Growth factors regulate the survival and fate of cells derived from human neurospheres

Abstract

Cells isolated from the embryonic, neonatal, and adult rodent central nervous system divide in response to epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF-2), while retaining the ability to differentiate into neurons and glia. These cultures can be grown in aggregates termed neurospheres, which contain a heterogeneous mix of both multipotent stem cells and more restricted progenitor populations. Neurospheres can also be generated from the embryonic human brain and in some cases have been expanded for extended periods of time in culture. However, the mechanisms controlling the number of neurons generated from human neurospheres are poorly understood. Here we show that maintaining cell-cell contact during the differentiation stage, in combination with growth factor administration, can increase the number of neurons generated under serum-free conditions from 8% to > 60%. Neurotrophic factors 3 and 4 (NT3, NT4) and platelet-derived growth factor (PDGF) were the most potent, and acted by increasing neuronal survival rather than inducing neuronal phenotype. Following differentiation, the neurons could survive dissociation and either replating or transplantation into the adult rat brain. This experimental system provides a practically limitless supply of enriched, non-genetically transformed neurons. These should be useful for both neuroactive drug screening in vitro and possibly cell therapy for neurodegenerative diseases.

A novel, neural potential of non-hematopoietic human umbilical cord blood stem cells

“…Upon attachment, in the presence of serum, surface cells of neurosphere start to migrate away from the center then differentiate into neurons (β-tubulin III -red), astrocytes (GFAPgreen) and oligodendrocytes (data not shown). These results are consistent with the pattern of differentiation in neurospheres (Caldwell et al, 2001) or human embryonic stem cells (Benzing et al, 2006). Neural character of HUCB-NSC line was further evaluated by comparative analysis of the transcriptional profile (Affymetrix DNA microarray probes HG-U133, covering the expression of about 40.000 genes), between the free-floating, non-differentiated HUCB-NSC and its starting, nonselected cell populationmononuclear cell (MNC) fraction of human cord blood.…”

Clusterin secreted by astrocytes enhances neuronal differentiation from human neural precursor cells

“…20 It has been shown to bind neurotrophic factors, such as NGF and NT3, which are secreted by astrocytes. 21 We have previously shown that human NPC's express the trk C-receptor, 22 so binding of a-2 macroglobulin to NT3 secreted by astrocytes may be having a role in the neurite outgrowth seen with hippocampal and midbrain ACM. In addition, extracellular matrix glycoprotein, falls into the family of neurite promoting factors, so this too may be having a role in neurite outgrowth.…”

Human peripheral blood-born hematosphere as a niche for hematopoietic stem cell expansion

“…Spheroid culture methods of stem cells from different tissues have been successfully used for expansion of cardiac and neural stem cells. These spheres sensitize target stem cells to growth factors and provide sufficient cell-to-cell and cell-to-matrix contacts, mimicking the in vivo stem cell niche [3,4]. Here we asked whether spheroid culture of blood mononuclear cells (MNCs) would potentiate the expansion of circulating blood HSPC.…”