Project description:It has been already reported that there are undifferentiated/proliferating neuroblasts in the postnatal sympathetic ganglia in TH-MYCN mice, a neuroblastoma model. We established suitable spheroid culture condition that selectively isolates undifferentiated neuroblasts from superior mesenteric ganligon (SMG) of TH-MYCN mice. In order to investigate the chromosomal alterations (gains or losses) of spheres derived from TH-MYCN mice, we carried out array comparative genomic hybridization. We investigated whether chromosomal alterations occured during early neuroblastoma tumorigenesis in TH-MYCN mice.
Project description:It has been already reported that there are undifferentiated/proliferating neuroblasts in the postnatal sympathetic ganglia in TH-MYCN mice. We established suitable spheroid culture condition that selectively isolates undifferentiated neuroblasts from superior mesenteric ganligon (SMG) of TH-MYCN mice. In the present study, in order to investigate the transcriptomic differences among 3-week-old WT SMG, TH-MYCN (hemizygote) SMG, and TH-MYCN (hemizygote) spheres, we carried out microarray gene expression analysis. We investigated whether or not undifferentiated cells observed in TH-MYCN SMG were selectively isolated and maintaned as spheres.
Project description:It has been already reported that there are undifferentiated/proliferating neuroblasts in the postnatal sympathetic ganglia in TH-MYCN mice. We established suitable spheroid culture condition that selectively isolates undifferentiated neuroblasts from superior mesenteric ganligon (SMG) of TH-MYCN mice. In the present study, in order to investigate the transcriptomic differences between embryonic day 13.5 (E13.5) WT derived primary spheres and E13.5 TH-MYCN derived passageable spheres, we carried out microarray gene expression analysis. We investigated critical molecular events in MYCN-transformed neuroblastoma cells in TH-MYCN mice.
Project description:It has been already reported that there are undifferentiated/proliferating neuroblasts in the postnatal sympathetic ganglia in TH-MYCN mice. Given the fact that neuroblastoma is derived from sympathoadrenal progenitors which are originated from neural crest stem cells, we attempted to use the well-established culture condition that has been utilized to maintain derivatives of neural crest stem cells. However, the medium contained retinoic acid (RA) which is known to induce neuroblastoma differentiation and is used in clinical treatment against high-risk patients. In the present study, the medium with RA (RA (+) medium) and without RA (RA (-) medium) were compared. Since undifferentiated neuroblasts are observable at one of the sympathetic ganglia, superior mesenteric ganglion (SMG), in 3-week-old TH-MYCN hemizygote (TH-MYCN+/-) mice with 129+Ter/SvJcl mice background, we dissected and dissociated the SMG to prepare single cells. These cells were cultured in either RA (+) or RA (-) media. Spheres formed in either RA (+) or RA (-) medium were subjected to microarray analysis.
Project description:Neuroblastoma is a pediatric tumor that accounts for more than 15% of cancer-related deaths in children. Survival chances for high-risk patients are less than 50%. Retinoic acid treatment is part of the maintenance therapy given to neuroblastoma patients; however, not all tumors respond to retinoic acid-mediated differentiation. Among neuroblastoma tumors, two phenotypically distinct cell types-adrenergic (ADRN) and mesenchymal (MES), have been identified based on their super-enhancer landscape and transcriptional core regulatory circuitries. We hypothesized that distinct super-enhancers in these different tumor cells could mediate differential response to retinoic acid. To this end, we treated four different neuroblastoma cell lines, comprising both ADRN (MYCN amplified and non-amplified) and MES subtypes, with retinoic acid and studied the super-enhancer landscape upon treatment and after removal of retinoic acid. Using H3K27ac ChIP-seq paired with RNA-seq, we compared the super-enhancers in cells that respond to retinoic acid-mediated differentiation versus those that fail to differentiate. We identified unique super-enhancers associated with cells differentiation; however, even among cells that respond to treatment, there was heterogeneity upon removal of retinoic acid, with MYCN amplified cells remaining differentiated whereas MYCN non-amplified cells reverted to a proliferative state. This study identifies regulatory super-enhancers as a plausible mechanism behind the differential response to retinoic acid-mediated differentiation.
Project description:Neuroblastoma is a pediatric tumor that accounts for more than 15% of cancer-related deaths in children. Survival chances for high-risk patients are less than 50%. Retinoic acid treatment is part of the maintenance therapy given to neuroblastoma patients; however, not all tumors respond to retinoic acid-mediated differentiation. Among neuroblastoma tumors, two phenotypically distinct cell types-adrenergic (ADRN) and mesenchymal (MES), have been identified based on their super-enhancer landscape and transcriptional core regulatory circuitries. We hypothesized that distinct super-enhancers in these different tumor cells could mediate differential response to retinoic acid. To this end, we treated four different neuroblastoma cell lines, comprising both ADRN (MYCN amplified and non-amplified) and MES subtypes, with retinoic acid and studied the super-enhancer landscape upon treatment and after removal of retinoic acid. Using H3K27ac ChIP-seq paired with RNA-seq, we compared the super-enhancers in cells that respond to retinoic acid-mediated differentiation versus those that fail to differentiate. We identified unique super-enhancers associated with cells differentiation; however, even among cells that respond to treatment, there was heterogeneity upon removal of retinoic acid, with MYCN amplified cells remaining differentiated whereas MYCN non-amplified cells reverted to a proliferative state. This study identifies regulatory super-enhancers as a plausible mechanism behind the differential response to retinoic acid-mediated differentiation.
Project description:Preclinical cancer drug discovery efforts have employed two-dimensional (2D)-cell-based assay models, which fail to forecast in vivo efficacy and contribute to a lower success rates of clinical approval. Three-dimensional (3D) cell culture models are recently expected to bridge the gap between 2D and in vivo models. We have developed a novel 3D culture method that is suitable for imaging analysis and improves the growth of spheroid-forming cancer cells under anchorage-independent condition by leveraging a feature of LA717, a seaweed-derived polysaccharide. Gene microarrays were used to observe the global gene expression in A549 cells cultured with adhesion condition (2D, control) or with low adhesion condition (LA717) and identified distinct classes of up or down-regulated genes.
Project description:Anchorage-independent spheroid cells of HCC possess stemness properties. Huh7 cells are cultured in ultra-low attachment surface plates with serum-free medium. RNA-sequencing (RNA-seq) was applied in spheroid culture cells and adherent culture cells.
Project description:Patient-derived cancer cells (PDCs) were established by three-dimensional (3D) spheroid culture from testicular germ cell tumor (GCT) specimens. Microarray expression analysis revealed that cancer stem-like cell-related genes were upregulated in 3D culture condition compared with two-dimensional (2D) culture condition.