Project description:In mammalian systems, extracellular small RNAs can operate in a paracrine manner to communicate information between cells, relying on transport within vesicles. “Foreign” small RNAs derived from bacteria, plants and parasites have also been detected in mammalian body fluids, sparking interest in whether these could mediate inter-species communication. However, there is no mechanistic framework for RNA-mediated interspecies communication and the active movement of RNA via vesicles has not been shown outside of mammals. Here we demonstrate that specific microRNAs and Y RNAs are packaged into vesicles secreted by a gastrointestinal nematode, Heligmosomoides polygyrus, which naturally infects mice. Total RNA was extracted from the serum of mice infected with Litomosoides sigmodontis at 60 days post infection
Project description:Increased expression of GLI1 is associated with poor prognosis for some breast cancer subtypes. A conditional transgenic GLI1 expressing mouse model, with or without heterozygous deletion of Trp53, was used to generate and study GLI1 induced mammary gland tumours. Tumour tissue was serially orthotopically transplanted for at least 10 generations in NSG mice.
Project description:Introduction: Pleural mesothelioma (PM) is known as one of the most aggressive malignancies among all cancers, despite of usually absent tumor-driver mutations in oncogenes. It develops in a unique inflammatory tumor microenvironment (TME), which has been postulated as major contributor of PM’s highly aggressive nature. Mesothelioma-associated fibroblasts (Meso-CAFs), a main component of the TME have recently been shown to substantially stimulate several aspects of PM aggressiveness and promote the malignant transformation of pleural mesothelial cells. However, respective cell models for TME research in PM are still very limited. The most commonly used pleural mesothelial cell line Met5A has been established decades ago and patient-derived Meso-CAFs have just recently been characterized for the first time. The aim of the current study was to generate and characterize pleural mesothelial and Meso-CAF cell models with an extended life span that closely resemble the primary cells isolated from human tissue. Methods: Pleural mesothelial cells and Meso-CAFs were isolated from human tissue of pneumothorax and PM patients, respectively. Retroviral transduction was used to induce stable expression of human telomerase reverse transcriptase (hTERT) and enhanced green fluorescent protein (EGFP) in the primary cells. The established cell models were evaluated by measuring their doubling times, gene expression and protein activity levels of hTERT, as well as the absolute lengths of telomeres. The transduced cells were compared to their primary counterparts on the protein level using proteome analysis and the impact of their conditioned media (CM) on tumor cell growth was investigated by videomicroscopy. Results: All transduced derivatives exhibit elevated hTERT gene expression and protein activity, increased hTERT protein amounts in the nucleus, and moderately higher absolute telomere lengths compared to their parental primary cells. The transduction with hTERT did not elicit marked changes in the morphology of the cells, as well as in their proteomes and secretomes. The CM of primary and hTERT-transduced Meso-CAFs comparably stimulated PM cell growth, while medium conditioned by normal pleural mesothelial cells including their hTERT-transduced derivatives was not able to induce a growth stimulating effect. Conclusion: The hTERT-transduced cells closely resemble their primary counterparts, while retaining telomerase activity, thus preventing replicative senescence. The new cell models provide valuable tools for the investigation of cellular interactions cellular interactions within the TME of PM and thus may help to identify novel biomarkers for early diagnosis and to develop new therapeutic strategies.
Project description:PCNA is an essential protein in all cells with multiple well-known function in DNA replication and DNA repair. Recently, PCNA was shown to be involved in regulation of cellular signalling and central carbon metabolism via stabilizing the metabolic enzymes ENO1 and 6PGD. Furthermore, ATX-101, a PCNA targeting peptide currently in Phase II, was shown to reduce metabolite pools in the glycolysis and the pentose phosphate pathway (PPP) in multiple haematological cells. Here integrated analysis of signalome and metabolome data from multiple myeloma (MM) cells reveals large effects of ATX-101 on signalling, metabolism and red-ox regulation. The sensitivity to ATX-101 varied between the eight MM cell lines studied and a metabolic shift was detected in the sensitive cell lines. These cell lines were characterized by low endogenous levels of the redox metabolites NAD+, NADH and GSH. Interestingly, 11 proteins were pulled down from untreated cells from the sensitive cell lines while not from 9 other cancer cell lines or primary monocytes from three donors. These proteins could be potential biomarkers for MM cells with low red-ox capacity and hypersensitivity for inhibitors of glycolysis or the PPP.
Project description:PCNA is an essential protein in all cells with multiple well-known function in DNA replication and DNA repair. Recently, PCNA was shown to be involved in regulation of cellular signalling and central carbon metabolism via stabilizing the metabolic enzymes ENO1 and 6PGD. Furthermore, ATX-101, a PCNA targeting peptide currently in Phase II, was shown to reduce metabolite pools in the glycolysis and the pentose phosphate pathway (PPP) in multiple haematological cells. Here integrated analysis of signalome and metabolome data from multiple myeloma (MM) cells reveals large effects of ATX-101 on signalling, metabolism and red-ox regulation. The sensitivity to ATX-101 varied between the eight MM cell lines studied and a metabolic shift was detected in the sensitive cell lines. These cell lines were characterized by low endogenous levels of the redox metabolites NAD+, NADH and GSH. Interestingly, 11 proteins were pulled down from untreated cells from the sensitive cell lines while not from 9 other cancer cell lines or primary monocytes from three donors. These proteins could be potential biomarkers for MM cells with low red-ox capacity and hypersensitivity for inhibitors of glycolysis or the PPP.
Project description:RNA-seq data characterising small cell lung cancer within circulating tumour cell (CTC) derived explants, generated within immune compromised mice. Both chemo naive and progression models were taken from patients and their drug sensitivity tested within the mouse models. Blood samples were taken from patients, and RosetteSep CTC Enrichment was used to implant these cells into mice. Tumours were allowed to grow, then extracted and re-implanted into second and third generations of mice, before finally being subjected to molecular profiling.
Project description:Malignant mesothelioma (MM) is an asbestos-related malignancy. Discrimination between MM and reactive mesothelial hyperplasia (RM) is often difficult. MM cells have a broad histological spectrum, and consist mainly of epithelioid, sarcomatoid, and biphasic cell types. The prognosis of MM is generally poor, but better prognosis has been reported with the epithelioid type of MM than the non-epithelioid type. We applied a genome-wide analysis to the identification of new markers that may aid in differentiating the epithelioid type of MM from other histological types and from RM cells. Array-based comparative genomic hybridization analysis was performed on malignant mesothelioma (MM) primary cell cultures, reactive mesothelial hyperplasia (RM) primary cell cultures; early passage of in vitro primary cell cultures to minimize acquisition of additional genomic changes. If available, matched peripheral blood was applied to analysis.
Project description:The aim of the experiment was to analyse gene expression profiles in Brca1 tumours arising from different mammary epithelial cell populations use a Cre-loxP based conditional knockout system. K14 promoter driving Cre expression caused Brca1 knockout in basal stem cells and thus stem cell origin tumours whereas Blg promoter driving Cre expression caused Brca1 knockout in luminal progenitor cells and thus progenitor origin tumours. Individual arrays were carried out on labelled cDNA made from RNA isolated from mouse mammary tumours. Only cDNA passing Almac diagnostics QC criteria were hybridised to arrays. Only arrays passing QC criteria after hybrisiation were subsequently analysed.