Hematopoietic stem cell/hematopoietic progenitor cell (HSC/HPC) homing to specific microenvironmental niches involves interactions between multiple receptor ligand pairs. Although CXCL12/CXCR4 plays a central role in these events, CXCR4 regulators that provide the specificity for such cells to lodge and be retained in particular niches are poorly defined. Here, we provide evidence that the sialomucin endolyn (CD164), an adhesion receptor that regulates the adhesion of CD34 ؉ cells to bone marrow stroma and the recruitment of CD34 ؉ CD38 lo/؊ cells into cycle, associates with CXCR4. The class II 103B2 monoclonal antibody, which binds the CD164 N-linked glycan-dependent epitope or CD164 knockdown by RNA interference, significantly inhibits the migration of CD133 ؉ HPCs toward CXCL12 in vitro. On presentation of CXCL12 on fibronectin, CD164 associates with CXCR4, an interaction that temporally follows the association of CXCR4 with the integrins VLA-4 and VLA-5. This coincides with PKC-and Akt signaling through the CXCR4 receptor, which was disrupted on the loss of CD164 though MAPK signaling was unaffected. We therefore demonstrate a novel association among 3 distinct families of cell-surface receptors that regulate cell migratory responses and identify a new role for CD164. We propose that this lends specificity to the homing and lodgment of these cells within the bone marrow niche. IntroductionAn important determinant of successful stem cell transplantation is the ability of transplanted cells to mobilize, home, migrate, and efficiently engraft and repair damaged tissues with functional cells. It is now standard practice to mobilize CD133 ϩ CD34 ϩ cells from bone marrow into the circulation by administering G-CSF or to collect such cells from umbilical cord blood and to use these for transplantation. [1][2][3][4] Once administered, these cells home to the bone marrow, where they engraft in specific stromal or vascular niches. 5 Homing begins with the chemoattraction of CD133 ϩ cells to the bone marrow and progresses to their extravasation across bone marrow sinusoidal endothelium and their transmigration through the basal lamina to specific hematopoietic stem cell/hematopoietic progenitor cell (HSC/HPC) niches. [5][6][7][8][9][10][11][12][13][14][15][16] The chemokine CXCL12 plays a central role as a chemoattractant for CD133 ϩ HSCs/HPCs, regulating their motility, homing to, and retention, survival, and proliferation in the bone marrow (for reviews, see Burger and Kipps,12 Broxmeyer et al, 17 Kollet et al, 18 and Kim et al 19 ). CXCL12 is the ligand for CXCR4, a 7-transmembrane G-protein-coupled receptor (for reviews, see Burger and Kipps 12 and Zou et al 14 ). Although lethal in the perinatal period, identical and significant reductions in B-lymphopoiesis and myelopoiesis have been described during fetal development in mice deficient in CXCL12 and CXCR4. 20,21 B-lymphopoiesis and myelopoiesis are both regulated by the interactions of their precursors, with specific microenvironmental niches within the bone mar...

Although the use of embryonic stem cells in the assisted repair of musculoskeletal tissues holds promise, a direct comparison of this cell source with adult marrow-derived stem cells has not been undertaken. Here we have compared the osteogenic differentiation potential of human embryonic stem cells (hESC) with human adult-derived stem cells in vivo. hESC lines H7, H9, the HEF-1 mesenchymal-like, telomerized H1 derivative, the human embryonic kidney epithelial cell line HEK293 (negative control), and adult human mesenchymal stem cells (hMSC) were either used untreated or treated with osteogenic factors for 4 days prior to injection into diffusion chambers and implantation into nude mice. After 11 weeks in vivo chambers were removed, frozen, and analyzed for evidence of bone, cartilage, and adipose tissue formation. All hESCs, when pretreated with osteogenic (OS) factors gave rise exclusively to bone in the chambers. In contrast, untreated hESCs (H9) formed both bone and cartilage in vivo. Untreated hMSCs did not give rise to bone, cartilage, or adipose tissue in vivo, while pretreatment with OS factors engendered both bone and adipose tissue. These data demonstrate that hESCs exposed to OS factors in vitro undergo directed differentiation toward the osteogenic lineage in vivo in a similar fashion to that produced by hMSCs. These findings support the potential future use of hESC-derived cells in regenerative medicine applications.

From 2006 to 2011, Roslin Cells Ltd derived 17 human embryonic stem cells (hESC) while developing (RCM1, RC-2 to -8, -10) and implementing (RC-9, -11 to -17) quality assured standards of operation in a facility operating in compliance with European Union (EU) directives and United Kingdom (UK) licensure for procurement, processing and storage of human cells as source material for clinical application, and targeted to comply with an EU Good Manufacturing Practice specification. Here we describe the evolution and specification of the facility, its operation and outputs, complementing hESC resource details communicated in Stem Cell Research Lab Resources.

The promise of human embryo stem cells (hESCs) for regenerative medicine is offset by the ethical and practical challenges involved in sourcing eggs and embryos for this objective. In this study we sought to isolate an hESC line from clinically failed eggs, the usage of which would not conflict with donor interests to conceive. A total of 8 blastocysts were allocated for hESC derivation from a pool of 579 eggs whose fertilization had been clinically assessed to have occurred abnormally (i.e., three pronuclei) or failed (i.e., no pronuclei) following in vitro insemination or intracytoplasmic sperm injection (ICSI). The latter were subjected to a recovery intervention consisting of either reinsemination by ICSI or parthenogenetic stimulation. One hESC line (RCM1) was obtained from a failed-to-fertilize inseminated egg recovered by parthenogenetic activation. Standard in vitro and in vivo characterization revealed this line to possess all of the properties attributed to a normal euploid hESC line. Whole-genome single-nucleotide polymorphism analysis further revealed that the line was biparental, indicating that sperm penetration had occurred, although parthenogenetic stimulation was required for activation. Our results demonstrate the viability of an alternative strategy to generate normal hESC lines from clinically failed eggs, thereby further minimizing the potential to conflict with donor reproductive interest to conceive.