scholarly journals Ten Fold Ex-vivo Expansion of Allostimulatory Dendritic Cells (DC) from Cord Blood by Flt-3 Ligand, GM-CSF, TNF-α, IL-4 and Autologous Plasma: Implications for Strategies for Neonatal Vaccinations and/or Post Cord Blood Transplant Immunotherapy ♦ 1378

1998 ◽  
Vol 43 ◽  
pp. 236-236
Author(s):  
Francisco A Bracho ◽  
Carmella van de Ven ◽  
John X Qian ◽  
Stephen Hou ◽  
Mitchell S Cairo
Keyword(s):  
Dendritic Cells ◽  
Cord Blood ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Autologous Plasma ◽  
Gm Csf ◽  
Cord Blood Transplant ◽  
Tnf Α

Attenuation of Surface CXCR4 Down-Regulation in Human Cord Blood HSC during Ex Vivo Expansion Using the HUBEC Co-Culture System.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1068-1068
Author(s):  
Naoko Takebe ◽  
Thomas MacVittie ◽  
Xiangfei Cheng ◽  
Ann M. Farese ◽  
Emily Welty ◽  
...  
Keyword(s):  
Cord Blood ◽  
Culture System ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Receptor Internalization ◽  
Down Regulation ◽  
Human Cord Blood ◽  
Hematopoietic Stem ◽  
Gm Csf

Abstract Down-modulation of surface CXCR4, a G-protein-coupled receptor, in hematopoietic stem cells (HSCs) undergoing ex vivo expansion culturing is considered to be one of the major causes of marrow reconstitution failure, possibly due to an HSC homing defect. Recently, it has been reported that severe combined immunodeficiency (SCID)-repopulating cells (SRC) were expanded from the CD34-enriched human adult bone marrow (ABM) or cord blood (CB) hematopoietic stem cells (HSC) using a human brain endothelial cell (HUBEC) co-culture system. We found that primitive cord blood cells expressing surface CXCR4 (82+5%) lost this capability significantly during 7 days of ex vivo expansion in the HUBEC co-culture containing the cytokines stem cell factor (SCF), flt-3, interleukin (IL)-6, IL-3, and granulocyte macrophage colony stimulating factor (GM-CSF). Expression levels of other surface proteins relevant to HSC homing, such as CD49d, CD95, CD26, or CD11a, were not down-modulated. We hypothesized that CXCR4 down-regulation was caused by a receptor internalization and tested several methods to reverse CXCR4 internalization back to the surface, such as elimination of GM-CSF in the culture media, performing a non-contact culture using the transwell, or adding either 0.3Mor 0.4M sucrose, or 25μg/ml chlorpromazine (CPZ), 24 hours prior to the analysis. CPZ and sucrose are known inhibitors of the cytokine-induced endocytosis of CXCR4 in neutrophils (Bruhl H. et al. Eur J Immunol 2003). Interestingly, 0.4M sucrose showed approximately a 2-fold increase of surface CXCR4 expression on CB CD34+ cells by flow cytometry analysis. CPZ and 0.3M sucrose showed a moderate increase expression of CXCR4. Using a transwell HUBEC co-culture system, CXCR4 surface expression on CD34+ cells was down-regulated during the ex vivo culture. In vitro HSC migration test showed 3.1-fold increase in migration compared to the control after incubation of HSC with 0.1M sucrose for 16 hours prior to the in vitro migration study. Eliminating GM-CSF from the cytokine cocktail or adding MG132 increased migration 1.36- and 1.2-fold compared to the control. We are currently performing an in vivo homing assay using nonobese diabetic (NOD)-SCID mice. In conclusion, the HUBEC ex vivo culture system down-regulates surface CXCR4 in human cord blood HSC. The mechanism of CXCR4 surface down regulation may be receptor internalization by cytokines. Sucrose may be useful in attenuation of CXCR4 surface expression in CD34+ HSC by inhibition of receptor internalization via clathrin-coated pits.

