Factors Influencing Percentage Yield of Side Population Isolated in Ovarian Cancer Cell LineSK-OV-3
DOI:
https://doi.org/10.6000/1929-2279.2014.03.01.5Keywords:
Side population (SP), SK-OV-3, cancer stem cells (CSCs), flow cytometry (FCM), Hoechst 33342.Abstract
Isolation of side population (SP) cells has been recognized as a useful technique for the isolation and identification of hematopoietic stem cells or cancer stem cells (CSCs). Thus the yield and purity of isolated SP cells would have a profound influence on the research outcomes in these two important areas. Hoechst 33342 exclusion assay technique has been used for the identification of SP cells. However, diverse Hoechst staining protocols giving different SP yields even from the same tissue type or same cell line have been reported in different laboratories. In this study we systematically investigated the underlying factors influencing the SP yield using Hoechst dye staining and a robust platform of flow cytometric analysis of the human ovarian cancer cell line SK-OV-3. Our study revealed that SP yield was not only affected by the Hoechst 33342 concentration, staining cell density, staining cell viability, staining duration, staining medium, flow cytometric setting and SP gating strategy, but was also affected by the cell passage number in SK-OV-3. This is the first systematic study on the factors affecting SP yield in adherent cells that mimic many solid tumour tissues. Our results provide important technical guidelines to help ensure reproducible and comparable results in SP and CSCs study.
References
Goodell MA, Brose K, Paradis G, Conner AS, Mulligan RC. Isolation and functional properties of murine hematopoietic stem cells that are replicating in vivo. J Exp Med 1996; 183(4): 1797-806. http://dx.doi.org/10.1084/jem.183.4.1797
Wulf GG, Wang RY, Kuehnle I, Weidner D, Marini F, Brenner MK, et al. A leukemic stem cell with intrinsic drug efflux capacity in acute myeloid leukemia. Blood 2001; 98(4): 1166-73. http://dx.doi.org/10.1182/blood.V98.4.1166
Han JS, Crowe DL. Tumor initiating cancer stem cells from human breast cancer cell lines. Int J Oncol 2009; 34(5): 1449-53.
Hiraga T, Ito S, Nakamura H. Side population in MDA-MB-231 human breast cancer cells exhibits cancer stem cell-like properties without higher bone-metastatic potential. Oncol Rep 2011; 25(1): 289-96.
Nakanishi T, Chumsri S, Khakpour N, Brodie AH, Leyland-Jones B, Hamburger AW, et al. Side-population cells in luminal-type breast cancer have tumour-initiating cell properties, and are regulated by HER2 expression and signalling. Br J Cancer 2010; 102(5): 815-26. http://dx.doi.org/10.1038/sj.bjc.6605553
Engelmann K, Shen H, Finn OJ. MCF7 side population cells with characteristics of cancer stem/progenitor cells express the tumor antigen MUC1. Cancer Res 2008; 68(7): 2419-26. http://dx.doi.org/10.1158/0008-5472.CAN-07-2249
Fukaya R, Ohta S, Yamaguchi M, Fujii H, Kawakami Y, Kawase T, et al. Isolation of cancer stem-like cells from a side population of a human glioblastoma cell line, SK-MG-1. Cancer Lett 2010; 291(2): 150-7. http://dx.doi.org/10.1016/j.canlet.2009.10.010
Ho MM, Ng AV, Lam S, Hung JY. Side population in human lung cancer cell lines and tumors is enriched with stem-like cancer cells. Cancer Res 2007; 67(10): 4827-33. http://dx.doi.org/10.1158/0008-5472.CAN-06-3557
Shi Y, Fu X, Hua Y, Han Y, Lu Y, Wang J. The side population in human lung cancer cell line NCI-H460 is enriched in stem-like cancer cells. PLoS One 2012; 7(3): e33358. http://dx.doi.org/10.1371/journal.pone.0033358
Wang J, Guo LP, Chen LZ, Zeng YX, Lu SH. Identification of cancer stem cell-like side population cells in human nasopharyngeal carcinoma cell line. Cancer Res 2007; 67(8): 3716-24. http://dx.doi.org/10.1158/0008-5472.CAN-06-4343
Chiba T, Kita K, Zheng YW, Yokosuka O, Saisho H, Iwama A, et al. Side population purified from hepatocellular carcinoma cells harbors cancer stem cell-like properties. Hepatology 2006; 44(1): 240-51. http://dx.doi.org/10.1002/hep.21227
Kamohara Y, Haraguchi N, Mimori K, Tanaka F, Inoue H, Mori M, et al. The search for cancer stem cells in hepatocellular carcinoma. Surgery 2008; 144(2): 119-24. http://dx.doi.org/10.1016/j.surg.2008.04.008
