ReNeuron Limited.

BackgroundNeural stem cells (NSCs) are powerful research tools for the design and discovery of new approaches to neurodegenerative disease. Overexpression of the myc family transcription factors in human primary cells from developing cortex and mesencephalon has produced two stable multipotential NSC lines (ReNcell VM and CX) that can be continuously expanded in monolayer culture.ResultsIn the undifferentiated state, both ReNcell VM and CX are nestin positive and have resting membrane potentials of around -60 mV but do not display any voltage-activated conductances. As initially hypothesized, using standard methods (stdD) for differentiation, both cell lines can form neurons, astrocytes and oligodendrocytes according to immunohistological characteristics. However it became clear that this was not true for electrophysiological features which designate neurons, such as the firing of action potentials. We have thus developed a new differentiation protocol, designated 'pre-aggregation differentiation' (preD) which appears to favor development of electrophysiologically functional neurons and to lead to an increase in dopaminergic neurons in the ReNcell VM line. In contrast, the protocol used had little effect on the differentiation of ReNcell CX in which dopaminergic differentiation was not observed. Moreover, after a week of differentiation with the preD protocol, 100% of ReNcell VM featured TTX-sensitive Na+-channels and fired action potentials, compared to 25% after stdD. Currents via other voltage-gated channels did not appear to depend on the differentiation protocol. ReNcell CX did not display the same electrophysiological properties as the VM line, generating voltage-dependant K+ currents but no Na+ currents or action potentials under either stdD or preD differentiation.ConclusionThese data demonstrate that overexpression of myc in NSCs can be used to generate electrophysiologically active neurons in culture. Development of a functional neuronal phenotype may be dependent on parameters of isolation and differentiation of the cell lines, indicating that not all human NSCs are functionally equivalent.

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Stroke remains one of the most promising targets for cell therapy. Thorough preclinical efficacy testing of human neural stem cell (hNSC) lines in a rat model of stroke (transient middle cerebral artery occlusion) is, however, required for translation into a clinical setting. Magnetic resonance imaging (MRI) here confirmed stroke damage and allowed the targeted injection of 450,000 hNSCs (CTX0E03) into peri-infarct tissue, rather than the lesion cyst. Intraparenchymal cell implants improved sensorimotor dysfunctions (bilateral asymmetry test) and motor deficits (footfault test and rotameter). Importantly, analyses based on lesion topology (striatal vs. striatal 1 cortical damage) revealed a more significant improvement in animals with a stroke confined to the striatum. However, no improvement in learning and memory (water maze) was evident. An intracerebroventricular injection of cells did not result in any improvement. MRI-based lesion, striatal and cortical volumes were unchanged in treated animals compared to those with stroke that received an intraparenchymal injection of suspension vehicle. Grafted cells only survived after intraparenchymal injection with a striatal 1 cortical topology resulting in better graft survival (16,026 cells) than in animals with smaller striatal lesions (2,374 cells). Almost 20% of cells differentiated into glial fibrillary acidic protein1 astrocytes, but <2% turned into FOX31 neurons. These results indicate that CTX0E03 implants robustly recover behavioral dysfunction over a 3-month time frame and that this effect is specific to their site of implantation. Lesion topology is potentially an important factor in the recovery, with a stroke confined to the striatum showing a better outcome compared to a larger area of damage. STEM CELLS 2012;30:785-796 Disclosure of potential conflicts of interest is found at the end of this article.

. This study found that the implantation of CTX0E03 human neural stem cells in rats after MCAO stroke promoted significant behavioral recovery depending on cell dose. The authors propose a paracrine trophic mechanism, which is triggered early after CTX0E03 cell implantation, and which in turn targets restoration of neurogenesis in the SVZ of MCAO rats.