scholarly journals TGF-β combined with M-CSF and IL-4 induces generation of immune inhibitory cord blood dendritic cells capable of enhancing cytokine-induced ex vivo expansion of myeloid progenitors

Blood ◽  
2007 ◽  
Vol 110 (8) ◽  
pp. 2872-2879 ◽  
Author(s):  
Geling Li ◽  
Saeid Abediankenari ◽  
Young-June Kim ◽  
Timothy B. Campbell ◽  
Shigeki Ito ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Cord Blood ◽  
Ex Vivo ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Myeloid Progenitor Cells ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Myeloid Progenitors

Abstract Tolerogenic dendritic cells (DCs) may be valuable in transplantation for silencing immune reaction. Macrophage colony-stimulating factor (M-CSF)/IL-4 induces differentiation of cord blood (CB) monocytes into DCs (M-DCs) with tolerogenic phenotype/function. We assessed whether factors produced by tolerogenic DCs could modulate hematopoiesis. TGF-β1 added to CB M-DC cultures induced bona fide DC morphology (TGF-M-DCs), similar to that of DCs generated with TGF-β and granulocyte-macrophage colony-stimulating factor (GM-CSF)/IL-4 (TGF-GM-DCs). Of conditioned media (CM) produced from TGF-M-DCs, TGF-GM-DCs, M-DCs, and GM-DCs, TGF-M-DC CM was the only one that enhanced SCF, Flt3 ligand, and TPO expansion of myeloid progenitor cells ex vivo. This effect was blocked by neutralizing anti–M-CSF Ab, but protein analysis of CM suggested that M-CSF alone was not manifesting enhanced expansion of myeloid progenitors. LPS-stimulated TGF-M-DCs induced T-cell tolerance/anergy as effectively as M-DCs. TGF-M-DCs secreted significantly lower concentrations of progenitor cell inhibitory cytokines and were less potent in activating T cells than TGF-GM-DCs. Functional differences between TGF-M-DCs and TGF-GM-DCs included enhanced responses to LPS-induced ERK, JNK, and P38 activation in TGF-M-DCs and their immune suppressive–skewed cytokine release profiles. TGF-M-DCs appear unique among culture-generated DCs in their capability for silencing immunity while promoting expansion of myeloid progenitors, events that may be of therapeutic value.

scholarly journals Ex Vivo Expansion Does Not Alter the Capacity of Umbilical Cord Blood CD34+Cells to Generate Functional T Lymphocytes and Dendritic Cells

Stem Cells ◽  
2006 ◽  
Vol 24 (9) ◽  
pp. 2150-2157 ◽  
Author(s):  
Ladan Kobari ◽  
Marie C. Giarratana ◽  
Jean C. Gluckman ◽  
Luc Douay ◽  
Michelle Rosenzwajg
Keyword(s):  
Dendritic Cells ◽  
T Lymphocytes ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Generate Functional ◽  
Cd34 Cells ◽  
Cord Blood Cd34

Incidence of Ex Vivo Expansion on the Capacity of Cord Blood Graft To Generate Immune Cells: Rational for Co-Infusion of Expanded and Non Expanded Fractions?.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 407-407
Author(s):  
Michelle Rosenzwajg ◽  
Marie-Catherine Giarratana ◽  
Jean-Claude Gluckman ◽  
Luc Douay ◽  
Ladan Kobari
Keyword(s):  
Cord Blood ◽  
Immune Cells ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Th2 Response ◽  
Gm Csf ◽  
Hla Dr ◽  
Cd34 Cells ◽  
Significant Difference ◽  
And Function