Li XX, Wang J, Wang HL, Wang W, Yin XB, Li QW, et al. Characterization of cancer stem-like cells derived from a side population of a human gallbladder carcinoma cell line, SGC-996. Biochem Biophys Res Commun 2012; 419(4): 728-34. http://dx.doi.org/10.1016/j.bbrc.2012.02.090
Zhang Z, Zhu F, Xiao L, Wang M, Tian R, Shi C, et al. Side population cells in human gallbladder cancer cell line GBC-SD regulated by TGF-beta-induced epithelial-mesenchymal transition. J Huazhong Univ Sci Technolog Med Sci 2011; 31(6): 749-55. http://dx.doi.org/10.1007/s11596-011-0671-1
Xu XT, Xu Q, Tong JL, Zhu MM, Nie F, Chen X, et al. MicroRNA expression profiling identifies miR-328 regulates cancer stem cell-like SP cells in colorectal cancer. Br J Cancer 2012; 106(7): 1320-30. http://dx.doi.org/10.1038/bjc.2012.88
Geng S, Wang Q, Wang J, Hu Z, Liu C, Qiu J, et al. Isolation and identification of a distinct side population cancer cells in the human epidermal squamous cancer cell line A431. Arch Dermatol Res 2011; 303(3): 181-9. http://dx.doi.org/10.1007/s00403-010-1100-1
Friel AM, Sergent PA, Patnaude C, Szotek PP, Oliva E, Scadden DT, et al. Functional analyses of the cancer stem cell-like properties of human endometrial tumor initiating cells. Cell Cycle 2008; 7(2): 242-9. http://dx.doi.org/10.4161/cc.7.2.5207
Zhang P, Zhang Y, Mao L, Zhang Z, Chen W. Side population in oral squamous cell carcinoma possesses tumor stem cell phenotypes. Cancer Lett 2009; 277(2): 227-34. http://dx.doi.org/10.1016/j.canlet.2008.12.015
Huang D, Gao Q, Guo L, Zhang C, Jiang W, Li H, et al. Isolation and identification of cancer stem-like cells in esophageal carcinoma cell lines. Stem Cells Dev 2009; 18(3): 465-73. http://dx.doi.org/10.1089/scd.2008.0033
Kato K, Yoshimoto M, Adachi S, Yamayoshi A, Arima T, Asanoma K, et al. Characterization of side-population cells in human normal endometrium. Hum Reprod 2007; 22(5): 1214-23. http://dx.doi.org/10.1093/humrep/del514
Storms RW, Goodell MA, Fisher A, Mulligan RC, Smith C. Hoechst dye efflux reveals a novel CD7(+)CD34(-) lymphoid progenitor in human umbilical cord blood. Blood 2000; 96(6): 2125-33.
Yanamoto S, Kawasaki G, Yamada S, Yoshitomi I, Kawano T, Yonezawa H, et al. Isolation and characterization of cancer stem-like side population cells in human oral cancer cells. Oral Oncol 2011; 47(9): 855-60. http://dx.doi.org/10.1016/j.oraloncology.2011.06.501
Kondo T, Setoguchi T, Taga T. Persistence of a small subpopulation of cancer stem-like cells in the C6 glioma cell line. Proc Natl Acad Sci USA 2004; 101(3): 781-6. http://dx.doi.org/10.1073/pnas.0307618100
Kobayashi I, Saito K, Moritomo T, Araki K, Takizawa F, Nakanishi T. Characterization and localization of side population (SP) cells in zebrafish kidney hematopoietic tissue. Blood 2008; 111(3): 1131-7. http://dx.doi.org/10.1182/blood-2007-08-104299
Feuring-Buske M, Hogge DE. Hoechst 33342 efflux identifies a subpopulation of cytogenetically normal CD34(+)CD38(-) progenitor cells from patients with acute myeloid leukemia. Blood 2001; 97(12): 3882-9. http://dx.doi.org/10.1182/blood.V97.12.3882
Moshaver B, van Rhenen A, Kelder A, van der Pol M, Terwijn M, Bachas C, et al. Identification of a small subpopulation of candidate leukemia-initiating cells in the side population of patients with acute myeloid leukemia. Stem Cells 2008; 26(12): 3059-67. http://dx.doi.org/10.1634/stemcells.2007-0861
Meeson AP, Hawke TJ, Graham S, Jiang N, Elterman J, Hutcheson K, et al. Cellular and molecular regulation of skeletal muscle side population cells. Stem Cells 2004; 22(7): 1305-20. http://dx.doi.org/10.1634/stemcells.2004-0077
Fujii T, Zen Y, Harada K, Niwa H, Masuda S, Kaizaki Y, et al. Participation of liver cancer stem/progenitor cells in tumorigenesis of scirrhous hepatocellular carcinoma--human and cell culture study. Hum Pathol 2008; 39(8): 1185-96. http://dx.doi.org/10.1016/j.humpath.2007.12.010
Goodell MA. Stem cell identification and sorting using the Hoechst 33342 side population (SP). Curr Protoc Cytom 2005; Chapter 9: Unit9 18.
