Umbilical cord blood (UCB) represents a readily available source of hematopoietic and endothelial precursors at early ontogeny. Understanding the proangiogenic functions of these somatic progenitor subtypes after transplantation is integral to the development of improved cell-based therapies to treat ischemic diseases. We used fluorescence-activated cell sorting to purify a rare (<0.5%) population of UCB cells with high aldehyde dehydrogenase (ALDH hi ) activity, a conserved stem/progenitor cell function. ALDH hi cells were depleted of mature monocytes and T-and B-lymphocytes and were enriched for early myeloid (CD33) and stem cell-associated (CD34, CD133, and CD117) phenotypes.

Transplanted murine bone marrow (BM) progenitor cells recruit to the injured pancreas and induce endogenous beta cell proliferation to improve islet function. To enrich for analogous human progenitor cell types that stimulate islet regeneration, we purified human BM based on high-aldehyde dehydrogenase activity (ALDH(hi)), an enzymatic function conserved in hematopoietic, endothelial, and mesenchymal progenitor lineages. We investigated the contributions of ALDH(hi) mixed progenitor cells or culture-expanded, ALDH-purified multipotent stromal cell (MSC) subsets to activate endogenous programs for islet regeneration after transplantation into streptozotocin-treated NOD/SCID mice. Intravenous injection of uncultured BM ALDH(hi) cells improved systemic hyperglycemia and augmented insulin secretion by increasing islet size and vascularization, without increasing total islet number. Augmented proliferation within regenerated endogenous islets and associated vascular endothelium indicated the induction of islet-specific proliferative and pro-angiogenic programs. Although cultured MSC from independent human BM samples showed variable capacity to improve islet function, and prolonged expansion diminished hyperglycemic recovery, transplantation of ALDH-purified regenerative MSC reduced hyperglycemia and augmented total beta cell mass by stimulating the formation of small beta cell clusters associated with the ductal epithelium, without evidence of increased islet vascularization or Ngn3(+) endocrine precursor activation. Thus, endogenous islet recovery after progenitor cell transplantation can occur via distinct regenerative mechanisms modulated by subtypes of progenitor cells administered. Further, understanding of how these islet regenerative and pro-angiogenic programs are activated by specific progenitor subsets may provide new approaches for combination cellular therapies to combat diabetes.

Transplanted human bone marrow (BM) and umbilical cord blood (UCB) progenitor cells activate islet-regenerative or revascularization programs depending on the progenitor subtypes administered. Using purification of multiple progenitor subtypes based on a conserved stem cell function, high aldehyde dehydrogenase (ALDH) activity (ALDH(hi)), we have recently shown that transplantation of BM-derived ALDH(hi) progenitors improved systemic hyperglycemia and augmented insulin secretion by increasing islet-associated proliferation and vascularization, without increasing islet number. Conversely, transplantation of culture-expanded multipotent-stromal cells (MSCs) derived from BM ALDH(hi) cells augmented total beta cell mass via formation of beta cell clusters associated with the ductal epithelium, without sustained islet vascularization. To identify paracrine effectors produced by islet-regenerative MSCs, culture-expanded BM ALDH(hi) MSCs were transplanted into streptozotocin-treated nonobese diabetic/severe combine immune deficient (SCID) mice and segregated into islet-regenerative versus nonregenerative cohorts based on hyperglycemia reduction, and subsequently compared for differential production of mRNA and secreted proteins. Regenerative MSCs showed increased expression of matrix metalloproteases, epidermal growth factor receptor (EGFR)-activating ligands, and downstream effectors of Wnt signaling. Regenerative MSC supernatant also contained increased levels of pro-angiogenic versus pro-inflammatory cytokines, and augmented the expansion of ductal epithelial but not beta cells in vitro. Conversely, co-culture with UCB ALDH(hi) cells induced beta cell but not ductal epithelial cell proliferation. Sequential transplantation of MSCs followed by UCB ALDH(hi) cells improved hyperglycemia and glucose tolerance by increasing beta cell mass associated with the ductal epithelium and by augmenting intra-islet capillary densities. Thus, combinatorial human progenitor cell transplantation stimulated both islet-regenerative and revascularization programs. Understanding the progenitor-specific pathways that modulate islet-regenerative and revascularization processes may provide new approaches for diabetes therapy.

Breast cancers expressing human embryonic stem cell (hESC)-associated genes are more likely to progress than well-differentiated cancers and are thus associated with poor patient prognosis. Elevated proliferation and evasion of growth control are similarly associated with disease progression, and are classical hallmarks of cancer. In the current study we demonstrate that the hESC-associated factor Nodal promotes breast cancer growth. Specifically, we show that Nodal is elevated in aggressive MDA-MB-231, MDA-MB-468 and Hs578t human breast cancer cell lines, compared to poorly aggressive MCF-7 and T47D breast cancer cell lines. Nodal knockdown in aggressive breast cancer cells via shRNA reduces tumour incidence and significantly blunts tumour growth at primary sites. In vitro, using Trypan Blue exclusion assays, Western blot analysis of phosphorylated histone H3 and cleaved caspase-9, and real time RT-PCR analysis of BAX and BCL2 gene expression, we demonstrate that Nodal promotes expansion of breast cancer cells, likely via a combinatorial mechanism involving increased proliferation and decreased apopotosis. In an experimental model of metastasis using beta-glucuronidase (GUSB)-deficient NOD/SCID/mucopolysaccharidosis type VII (MPSVII) mice, we show that although Nodal is not required for the formation of small (<100 cells) micrometastases at secondary sites, it supports an elevated proliferation:apoptosis ratio (Ki67:TUNEL) in micrometastatic lesions. Indeed, at longer time points (8 weeks), we determined that Nodal is necessary for the subsequent development of macrometastatic lesions. Our findings demonstrate that Nodal supports tumour growth at primary and secondary sites by increasing the ratio of proliferation:apoptosis in breast cancer cells. As Nodal expression is relatively limited to embryonic systems and cancer, this study establishes Nodal as a potential tumour-specific target for the treatment of breast cancer.

Peripheral arterial disease is a clinical problem in which mesenchymal stromal cell (MSC) transplantation may offer substantial benefit by promoting the generation of new blood vessels and improving limb ischemia and wound healing via their potent paracrine activities. MRI allows for the noninvasive tracking of cells over time using iron oxide contrast agents to label cells before they are injected or transplanted. However, a major limitation of the tracking of iron oxide-labeled cells with MRI is the possibility that dead or dying cells will transfer the iron oxide label to local bystander macrophages, making it very difficult to distinguish between viable transplanted cells and endogenous macrophages in the images. In this study, a severely immune-compromised mouse, with limited macrophage activity, was investigated to examine cell tracking in a system in which bystander cell uptake of dead, iron-labeled cells or free iron particles was minimized. MRI was used to track the fate of MSCs over 21 days after their intramuscular transplantation in mice with a femoral artery ligation. In all mice, a region of signal loss was observed at the injection site and the volume of signal hypointensity diminished over time. Fluorescence and light microscopy showed that iron-positive MSCs persisted at the transplant site and often appeared to be integrated in perivascular niches. This was compared with MSC transplantation in immune-competent mice with femoral artery ligation. In these mice, the regions of signal loss caused by iron-labeled MSC cleared more slowly, and histology revealed iron particles trapped at the site of cell transplantation and associated with areas of inflammation.