Impaired stem cell function of CD34+ cells selected by two different immunomagnetic beads systems

Kimura, Takafumi; Minamiguchi, Hitoshi; J, Wang; Kaneko, Hiroto; Nakagawa, Hitoshi; Fujii, Hideki; Sonoda, Yoshiaki

doi:10.1038/sj.leu.2403211

Cited by 9 publications

(9 citation statements)

References 30 publications

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“…cells were isolated by MiniMACS column with CD34 Isolation Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) from MNCs, as previously described [22,23]. Briefly, MNCs were incubated with a blocking agent and CD34 antibody (QBEND/10) from the Isolation Kit for 20 min at 4°C.…”

Section: Isolation Of Cd34 ? Cellsmentioning

confidence: 99%

Ex vivo expansion and long-term hematopoietic reconstitution ability of sorted CD34+CD59+ cells from patients with paroxysmal nocturnal hemoglobinuria

Han

Zhong

et al. 2010

Int J Hematol

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Autologous bone marrow transplantation (ABMT) for paroxysmal nocturnal hemoglobinuria (PNH) remains difficult so far. To expand residual normal CD34(+)CD59(+) cells isolated from patients with PNH and observe the long-term hematopoietic reconstruction ability of the expanded cells both ex vivo and in vivo, CD34(+)CD59(+) cells from 13 PNH patients and CD34(+) cells from 11 normal controls were separated from bone marrow mononuclear cells first by immunomagnetic microbeads and then by flow cytometry autoclone sorting. The cells were then cultivated under different conditions. The long-term hematopoietic supporting ability of expanded CD34(+)CD59(+) cells was evaluated by long-term culture in semi-solid medium in vitro and long-term engraftment in irradiated severe combined immunodeficiency (SCID) mice in vivo. The best combination of hematopoietic growth factors for ex vivo expansion was SCF + IL-3 + IL-6 + FL + Tpo + Epo. The most suitable time for harvest was on day 7. CD34(+)CD59(+) PNH cells retained strong colony-forming capacity even after expansion. The survival rate, complete blood cell count recovery on day 90, and human CD45 expression in different organs were similar between the irradiated SCID mice transplanted with expanded CD34(+)CD59(+) PNH cells and those with normal CD34(+) cells (P > 0.05) both in primary and secondary transplantation. These data provided a new potential way of managing PNH with ABMT.

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Section: Isolation Of Cd34 ? Cellsmentioning

confidence: 99%

Ex vivo expansion and long-term hematopoietic reconstitution ability of sorted CD34+CD59+ cells from patients with paroxysmal nocturnal hemoglobinuria

Han

Zhong

et al. 2010

Int J Hematol

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“…For gene therapy application, these cells are transduced ex vivo, and reinfused into patients [11][12][13][14]. One of the adverse effects of these time-consuming, technically demanding, and costly procedures is the loss of HSC repopulating capacity [15][16][17][18][19]. Reducing the in vitro handling of HSCs would help preserve their inherent pluripotency, and the ideal way to achieve this reduction in handling would be to directly transduce CD34+ progenitor cells in the mobilized peripheral blood mononuclear cells (mPBMCs), bypassing the process of purification.…”

Section: Introductionmentioning

confidence: 99%

Targeted transduction of CD34+ hematopoietic progenitor cells in nonpurified human mobilized peripheral blood mononuclear cells

Liang¹,

Pariente²,

Morizono³

et al. 2009

The Journal of Gene Medicine

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Background Conventional gene-therapy applications of hematopoietic stem cells (HSCs) involve purification of CD34+ progenitor cells from the mobilized peripheral blood, ex vivo transduction of the gene of interest into them, and reinfusion of the transduced CD34+ progenitor cells into patients. Eliminating the process of purification would save labor, time and money, while enhancing HSCs viability, transplantability and pluripotency. Lentiviral vectors have been widely used in gene therapy because they infect both dividing and nondividing cells and provide sustained transgene expression. One of the exceptions to this rule is quiescent primary lymphocytes, in which reverse transcription of viral DNA is not completed. Methods In the present study, we tested the possibility of targeting CD34+ progenitor cells within nonpurified human mobilized peripheral blood mononuclear cells (mPBMCs) utilizing vesicular stomatitis virus G (VSV-G) pseudotyped lentiviral vectors, based on the assumption that the CD34+ progenitor cells would be preferentially transduced. To further enhance the specificity of vector transduction, we also examined utilizing a modified Sindbis virus envelope (2.2) pseudotyped lentiviral vector, developed in our laboratory, that allows targeted transduction to specific cell receptors via antibody recognition. Results Both the VSV-G and 2.2 pseudotyped vectors achieved measurable results when they were used to target CD34+ progenitor cells in nonpurified mPBMCs. Conclusions Overall, the data obtained demonstrate the potential of ex vivo targeting of CD34+ progenitor cells without purification.

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“…Nowadays, MACS techniques have been widely used in cell biology, clinical diagnostics, environment protection, and food security area [3, 6, 10], especially in nucleic acid and protein separation, cell fast isolation and tumor cells depletion [8, 34]. Meanwhile, MACS techniques are also popular in hematological system cell sorting [14]. …”

Section: Introductionmentioning

confidence: 99%

Characterization of Rat Hair Follicle Stem Cells Selected by Vario Magnetic Activated Cell Sorting System

En-yi

Lian

Chen

et al. 2009

Acta Histochem. Cytochem.

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Hair follicle stem cells (HfSCs) play crucial roles in hair follicle morphogenesis and hair cycling. These stem cells are self-renewable and have the multi-lineage potential to generate epidermis, sebaceous glands, and hair follicle. The separation and identification of hair follicle stem cells are important for further research in stem cell biology. In this study, we report on the successful enrichment of rat hair follicle stem cells through vario magnetic activated cell sorting (Vario MACS) and the biological characteristics of the stem cells. We chose the HfSCs positive surface markers CD34, α6-integrin and the negative marker CD71 to design four isolation strategies: positive selection with single marker of CD34, positive selection with single marker of α6-integrin, CD71 depletion followed by CD34 positive selection, and CD71 depletion followed by α6-integrin positive selection. The results of flow cytometry analysis showed that all four strategies had ideal effects. Specifically, we conducted a series of researches on HfSCs characterized by their high level of CD34, termed CD34bri cells, and low to undetectable expression of CD34, termed CD34dim cells. CD34bri cells had greater proliferative potential and higher colony-forming ability than CD34dim cells. Furthermore, CD34bri cells had some typical characteristics as progenitor cells, such as large nucleus, obvious nucleolus, large nuclear:cytoplasmic ratio and few cytoplasmic organelles. Our findings clearly demonstrated that HfSCs with high purity and viability could be successfully enriched with Vario MACS.

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Impaired stem cell function of CD34+ cells selected by two different immunomagnetic beads systems

Cited by 9 publications

References 30 publications

Ex vivo expansion and long-term hematopoietic reconstitution ability of sorted CD34+CD59+ cells from patients with paroxysmal nocturnal hemoglobinuria

Ex vivo expansion and long-term hematopoietic reconstitution ability of sorted CD34+CD59+ cells from patients with paroxysmal nocturnal hemoglobinuria

Targeted transduction of CD34+ hematopoietic progenitor cells in nonpurified human mobilized peripheral blood mononuclear cells

Characterization of Rat Hair Follicle Stem Cells Selected by Vario Magnetic Activated Cell Sorting System

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