Striatal Transplantation of Human Dopaminergic Neurons Differentiated From Induced Pluripotent Stem Cells Derived From Umbilical Cord Blood Using Lentiviral Reprogramming.

Human induced pluripotent stem cells (hiPSCs) are promising sources for regenerative therapies like the replacement of dopaminergic neurons in Parkinson's disease. They offer an unlimited cell source that can be standardized and optimized to produce applicable cell populations to gain maximal functional recovery. In the present study, human cord blood-derived iPSCs (hCBiPSCs) were differentiated into dopaminergic neurons utilizing two different in vitro protocols for neural induction: (protocol I) by fibroblast growth factor (FGF-2) signaling, (protocol II) by bone morphogenetic protein (BMP)/transforming growth factor (TGF-ß) inhibition. After maturation, in vitro increased numbers of tyrosine hydroxylase (TH)-positive neurons (7.4% of total cells) were observed by protocol II compared to 3.5% in protocol I. Furthermore, 3 weeks after transplantation in hemiparkinsonian rats in vivo, a reduced number of undifferentiated proliferating cells was achieved with protocol II. In contrast, proliferation still occurred in protocol I-derived grafts, resulting in tumor-like growth in two out of four animals 3 weeks after transplantation. Protocol II, however, did not increase the number of TH(+) cells in the striatal grafts of hemiparkinsonian rats. In conclusion, BMP/TGF-ß inhibition was more effective than FGF-2 signaling with regard to dopaminergic induction of hCBiPSCs in vitro and prevented graft overgrowth in vivo.

Electrophysiological Characterization of eGFP-Labeled Intrastriatal Dopamine Grafts.

Substitution of degenerated dopaminergic (DA) neurons by intrastriatally transplanted ventral mesencephalon (VM)-derived progenitor cells has been shown to improve motor functions in parkinsonian patients and animal models, whereas investigations of electrophysiological properties of the grafted DA neurons have been rarely performed. Here we show electrophysiological properties of grafted VM progenitor cells at different time intervals up to 12 weeks after transplantation measured in acute brain slices using eGFP-Flag transfection to identify the graft. We were able to classify typical DA neurons according to the biphasic progression (voltage "sag") to hyperpolarizing current injections. Two types of DA-like neurons were classified. Whereas type 1 neurons were characterized by delayed action potentials after hyperpolarization and irregular spontaneous firing, type 2 neurons displayed burst firing after hyperpolarization, spontaneous bursts, and regular firing. Comparison to identified DA neurons in vitro indicates a high integration of the intrastriatally grafted neurons, since in vitro cultures displayed regular firing spontaneously, whereas grafted identified DA neurons showed irregular firing. Additionally, type 1 and type 2 neurons exhibited a slight increase in the spontaneous firing frequency over time intervals after grafting, which might reflect a progressive integration of the grafted DA neurons. Our results provide evidence of the differentiation of grafted VM progenitor cells into mature integrated DA neurons, which are shown to replace the missing DA neurons functionally early after grafting.

Adult hemiparkinsonian rats do not benefit from tactile stimulation.

Tactile stimulation (TS) applied to adult rats after cortical injury (medial frontal cortex aspiration or sensorimotor pial stripping stroke model) has been previously shown to ameliorate behavioral impairments and to improve morphological parameters like dendritic length of prefrontal cortical neurons (Gibb et al., 2010). The purpose of this study was to examine the effect of TS on healthy and hemiparkinsonian adult rats. Therefore, the animals received TS for 14 days and 15 min three times daily. At different time points rats were tested in various behavioral tests (amphetamine-induced rotation, cylinder test, staircase test). Finally, rats were sacrificed, their brains removed, and processed for Golgi-Cox analyses, tyrosine hydroxylase immunohistochemistry and quantitative RT-PCR. We found that the striatal 6-OHDA lesion itself induced a long-term increase of astroglial Fgf2 transcript levels, but was not further increased by TS. In contrast TS applied to healthy rats elicited a transient short-term increase of Fgf2 in the striatum and Bdnf, Grin1, and Fgf2 in the hippocampus. Moreover, behavioral and histological analyses do not support a beneficial effect of TS for hemiparkinsonian rats, applied for two weeks starting one day after partial striatal 6-OHDA lesion.