Neural stem cells transplanted into intact brains as neurospheres form solid grafts composed of neurons, astrocytes and oligodendrocyte precursors

Neural stem cells (NSCs) are tissue-specific stem cells with self-renewal potential that can give rise to neurons and glia in vivo and in vitro. The aim of this study was to transplant NSCs as whole neurospheres into intact brain and assess the fate and phenotype of their progeny generated in vivo. We isolated NSCs from E14 foetal rat forebrains and cultured them in basic fibroblast and epidermal growth factor-supplemented serum-free medium in the form of neurospheres in vitro. Neurospheres were transplanted into the intact brains of 2 Wistar rats and after a period of 3 weeks, grafted brains were examined immunohistochemically. Neurospheres formed solid grafts that were found in the lateral ventricle and in the velum interpositum under the hippocampus. The majority of cells in the transplanted tissue were identified as beta-III-tubulin(+), NeuN(+), PanNF(+) and synaptophysin(+) neurons and were accumulated throughout the graft centre. GFAP(+) astrocytes were scattered throughout the entire graft and astrocyte processes delimited the outer and perivascular surfaces. A great number of NG2(+) oligodendrocyte precursors was detected. Nestin(+) endothelial cells were found to line capillaries growing in the transplant. These data indicate that nestin(+) NSCs prevailing in neurospheres differentiate following transplantation into nestin(-) neuronal and glial cells which confirms the multipotency of NSCs. Three weeks posttransplantation neuronal and astrocyte cells reached terminal differentiation (formation of synaptic vesicles and superficial and perivascular limiting membranes) while elements of oligodendroglial cell lineage remained immature. Grafting stem cells as non-dissociated neurospheres provide cells with favourable conditions which facilitate cell survival, proliferation and differentiation. However, in the intact brain, grafted neurosphere cells were not found to integrate with the brain parenchyma and formed a compact structure demarcated from its surroundings.     

Differentiation of neurosphere-derived rat neural stem cells into oligodendrocyte-like cells by repressing PDGF-α and Olig2 with triiodothyronine

One of the approaches for treating demyelination diseases is cytotherapy, and adult stem cells are potential sources. In this investigation, we tried to increase the yield of oligodendrocyte-like cells (OLCs) by inducing neural stem cells generated from BMSCs-derived neurospheres, which were used for deriving the neural stem cells (NSCs). The latter were induced into OLCs by heregulin, PDGF-AA, bFGF and triiodothyronine (T3). The BMSCs, NS, NSCs and OLCs were characterized by using immunocytochemistry for fibronectin, CD44, CD90, CD45, Oct-4, O4, Olig2, O1 and MBP markers. PDGF receptor α (PDGFR-α), Olig2 and MOG expression were evaluated by RT-PCR. The BMSCs expressed CD44, CD90, CD106 and Oct-4; the NSCs were immunoreactive to nestin and neurofilament 68. Incubation of the NSCs for 4 days with heregulin, PDGF-AA and bFGF resulted in their induction into oligodendrocyte progenitor-like cells (OPLCs), which immunoreacted to O4, Olig2 and O1, while Olig2 and PDGFR-α were detected by RT-PCR. Replacing heregulin, PDGF-AA and bFGF with T3 for 6 days resulted in repression of O4, O1, Olig2 and PDGFR-α. The OLCs were co-cultured with motoneurons resulted in induction of MOG and MBP, which were expressed in functional OLCs. The latter can be generated from BMSCs-derive NS with high yield.     

Cultured Rat Astrocytes Give Rise to Neural Stem Cells

Previously, we reported the occurrence of neural stem cells (NSCs) around an area of damage after rat traumatic brain injury (TBI), but it was unclear if this was due to blastgenesis in astrocytes, or to NSCs migrating from the subventricular zone (SVZ). In this study, NSCs were isolated and cultured from cultured type 1 astrocytes taken from newborn rat cortex in which the subventricular zone and hippocampus had been discarded. All cultured type 1 astrocytes showed glial fibrillary acidic protein (GFAP) immunopositivity. Nestin immunopositive spheres were isolated from type 1 astrocytes and cultured in the presence of bFGF and EGF in the medium. Neurospheres differentiated into Tuj1-, GFAP- and A2B5-positive cells after 4 days of culture without bFGF and EGF. These results indicate that isolated neurospheres from brain cortex astrocytes can differentiate into neurons and glia and might contribute to neurogenesis and neuroplasticity.     

