Neurosphere

About: Neurosphere is a research topic. Over the lifetime, 5145 publications have been published within this topic receiving 321088 citations.

MUC1* Mediates the Growth of Human Pluripotent Stem Cells

Abstract: The MUC1 protein is aberrantly expressed on an estimated 75% of all human solid tumor cancers. We recently reported that a transmembrane cleavage product, MUC1*, is the predominant form of the protein on cancer cells [1]. Further, our evidence indicated that MUC1* functions as a growth factor receptor on tumor cells, while the full-length protein appeared to have no growth promoting activity. Here, we report that MUC1* acts as a growth factor receptor on undifferentiated human embryonic stem cells (hESCs). Cleavage of the full-length ectodomain to form MUC1*, a membrane receptor, appears to make binding to its ligand, NM23, possible. Unexpectedly, we found that newly differentiated cells no longer express the cleaved form, MUC1*, or its ligand, NM23. Newly differentiated stem cells exclusively present full-length MUC1. Antibody-induced dimerization of the MUC1* receptor on hESCs stimulated cell growth to a far greater degree than currently used methods that require the addition of exogenous basic fibroblast growth factor (bFGF) as well as factors secreted by fibroblast “feeder cells”. Further, MUC1* mediated growth was shown to be independent of growth stimulated by bFGF or the milieu of factors secreted by feeder cells. Stimulating the MUC1* receptor with either the cognate antibody or its ligand NM23 enabled hESC growth in a feeder cell-free system and produced pluripotent colonies that resisted spontaneous differentiation. These findings suggest that this primal growth mechanism could be utilized to propagate large numbers of pluripotent stem cells for therapeutic interventions.

Suppression of stroke-induced progenitor proliferation in adult subventricular zone by tumor necrosis factor receptor 1.

Abstract: Stroke induced by middle cerebral artery occlusion leads to transiently increased progenitor proliferation in the subventricular zone (SVZ) and long-lasting striatal neurogenesis in adult rodents. Tumor necrosis factor-alpha (TNF-alpha) is upregulated in stroke-damaged brain. Whether TNF-alpha and its receptors influence SVZ progenitor proliferation after stroke is unclear. Here we show that the increased proliferation 1 week after stroke occurred concomitantly with elevated microglia numbers and TNF-alpha and TNF receptor-1 (TNF-R1) gene expression in the SVZ of wild-type mice. TNF receptor-1 was expressed on sorted SVZ progenitor cells from nestin-green fluorescent protein reporter mice. In animals lacking TNF-R1, stroke-induced SVZ cell proliferation and neuroblast formation were enhanced. In contrast, deletion of TNF-R1 did not alter basal or status epilepticus-stimulated cell proliferation in SVZ. Addition of TNF-alpha reduced the size and numbers of SVZ neurospheres through a TNF-R1-dependent mechanism without affecting cell survival. Our results provide the first evidence that TNF-R1 is a negative regulator of stroke-induced SVZ progenitor proliferation. Blockade of TNF-R1 signaling might be a novel strategy to promote the proliferative response in SVZ after stroke.

Engineering of midbrain organoids containing long-lived dopaminergic neurons.

Abstract: The possibility to generate dopaminergic (DA) neurons from pluripotent stem cells represents an unlimited source of material for tissue engineering and cell therapy for neurodegenerative disease. We set up a protocol based on the generation of size-calibrated neurospheres for a rapid production (3 weeks) of a high amount of DA neurons (>60%) oriented toward a midbrain-like phenotype, characterized by the expression of FOXA2, LMX1A, tyrosine hydroxylase (TH), NURR1, and EN1. By using γ-secretase inhibitors and varying culture time of neurospheres, we controlled maturation and cellular composition of a three-dimensional (3D) engineered nervous tissue (ENT). ENT contained neurons and glial cells expressing various markers of maturity, such as synaptophysin, neuronal nuclei-specific protein (NeuN), and glial fibrillary acidic protein (GFAP), and were electrophysiologically active. We found that 3-week-old neurospheres were optimal to generate 3D tissue containing DA neurons with typical A9 morphology. ENT generated from 4-week-old neurospheres launched glial cell type since astrocytes and myelin could be detected massively at the expense of TH-immunoreactive neurons. All γ-secretase inhibitors were not equivalent; compound E was more efficient than DAPT in generating DA neurons. This DA tissue provides a tool for drug screening, and toxicology. It should also become a useful biomaterial for studies on Parkinson's disease.

Enhancing the Reliability and Throughput of Neurosphere Culture on Hydrogel Microwell Arrays

Abstract: The neurosphere assay is the standard retrospective assay to test the self-renewal capability and multipotency of neural stem cells (NSC) in vitro. However, it has recently become clear that not all neurospheres are derived from a NSC and that on conventional cell culture substrates, neurosphere motility may cause frequent neurosphere 'merging' (Singec et al., Nature Methods, 2006; Jessberger et al., Stem Cells, 2007). Combining biomimetic hydrogel matrix technology with microengineering, we developed a microwell array platform on which NSC fate and neurosphere formation can be unequivocally attributed to a single founding cell. Using time-lapse microscopy and retrospective immunostaining, the fate of several hundred single NSCs was quantified. Compared to conventional neurosphere culture methods on plastic dishes, we detected a more than 100% increase in single NSC viability on soft hydrogels. Effective confinement of single proliferating cells to microwells led to neurosphere formation of vastly different sizes, a high percentage of which showed stem cell phenotypes after one week in culture. The reliability and increased throughput of this platform should help to elucidate better the function of sphere-forming stem/progenitor cells independent of their proliferation dynamics.