Abstract The aim of our study was to determine whether ex vivo expansion of umbilical cord blood (UCB) progenitor cells induces changes in their capacity to generate immune cells. CD34+ CB cells were cultured for 14 days with SCF, FLT3-l, TPO and G-CSF, inducing a total cells, CD34+ and LTC-ICs increase of 1500, 120 and 8 fold respectively. Non expanded (d0) and 14-day expanded (d14) CD34+ cells were compared for their capacity to produce T lymphocytes (TLs) using the fetal thymus organ culture system and DCs generated from d0 and d14 CD34+ cells were compared for their differentiation, phenotype and function. Total percentages of CD4+, CD4+CD8− and CD4+CD8+ TLs obtained from d0 and d14 CD34+ cells were comparable. In both fractions, most of the CD4+ T cells co-expressed iCD3 but a lower proportion of d14 derived TLs expressed sCD3. However, there was no significant difference between d0 and d14 derived TLs in term of Vb chain representation, all TCR-Vb chains examined being represented in each case. These data indicate that d0 and d14 CD34+ cells have a similar capacity to generate TLs and that expansion does not induced any skewing of the TCR-Vb repertoire. D0 and d14 CD34+ cells were next cultured with SCF, FL, GM-CSF and TNF-a to compare their capacity to differentiate into DCs. Similar percentages of CD1a+ DCs expressing the same levels of HLA-DR and co stimulatory molecules were obtained. DCs derived from d14 CD34+ cells were less potent to stimulate allogeneic TLs, but the pattern of cytokines produced by stimulated TLs was similar and no shift towards a predominant Th1 or Th2 response was observed. Moreover, in spite of a quantitative increase (15 fold) related to the CD34 pool amplification, we observed a decreased capacity (13-fold) of d14 cells to generate DCs compared to d0 CD34+ cells. Overall, these results indicate that ex vivo expansion of CD34+ cells doesn’t induce any major modification in T Lymphopoiesis capacity while alters somehow the capacity of the graft to generate DCs. We discuss in the context of UCB transplantation, the putative interest of co-infusion of expanded and non expanded fractions in view of improving myelopoiesis in the graft without subverting the immune reconstitution.

TGF-β1, in Synergy with M-CSF Induce Generation of a Novel Type of Human Cord Blood Dendritic Cell That Is Both Anti-Inflammatory and Capable of Ex-Vivo Expansion of Myeloid Progenitor Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2380-2380
Author(s):  
Geling Li ◽  
Timothy B. Campbell ◽  
Barbara Graham-Evans ◽  
Scott Cooper ◽  
Hal E. Broxmeyer
Keyword(s):  
Dendritic Cells ◽  
Growth Factor ◽  
Immune Responses ◽  
Cord Blood ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Cd34 Cells ◽  
Anti Inflammatory ◽  
Induced Apoptosis ◽  
Tgf Β1

