Multipotent neural precursors express neural and hematopoietic factors, and enhance ex vivo expansion of cord blood CD34+ cells, colony forming units and NOD/SCID-repopulating cells in contact and noncontact cultures

Abstract

In view of the possible crosstalks between hematopoiesis and neuropoiesis, we evaluated two microenvironments, murine neonatal neural cell line C17.2 and primary embryonic aortagonad-mesonephros (AGM) stromal cells, on the ex vivo expansion of CD34 þ cells from human cord blood. In a contact culture system, C17.2 or AGM cells significantly enhanced the expansion of CD34 þ cells to a panel of early and committed hematopoietic progenitor cells. In a noncontact transwell system, pre-established C17.2 cells significantly increased the expansion of total nucleated cells, CD34 þ cells and multilineage colony forming cells (Po0.01). Expanded cells were infused into nonobese diabetic/severe-combined immunodeficient mice. The engraftment of human (hu)CD45 þ cells in the bone marrow of these mice was consistently higher in all the 10 experiments conducted with the support of C17.2 cells when compared with those in respective control groups (11.9 vs 2.43%, P ¼ 0.03). Using RT-PCR and Southern blot analysis, we showed that AGM and C17.2 cells expressed a panel of hematopoietic, bone morphogenetic and neurotrophic factors. Our data provided the first evidence on the promoting effects of a neural progenitor cell line on hematopoiesis at a noncontact condition. The mechanism could be mediated by the expression of multilineage regulatory factors.

Promoting Effects of Serotonin on Hematopoiesis: Ex Vivo Expansion of Cord Blood CD34+Stem/Progenitor Cells, Proliferation of Bone Marrow Stromal Cells, and Antiapoptosis

“…Colonies were scored in a blinded manner. Colony‐forming unit‐megakaryocytes (CFU‐MK) were assayed using the plasma clot system as described previously [33, 34]. CFU‐MK was identified as a cluster of three or more strongly stained CD61‐FITC (Dako) positive cells examined by fluorescence microscopy.…”

“…In each independent cord blood experiment ( n = 16), cells expanded in the presence or absence of serotonin were infused into sex‐ and age‐matched mice (progenies of 3 × 10 4 CD34 + cells at day 0 per mouse). To prevent the loss of data due to animal mortality, two mice were assigned to each treatment group, and the engraftment parameters averaged as a single datum for analysis, as described previously [33, 34]. These animals were sacrificed 6 weeks post‐transplantation.…”

Expansion of hematopoietic stem/progenitor cells

“…A report by Jang et al showed that MSC derived from human umbilical cord, human placenta, and lung could support ex vivo expansion of committed HPCs collected from the UCB in the presence of recombinant cytokines [7]. Li et al reported that murine neonatal neural cell line C17.2 significantly increased the expansion of total nucleated cells, CD341 cells, and multilineage colony forming cells at a noncontact condition [8]. Moreover, porcine microvascular endothelial cell [9], human yolk sac-derived endothelial cells (hYSECs) [10], umbilical vein endothelial cells (HUVEC) [11], osteoblasts [12], and extracellular matrix [13] have also been reported to play a significant role in HSPCs expansion.…”

The plant mannose‐binding lectin NTL preserves cord blood haematopoietic stem/progenitor cells in long‐term culture and enhances their ex vivo expansion

“…Enriched CD34 + cells or expanded cells at 3 · 10 3 /ml were seeded in triplicate. Colonies were scored after culturing for 14 d. Megakaryocyte colony forming units (CFU-MK) were assayed using the plasma clot system as described previously (Li et al, 2005(Li et al, , 2006. CFU-MK was identified as a cluster of three or more strongly stained CD61-FITC (Dako) positive cells examined by fluorescence microscopy.…”

“…CD34 + cells were enriched using the VarioMACS Isolation Kit (Miltenyi Biotec Inc., Gladbach, Germany) as described previously (Li et al, 2005(Li et al, , 2006 and respective isotype controls for 20 min. All antibodies and cytometric reagents were purchased from Becton DickinsonPharmingen (BD, San Jose, CA, USA), unless specified otherwise.…”