GSE111540
Transcriptional profiling of pancreatic acinar to ductal metaplasia using laser capture microdissection
Pancreatic ductal adenocarcinoma (PDAC) is a deadly disease when diagnosed at a late stage, however patient survivorship significantly increases when the disease is detect prior to metastasis. To study the earliest events leading to PDAC initiation, we developed a genetically engineered mouse model of PDAC utilizing a tamoxifen-inducible Cre Recombinase knocked into the transcription factor Ptf1a locus to induce expression of oncogenic KrasG12D and Trp53R270H alleles in adult pancreatic acinar cells. Mice of the genotype KrasLSL-G12D/+; Trp53LSL-R270H/+; Ptf1aCreERTM/+ (KPT) developed PDAC following tamoxifen injection while control Cre recombinase negative KrasLSL-G12D/+; Trp53LSL-R270H/+; (KP) mice injected with tamoxifen did not develop PDAC. Acinar cells comprising the pancreata of tamoxifen treated KPT mice we observed to undergo acinar to ductal metaplasia (ADM), and formed precancerous lesions. We used laser capture microdissection (LCM) and RNA sequencing to generate, to our knowledge, the first transcriptional profile of an enriched population of metaplastic acinar cells in situ. Comparing the transcriptional profile of metaplastic acinar cells with the transcriptional profile of healthy pancreatic tissue identified differentially expressed genes associated with ADM. Ingenuity pathway analysis revealed transcriptional regulators and canonical signaling pathways involved in ADM. LCM was used to generate a transactional profile of cancer cells isolated from pancreatic tumors, and differential gene expression analysis revealed a subset of genes which are overexpressed in both ADM and PDAC relative to healthy pancreas. Further analysis of genes expressed in both pancreatic cancer precursor ADM lesions and invasive PDAC may lead to the identification of novel biomarkers of PDAC. Overall design: Total RNA was extracted from metaplastic epithelial cells isolated using LCM from the pancreata of 3 unique tamoxifen treated KPT mice. Total RNA was extracted from cancer epithelium which was isolated from PDAC tumors using LCM. Tumors were resected from 3 unique tamoxifen treated KPT mice. Healthy pancreatic control total RNA was extracted from the pancreata of 3 unique tamoxifen treated KP mice. cDNA libraries were prepared from extracted RNA using the Kapa RNA HyperPrep kit with RiboEase (Kapa Biosystems), following the manufacturer's protocol. Sequencing was performed using the Illumina HiSeq 2500 platform using the HiSeq SBS V4 Kit. Real-time analysis (RTA) software version 2.2.38 was used to process the resulting sequencing images and perform base calling.