Christgen M, Ballmaier M, Lehmann U, Kreipe H. Detection of putative cancer stem cells of the side population phenotype in human tumor cell cultures. Methods Mol Biol 2012; 878: 201-15. http://dx.doi.org/10.1007/978-1-61779-854-2_13
Kim M, Turnquist H, Jackson J, Sgagias M, Yan Y, Gong M, et al. The multidrug resistance transporter ABCG2 (breast cancer resistance protein 1) effluxes Hoechst 33342 and is overexpressed in hematopoietic stem cells. Clin Cancer Res 2002; 8(1): 22-8.
Patrawala L, Calhoun T, Schneider-Broussard R, Zhou J, Claypool K, Tang DG. Side population is enriched in tumorigenic, stem-like cancer cells, whereas ABCG2+ and ABCG2- cancer cells are similarly tumorigenic. Cancer Res 2005; 65(14): 6207-19. http://dx.doi.org/10.1158/0008-5472.CAN-05-0592
Mitsutake N, Iwao A, Nagai K, Namba H, Ohtsuru A, Saenko V, et al. Characterization of side population in thyroid cancer cell lines: cancer stem-like cells are enriched partly but not exclusively. Endocrinology 2007; 148(4): 1797-803. http://dx.doi.org/10.1210/en.2006-1553
Christgen M, Ballmaier M, Bruchhardt H, von Wasielewski R, Kreipe H, Lehmann U. Identification of a distinct side population of cancer cells in the Cal-51 human breast carcinoma cell line. Mol Cell Biochem 2007; 306(1-2): 201-12. http://dx.doi.org/10.1007/s11010-007-9570-y
Montanaro F, Liadaki K, Schienda J, Flint A, Gussoni E, Kunkel LM. Demystifying SP cell purification: viability, yield, and phenotype are defined by isolation parameters. Exp Cell Res 2004; 298(1): 144-54. http://dx.doi.org/10.1016/j.yexcr.2004.04.010
Mo SL, Li J, Loh YS, Brown RD, Smith AL, Chen Y, et al. Factors influencing the abundance of the side population in a human myeloma cell line. Bone Marrow Res 2011; 2011: 524845.
