Multipotent Differentiation Potential Research Articles

Glioblastoma stem cells produce vascular endothelial growth factor by activation of a G‐protein coupled formylpeptide receptor FPR

Glioma stem cells (GSCs), or stem cell-like glioma cells, isolated from malignant glioma cell lines, were capable of producing vascular endothelial growth factor (VEGF). However, the exact role of such tumour cells in angiogenesis remains unknown. In this study, we isolated a small proportion of CD133+ GSCs from the human glioblastoma cell line U87 and found that these GSCs possessed multipotent differentiation potential and released high levels of VEGF as compared with CD133(-) tumour cells. The CD133+ GSCs also formed larger xenograft tumours that contained higher VEGF immunoreactivity and denser microvessels. Moreover, GSCs expressed a functional G protein-coupled formylpeptide receptor FPR, which was activated by a chemotactic peptide ligand, N-formylmethionyl-leucyl-phenylalanine (fMLF), to mediate calcium flux and the production of VEGF by GSCs. Our results indicate that FPR expressed by human GSCs may play an important role in glioma angiogenesis.

Neural Induction from ES Cells Portrays Default Commitment but Instructive Maturation

The neural induction has remained a debatable issue pertaining to whether it is a mere default process or it involves precise instructive cues. We have chosen the embryonic stem (ES) cell model to address this issue. In a devised monoculture strategy, the cell–cell interaction availed through optimum cell plating density could define the niche for the attainment of efficient in vitro neurogenesis from the ES cells. The medium plating density was found ideal in generating optimum number of progenitors and also yielded about 80% mature neurons in a serum free culture set up barring any exogenous inducers. We could also demarcate and quantify the neural stem cells/progenitors among the heterogeneous cell population of differentiating ES cells using nestin intron II driven EGFP expression as a tool. The one week post-plating was determined to be the critical time window for optimum neural progenitor generation from ES cells that helped us further in purifying these cells and in demonstrating their proliferation and multipotent differentiation potential. Seeding cells at varying densities, we could decipher an interesting paradoxical scenario that interlinked both commitment and maturation with the initial plating density having a vital influence on neuronal maturation but not specification and the secretory factors were apparently playing a key role during this process. Thus it was comprehended that, the neural specification was a default process independent of exogenous factors and cellular interaction. Conversely, a defined number of cells at the specification stage itself seemed critical to provide an auto-/paracrine means of signaling threshold for the maturation process to materialize.

"In vitro" and multicolor phenotypic characterization of cell subpopulations identified in fresh human adipose tissue stromal vascular fraction and in the derived mesenchymal stem cells

BackgroundThe stromal vascular fraction (SVF) is a heterogeneous cell population derived from the adipose tissue. There is still a lack of information concerning the characterization of the cell subpopulations constituting the SVF as well as its mesenchymal and haematopoietic potential. Furthermore there are great variations in its phenotypical characterization.MethodsComposition of SVF was investigated by FACS analysis, cytological and "in vitro" assays. We studied CD34+ population by combining FACS with human CFC (colony-forming-cell haematopoietic assay). The endothelial fraction was investigated by quantifying the co-expression of specific markers (CD146, CD105, CD31 and UEA-1). Mesenchymal potential was assessed by CFU-F assay and cultured AT-MSC were characterized by a 5-color FACS analysis. The multipotent differentiation potential (osteogenic, adipogenic and chondrogenic) was investigated both at cellular and molecular level.ResultsWe identified in the SVF two CD34+ populations with a marked difference in the intensity of antigen expression, the majority of the cells expressing CD34 at low intensity. Moreover, two CD146+ cell populations were clearly distinguishable in the SVF:a CD146 dim accounting for 9.9% of the total SVF cells and a CD146+ bright cell population accounting for about 39.3%. The frequency of CFC clones was comparable with the one reported for peripheral blood. Endothelial cells account for about 7.7% of the SVF cells. AT-MSC differenced in the osteogenic adipogenic and chondrogenic lineage.ConclusionThe SVF is not a homogeneous cell population, and its final composition could be influenced both by the flow cytometric technique analysis and the SVF extraction steps. The CFU-F frequency in the SVF was 1/4880, a value about seven times greater than the data reported for bone marrow. The antigenic profile of AT-MSC was comparable with bone-marrow derived MSC. AT-MSC were able to differentiate along the osteogenic adipogenic and chondrogenic lineages. The data here reported, further contribute to the characterization of SVF, a tissue providing an alternative as a source of MSC for clinical applications.

