Expression data from lung tumors of KrasTgfbr2 -/- mouse model
ABSTRACT: Recent data suggests that repression of the Type II TGF-B Receptor (Tgfr2) repression in human lung adenocarcinoma is important for progression from noninvasive to invasive adenocarcinoma. To test this hypothesis in a animal model of non-invasive lung cancer, we generated an inducible, lung specific Tgfbr2 knockout model in the oncogenic Kras mouse. LSL-KrasG12D positive mice were simultaneously backcrossed to C57/Bl6 mice and to the Tgfbr2 flox/flox mice. To induce tumors, 100 _l of saline containing 3x10e10 particles of an adenovirus containing the Cre recombinase (Ad.Cre) was administered to each LSL-KrasG12D mouse intra-nasally. Mice were sacrificed at 7 weeks after administration of Adeno-Cre. We used laser capture microdissection to acquire tumor cells from KrasTgfbr2-/- and KrasTgfbr2 wt mouse tumors.
Project description:Recent data suggests that repression of the Type II TGF-B Receptor (Tgfr2) repression in human lung adenocarcinoma is important for progression from noninvasive to invasive adenocarcinoma. To test this hypothesis in a animal model of non-invasive lung cancer, we generated an inducible, lung specific Tgfbr2 knockout model in the oncogenic Kras mouse. LSL-KrasG12D positive mice were simultaneously backcrossed to C57/Bl6 mice and to the Tgfbr2 flox/flox mice. To induce tumors, 100 ul of saline containing 3x10e10 particles of an adenovirus containing the Cre recombinase (Ad.Cre) was administered to each LSL-KrasG12D mouse intra-nasally. We evaluated the tumor microenvironment response to Tgfbr2 deficient tumor cells. We compared lung tumor cell and stromal cell transcriptional profiles from five-week KrasTgfbr2 -/- and nine-week KrasTgfbr2 WT mice. We used mice at these time points to allow comparison of the stromal compartment of similarly advanced tumors.
Project description:To identify downstream genes involved in cholangiocarcinoma development by genetic manipulations of Kras, TGFbR2, and CDH1, we cultured the biliary organoid from the extrahepatic bile duct of Cre-negative LSL-KrasG12D;Tgfbr2flox/flox;CDH1flox/flox (KTC) mice, and induced gene recombination using a lentivirus expressing Cre-recombinase (Cre+ KTC organoid) or control lentivirus (Cre- KTC organoid). Then, we compared gene expression signatures between Cre- KTC organoid organoid and Cre+ KTC organoid. Overall design: Biliary organoids isolated from KTC mice were infected with Cre-expressing or control lentivirus, and transduced cells were selected using puromycin. Two weeks later, we compared gene expression signatures between Cre- KTC organoid organoid and Cre+ KTC organoid.
Project description:Triple transgenic mice harboring a conditional Pten allele were developed and Pten was conditionally deleted in vivo from the lung epithelial cells using the doxycycline dependent Cre/LoxP approach. Microarray analysis of lung RNA isolated from Pten-deleted lung epithelial cells (PtenΔ/Δ) and control mice was performed to identify potential PTEN responsive genes. Overall design: Compound-transgenic mice harboring Pten gene with loxP-flanked exon-V (Ptenflox/flox), SP-C–rtTAtg/−; and TetO-Cretg/- were generated. Mice harboring SPC-rtTA/Ptenflox/flox, TetO-Cretg/-/Ptenflox/flox, SPC-rtTA, or TetO-Cretg/- as controls. Likewise, Dox treatment (E6.5 to E14.5) induced tumors in CCSP-rtTA/TetO-Cretg/-/LSL-KrasG12D/PtenΔ/Δ mice between 10-12 weeks of age. Mice expressing rtTA, or bearing TetO-Cretg/- alone, were normal controls
Project description:Pooled KRC (LSL-KrasG12D; Rb1L/L; Pdx1-Cre: oncogenic Kras and deleted Rb1 in the pancreas) cells derived from 2 month old mice were compared to pooled KC (LSL-KrasG12D; Pdx1-Cre: oncogenic Kras in the pancreas) cells derived from 8 month old mice.
Project description:These experiments aimed to determine the global gene expression patterns in p120WT, p120HET, and p120NULL cells in the context on oncogenic mutant KrasG12D Overall design: p120ctn was deleted using in vitro lentiviral Cre recombinase in pancreatic ductal cells isolated from LSL-KrasG12D;Ctnnd1wt/wt (KC-p120ctnWT), LSL-KrasG12D;Ctnnd1fl/wt (KC-p120ctnHET) , and LSL-KrasG12D;Ctnnd1fl/fl (KC-p120ctnNULL) mice.
