Prolonged ex vivo culture of cord blood CD34⁺ cells facilitates myeloid and megakaryocytic engraftment in the non‐obese diabetic severe combined immunodeficient mouse model

Abstract

Summary. A clinical goal for ex vivo expansion of cord blood (CB) CD341 cells is to shorten the period of neutropenia and thrombocytopenia following myeloablative therapy and transplantation. Prolongation of cytokine expansion leads to the production of greater numbers of cells, and should have an impact on neutrophil and platelet recovery. Furthermore, expansion of CD34 1 cells should support the continued production of neutrophils and platelets in the 6-week period following transplantation. We tested these hypotheses by characterization of the kinetics (human CD45 1 cells in the blood) and phenotype (CD45, CD34, CD61, CD33, CD19 and CD3) of human engraftment in the non-obese diabetic severe combined immunodeficient mouse (NOD±SCID

The nonobese diabetic/severe combined immunodeficient (NOD/SCID) mouse model of childhood acute lymphoblastic leukemia reveals intrinsic differences in biologic characteristics at diagnosis and relapse

“…12 Our previous studies show that this method reliably detects 0.01% human CD45 ϩ cells in murine peripheral blood. 11,13 At the first indication of morbidity (weight loss, lethargy, ruffled fur), or no more than 28 weeks following inoculation, mice were killed by cervical dislocation. Cell suspensions of spleens and other organs were prepared by mincing the tissues and filtering through 70-m cell strainers (BD Labware, Franklin Lakes, NJ).…”

Evaluation of different protocols for gene transfer into non-obese diabetes/severe combined immunodeficiency disease mouse repopulating cells

“…Other factors which may contribute as the induction of proapoptotic surface molecules such as the CD95/Fas antigen or the loss of adhesion receptors necessary for engraftment, such as VLA-4 or CXCR4, during cytokine stimulation (Denning-Kendall et al 2003;Liu et al 2003;Ma et al 2002). Noteworthy and controversial to our data, several authors have described preservation of long-term engraftment capacity (Lewis et al 2001;Novelli et al 1999) or even a beneWcial eVect on engraftment capacity (Bjorgvinsdottir et al 2002;Piacibello et al 2002;Rice et al 2001) during ex vivo culture.…”

“…Nevertheless, a number of questions remain. For example, several groups have described successful transduction of primitive hematopoietic cells with transduction protocols lasting 4-5 days (Barquinero et al 2000;Leurs et al 2003) but the eVect of prolonged in vitro culture on the transduction eYciency and repopulating potential of human progenitor and stem cells has remained controversial (Ballen et al 2000;Dorrell et al 2000;van Hennik et al 1998;Novelli et al 1999;Rice et al 2001). Therefore, we have utilized the NOD/SCID model to address these issues by comparing transduction protocols with standard or prolonged ex vivo culture periods and analyzed the eVect on transduction eYciency, engraftment, stem and progenitor cell diVerentiation, as well as gene expression from various diVerentiated lympho-hematopoietic cell populations in this model.…”

Hematopoietic reconstitution of CD34+ cells derived from short-term cultured cord blood mononuclear cells

“…A number of previous studies had sought to optimize culture duration of ex vivo cultures initiated from CD34 + cells [10][11]19]. However, to our knowledge, there have been few reports about the optimization of culture duration of ex vivo cultures started from MNC.…”

“…Therefore, a few studies have been performed to determine the optimum culture duration to expand functional HSCs with xenogeneic systems. Rice et al [10] reported that mice transplanted with fourteenday cultured CD34 + cells showed more rapid engraftment and higher engraftment levels than those transplanted with unexpanded cells or seven-day cultured CD34 + cells. Bhatia et al [11] reported that SCID-repopulating cells (SRC) increased fourfold after 4 days' culture of CD34 + CD38 -cells but no SRC were detected when CD34 + CD38 -cells were cultured for 9 days.…”