Project description:We performed single-cell RNA-sequencing (scRNA-seq) to elucidate osteoblastic transcriptional programs during zebrafish caudal fin regeneration. We show that osteoprogenitors are enriched with components associated with epithelial-to-mesenchymal transition (EMT) and its reverse, mesenchymal-to-epithelial transition (MET). Trajectory analyses indicate osteoblastic cells solely expressed EMT components, or transiently expressed components for MET before EMT. We provide evidence that the EMT markers cdh11 and twist2 are co-expressed in dedifferentiating cells at the amputation stump, and in differentiating osteoblastic cells in the regenerate, the latter of which are enriched in EMT signatures. We also show that esrp1, a regulator of alternative splicing in epithelial cells that is associated with MET, is expressed in a subset of osteoprogenitors during outgrowth. This study provides a single cell resource for the study of osteoblastic cells during zebrafish fin regeneration, and supports the contribution of MET- and EMT-associated components to this process.
Project description:Zebrafish regenerate fin rays following amputation through epimorphic regeneration, a process that has been proposed to involve the epithelial-to-mesenchymal transition (EMT). We performed single-cell RNA sequencing (scRNA-seq) to elucidate osteoblastic transcriptional programs during zebrafish caudal fin regeneration. We show that osteoprogenitors are enriched with components associated with EMT and its reverse, mesenchymal-to-epithelial transition (MET), and provide evidence that the EMT markers cdh11 and twist2 are co-expressed in dedifferentiating cells at the amputation stump at 1 dpa, and in differentiating osteoblastic cells in the regenerate, the latter of which are enriched in EMT signatures. We also show that esrp1, a regulator of alternative splicing in epithelial cells that is associated with MET, is expressed in a subset of osteoprogenitors during outgrowth. This study provides a single cell resource for the study of osteoblastic cells during zebrafish fin regeneration, and supports the contribution of MET- and EMT-associated components to this process.
Project description:This study investigated the cell composition and lineage relationships of FACS (fluorescence activated cell sorting)-enriched epidermal, bone forming (osteoblast) and (non-osteoblast) blastemal fin regenerate cells by single cell (sc) RNA sequencing. The repetitive cell harvesting in four-week intervals revealed a constant gene regulation over four successive regeneration rounds in the early outgrowth stage. In addition, the sc RNA dataset uncovered a potential lineage relationship between distal blastema cells and osteoblasts during fin regeneration. These findings, together with the identification of novel fin regenerate markers, advance our understanding of complex tissue regeneration in zebrafish.
Project description:We studied MET-transformed human primary osteoblasts (MET-HOBs), which we previously turned into osteosarcoma cells by LV driven over-expression of MET oncogene. We obtained distinct MET transformed HOB clones derived from independent events of transgene integration. To characterise the phenotype of the MET-HOB clones we used oligonucleotide microarrays. Expression profiles of MET-HOBs and parental HOBs were compared.
Project description:We studied MET-transformed human primary osteoblasts (MET-HOBs), which we previously turned into osteosarcoma cells by LV driven over-expression of MET oncogene. We obtained distinct MET transformed HOB clones derived from independent events of transgene integration. To characterise the phenotype of the MET-HOB clones we used oligonucleotide microarrays. Expression profiles of MET-HOBs and parental HOBs were compared. To characterise the phenotype of the MET-HOB clones we used oligonucleotide microarrays
Project description:NMuMG is an epithelial cell line that can be induced into EMT by TGF-β treatment or MET by TGF-β withdrawl. During EMT, several marker genes were downregulated/upregulated, which is consistent with its mesenchymal phenotype. Transcription factors that are regulated during EMT and its reverse process MET are candidate genes for the regulations of the EMT marker genes.
Project description:NMuMG is an epithelial cell line that can be induced into EMT by TGF-β treatment or MET by TGF-β withdrawl. During EMT, several marker genes were downregulated/upregulated, which is consistent with its mesenchymal phenotype. Transcription factors that are regulated during EMT and its reverse process MET are candidate genes for the regulations of the EMT marker genes. NMuMG cells treated with vehicle, TGF-β for 11 days, or 11days of TGF-β treatment followed by TGF-β withdrawl for another 13 days. RNA from these 3 conditions of NMuMG were extracted and subject to microarray analysis
Project description:Epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) facilitate breast cancer (BC) metastasis, however stable molecular changes that result as a consequence of these processes remain poorly defined. Therefore, we sought to identify molecular markers that could distinguish tumor cells that had completed the EMT:MET cycle in the hopes of identifying and targeting unique aspects of metastatic tumor outgrowth.Therefore, normal murine mammary gland (NMumG) cells transformed by overexpression of EGFR (NME) cells were cultured in the presence of TGF-beta1 (5 ng/ml) for 4 weeks, at which point TGF-beta1 supplementation was discontinued and the cells were allowed to recover for an additional 4 weeks (Post-TGF-Rec). Total RNA was prepared from unstimulated cells (Pre-TGF) of similar passage and compared by microarray analysis. The two groups were analyzed in triplicate, three Pre-TGF samples and three Post-TGF-Rec samples.
Project description:Epithelial-mesenchymal transition (EMT) is a process by which epithelial cells lose cell-cell contact and gain cancer malignancy such as invasion, stemness, chemoresistance and metastasis. Reverse precess, mesenchymal-epithelial transition (MET) is also important for colonization. Extracellular vesicles (EVs) secreted from cancer cells are also important for cancer malignancy. To analyze RNAs from cells and EVs during EMT and MET, RNA sequencing was performed using E-cadherin-RFP/Py2T reporter system.
Project description:Epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) facilitate breast cancer (BC) metastasis, however stable molecular changes that result as a consequence of these processes remain poorly defined. Therefore, we sought to identify molecular markers that could distinguish tumor cells that had completed the EMT:MET cycle in the hopes of identifying and targeting unique aspects of metastatic tumor outgrowth.Therefore, normal murine mammary gland (NMumG) cells transformed by overexpression of EGFR (NME) cells were cultured in the presence of TGF-beta1 (5 ng/ml) for 4 weeks, at which point TGF-beta1 supplementation was discontinued and the cells were allowed to recover for an additional 4 weeks (Post-TGF-Rec). Total RNA was prepared from unstimulated cells (Pre-TGF) of similar passage and compared by microarray analysis.