Petersen TW, Ibrahim SF, Diercks AH, van den Engh G. Chromatic shifts in the fluorescence emitted by murine thymocytes stained with Hoechst 33342. Cytometry A 2004; 60(2): 173-81. http://dx.doi.org/10.1002/cyto.a.20058
BD. BD LSR II User's Guide. [cited 2013 November 28]; Available from: http: //flowcytometry.sysbio.med.harvard.edu/ documents/BD%20LSRII%20User's%20Guide.pdf
ATCC. Passage number effects in cell lines. [cited 2013 November 28]; Available from: http: //www.atcc.org/~/media/ PDFs/Technical%20Bulletins/tb07.ashx
Pearce DJ, Ridler CM, Simpson C, Bonnet D. Multiparameter analysis of murine bone marrow side population cells. Blood 2004; 103(7): 2541-6. http://dx.doi.org/10.1182/blood-2003-09-3281
Guo Y, Follo M, Geiger K, Lubbert M, Engelhardt M. Side-population cells from different precursor compartments. J Hematother Stem Cell Res 2003; 12(1): 71-82. http://dx.doi.org/10.1089/152581603321210154
Parmar K, Sauk-Schubert C, Burdick D, Handley M, Mauch P. Sca+CD34- murine side population cells are highly enriched for primitive stem cells. Exp Hematol 2003; 31(3): 244-50. http://dx.doi.org/10.1016/S0301-472X(02)01074-3
Alvi AJ, Clayton H, Joshi C, Enver T, Ashworth A, Vivanco MM, et al. Functional and molecular characterisation of mammary side population cells. Breast Cancer Res 2003; 5(1): R1-8. http://dx.doi.org/10.1186/bcr563
Watanabe K, Nishida K, Yamato M, Umemoto T, Sumide T, Yamamoto K, et al. Human limbal epithelium contains side population cells expressing the ATP-binding cassette transporter ABCG2. FEBS Lett 2004; 565(1-3): 6-10. http://dx.doi.org/10.1016/j.febslet.2004.03.064
Umemoto T, Yamato M, Nishida K, Kohno C, Yang J, Tano Y, et al. Rat limbal epithelial side population cells exhibit a distinct expression of stem cell markers that are lacking in side population cells from the central cornea. FEBS Lett 2005; 579(29): 6569-74. http://dx.doi.org/10.1016/j.febslet.2005.10.047
Iwatani H, Ito T, Imai E, Matsuzaki Y, Suzuki A, Yamato M, et al. Hematopoietic and nonhematopoietic potentials of Hoechst(low)/side population cells isolated from adult rat kidney. Kidney Int 2004; 65(5): 1604-14. http://dx.doi.org/10.1111/j.1523-1755.2004.00561.x
Challen GA, Bertoncello I, Deane JA, Ricardo SD, Little MH. Kidney side population reveals multilineage potential and renal functional capacity but also cellular heterogeneity. J Am Soc Nephrol 2006; 17(7): 1896-912. http://dx.doi.org/10.1681/ASN.2005111228
Kawanabe N, Murakami K, Takano-Yamamoto T. The presence of ABCG2-dependent side population cells in human periodontal ligaments. Biochem Biophys Res Commun 2006; 344(4): 1278-83. http://dx.doi.org/10.1016/j.bbrc.2006.03.237
Moserle L, Indraccolo S, Ghisi M, Frasson C, Fortunato E, Canevari S, et al. The side population of ovarian cancer cells is a primary target of IFN-alpha antitumor effects. Cancer Res 2008; 68(14): 5658-68. http://dx.doi.org/10.1158/0008-5472.CAN-07-6341
Britton KM, Eyre R, Harvey IJ, Stemke-Hale K, Browell D, Lennard TW, et al. Breast cancer, side population cells and ABCG2 expression. Cancer Lett 2012; 323(1): 97-105. http://dx.doi.org/10.1016/j.canlet.2012.03.041
Gao Q, Geng L, Kvalheim G, Gaudernack G, Suo Z. Identification of cancer stem-like side population cells in ovarian cancer cell line OVCAR-3. Ultrastruct Pathol 2009; 33(4): 175-81.
Bar EE, Chaudhry A, Lin A, Fan X, Schreck K, Matsui W,
et al. Cyclopamine-mediated hedgehog pathway inhibition depletes stem-like cancer cells in glioblastoma. Stem Cells 2007; 25(10): 2524-33. http://dx.doi.org/10.1634/stemcells.2007-0166
Weber K, Paulus W, Senner V. The side population of gliomas exhibits decreased cell migration. J Neuropathol Exp Neurol 2010; 69(6): 623-31. http://dx.doi.org/10.1097/NEN.0b013e3181e04739
Hirschmann-Jax C, Foster AE, Wulf GG, Nuchtern JG, Jax TW, Gobel U, et al. A distinct "side population" of cells with high drug efflux capacity in human tumor cells. Proc Natl Acad Sci USA 2004; 101(39): 14228-33. http://dx.doi.org/10.1073/pnas.0400067101
Chen JS, Pardo FS, Wang-Rodriguez J, Chu TS, Lopez JP, Aguilera J, et al. EGFR regulates the side population in head and neck squamous cell carcinoma. Laryngoscope 2006; 116(3): 401-6. http://dx.doi.org/10.1097/01.mlg.0000195075.14093.fb
Loh YS, Mo S, Brown RD, Yamagishi T, Yang S, Joshua DE, et al. Presence of Hoechst low side populations in multiple myeloma. Leuk Lymphoma 2008; 49(9): 1813-6. http://dx.doi.org/10.1080/10428190802272676
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