Project description:Tamoxifen is an anti-estrogen drug used in the treatment of Estrogen Receptor (ER) positive breast cancer. Effects and side effects of tamoxifen is the sum of tamoxifen and all its metabolites. Using concentrations that mimic the clinical situation we examined effects of 4OHtam, 4OHNDtam and NDtam on global gene expression in 17beta-estradiol (E2) treated MCF-7 cells.
Project description:Determine differentially expressed human genes in A549 tumors when co-cultured in spheroids with mouse fibroblast harbouring a mutation in the heparan sulphate elongation enzyme Ext1 compared to corresponding wild-type fibroblast/A549-containing spheroids.
Project description:The aim was to compare the global gene expression of epididymal white adipose tissue (eWAT) of WT and Irx5-KO mice. Mice 10 weeks of age were fed a high-fat diet for 10 weeks before eWAT was dissected out, RNA extracted and microarray performed
Project description:The aim of the project was to compare global gene expression in adipocytes from obese patients and lean controls. Subcutaneous adipose tissue was collected from severely obese patients undergoing bariatric surgery (average body-mass index (BMI) of 45.5 kg/m2 (n = 12, thereof 4 men) and healthy lean patients undergoing hernia repairs (average BMI of 24.2 kg/m2 (n = 12, thereof 7 men), between 27 and 56 years of age. Adipocytes were isolated by collagenase treatment of adipose tissue, followed by filtering and centrifugation. Floating adipocytes were lysed in Qiazol before RNA purification and microarray analysis.
Project description:B-cell chronic lymphocytic leukemia (CLL) is a common type of leukemia, characterized by the progressive accumulation of CD5+ “mature” monoclonal B lymphocytes in peripheral blood, bone marrow and lymphoid tissues. Although circulating CLL cells are non-dividing cells, prone to spontaneous apoptosis, their progressive accumulation is the result of a dynamic balance between cell death and proliferation and a high turn-over rate has been related to a poor prognosis. Indeed, CLL cells are protected from apoptosis and proliferate in specific niches within the lymphoid tissues and the bone marrow. CLL cells show variable expression of IL-21R that can be up-regulated by cell activation via CD40. CD40-activated CLL cells phosphorylate STAT-1 and STAT-3 and undergo apoptosis in response to IL-21 stimulation. By gene-expression profiling we found out that IL-21 modulates the expression of several genes including cytokine and chemokine genes and genes involved in cell survival and apoptosis in CD40-activated CLL cells. PBMCs were isolated from heparinized blood obtained from 13 untreated patients diagnosed with CLL on the basis of clinical and immunophenotypic criteria. PBMCs were isolated by Ficoll density gradient centrifugation and characterized by immunofluorescence and FACS analysis. When residual non B-cells exceeded 10%, B cells were enriched by negative selection with antibody-coated magnetic beads (CD2-beads, Dynal, Oslo, Norway) to obtain a >95% pure CD19+/CD5+ B cell population. B-CLL cells were pre-activated on adherent CD40L-transduced L cells for 48-36h and then stimulated with IL-21 or medium only for additional 18h. Total RNA was isolated from samples using TriZol (Invitrogen) reagent.
Project description:B-cell chronic lymphocytic leukemia (CLL) is a common type of leukemia, characterized by the progressive accumulation of CD5+ “mature” monoclonal B lymphocytes in peripheral blood, bone marrow and lymphoid tissues. Although circulating CLL cells are non-dividing cells, prone to spontaneous apoptosis, their progressive accumulation is the result of a dynamic balance between cell death and proliferation and a high turn-over rate has been related to a poor prognosis. Indeed, CLL cells are protected from apoptosis and proliferate in specific niches within the lymphoid tissues and the bone marrow. By gene-expression profiling we found out that IL-21 modulates the expression of several genes including cytokine and chemokine genes and genes involved in cell survival and apoptosis in CD40-activated CLL cells. To gain information of the possible mechanisms involved in the regulation of these genes we tested the possibility that IL-21 may act through specific miRNA regulation. PBMCs were isolated from heparinized blood obtained from 13 untreated patients diagnosed with CLL on the basis of clinical and immunophenotypic criteria. PBMCs were isolated by Ficoll density gradient centrifugation and characterized by immunofluorescence and FACS analysis. When residual non B-cells exceeded 10%, B cells were enriched by negative selection with antibody-coated magnetic beads (CD2-beads, Dynal, Oslo, Norway) to obtain a >95% pure CD19+/CD5+ B cell population. B-CLL cells were pre-activated on adherent CD40L-transduced L cells for 48-36h and then stimulated with IL-21 or medium only for additional 18h. Total RNA was isolated from samples using TriZol (Invitrogen) reagent. This Series represents 9 of the 13 cases.
