Abstract
Gene amplifications are an attribute of tumor cells and have for long time been overlooked in normal cells. A growing number of investigations describe gene amplifications in normal mammalian cells during development and differentiation. Possibly, tumor cells have rescued the gene amplification mechanism as a physiological attribute of stem cells. Here, we investigated human mesenchymal stem cells (hMSCs) for gene amplification using array-CGH, single cell fluorescence in situ hybridization and qPCR. Gene amplifications were detected in mesenchymal stem cells and in mesenchymal stem cells during differentiation towards adipocytes and osteoblasts. Undifferentiated hMSCs harbor 12 amplified chromosomal regions, hMSCs that differentiated towards adipocytes 18 amplified chromosome regions, and hMSCs that differentiate towards osteoblasts 19 amplified regions. Specifically, hMSCs that differentiated towards adipocytes or osteoblasts harbor CDK4 and MDM2 amplifications both of which frequently occur in osteosarcoma and liposarcoma that are both of same cell origin. Beside the amplifications, we identified 36 under-replicated regions in undifferentiated and in differentiating hMSC cells.
Highlights
Gene amplifications can be found in tumor cells, drug resistant cells and in cells essential for developmental processes in amphibians and flies [1]
HMSCs that differentiated towards adipocytes or osteoblasts harbor CDK4 and MDM2 amplifications both of which frequently occur in osteosarcoma and liposarcoma that are both of same cell origin
As previous results demonstrated an overlap of gene amplifications between differentiating cells and tumors derived from the same lineage, we investigated genes CDK4 and MDM2 for amplification because CDK4 and MDM2 amplifications were frequently reported in tumor cells e.g. in liposarcoma and in osteosarcoma [8, 9]
Summary
Gene amplifications can be found in tumor cells, drug resistant cells and in cells essential for developmental processes in amphibians and flies [1]. In addition ERBB2 gene amplification was reported in human trophoblast cells during differentiation [6] and amplification of placental genes in mouse giant trophoblast cells during differentiation [7]. These results leave the question to what extend gene amplifications during differentiation were a common attribute and if one can find gene amplifications in different progenitor cells including adipogenic progenitor cells or osteogenic progenitor cells during their differentiation towards adipocytes and osteoblasts. We set out to analyze gene amplification in human mesenchymal stem cells and in human mesenchymal stem cell differentiation induced towards adipocytes and osteoblasts. Amplification analysis was performed using three techniques: using array CGH we determined amplified chromosomal regions, using fluorescence in situ hybridization (FISH) we confirmed gene amplifications on single cell level and using qPCR we verified the results with an independent method
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