Project description:Transcriptional profiling of mouse Prostate cancer cells comparing Pbsn-Cre LSL-KrasG12D P53L/L cells with Pbsn-Cre LSL-BrafV600E P53L/L cells, and to determine the effects of Kras or Braf mutantion on murine PCa gene expression. Overall design: Three-condition experiment, Pbsn-Cre LSL-KrasG12D P53L/L cells vs. Pbsn-Cre LSL-BrafV600E P53L/L cells vs. Pbsn-Cre P53L/L cells: 2 wild type replicates, 2 KrasG12D mutant replicates and 2 BrafV600E mutant replicates.
Project description:To investigate the role of SHP2 (Ptpn11) in pancreatic carcinogenesis, murine pancreatic whole tissue RNA samples of 9 week old mice with the genotypes Ptf1a-Cre;LSL-KrasG12D (ID-labels Kxxx) and Ptf1a-Cre;LSL-KrasG12D;Ptpn11fl/fl (ID-labels Mxxxx) were analyzed by microarray.
Project description:A subset of human pancreatic ductal adenocarcinoma cells (PDACs) is characterized by high Fosl1 expression and Fosl1 is linked to the control of pro-inflammatory pathways and growth of PDAC cells. To mimick the human disease in mice (> 90% of PDAC patients harbour Kras mutations) the mutated LSL-KrasG12D allele was combined with the pancreas specific Cre recombinase Ptf1aCre (p48Cre). The two pancreatic cancer cell lines (Ptf1aCre, LSL-KrasG12D/+) were isolated from these mice and used for transcriptomics studies. The two different murine pancreatic cancer cell lines (Ptf1aCre, LSL-KrasG12D) were treated with two different Fosl1 siRNAs and one control siRNA, each. 72h after transfection a sufficient knockdown was tested by immunoblotting and qPCR. Total mRNA was isolated and checked for integrity. According to manufacture's recommendation the samples were subjected to microarray analysis using the Affymetrix Mouse Gene ST 1.0 array chip to discover differentially expressed genes.
Project description:Although mutations in Kras are present in 21% of lung tumors, there is a high level of heterogeneity in phenotype and outcomes amongst lung cancer patients suggesting the importance of other pathways. Wnt/β-catenin signaling is a known oncogenic pathway that plays a well defined role in colon and skin cancer but its role in lung cancer remains unclear. We show that activation of Wnt/β-catenin in the bronchiolar epithelium of the adult lung does not promote tumor development by itself. However, activation of Wnt/β- catenin signaling leads to a dramatic increase in tumor formation both in overall tumor number and size compared to KrasG12D alone. We show that activation of Wnt/β- catenin signaling significantly alters the KrasG12D tumor phenotype resulting in a phenotypic switch from bronchiolar epithelium to the highly proliferative distal progenitors found in the embryonic lung. This is associated with a decrease in E- cadherin expression at the cell surface which may increase metastasis in Wnt/β-catenin signaling positive tumors. Together, these data suggest that activation of Wnt/β-catenin signaling in combination with other oncogenic pathways in lung epithelium may lead to a more aggressive phenotype due to the imposition of an embryonic distal progenitor phenotype accompanied by decreased E-cadherin expression. We performed microarray analysis of control murine lung, CC10-cre:KrasG12D, and CC10-cre:Ctnnb1ex3flox:LSL-KrasG12D double mutant micro-dissected murine lung tumors to determine their transcriptional phenotype. Overall design: Lungs of five-month-old mice were PBS inflated and all the tumors in each lobe were dissected. The total number of tumors obtained from three out of the 5 pulmonar lobes of each animal was called a sample the other two lobes were saved in case there were problems and the array needed to be repeated. Trizol was used to isolate RNA for microarray analysis. Samples & Genotypes: control murine lung n=2 animals, CC10-cre:KrasG12D n=2 animals, and CC10-cre:Ctnnb1ex3flox:LSL-KrasG12D n=2 animals.
Project description:Analysis of differentially expressed genes in CAF associated with PDAC vs NF. Genetically engineered mice with spontaneous pancreas cancer were generated. Their genotype is Ptfa-cre/+:LSL KrasG12D/+;Tgfrb2flox/flox. Cancer associated fibroblasts were expanded in vitro from the tumors of these mice (CAF). Normal fibroblasts (NF) were also expanded from normal pancreas of mice. The experiement consists in comparing the expression profile of CAF vs. NF. Expanded cultures of fibroblasts in vitro from pancreas tumor and normal pancreas tissue were lyzed in TRIzol and RNA extracted from them.