Competent dendritic cells derived from CD34+ progenitors express CMRF-44 antigen early in the differentiation pathway

Differentiation of CD34(+) haematopoietic stem cells into functional dendritic cells (DC) was investigated using the mAb CMRF-44 and other mAb against DC-associated markers. GM-CSF mobilized peripheral blood stem cells were obtained from healthy donors by leukapheresis. CD34(+) cells were purified using CD34(+)-positive selection,and subsequent immunomagnetic depletion of CD14 and CD2 cells. CD34(+) cells were cultured in medium supplemented with one or more of GM-CSF,TNF-alpha, IL-4 or IL-6. CMRF-44 Ag expression was monitored by flow cytometry, and DC function by allogeneic MLR and tetanus toxoid(TT) presentation assays. CD34(+) cells quickly acquired the CMRF-44 Ag when cultured in the presence of TNF-alpha.By day 3, more than 50% of the cells were double-positive for CD34 and CMRF-44. CD34 expression was gradually lost, so that by day 9, the majority of the cells were CD34(-)/CMRF-44(+).GM-CSF and TNF-alpha also induced CD40 expression, and up-regulation of CD54 and MHC class II on CD34(+) cells; their expression was correlated to the CMRF-44 Ag. Day 3 CD34(+)/CMRF-44(+) cells,but not CD34(+)/CMRF-44(-) cells, become potent APC when cultured further with GM-CSF plus TNF-alpha. These CMRF-44(+) cells were potent inducers of Th1-type immune response in the primary allogeneic MLR and present TT to autologous CD4(+) T cells. TNF-alpha alone is sufficient to induce CMRF-44 expression on CD34(+) cells, but in combination with GM-CSF expands the CMRF-44(+) population. CMRF-44 expression correlates with DC function and may be a useful early marker for commitment of CD34(+) cells to the DC differentiation pathway.     

Distinct hemogenic potential of endothelial cells and CD41+ cells in mouse embryos

Definitive hematopoietic progenitor cells have been thought to develop from the vascular endothelium located in the aorta-gonad-mesonephros region of the mouse embryo. However, several recent findings have suggested that most hematopoietic progenitors are derived from non-endothelial precursor cells expressing CD41. We characterized two distinct precursor populations of definitive hematopoietic cell lineages, vascular endothelial (VE)-cadherin(+) CD41(-) CD45(-) endothelial cells and CD41(+) CD45(-) non-endothelial progenitors, both of which are derived from lateral mesoderm. VE-cadherin(+) endothelial cells obtained from cultures of differentiating embryonic stem cells possessed hematopoietic potential encompassing erythroid, myeloid and B lymphoid lineages, whereas CD41(+) progenitors lacked the B lymphopoietic potential. VE-cadherin(+) endothelial cells in the lower trunk of the embryo proper showed a significant potential for initiating B lymphopoiesis in cultures, while endothelial cells in the yolk sac appeared to have a bias for myeloerythropoietic differentiation. CD41(+) progenitors isolated from yolk sac and embryo proper were capable of generating multiple hematopoietic lineages, although mast cell precursors were exclusively enriched in CD41(+) progenitors in the yolk sac. These results suggest that hemogenic endothelial cells and CD41(+) progenitors possess distinct hematopoietic potential depending on the tissues in which they reside.     

A comparison of CFU-GM, BFU-E and endothelial progenitor cells using ex vivo expansion of selected cord blood CD133(+) and CD34(+) cells

BACKGROUND: CD133 is a newly developed hematopoietic stem cell marker but little is known about its function. Whether CD133(+) cell selection provides any advantage over CD34(+) selection for hematopoietic stem cell isolation and transplantation is unclear. The present study compared colony formation and endothelial cell differentiation of these two cell types from umbilical cord blood (UCB). METHODS: Mononuclear cells from the same UCB samples were used for both CD133(+) and CD34(+) cell selection. Cells with 97.1% purity were incubated in semi-solid culture medium containing stem cell growth factor (SCGF) and G-CSF or erythropoietin (EPO). Purified cells were also cultured in M199 containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and insulin-like growth factor-1 (IGF-1). RESULTS: CD34(+) and CD133(+) cells produced similar numbers of CFU-GM colonies (median 43.25 and 30.5, respectively; P>0.2). However, a greater than four-fold difference in BFU-E colony formation was observed from CD34(+) cells compared with CD133(+) cells (median 35 and 8, respectively; P<0.04). CD34(+) cells gave rise to endothelial-like cells when stimulated with VEGF, bFGF and IGF-1. CD133(+) cells were unable produce this cell type under the same conditions. DISCUSSION: CD133(+) cells produced smaller BFU-E colonies and were unable to differentiate into mature endothelial cells. CD34(+) cells contained endothelial progenitors that could differentiate into mature cells of this lineage. Based on these data, it appears that CD133 offers no distinct advantage over CD34 as a selective marker for immunoaffinity-based isolation of hematopoietic stem cells and endothelial progenitor cells.     