Abstract Tolerogenic dendritic cells (DCs) could be a promising cellular therapeutic tool in human transplantation due to their potential for silencing unwanted immune reactions. However, it is not known whether tolerogenic DCs are capable of regulating human hematopoiesis. We recently reported that M-CSF, in combination with IL-4, induced cord blood monocyte differentiation into a population of dendritic cells (DC) with tolerogenic potential (M-DC) (Li et.al. J. Immol.174: 4706, 2005.). TGF-β1 has potent immune regulatory functions on DCs. In this context, we investigated effects of TGF-β1 on cord blood M-DC differentiation by characterizing phenotype, cytokine production and allogeneic mixed lymphocyte reaction (MLR) of the resulting DC population (TGF/M-DC). We also evaluated effects of the resultant cells on expansion and growth factor withdrawal-induced apoptosis of human CD34+ progenitors isolated from cord blood. The effects were compared to those of TGF-β1, GM-CSF and IL-4-induced DCs (TGF/DC). Addition of TGF-β1 to highly purified cord blood monocytes cultured with M-CSF and IL-4 changed M-DC morphology to that of a homogeneous DC morphology (TGF/M-DC). Similar to M-DC, TGF/M-DC negatively stained for monocyte/macrophage markers CD14 and CD16, and positively stained for CD86 and HLA-DR, molecules involved in antigen presentation. It appeared that TGF/M-DC were bona fide DC with typical DC morphology and phenotype. In comparison with TGF-DC, TGF/M-DC produced high amounts of IL-10, but displayed reduced capacity in induction of allogeneic MLR, features categorizing them more appropriately into an anti-inflammatory DC subset. We generated conditioned medium (CM) from both types of DC by stimulation of these DC with LPS and studied effects of their CM on ex-vivo expansion of CD34+ progenitors isolated from cord blood. In the absence of DC CM, the combination of SCF, Flt3 ligand and TPO (SFT, a cytokine combination widely used for cell expansion) induced a 10.5 ± 1.8 fold expansion of myeloid progenitors (CFU-GM) in 1 week culture. TGF/M-DC CM (25% v/v), but not TGF DC CM, in synergy with SFT further expanded myeloid progenitors (CFU-GM) 19.4 ± 2.7 fold beyond that seen with SFT alone. The effects of TGF/M-DC CM was accompanied with a significant increase of CD34+ cell numbers after 1 week (TGF/M-DC CM in combination with SFT, TGF-DC CM in combination with SFT, and SFT alone expanded CD34+ cells by 8.4 ± 2.8 fold, 3.8 ± 2.8 fold and 5.9 ± 1.1 fold respectively). TGF/M-DC CM and TGF/DC CM prevented growth factor withdrawal-induced apoptosis of CD34+ cells, as analyzed by annexin-V staining. Therefore, TGF/M-DC might be capable of in vivo silencing of unwanted immune responses while promoting myeloid recovery. These features of TGF/M-DC make them an interesting cellular therapeutic candidate to potentially facilitate hematopoietic recovery as well as down-modulate unwanted immune responses after hematopoietic stem cell transplantation.

scholarly journals Human amniotic mesenchymal stromal cells support the ex vivo expansion of cord blood hematopoietic stem cells

2021 ◽  
Author(s):  
Valentina Orticelli ◽  
Andrea Papait ◽  
Elsa Vertua ◽  
Patrizia Bonassi Signoroni ◽  
Pietro Romele ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Cord Blood ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Hematopoietic Stem

scholarly journals CD14 Expressing Precursors Give Rise to Highly Functional Conventional Dendritic Cells for Use as Dendritic Cell Vaccine

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3818
Author(s):  
Maud Plantinga ◽  
Denise A. M. H. van den Beemt ◽  
Ester Dünnebach ◽  
Stefan Nierkens
Keyword(s):  
Dendritic Cells ◽  
T Cell ◽  
Cord Blood ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ex Vivo ◽  
Dendritic Cell Vaccine ◽  
Cell Vaccine ◽  
Activated T Cells

Induction of long-lasting immunity by dendritic cells (DCs) makes them attractive candidates for anti-tumor vaccination. Although DC vaccinations are generally considered safe, clinical responses remain inconsistent in clinical trials. This initiated studies to identify subsets of DCs with superior capabilities to induce effective and memory anti-tumor responses. The use of primary DCs has been suggested to overcome the functional limitations of ex vivo monocyte-derived DCs (moDC). The ontogeny of primary DCs has recently been revised by the introduction of DC3, which phenotypically resembles conventional (c)DC2 as well as moDC. Previously, we developed a protocol to generate cDC2s from cord blood (CB)-derived stem cells via a CD115-expressing precursor. Here, we performed index sorting and single-cell RNA-sequencing to define the heterogeneity of in vitro developed DC precursors and identified CD14+CD115+ expressing cells that develop into CD1c++DCs and the remainder cells brought about CD123+DCs, as well as assessed their potency. The maturation status and T-cell activation potential were assessed using flow cytometry. CD123+DCs were specifically prone to take up antigens but only modestly activated T-cells. In contrast, CD1c++ are highly mature and specialized in both naïve as well as antigen-experienced T-cell activation. These findings show in vitro functional diversity between cord blood stem cell-derived CD123+DC and CD1c++DCs and may advance the efficiency of DC-based vaccines.

Sign in / Sign up

Export Citation Format

Share Document