Isolation of cancer stem-like cells from hepatocellular carcinoma cell line HepG2 by methods of magnetic-activated cell sorting, spheroid culture, and anti-tumor drug-resistant selection: A primary evaluation

Abstract

Introduction:Recently reported data have suggested that only a small subset of cancer cells possess the capability to initiate malignancies. These observations were based on investigation of cells within the primary tumors displaying a distinct surface marker pattern. CD133 marker is a putative hematopoietic and neuronal stem cell marker, which is also considered to be a tumorigenic marker in brain, prostate and liver. Recent studies have shown that a small population of CD133-positive cells, indeed, exists in human hepatocellular carcinoma (HCC) cell lines and primary HCC tissues. This study was aimed at isolating the cancer stem-like cells from hepatocellular carcinoma cell line HepG2 using three different methods: magnetic-activated cell sorting (MACS), spheroid culture (SC), and anti-tumor drug (ATD) resistant selection. Methods: HepG2 hepatocellular carcinoma cells were expanded to yield enough cells that could be used to isolate cancer stem-like cells by these three methods. For MACS, cancer stem-like cells were sorted using anti-CD133 monoclonal antibody. For the second method, cancer stem-like cells were enriched by selection of anti-tumor drug resistance property. Lastly, for the third method, three-dimensional (3D) culture was used to enrich for the cancer stem-like cells. The cells obtained by the three methods were expanded to obtain an adequate number of cells for confirmation of CD133 expression. Results: The expression of CD133 + cells in the three methods was found to be different. In the MACS method, the expanded CD133 + sorted cells cultured through 2 passages only contained 0.40 % CD133 + cells. In the 3D spheroid cell culture, of the population of cells there were 38.39 % that were CD133 + cells. Lastly, in the anti-tumor drug (doxorubicin at 150 nM) resistant selection, 66.22 % were CD133 + cells. Conclusion: This study shows that isolation of HepG2 derived CD133 + population by culture with doxorubicin (150 nM) yields the highest efficiency and purity of the 3 methods studied. P. Isolation of cancer stem-like cells from hepatocellular carcinoma cell lineHepG2 by methods of magneticactivated cell sorting, spheroid culture, and anti-tumor drug-resistant selection: A primary evaluation. Progress. Stem. Cell.; 7(1):279-289. 279 History •

Inhibition of Histone Deacetylase Activity Increases Cisplatin Efficacy to Eliminate Metastatic Cells in Pediatric Liver Cancers

“…Figure 1C reveals that HepG2 cells with the shape of neurons are resistant to cisplatin. This result agrees with a previous study that detected chemoresistant HepG2 cells with the shape of neurons which are positive for a neuronal stem marker CD133 [24]. Western blot analyses with β-III-tubulin and NeuN confirmed that the cells resistant to cisplatin treatments are positive for markers of neurons (Figure 1D).…”

“…The inhibition of DNA replication by anti-cancer drug gemcitabine parallels the inhibition of CAFs and blocks the formation of tumor clusters in cells derived from lung metastases. Gemcitabine is an FDA-approved inhibitor of DNA replication and effectively kills cancer cells by increasing apoptosis [24][25][26]. We found that in a low-density setting, LM81 cells treated with gemcitabine (5 nM) do not form tumor clusters, compared to DMSO-treated cells (Figure 9A).…”