Immunocytochemical analysis of valproic acid induced histone H3 and H4 acetylation during differentiation of rat adipose derived stem cells into neuron-like cells

Valproic acid (VPA) is an inhibitor of histone deacetylases (HDACs) that can regulate differentiation and proliferation of stem cells by epigenetic mechanisms. We investigated VPA induced histone H3 and H4 acetylation in adipose derived stem cells (ADSCs) transdifferentiated into neuron-like cells (NLCs). Rat ADSCs were transdifferentiated into neural stem cells (NSCs) that had been generated from neurospheres. The NSCs were differentiated into NLCs by induction with different concentrations of VPA at 24, 48 and 72 h. The NLCs were evaluated using anti-H3 and -H4 antibodies, and ADSCs, NSCs and NLCs were evaluated using immunofluorescence. The ADSCs were immunoreactive to CD90 and CD49d, but not to CD45 and CD31. Both the neurospheres and NSCs were immunostained with nestin and neurofilament 68. The neurospheres expressed Musashi1, Sox2 and Neu N genes as determined by RT-PCR. Our dose-response study indicated that the optimal concentration of VPA was 1 mM at 72 h. Histone acetylation levels of H3 and H4 immunostaining intensities in NLCs were significantly greater than for ADSCs and NSCs. VPA alters H4 and H3 acetylation immunoreactivities of ADSCs transdifferentiated into NLCs.     

Isolation and characterization of the CD133+ precursors from the ventricular zone of human fetal brain by magnetic affinity cell sorting

A fast and effective method to enrich large number of neural precursors from the ventricular zone of human fetus by magnetic affinity cell sorting (MACS) is reported. After incubation with phycoerythrin (PE)-conjugated anti-CD133 antibodies and anti-PE magnetic beads followed by one cycle of MACS, CD133(+) cells were harvested at 85% purity as confirmed by flow-cytometry and immunocytochemistry. In contrast to CD133(-) cells, these CD133(+) cells initiated primary and secondary neurospheres in culture, and the progeny of sorted cells could be differentiated into both neurons and glia, indicating that these highly enriched cells are capable of self-renewal and multi-lineage potential.     

Characterization of CD133+ hepatocellular carcinoma cells as cancer stem/progenitor cells

The CD133 antigen, identified as a hematopoietic stem cell marker, appears in various human embryonic epithelia including the neural tube, gut, and kidney. We herein investigated whether CD133(+) cells isolated from human hepatocellular carcinoma cell lines possess cancer stem/progenitor cell-like properties. Among the three cell lines studied, the CD133 antigen was found to be expressed only on the surface of Huh-7 cells. CD133(+) cells from Huh-7 performed a higher in vitro proliferative potential and lower mRNA expressions of mature hepatocyte markers, glutamine synthetase and cytochrome P450 3A4, than CD133(-) population of Huh-7 cells. When either CD133(+) or CD133(-) cells were subcutaneously injected into SCID mice, CD133(+) cells formed tumors, whereas CD133(-) cells induced either a very small number of tumors or none at all. Taken together, the identification of CD133(+) cells could thus be a potentially powerful tool to investigate the tumorigenic process in the hepatoma system and to also develop effective therapies targeted against hepatocellular carcinoma.     

Human CD34+/CD90+ ASCs Are Capable of Growing as Sphere Clusters,Producing High Levels of VEGF and Forming Capillaries

Background

Human adult adipose tissue is an abundant source of mesenchymal stem cells (MSCs). Moreover, it is an easily accessible site producing a considerable amount of stem cells.

