Isolation, Culture and Characterization of Cancer Stem Cells from Primary Osteosarcoma release_kshqh2l5efgkvjqyneuhuocm2i

by Achmad Fauzi Kamal, Deded Yudha Pranatha, Waluyo Sugito, Faisal Rahman, Eka Susanto, Silmi Mariya, Wei Ming Chen

Published in The Open Stem Cell Journal by Bentham Science Publishers Ltd..

2018   p1-13

Abstract

<jats:sec> <jats:title>Background:</jats:title> Osteosarcoma cancer stem cells (CSCs) are defined as a subpopulation of osteosarcoma cells, which have the ability of self-renewal, proliferation and differentiation. This study aimed to identify CSCs from human osteosarcoma<jats:italic> in vitro </jats:italic>. </jats:sec> <jats:sec> <jats:title>Methods:</jats:title> Osteosarcoma CSCs were isolated and cultured with sphere-forming assay technique on an ultra-low well attachment surface plate. After sarcosphere colonies were formed, we conducted reverse transcriptase-polymerase chain reaction (RT-PCR) to detect the expression of genes of embryonic stem cells such as <jats:italic>NANOG</jats:italic>, Oct3/4, STAT3 and gene of MSC CD133. Immunofluorescence analysis (IFA) of alkaline phosphatase (ALP), osteocalcin, and CD 133 was also performed to see the expression of osteosarcoma CSC surface protein with immuno-enzymatic staining principle. We also performed alizarin red staining to evaluate calcification in osteosarcoma CSCs. </jats:sec> <jats:sec> <jats:title>Results:</jats:title> The culture sphere-of the osteosarcoma cells showed three dimension round shaped colonies (sarcospheres) in slightly hypoxicand serum free condition which was not attached to the substrate with tight density. RT-PCR demonstrated that sarcospheres expressed genes which encode <jats:italic>NANOG</jats:italic>, Oct3/4 STAT 3, but not for CD 133. IFA showed positive protein expression of ALP, osteocalcin and CD 133 which was moderate, strong, and weak positive respectively. Sarcospheres also had a positive reaction toward alizarin red staining. </jats:sec> <jats:sec> <jats:title>Conclusion:</jats:title> Osteosarcoma CSCs could be isolated from human osteosarcoma by sphere-forming assay technique and characterized by the expression of genes of embryonic stem cells,such as <jats:italic>NANOG</jats:italic>, Oct3/4, STAT3 and IFA of ALP, osteocalcin, and CD 133. </jats:sec>
In application/xml+jats format

Archived Files and Locations

application/pdf  1.1 MB
file_yglnaslel5b4dfenfcik3uqvda
openstemcelljournal.com (web)
web.archive.org (webarchive)
Read Archived PDF
Preserved and Accessible
Type  article-journal
Stage   published
Date   2018-09-28
Language   en ?
Container Metadata
Not in DOAJ
In Keepers Registry
ISSN-L:  1876-8938
Work Entity
access all versions, variants, and formats of this works (eg, pre-prints)
Catalog Record
Revision: 81da10e1-aa03-4fa5-861e-76780871f08c
API URL: JSON