Objective: To establish of a bone cancer stem cell (CSC) line from a rare human bone disease, the high grade osteoblastic osteosarcoma (OS). Material and Methods: Central high grade osteoblastic OS samples were collected at the “Unit Ortopedia Oncologica e Ricostruttiva”, AOUC Careggi, Florence, with informed consent approved by the local Ethical Committee. First, primary human cancer cell cultures of osteoblastic OS have been established. After that, the subpopulations of CSCs have been isolated from these, with the sphere-formation assay. Hence, the cancer stem cell phenotype has been evaluated by several cellular assays/stainings, by flow cytometry analysis and by analyzing the pro-files expressions of a few genes related to CSCs. Results: We have established primary cell cultures of high grade osteoblastic OS from each samples collected. Consequently, from these we have started to isolate CSCs. Nowadays we have established a OS-CSC line, called as OSA5-CSC. The cancer stem cell phenotype from OSA5-CSC line was confirmed by observing the capacity of the OSA5-CSCs to differentiate into osteoblasts and into adipocytes, by showing the positive presence of the MSCs and ofthe ESCs markers into the cell line, and by evaluating a good rate (14 %) as clonogenic capacity. The presence of the ESC markers was confirmed also by their gene expression together with the expression of CD133. On the other hand, the malignant phenotype was verify by demonstrating the presence of the expression of genes involved in invasion/migration process (i.e., AXL and EZR) and in the pluripotency of CSC (i.e., MYC), by the agar soft and bythe ALDH1A1 assays. Conclusions: In conclusion, we have collected a few biopsies of this rare bone disease. We have established primary cell cultures from these. We have completely characterized and studied a OS-CSC line at cellular and molecular level. Nowadays we are concluding the characterization of the OS-CSCs isolated from the relative cell cultures and we are starting the study of micro-RNAs related to their phenotype.
ESTABLISHMENT AND CHARACTERIZATION OF A CANCER STEM CELL LINE FROM A RARE BONE SARCOMA
FRANCHI, ALESSANDRO;Capanna, R;
2016-01-01
Abstract
Objective: To establish of a bone cancer stem cell (CSC) line from a rare human bone disease, the high grade osteoblastic osteosarcoma (OS). Material and Methods: Central high grade osteoblastic OS samples were collected at the “Unit Ortopedia Oncologica e Ricostruttiva”, AOUC Careggi, Florence, with informed consent approved by the local Ethical Committee. First, primary human cancer cell cultures of osteoblastic OS have been established. After that, the subpopulations of CSCs have been isolated from these, with the sphere-formation assay. Hence, the cancer stem cell phenotype has been evaluated by several cellular assays/stainings, by flow cytometry analysis and by analyzing the pro-files expressions of a few genes related to CSCs. Results: We have established primary cell cultures of high grade osteoblastic OS from each samples collected. Consequently, from these we have started to isolate CSCs. Nowadays we have established a OS-CSC line, called as OSA5-CSC. The cancer stem cell phenotype from OSA5-CSC line was confirmed by observing the capacity of the OSA5-CSCs to differentiate into osteoblasts and into adipocytes, by showing the positive presence of the MSCs and ofthe ESCs markers into the cell line, and by evaluating a good rate (14 %) as clonogenic capacity. The presence of the ESC markers was confirmed also by their gene expression together with the expression of CD133. On the other hand, the malignant phenotype was verify by demonstrating the presence of the expression of genes involved in invasion/migration process (i.e., AXL and EZR) and in the pluripotency of CSC (i.e., MYC), by the agar soft and bythe ALDH1A1 assays. Conclusions: In conclusion, we have collected a few biopsies of this rare bone disease. We have established primary cell cultures from these. We have completely characterized and studied a OS-CSC line at cellular and molecular level. Nowadays we are concluding the characterization of the OS-CSCs isolated from the relative cell cultures and we are starting the study of micro-RNAs related to their phenotype.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