Methodology/Principal Findings

In this study, we have selected and characterized stem cells within the stromal vascular fraction (SVF) of human adult adipose tissue with the aim of understanding their differentiation capabilities and performance. We have found, within the SVF, different cell populations expressing MSC markers – including CD34, CD90, CD29, CD44, CD105, and CD117 – and endothelial-progenitor-cell markers – including CD34, CD90, CD44, and CD54. Interestingly, CD34⁺/CD90⁺ cells formed sphere clusters, when placed in non-adherent growth conditions. Moreover, they showed a high proliferative capability, a telomerase activity that was significantly higher than that found in differentiated cells, and contained a fraction of cells displaying the phenotype of a side population. When cultured in adipogenic medium, CD34⁺/CD90⁺ quickly differentiated into adipocytes. In addition, they differentiated into endothelial cells (CD31⁺/VEGF⁺/Flk-1⁺) and, when placed in methylcellulose, were capable of forming capillary-like structures producing a high level of VEGF, as substantiated with ELISA tests.

Conclusions/Significance

Our results demonstrate, for the first time, that CD34⁺/CD90⁺ cells of human adipose tissue are capable of forming sphere clusters, when grown in free-floating conditions, and differentiate in endothelial cells that form capillary-like structures in methylcellulose. These cells might be suitable for tissue reconstruction in regenerative medicine, especially when patients need treatments for vascular disease.     

CD90 is identified as a candidate marker for cancer stem cells in primary high-grade gliomas using tissue microarrays

Although CD90 has been identified as a marker for various kinds of stem cells including liver cancer stem cells (CSCs) that are responsible for tumorigenesis, the potential role of CD90 as a marker for CSCs in gliomas has not been characterized. To address the issue, we investigated the expression of CD90 in tissue microarrays containing 15 glioblastoma multiformes (GBMs), 19 WHO grade III astrocytomas, 13 WHO grade II astrocytomas, 3 WHO grade I astrocytomas and 8 normal brain tissues. Immunohistochemical analysis showed that CD90 was expressed at a medium to high level in all tested high-grade gliomas (grade III and GBM) whereas it was barely detectable in low-grade gliomas (grade I and grade II) and normal brains. Double immunofluorescence staining for CD90 and CD133 in GBM tissues revealed that CD133(+) CSCs are a subpopulation of CD90(+) cells in GBMs in vivo. Flow cytometry analysis of the expression of CD90 and CD133 in GBM-derived stem-like neurospheres further confirmed the conclusion in vitro. The expression levels of both CD90 and CD133 were reduced along with the loss of stem cells after differentiation. Furthermore, the limiting dilution assay demonstrated that the sphere formation ability was comparable between the CD90(+)/CD133(+) and the CD90(+)/CD133(-) populations of GBM neurospheres, which is much higher than that of the CD90(-)/CD133(-) population. We also performed double staining for CD90 and a vascular endothelial cell marker CD31 in tissue microarrays which revealed that the CD90(+) cells were clustered around the tumor vasculatures in high-grade glioma tissues. These findings suggest that CD90 is not only a potential prognostic marker for high-grade gliomas but also a marker for CSCs within gliomas, and it resides within endothelial niche and may also play a critical role in the generation of tumor vasculatures via differentiation into endothelial cells.     

CD31 (PECAM-1)-bright cells derived from AC133-positive cells in human peripheral blood as endothelial-precursor cells

To clarify the process of endothelial differentiation, we isolated AC133(+) cells and induced the in vitro differentiation of these cells into endothelial cells. AC133(+) cells efficiently differentiated into endothelial cells when the cells were cultured on fibronectin-coated dishes in the presence of vascular endothelial growth factor. Time-course analysis of the alteration of endothelial markers on cultured AC133(+) cells revealed that the expression of CD31 (PECAM-1) on AC133(+) cells was the earliest marker among all of the tested markers. Based on the hypothesis that CD31 is an early indicator during the endothelial differentiation, we examined the relationship between CD31 expression and the ability to differentiate into endothelial cells in cells derived from AC133(+) cells. CD31-bright cells, which were sorted from cultured AC133(+) cells, differentiated more efficiently into endothelial cells than had CD31-positive or CD31-negative cells, suggesting that CD31-bright cells may be precursor cells for endothelial cells. In the present study, we identified CD31(+) cells derived from cultured AC133(+) cells that are able to differentiate to endothelial cells as precursor cells.     