Multipotent stem cells from the young rat inner ear

The terminal mitosis of hair cells (HCs) and supporting cells (SCs) in mammalian cochlea occurred during middle embryonic development. Most hearing loss results from the incapacity of the cochlear sensory epithelium to replace lost hear cells. Deafness due to hair cells loss is normally irreversible. The present study showed that cells acutely dissociated from the cochlea of young rat, cultured with EGF and FGF2, developed into otospheres that showed expression of nestin and incorporation of 5′-Bromo-2-deoxyuridine (BrdU). The subcultured otospheres maintained for up to 10 passages. In addition, the cochlea sphere-derivatives contributed to a variety of cell types. They were found to differentiate to neuron, glia, hair cell and supporting cell phenotypes. The results suggest that the young rat inner ear cells have self-renewal capability and multipotent differentiation potential. This work raises the possibility that inner ear cells in the early post-natal rat have the character of pluripotent stem cells and might be a source for cell replacement therapy in the inner ear.

Isolation and characterization of mesenchymal cells from human fetal membranes

Bone marrow (BM) multipotent mesenchymal stromal cells (MSCs) present with multipotent differentiation potential and immunomodulatory properties. As an alternative to bone marrow, we have examined fetal membranes, amnion and chorion, of term human placenta as a potential source of multipotent MSCs. Here we show that amnion mesenchymal cells (AMCs) and chorion mesenchymal cells (CMCs), isolated by mechanical separation and subsequent enzymatic digestion, demonstrate plastic adherence and fibroblast-like morphology and are able to form colonies that could be expanded for at least 15 passages. By FACS analysis, AMCs and CMCs were shown to be phenotypically similar to BM-MSCs and, when cultured in differentiation media, they demonstrated high morphogenetic plasticity by differentiating into osteocytes, chondrocytes and adipocytes. In an attempt to isolate cells with MSC characteristics from human fetal membranes, AMCs and CMCs expressing CD271 were enriched by immunomagnetic isolation and were demonstrated to possess higher clonogenic and osteogenic differentiation potential than CD271-depleted fractions. Based on these findings, amnion and chorion can be considered as a novel and convenient source of adult MSCs.

25. Multipotent adult stem cells

The quintessential stem cell is the embryonal stem (ES) cell which has unlimited self-renewal and multipotent differentiation potential. Stem cells have also been identified in most tissues. Compared with ES cells, tissue specific stem cells have less self-renewal ability and, although they differentiate into multiple lineages, they are not multipotent. A large number of recent published studies have suggested that tissue specific stem cells may have the ability to generate cells of tissues from unrelated organs.

Multipotent Stem Cells in Human Corneal Stroma

Keratocytes of the corneal stroma secrete a specialized extracellular matrix essential for vision. These quiescent cells exhibit limited capacity for self-renewal and after cell division become fibroblastic, secreting nontransparent tissue. This study sought to identify progenitor cells for human keratocytes. Near the corneal limbus, stromal cells expressed ABCG2, a protein present in many adult stem cells. The ABCG2-expressing cell population was isolated as a side population (SP) by cell sorting after exposure to Hoechst 33342 dye. The SP cells exhibited clonal growth and continued to express ABCG2 and also PAX6, product of a homeobox gene not expressed in adult keratocytes. Cloned SP cells cultured in medium with fibroblast growth factor-2 lost ABCG2 and PAX6 expression and upregulated several molecular markers of keratocytes, including keratocan, aldehyde dehydrogenase 3A1, and keratan sulfate. Cloned corneal SP cells under chondrogenic conditions produced matrix staining with toluidine blue and expressed cartilage-specific markers: collagen II, cartilage oligomatrix protein, and aggrecan. Exposure of cloned SP cells to neurogenic culture medium upregulated mRNA and protein for glial fibrillary acidic protein, neurofilament protein, and beta-tubulin II. These results demonstrate the presence of a population of cells in the human corneal stroma expressing stem cell markers and exhibiting multipotent differentiation potential. These appear to be the first human cells identified with keratocyte progenitor potential. Further analysis of these cells will aid elucidation of molecular mechanisms of corneal development, differentiation, and wound healing. These cells may be a resource for bioengineering of corneal stroma and for cell-based therapeutics.

Unexpected potential of adult stem cells

We have identified a population of primitive cells in normal human (h) as well as murine (m) and rat (r), postnatal bone marrow (BM) that have, at the single cell level, multipotent differentiation and extensive proliferation potential, which we named Multipotent Adult Progenitor Cell or MAPC. MAPC differentiate in vitro into most mesodermal cell types (cells with characteristics of osteoblasts, chondroblasts, fibroblasts, adipocytes, skeletal, smooth and cardiac myoblasts, endothelial cells), as well as cells with neuroectodermal and with endodermal features. Using retroviral marking we have shown that multi-lineage differentiation is derived from single MAPC. MAPC express active telomerase and can undergo 100+ cell doublings without telomere shortening, suggesting that they do not senesce. MAPC express oct-4 mRNA. MAPC engraft in vivo and persist for 6+ months, differentiate into hematopoietic and epithelial cells in response to local “cues”, and contribute to all somatic cell types when injected in the blastocyst. The finding that stem cells exist in post-natal tissues with previously unknown proliferation and differentiation potential opens up the possibility of using autologous stem cells to treat a host of degenerative, traumatic or congenital diseases.