4) the pancreatic lesion of PDAC of Pdx1-cre;KrasLSL-G12D mice. Each group included three mice. RNA samples from mouse pancreas were hybridized on GeneChip Mouse Gene 1.0 ST arrays (Affymetrix). Group (1) and group (2) were compared, and group (2), group (3) and group (4) were compared.
Project description:Ablative RT results in increased expression of CCL2 within the tumor microenvironment of pancreatic ductal adenocarcinoma (PDAC) and also increased recruitment of CD45+CD11b+Ly6Chi inflammatory monocytes/macrophages. This increase in CCL2 expression and recruitment of inflammatory monocytes/macrophages is a mechanism of resistance to the anti-tumor effects of ablative radiotherapy (RT). We used microarrays to study changes in gene expression patterns of inflammatory monocytes/macrophages sorted from the tumor microenvironment after ablative RT in a subcutenous model of pancreatic adenocarcinoma. From this, we identified 8 genes with an absolute fold change of expression equal to or greater than 2 with a false discovery rate equal to or less than 25 %. A pancreatic cancer tumor cell line derived from spontaneously arising tumors in KrasLSL-G12D/+, Trp53LSL-R172H/+, Pdx1-Cre (KPC) mice was subcutaneously implanted into 8 week old female C57BL/6 and allowed to grow for 14 days. After 14 days, 4 mice received 20 Gy of radiation, and 4 mice received a sham treatment. One day post treatment, tumors were harvested, and inflammatory monocytes/macrophages were isolated using flow sorting based on a surface expression phenotype of CD45+ CD11b+ Ly6Chi. From this cell population, total RNA was extracted for creation of cDNA and hybridization on Affymetrix microarrays. From the microarrays, a set of genes associated with radiation treatment of PDAC was identified.
Project description:The goal of this study is to investigate the molecular mechanisms of LIFR signaling in pancreatic cancer cells isolated from the classical pancreatic ductal adenocarcinoma mouse model KrasLSL-G12D;Tp53f/f;Rosa26LSL-Luc;Pdx1-Cre mice EpCAM+ pancreatic cancer cells were isolated by FACS from pancreatic tumors developed in Lifrf/f;KrasLSL-G12D/+;Trp53f/f;Rosa26LSL-Luc;Pdx1-Cre or LifrWT;KrasLSL-G12D/+;Trp53f/f;Rosa26LSL-Luc;Pdx1-Cre mice respectively and directly lysed for RNA extraction
Project description:Murine pancreatic ductal adenocarcinoma tumor organoids (T6 and T23) were generated from primary tumors from KrasLSL-G12D/+; Trp53LSL-R172H/+; Pdx1-Cre mice. At the first passage, cells in Matrigel were overlaid with mouse complete medium alone or supplemented with 10 uM Nutlin-3a to select for cells that have undergone loss-of-heterozygosity (LOH) for the wild-type (wt, "+") allele of Trp53. Organoids were cultured for 3 passages in the presence or absense of Nutlin-3a, and then culturing was continued in unsupplemented mouse complete medium. H3K27ac ChIP-sequencing (ChIP-seq) was performed to identify regions differentially enriched for H3K27ac binding in Trp53-LOH vs. wt tumor organoids.
Project description:The goal of this study is to investigate the molecular mechanisms of LIF action on pancreatic cancer cells in the classical pancreatic ductal adenocarcinoma mouse model KrasLSL-G12D;Tp53f/f;Rosa26LSL-Luc;Pdx1-Cre mice EpCAM+ pancreatic cancer cells were isolated from pancreatic tumors developped in KrasLSL-G12D;Tp53f/f;Rosa26LSL-Luc;Pdx1-Cre mice treated with either gemcitabine plus control IgG or gemcitabine plus anti-LIF antibody by FACS
Project description:This study used Illumina strand-specific, paired-end RNA-sequencing to examine gene expression differences between normal and neoplastic mouse pancreatic ductal cells grown as three-dimensional, organoid cultures. The study analyzed 19 independently derived organoid lines. These 19 lines included 7 biological replicate "mN," or mouse normal, organoid lines, derived from the pancreatic ductal cells of wild-type C57Bl/6J or C57Bl/6N mice. In addition, the study analyzed 6 biological replicate "mP," or mouse PanIN, organoid lines, derived from PanIN-containing mouse pancreata from the KrasLSL-G12D; Pdx1-Cre mouse model. (The presence of PanIN in the pancreata from which these organoid lines were derived was confirmed by examining histological sections from the same pancreata). The final 6 lines examined in this study were biological replicate "mT," or mouse tumor, organoid lines, derived from mouse pancreata containing pancreatic ductal adenocarcinoma (PDAC) from the KrasLSL-G12D; Trp53LSL-R162H; Pdx1-Cre mouse model. We genes differentially expressed in the PanIN- or tumor-derived mouse organoid lines, relative to the mouse normal organoids, that may reflect expression changes required for pancreatic tumorigenesis.
Project description:Pancreatic ductal adenocarcinoma (PDAC) causes involuntary wasting of adipose and muscle tissue, also known as cachexia. Cachexia is a major cause of cancer-related deaths, particularly among patients with PDAC. Here we profiled gene expression in adipose tissue and skeletal muscle in normal/sham control mice and in mice bearing orthotopic PDAC tumors. PDAC tumors were initiated by intra-pancreatic injection of a cell line derived from the KPC (Kras-G12D;Trp-R172H;Pdx1::Cre) genetically engineered mouse model of pancreatic cancer, or by injection of the same cell line deleted for the IL6 gene using CRISPR/Cas9. KPC-IL6 knockout (ko) cells caused less adipose wasting and no muscle loss compared with KPC-wildtype (wt) cells.