Cell surface antigens on rat neural progenitors and characterization of the CD3 (+)/CD3 (-) cell populations

While the hematopoietic lineage has been extensively studied using cluster of differentiation (CD) antibodies, very few data are available on the extracellular epitopes expressed by rat neural progenitors (rNPC) and their derivatives. In the present study, we used flow cytometry to screen 47 cell surface antigens, initially known as immune markers. The quantitative analyses were performed on rat neurospheres and compared with primary cultures of astroglial cells or cerebellar neurons. Several antigens such as CD80 or CD86 were clearly undetectable while others, like CD26 or CD161, showed a weak expression. Interestingly, 10% and 15% of the cells were immunopositive for CD172a and CD200, two immunoglobulin superfamily members preferentially expressed by glial or neuronal cells, respectively. Over 40% of the cells were immunopositive for CD3, CD71, or MHCI. The biological significance of the latter markers in rNPC remains to be determined but analyses of the CD3(-)/CD3(+) populations isolated by magnetic cell separation revealed differences in their cell fate. Indeed, CD3(+) cells did not establish neurospheres and differentiated mostly into GFAP(+) cells while CD3(-) cells were able to generate neurospheres upon mitogen treatment and gave rise to GFAP(+), A2B5(+), Tuj-1(+), and RIP(+) cells under differentiating conditions. In contrast, CD71(-)/CD71(+) cells did not show any significant difference in their proliferating and differentiating potentials. Finally, it is worth noting that an subpopulation of cells in rat neurospheres exhibit an immunoreactivity against anti-CD25 (IL2 receptor) and anti-CD62L (L-selectin) antibodies. The results reveal particular surface antigen profiles, giving new perspectives on the properties of rat brain-derived cells.     

BM cells giving rise to MSC in culture have a heterogeneous CD34 and CD45 phenotype

BACKGROUND: Mesenchymal stromal cells (MSC) isolated from adult human BM are characterized by their fibroblast-like morphology, adherent growth and capacity to differentiate into adipocytes, osteocytes, chondrocytes, cardiomyocytes and neuroprogenitors. After culturing these cells in vitro, they express the cell-surface molecules CD44, CD90, SH2 and SH3, and are negative for CD34 and the hematopoietic marker CD45. The aim of this study was to characterize the in vivo phenotype of MSC relative to the expression of CD34 and CD45. METHODS: BM mononuclear cells were stained with Ab against both molecules and separated into the CD34(+), CD34(-), CD45(+) CD34(+), CD45(high+) CD34(-), CD45(med,low+) CD34(-) and CD45(-) CD34(-) subpopulations, which were then cultured under the same conditions and analyzed for growth of MSC. RESULTS: A small population of MSC arose from the CD45(+) CD34(+) fraction, although the majority was obtained from the CD45(-) CD34(-) subpopulation. MSC from all fractions could be differentiated into adipocytes and osteocytes. In addition, MSC from the CD34(+) and CD34(-) fractions were shown to differentiate into chondrocytes. After in vitro culture, MSC from both fractions possessed the same phenotype, which was negative for CD34 and CD45. DISCUSSION: MSC from the CD45(+) CD34(+) fraction change their phenotype under in vitro conditions.     

Adiponectin promotes endothelial cell differentiation from human peripheral CD14+ monocytes in vitro

Adiponectin was revealed to have anti-atherogenic and anti-inflammatory properties and has been recently found to stimulate angiogenesis in vivo and in vitro. However, the role of adiponectin in endothelial differentiation remains unclear. The objective of this study was to investigate whether adiponectin can promote peripheral CD14(+) monocytes differentiation into endothelial cells (ECs). Human peripheral blood CD14(+) monocytes were cultured with or without adiponectin (10 microg/ml) for 10 days. Adiponectin significantly promoted EC morphology formation from CD14(+) monocytes. By flow cytometery analysis, cells treated with adiponection substantially increased mean fluorescence intensity of vascular endothelial growth factor receptor-2 (VEGFR-2) and endothelial nitric oxide synthase (eNOS), two specific endothelial markers, by 49.2 % and 53.9 %, respectively, as compared to control cells. By real time PCR analysis, the mRNA level of eNOS in adiponectin-treated cells was also increased by 31.9 % of that of the control cells. However, the mRNA levels of calponin and SMMHC, two specific SMC markers, in adiponectin-treated cells were decreased by 81.1 % and 79.7 % of that of the control cells, respectively. These data demonstrated that adiponectin could promote endothelial differentiation from peripheral blood CD14(+) monocytes by morphology change, upregulation of EC markers and downregulation of SMC markers. Adiponectin-promoted EC differentiation may contribute to vascular healing and angiogenesis.     

