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Series GSE33020 Query DataSets for GSE33020
Status Public on Feb 27, 2012
Title CD20 positive cells are undetectable in the majority of multiple myeloma cell lines and are not associated with a cancer stem cell phenotype
Organism Homo sapiens
Experiment type Expression profiling by array
Summary Although new therapies have doubled the survival of multiple myeloma (MM) patients, this remains an incurable disease. It has been postulated that the so-called MM Cancer Stem Cells (MM-CSC) would be responsible for tumor initiation and relapse but their unequivocal identification remains unclear. Here, we investigated in a panel of MM cell lines the presence of CD20+ cells harboring a MM-CSC phenotype. Among the multiple cell lines investigated, only a small population of CD20dim+ cells (0.3%) in the RPMI-8226 cell line was found. CD20dim+ RPMI-8226 cells expressed the plasma cell markers CD38 and CD138 and were CD19-CD27-. Additionally, CD20dim+ RPMI-8226 cells did not exhibit stem-cell markers as shown by gene expression profiling and the aldehyde dehydrogenase (ALDH) assay. Moreover, we demonstrated that CD20dim+ RPMI-8226 cells are not essential for CB17-SCID mice engraftment and show lower self-renewal potential than the CD20- RPMI-8226 cells. These results do not support CD20+ expression for the identification of MM-CSC.
 
Overall design For detailed description of Material and Methods, see supplementary Methods.
The human MM cell lines RPMI-8226, U266, MM1S, MM1R, NCI-H929, RPMI-LR5, U266-LR7 and U266-Dox4 were studied. Cells were cultured as previously described [Ocio, 2009, 3781].
MM cell lines were immunophenotyped using a 7-color immunofluorescence technique [Paiva, 2011, 697], with the following combination of monoclonal antibodies (Pacific Blue (PB)/ anemonia majano cyan (AmCyan)/ fluorescein isothiocyanate (FITC)/ peridinin chlorophyll protein-cyanin 5.5 (PerCP-Cy5.5)/ PE-cyanin 7 (PE-Cy7)/ allophycocyanin (APC)/ alexafluor 700 (AF700)): CD19/CD45/CD20/CD138/CD27/CD56/CD38. Data were stored for a minimum of 3x105 events. CD20dim+ and CD20- RPMI-8226 cells were sorted after incubation with CD20-APC/7AAD and acquisition on a FACSAria cytometer (Becton Dickison Biosciences). Sorting was performed only for viable cells (7AAD-) and debris were excluded by scatter properties. The CD20- and CD20dim+ RPMI-8266 sorted cells had a final purity of 98% and 88%, respectively.
CD20dim+ and CD20- RPMI-8226 cells were extensively characterized. Morphological characterization was done with May-Grünwald-Giemsa staining. Characterization of VDJH and IGKappa rearrangements was performed as described elsewhere [BIOMED-2, BMH4-CT98-3936]. The expression of aldehyde dehydrogenase (ALDH) was assessed using the Aldefluor Kit (StemCell Technologies) following manufacturer’s instructions with further staining with a CD20-APC antibody. For gene expression profile studies, RNA was isolated, labeled and hybridized to Human Gene 1.0 ST array (Affymetrix) according to Affymetrix protocols [Gutiérrez, 2005, 402]. The arrays were analyzed using the DNA-Chip Analyzer software (DChip). Fold change ≥2 was considered significant.
 
Contributor(s) Paíno T, Ocio EM, Paiva B, San-Segundo L, Garayoa M, Gutiérrez NC, Sarasquete ME, Pandiella A, Orfao A, San Miguel JF
Citation(s) 22315496
Submission date Oct 17, 2011
Last update date Jul 26, 2018
Contact name Teresa Paíno
E-mail(s) tpaino@usal.es
Phone +34 923294812
Organization name Centro de investigación del cáncer
Street address Campus Miguel de Unamuno
City Salamanca
ZIP/Postal code 37007
Country Spain
 
Platforms (1)
GPL6244 [HuGene-1_0-st] Affymetrix Human Gene 1.0 ST Array [transcript (gene) version]
Samples (6)
GSM818466 CD20- RPMI-8226 REPLICATE 1
GSM818471 CD20dim+ RPMI-8226 REPLICATE 1
GSM818474 CD20dim+ RPMI-8226 REPLICATE 2
Relations
BioProject PRJNA148017

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Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE33020_RAW.tar 26.7 Mb (http)(custom) TAR (of CEL)
Processed data included within Sample table

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