Project description:Cellular directed migration is critical to invasion-metastasis cascade of cancer cells. We used in vitro transwell model to screen two esophageal cancer cell lines (KYSE30/180) and obtained two pairs of subpopulations with distinc motiliyt ability. Then we used microarrays to detail the differentially expressed genes and microRNAs between these two cell sublines (30U/D and 180U/D) to identify those responsible for ESCC motility.
Project description:Cellular directed migration is critical to invasion-metastasis cascade of cancer cells. We used in vitro transwell model to screen two esophageal cancer cell lines (KYSE30/180) and obtained two pairs of subpopulations with distinct motility ability. Then we used microarrays to detail the differentially expressed genes and microRNAs between these two cell sublines (30U/D and 180U/D) to identify those responsible for ESCC motility.
Project description:Cellular directed migration is critical to invasion-metastasis cascade of cancer cells. We used in vitro transwell model to screen two esophageal cancer cell lines (KYSE30/180) and obtained two pairs of subpopulations with distinc motiliyt ability. Then we used microarrays to detail the differentially expressed genes and microRNAs between these two cell sublines (30U/D and 180U/D) to identify those responsible for ESCC motility. KYSE30/180 cells were subject to four successive selections using transwell (CORNING, USA). Subpopulations penetrated through membrane (D) or not (U) were harvested respectively for RNA extraction and hybridization on Affymetrix genome and LC Sciences microRNA microarrays
Project description:Cellular directed migration is critical to invasion-metastasis cascade of cancer cells. We used in vitro transwell model to screen two esophageal cancer cell lines (KYSE30/180) and obtained two pairs of subpopulations with distinct motility ability. Then we used microarrays to detail the differentially expressed genes and microRNAs between these two cell sublines (30U/D and 180U/D) to identify those responsible for ESCC motility. KYSE30/180 cells were subject to four successive selections using transwell (CORNING, USA). Subpopulations penetrated through membrane (D) or not (U) were harvested respectively for RNA extraction and hybridization on Affymetrix genome and LC Sciences microRNA microarrays simultaneously.
Project description:In vivo and in vitro analyses revealed distinct and shared characteristics of the metastatic D2A1-m1 and D2A1-m2 sublines. In particular, D2A1-m1 cells are more aggressive in experimental metastasis assays, while D2A1-m2 cells are more efficient at disseminating from the primary tumour in spontaneous metastasis assays. Surprisingly, classical metastasis-associated in vitro phenotypes such as enhanced proliferation, migration and invasion are reduced in the sublines compared to the parental cell line. Further, evasion of immune control cannot fully explain their enhanced metastatic properties. By contrast, both sublines show increased resistance to apoptosis when cultured in non-adherent conditions and, for the D2A1-m2 subline, increased 3D tumour spheroid growth. Moreover, the enhanced spontaneous metastatic phenotype of the D2A1-m2 sublines is associated with the ability to recruit an activated tumour stroma and promote directional migration of fibroblasts. Gene expression profiling revealed that the two metastatic sublines are distinct but more closely related to each other than the parental D2A1 cells.
Project description:Human embryonic stem (ES) cell lines acquire recurrent karyotypic abnormalities that may promote growth and survival in long-term culture. Cells with these abnormalities are often present at low levels in human ES cell cultures, requiring subcloning for isolation and analyses. We have developed a rapid method to isolate either abnormal or normal clones from karyotypically mosaic human ES cell cultures that we call Genetic Diagnosis at Passage (G-DAP). We have used this technique to isolate three distinct abnormal sublines from H9 human ES cells in continuous culture. These sublines were characterized by G-banding, fluorescence in situ hybridization (FISH), spectral karyotyping (SKY), and array comparative genomic hybridization (aCGH). One subline (H9.DT) has, as it’s only abnormality, a duplication of 12p capped by a 5q subtelomere. A related subline (H9.TSF) has an additional copy of the derivative chromosome 12, partial trisomy of 17q, and trisomy 14. The third subline (H9.DE) has an interstitial deletion of 18q as the sole anomaly. Despite these karyotypic abnormalities, all three sublines retain markers of the undifferentiated state. The subline H9.TSF demonstrated the ability to form embryoid bodies consisting of all three germ layers. Significantly, H9.DT and H9.TSF sublines show a classic karyotype progression that mirrors that seen in both hematopoietic and solid tumors. This progression was accompanied by an impressive increase in growth potential. The isolation of human ES cell sublines with clonal cytogenetic abnormalities provides a valuable tool for study of factors that promote in vitro genetic changes and for analysis of mechanisms of aneuploidy related to genetic progression found in human cancers.