The critical role of CD133(+)CD44(+/high) tumor cells in hematogenous metastasis of liver cancers

Metastatic hepatocellular carcinoma (HCC) is one of the most lethal cancers worldwide. However, the cell population responsible for its metastasis remains largely unknown. Here, we reported that CD133(+)CD44(+/high) defined a subgroup of tumor cells that was responsible for hematogenous metastasis of liver cancers. Immunohistochemical investigation of human HCC specimens revealed that the number of CD133(+) and CD44(+) HCC cells was increased and was associated with portal vein invasion. Purified CD133(+) or CD44(high) HCC cells were superior in clonogenic growth and vascular invasion, respectively. Thus, the combination of CD133 and CD44 was used to define a novel HCC sub-population. CD133(+)CD44(high), but not CD133(+)CD44(low/-), CD133(-)CD44(high) or CD133(-)CD44(low/-) xenografts, produced intrahepatic or lung metastasis in nude mice. Further analysis of human HCC samples by flow cytometry showed that the number of CD133(+)CD44(+) tumor cells was associated with portal vein metastasis. The cDNA microarray analysis of CD133(+)CD44(+) and CD133(+)CD44(-) tumor cells isolated from metastatic HCC patients revealed that these cells comprised of two different populations possessing distinct gene expression profiles. Our results suggest that CD133(+)CD44(+) tumor cells are a particular population responsible for hematogenous metastasis in liver cancers and that these cells might be targets for treatment of HCC metastasis.     

Successful elimination of memory-type CD8+ T cell subsets by the administration of anti-Gr-1 monoclonal antibody in vivo

During investigating the expression of Gr-1 antigen on various subsets of mouse spleen cells, we found that Gr-1 was expressed on memory-type CD8(+)CD44(high)CD62L(high) T cells in addition to granulocytes. Intraperitoneal administration of anti-Gr-1 mAb caused almost complete elimination of Ly-6C(+) memory-type CD8(+) T cells as well as Ly-6G(+) granulocytes. Anti-Gr-1 mAb-treated mouse spleen cells exhibited greatly reduced IFN-gamma production in response to anti-CD3 mAb both in vitro and in vivo. This reduced cytokine production appeared to be derived from elimination of IFN-gamma-producing Gr-1(+)CD8(+) T cells. Indeed, CD8(+) T cells with IFN-gamma-producing activity and cytotoxicity were generated from isolated Gr-1(+)CD8(+) cells but not from Gr-1(-)CD8(+) T cells. We also demonstrated that therapeutic effect of MBL-2 tumor-immunized spleen cells was greatly reduced by anti-Gr-1 mAb-treatment. Thus, we initially demonstrated that anti-Gr-1 mAb might become a good tool to investigate a precise role for memory-type CD8(+) T cells in vivo.     

Vascular wall-resident CD44+ multipotent stem cells give rise to pericytes and smooth muscle cells and contribute to new vessel maturation

Here, we identify CD44(+)CD90(+)CD73(+)CD34(−)CD45(−) cells within the adult human arterial adventitia with properties of multipotency which were named vascular wall-resident multipotent stem cells (VW-MPSCs). VW-MPSCs exhibit typical mesenchymal stem cell characteristics including cell surface markers in immunostaining and flow cytometric analyses, and differentiation into adipocytes, chondrocytes and osteocytes under culture conditions. Particularly, TGFß1 stimulation up-regulates smooth muscle cell markers in VW-MPSCs. Using fluorescent cell labelling and co-localisation studies we show that VW-MPSCs differentiate to pericytes/smooth muscle cells which cover the wall of newly formed endothelial capillary-like structures in vitro. Co-implantation of EGFP-labelled VW-MPSCs and human umbilical vein endothelial cells into SCID mice subcutaneously via Matrigel results in new vessels formation which were covered by pericyte- or smooth muscle-like cells generated from implanted VW-MPSCs. Our results suggest that VW-MPSCs are of relevance for vascular morphogenesis, repair and self-renewal of vascular wall cells and for local capacity of neovascularization in disease processes.