Neural Differentiation of Brain-Derived Neurotrophic Factor-Expressing Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells in Culture via TrkB-Mediated ERK and β-Catenin Phosphorylation and following Transplantation into the Developing Brain

Abstract

The ability of mesenchymal stem cells (MSCs) to differentiate into neural cells makes them potential replacement therapeutic candidates in neurological diseases. Presently, overexpression of brain-derived neurotrophic factor (BDNF), which is crucial in the regulation of neural progenitor cell differentiation and maturation during development, was sufficient to convert the mesodermal cell fate of human umbilical cord bloodderived MSCs (hUCB-MSCs) into a neuronal fate in culture, in the absence of specialized induction chemicals. BDNF overexpressing hUCB-MSCs (MSCs-BDNF) yielded an increased number of neuron-like cells and, surprisingly, increased the expression of neuronal phenotype markers in a time-dependent manner compared with control hUCB-MSCs. In addition, MSCs-BDNF exhibited a decreased labeling for MSCs-related antigens such as CD44, CD73, and CD90, and decreased potential to differentiate into mesodermal lineages. Phosphorylation of the receptor tyrosine kinase B (TrkB), which is a receptor of BDNF, was increased significantly in MSC-BDNF. BDNF overexpression also increased the phosphorylation of β-catenin and extracellular signal-regulated kinases (ERKs). Inhibition of TrkB availability by treatment with the TrkBspecific inhibitor K252a blocked the BDNF-stimulated phosphorylation of β-catenin and ERKs, indicating the involvement of both the β-catenin and ERKs signals in the BDNF-stimulated and TrkB-mediated neural differentiation of hUCB-MSCs. Reduction of β-catenin availability using small interfering RNA-mediated gene silencing inhibited ERKs phosphorylation. However, β-catenin activation was maintained. In addition, inhibition of β-catenin and ERKs expression levels abrogated the BDNF-stimulated upregulation of neuronal phenotype markers. Furthermore, MSC-BDNF survived and migrated more extensively when grafted into the lateral ventricles of neonatal mouse brain, and differentiated significantly into neurons in the olfactory bulb and periventricular astrocytes. These results indicate that BDNF induces the neural differentiation of hUCB-MSCs in culture via the TrkB-mediated phosphorylation of ERKs and β-catenin and following transplantation into the developing brain.

Stably BDNF-GFP expressing embryonic stem cells exhibit a BDNF release-dependent enhancement of neuronal differentiation

“…As described previously for MSCs or adult NSCs overexpressing untagged BDNF (Lim et al, 2011;Ma et al, 2012), or as revealed by application of BDNF to EBs (Kang et al, 2001) or to NSCs (Silva et al, 2009), neuronal differentiation of our BDNF-GFPoverexpressing ESCs was strongly enhanced (Figs 4,5). Given that this enhancement was critically dependent on the extracellular availability of BDNF (see inhibition by TrkB-Fc receptor bodies; Fig.…”

“…To put the present level of overexpression into context, relatively high levels of virusmediated BDNF expression in bone marrow stromal cells, MSCs and adult NSCs have been previously shown to exert beneficial effects, for example after grafting into lesioned tissue (see e.g. Lim et al, 2011;Ma et al, 2012;Makar et al, 2008;Makar et al, 2009). Furthermore, modest endogenous levels of BDNF protein in mesenchymal and neural stem cells (Chekhonin et al, 2011;Li et al, 2006;Mauri et al, 2012) have also been shown to be beneficial on co-cultured neurons (Mauri et al, 2012).…”

“…Because BDNF enhances differentiation, these stem cells potentially show the capacity to support their own differentiation in an autocrine fashion. Viral transduction of MSCs and adult NSCs in order to produce BDNF has recently been shown to improve neuronal survival of host neurons as well as neuronal differentiation of grafted stem cells (Lim et al, 2011;Ma et al, 2012). Given that ESCs display a higher potential for differentiation than, for example, MSCs, and are easier to access and amplify than adult NSCs, ESCs genetically engineered to stably overexpress BDNF could be a very suitable tool to provide a pluripotent cell source for long-lasting expression and secretion of BDNF after grafting.…”

Transplantation of umbilical cord–derived mesenchymal stem cells as a novel strategy to protect the central nervous system: technical aspects, preclinical studies, and clinical perspectives

“…Lim et al transplanted BDNF-expressing hUCB-MSCs into the lateral ventricle of healthy mouse pups on first postnatal day (101). These genetically modified hUCB-MSCs were found to be able to survive, migrate, and differentiate into periventricular astrocytes and olfactory bulb neurons, and performed better than non-engineered hUCB-MSCs under the same conditions.…”

Neural Fate of Mesenchymal Stem Cells and Neural Crest Stem Cells: Which Ways to Get Neurons for Cell Therapy Purpose?

“…Few years later, after genetically modifying UCB-MSCs through transfection with a BDNF-expressing plasmid, they showed the expression of βIII-tubulin, NeuN, GFAP and myelin basic protein (MBP) in differentiated cells, associated with an up-regulated phosphorylation level of TrkB, Raf-1 and ERKs [77].…”