Project description:Mst1 and Mst2 were conditionally deleted from non-ciliated bronchiolar epithelial cells in the mature lung. Bronchiolar epithelial cells from control and Mst1/2 deleted mice were isolated by cell sorting and used for RNA-seq analysis. Scgb1a1-rtTA/tetO-Cre/Mst1;2-flx/flx (Mst1/2 D/D) mice were generated to conditionally delete Mst1 and Mst2 from non-ciliated, secretory bronchiolar epithelial cells. Adult mice were maintained on doxycycline food for 16 days to induce deletion of Mst1/2. Lin-/CD326+/CD24-intermediate cells were isolated by fluorescence cell sorting to enrich for the targeted airway epithelial cells. mRNA isolated from Lin-/CD326+/CD24-intermediate cells from control and Mst1/2 D/D mice was pooled and analyzed by RNA-seq to identify transcriptional changes following deletion of Mst1 and Mst2 from mature lung bronchiolar epithelial cells.
Project description:IL-17 receptor plays a major role in the antibacterial defense and regulation of host-microbiota interaction. However its impact on Paneth cells, cell-type specific for production of antibacterial peptides, is still not clear. Here, we used genetic depletion of IL-17 receptor specific for Paneth cell population (Defa6-iCre x Il17ra-flox) and performed the RNA sequencing on FACS-sorted Paneth cells and also complete ileum tissue, comparing Cre+ and Cre- littermates.
Project description:Characterisation and identification of putative human extra-thymic AIRE-expressing cells from paediatric tonsillitis samples using single -cell transcriptome profiling.
Project description:Primary human bronchial epithelial cells were transduced with control or hYAP(S127A) lentivirus in sphere forming conditions. Bronchospheres were harvested on day 18-20 for RNAseq analysis Passage 1 Primary HBECs from 2 independent donors were transduced with control or hYAP lentivirus. 48 hours post infection, cells were plated on transwell inserts in a 50-50 mixture of ALI medium-Cultrex BME reduced growth factor (RGF) to form spheres. Well differentiated bronchospheres were harvested for RNA-seq analysis on day 18-20 by combining 3 wells of each group for each donor.
Project description:Procr, also known as EPCR, is a protein C receptor known to function in the platelet coagulation cascade. We have used RNA-seq to examine RNA differential expression between Procr+ and Procr- myoepithelium in mouse mammary gland. Examine RNA differential expression among subsets of freshly sorted mouse mammary gland myoepithelium.
Project description:Breast cancers contain a minority population of cancer cells characterized by CD44 expression but low or undetectable levels of CD24 (CD44+CD24-/low) that have higher tumorigenic capacity than other subtypes of cancer cells. METHODS: We compared the gene-expression profile of CD44+CD24-/low tumorigenic breast-cancer cells with that of normal breast epithelium. Differentially expressed genes were used to generate a 186-gene invasiveness gene signature (IGS), which was evaluated for its association with overall survival and metastasis-free survival in patients with breast cancer or other types of cancer. RESULTS: There was a significant association between the IGS and both overall and metastasis-free survival (P<0.001, for both) in patients with breast cancer, which was independent of established clinical and pathological variables. When combined with the prognostic criteria of the National Institutes of Health, the IGS was used to stratify patients with high-risk early breast cancer into prognostic categories (good or poor); among patients with a good prognosis, the 10-year rate of metastasis-free survival was 81%, and among those with a poor prognosis, it was 57%. The IGS was also associated with the prognosis in medulloblastoma (P=0.004), lung cancer (P=0.03), and prostate cancer (P=0.01). The prognostic power of the IGS was increased when combined with the wound-response (WR) signature. CONCLUSIONS: The IGS is strongly associated with metastasis-free survival and overall survival for four different types of tumors. This genetic signature of tumorigenic breast-cancer cells was even more strongly associated with clinical outcomes when combined with the WR signature in breast cancer. Expression profling was performed on 6 tumorigenic, 3 non tumorigenic samples of breast tumors and 3 normal breast samples on two different platforms GPL96 and GPL97. A gene signature was derived by comparing the gene expressions of 6 tumorigenic samples with 3 normal breast samples.
Project description:Genomic instability is a prominent driver of tumorigenesis. However, a significant fraction of human cancers display few genomic aberrations, suggesting alternative roads towards malignancy. Here, we show that the differentiation status of normal human mammary epithelial cells influences the early response to an oncogenic activation and determines the genetic routes towards tumorigenesis. Following an oncogenic insult, luminal progenitors and differentiated luminal cells undergo oxidative and DNA replication stress, initiating genomic instability. In contrast, mammary stem cells exhibit the innate capacity to withstand aberrant mitogenic activation, fostering malignant transformation. This property relies upon a pre-emptive program driven by the ZEB1 transcription factor and the methionine sulfoxide reductase MSRB3. The ZEB1-MSRB3 axis governs cellular pliancy and prevents the continuous formation of oncogene-induced DNA damage, leading to neoplasms with unique pathological features. These gene expression data correspond to the different subpopulations that compose the hierarchy of normal human mammary epithelial cells. These subpopulations were freshly isolated from mammary tissue, originating from reduction mammoplasties and flow sorted using four markers (EpCAM, CD10, CD49f, ALDH). Three subpopulations enriched in mammary stem cells (MaSCs) are designed as MaSC1, 2, 3; the luminal progenitor are designed as LP, and the mature luminal cells as mL1 and mL2.
Project description:Analysis of stage-specific gene expression in Zbtb46GFP/+ pre-CD8 DCs, pre-CD4 DCs, CD24 cDCs and CD172a cDCs Bone Marrow and Splenocytes were harvested from 8-10 littermate Zbtb46GFP/+ mice and sorted to >95% purity on the FACS AriaFusion.
Project description:The goal of this study was to compare the transcriptional changes evoked by Csnk2b-deficiency in Foxp3+ regulatory T cells Messenger RNA profiles of CD4+Foxp3+ regulatory T cells isolated from 8 week old wild type (WT) and Csnk2b-deficient mice were generated by deep sequencing in duplicate, using Illumina MiSeq
Project description:Subset-specific and progenitor gene expression analysis of Klf4+/+ and Klf4-/- DCs. The two major lineages of classical dendritic cells (cDCs) express and require either IRF8 or IRF4 transcription factors for their development and function. IRF8-dependent cDCs promote anti-viral and T-helper 1 (Th1) cell responses, whereas IRF4-expressing cDCs have been implicated in controlling both Th2 and Th17 cell responses. Here, we have provided evidence that Kruppel-like factor 4 (Klf4) is required in IRF4-expressing cDCs to promote Th2 but not Th17 cell responses in vivo. Conditional Klf4 deletion within cDCs impaired Th2 cell responses during Schistosoma mansoni infection, Schistosoma egg antigen (SEA) immunization, and house dust mite challenge (HDM), without affecting cytotoxic T lymphocyte (CTL), Th1 and Th17 cell responses to herpes simplex virus, Toxoplasma gondii and Citrobacter rodentium infections. Further, Klf4 deletion reduced IRF4 expression in pre-cDCs and resulted in selective loss of IRF4-expressing cDCs subsets in several tissues. These results indicate that Klf4 guides a transcriptional program promoting IRF4-expressing cDCs heterogeneity. Bone marrow progenitors and skin draining LN subsets were harvested from 4 Klf4fl/fl cre negative or Vav1-icre mice and were sorted to >95% purity on the FACS Aria 3.