Neural Stem/Progenitor Cells Transplanted to the Hypoglossal Nucleus Integrates with the Host CNS in Adult Rats and Promotes Motor Neuron Survival

Abstract

Transplantation of neural stem cells and the mobilization of endogenous neuronal precursors in the adult brain have been proposed as therapeutic strategies for central nervous system disorders and injuries. The aim of the present study was to investigate the possible survival and integration of grafted neural progenitor cells (NPCs) from the subventricular zone (SVZ) in a hypoglossal nerve avulsion model with substantial neuronal loss. Adult neural progenitor cells (NPCs) from the subventricular zone (SVZ) were cultured from inbred transgenic eGFP Lewis rats and transplanted to the hypoglossal nucleus of inbred Lewis rat from the same family but that were not carrying the eGFP strain after avulsion of the hypoglossal nerve. Grafted cells survived in the host more than 3 months and differentiated into neurons [βIII tubulin (Tuj-1 staining)] with fine axon- and dendrite-like processes as well as astrocytes (GFAP) and oligodendrocytes (O4) with typical morphology. Staining for synaptic structures (synaptophysin and bassoon) indicated integration of differentiated cells from the graft with the host CNS. Furthermore, transplantation of NPCs increased the number of surviving motoneurons in the hypoglossal nucleus after nerve avulsion that, if untreated, result in substantial neuronal death. The NPCs used in this study expressed VEGF in vitro as well as in vivo following transplantation that may mediate the rescue effect of the axotomized motoneurons.

Amelioration of motor/sensory dysfunction and spasticity in a rat model of acute lumbar spinal cord injury by human neural stem cell transplantation

“…Thus, the use of treatment strategies that can replace or supplement the loss of the local neurotrophic activity and are initiated during this acute period should thus lead to a measurable treatment effect. Previous studies have demonstrated that neural stem cells of mouse, rat or human origin are a rich source of extracellularly released trophic factors (such as NGF, BDNF, GDNF, EGF, IGF-1, and VEGF) in in vitro cultured cells and that these cell populations retain a high level of neurotrophin expression after in vivo grafting in naïve animals and in a variety of neurodegenerative models including spinal injury and transgenic ALS models [56-61]. In addition, using long-term post-grafting survival periods, it was shown that following in vivo grafting of neural precursors with neurogenic potential into either the spinal cord at nine days post spinal cord injury, the brain at three days post ischemic insult, or the central nervous system (brain or spinal cord) of adult or developing rats, there is the development of functionally and morphologically-defined synaptic contacts between grafted neurons and the neurons of the host [62-64].…”

Neural Progenitor Cell Implants in the Lesioned Medial Longitudinal Fascicle of Adult Cats Regulate Synaptic Composition and Firing Properties of Abducens Internuclear Neurons

“…Coronal sections were cut on a Vibratome to a thickness of 50 m. Sections containing the abducens nucleus were washed in PBS-T and, after blocking unspecific sites with 10% normal donkey serum in PBS-T for 1 h, were left overnight at room temperature in the primary antibody solution prepared in PBS-T with 5% normal donkey serum and 0.05% sodium azide. Double, occasionally triple, immunofluorescence labeling was undertaken for the following: calretinin (a marker for abducens internuclear neurons; de la Cruz et al, 1998), along with either synaptophysin (a general synaptic marker), vesicular GABA and glycine amino acid transporter (VGAT) for inhibitory synaptic boutons (González-Forero et al, 2004), or glial fibrillary acidic protein (GFAP) to identify astrocytes. The primary antibodies used were the following: calretinin [goat polyclonal antibody (pAb), 1:500; Swant], synaptophysin (mouse mAb, 1:2000; Millipore), VGAT (rabbit pAb, 1:500; Millipore), and GFAP (rabbit pAb, 1:500; DAKO).…”

“…1F ). Before transplantation, neurospheres were prelabeled with either a fluorescent lipophilic tracer (PKH26; Sigma; Gudiño-Cabrera et al, 2000) or transfected overnight with a lentiviral vector containing a reporter gene for green fluorescent protein (GFP) expressed under the control of a cytomegalovirus promoter (titer, 10 7 pfu/ml) with a multiplicity of infection of 3. The virus-containing culture medium was removed and replaced with fresh DMEM/F12 supplemented with B-27, EGF, and FGF-2.…”

“…They have been useful for the re-establishment of alterations produced by a variety of neuronal lesions (Oka et al, 2004;Fagerlund et al, 2012). Since abducens neurons are endowed with the high-affinity receptors for neurotrophins trkA, trkB, and trkC , they can retrogradely transport these factors from implanted cells.…”

“…As a proof of concept, we sought to deliver at the site of a central lesion in the medial longitudinal fascicle (MLF) neural progenitor cells that have the potential not only to integrate within the scaffold of cells in the host tissue (Cusimano et al, 2012;Fagerlund et al, 2012), but also to deliver trophic factors such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and vascular endothelial growth factor (VEGF;Tonchev et al, 2007). The synaptotrophic action of these factors is supported by the fact that their delivery modifies synaptic and discharge characteristics of neurons (Zhou et al, 2005;Boyce et al, 2012).…”