Project description:siRNA-mediated knockdown of MIF expression in HEK293 cells resulted in inhibition of cell proliferation and cell cycle progress. The microarray study of MIF KD cells reveald knockdown of MIF would lead to extensive changes in gene expression profiles which may elucidate the molecular mechanism of MIF siRNA-mediated inhibition of cell cycle and cell proliferation. Keywords: The whole-genome expression analysis in MIF knockdown cells and the control cells HEK293 cells were transfected MIF siRNA (50pmol/ml & 100pmol/ml) or control RNA using LipofectamineTM 2000 reagent and re-incubated for 24 hours.At the designed time point, totol RNA was isolated from the different treated cells.The normal cells were used as control.
Project description:Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy that resists current treatments. To test epigenetic therapy against this cancer we used the DNA demethylating drug 5-aza-2’-deoxycytidine (DAC) in a KrasLSL-G12D; p53LSL-R270H/+; Pdx1-cre; Brca1flex2/flex2 (KPC-Brca1) mouse model of aggressive stroma-rich PDAC. In untreated tumors, we found globally decreased 5-methyl-cytosine (5mC) in malignant epithelial cells and in cancer-associated myofibroblasts (CAFs), and increased amounts of 5-hydroxymethyl-cytosine (5HmC) in CAFs, in progression from pancreatic intraepithelial neoplasia (PanIN) to PDAC. DAC further reduced DNA methylation and slowed PDAC progression, markedly extending survival in an early treatment protocol and significantly though transiently inhibiting tumor growth when initiated later, without adverse side effects. Escaping tumors contained areas of sarcomatoid transformation with disappearance of CAFs. Mixing-allografting experiments and proliferation indices showed that DAC efficacy was due to inhibition of both the malignant epithelial cells and the stromal CAFs. Expression profiling and immunohistochemistry highlighted DAC-induction of STAT1 in the tumors, and DAC plus gamma-interferon produced an additive anti-proliferative effect on PDAC cells. DAC induced strong expression of the testis antigen DAZL in CAFs. These data show that DAC is effective against PDAC in vivo and provide a rationale for future studies combining hypomethylating agents with cytokines and immunotherapy. Treatment of a short-term explant culture of malignant epithelial cells from a KPC-Brca1 mouse pancreatic carcinoma, with 0.5 micromolar 5-aza-dC (decitabine; DAC) for 48 hours. The experiment includes 3 replicate plates untreated and 3 replicates treated.
Project description:Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy that resists current treatments. To test epigenetic therapy against this cancer we used the DNA demethylating drug 5-aza-2’-deoxycytidine (DAC) in a KrasLSL-G12D; p53LSL-R270H/+; Pdx1-cre; Brca1flex2/flex2 (KPC-Brca1) mouse model of aggressive stroma-rich PDAC. In untreated tumors, we found globally decreased 5-methyl-cytosine (5mC) in malignant epithelial cells and in cancer-associated myofibroblasts (CAFs), and increased amounts of 5-hydroxymethyl-cytosine (5HmC) in CAFs, in progression from pancreatic intraepithelial neoplasia (PanIN) to PDAC. DAC further reduced DNA methylation and slowed PDAC progression, markedly extending survival in an early treatment protocol and significantly though transiently inhibiting tumor growth when initiated later, without adverse side effects. Escaping tumors contained areas of sarcomatoid transformation with disappearance of CAFs. Mixing-allografting experiments and proliferation indices showed that DAC efficacy was due to inhibition of both the malignant epithelial cells and the stromal CAFs. Expression profiling and immunohistochemistry highlighted DAC-induction of STAT1 in the tumors, and DAC plus gamma-interferon produced an additive anti-proliferative effect on PDAC cells. DAC induced strong expression of the testis antigen DAZL in CAFs. These data show that DAC is effective against PDAC in vivo and provide a rationale for future studies combining hypomethylating agents with cytokines and immunotherapy. Treatment of a short-term explant culture of cancer-associated fibroblasts (CAFs) from a KPC-Brca1 mouse pancreatic carcinoma, with 2 micromolar 5-aza-dC (decitabine; DAC) for 48 hours. The experiment includes 3 replicate plates untreated and 3 replicates treated.