ESC-Derived BDNF-Overexpressing Neural Progenitors Differentially Promote Recovery in Huntingtons Disease Models by Enhanced Striatal Differentiation

摘要

class="head no_bottom_margin" id="sec1title">IntroductionThe neurodegenerative disease Huntington's disease (HD) is characterized by dramatic motor dysfunction, cognitive decline, and psychiatric symptoms, which lead to progressive dementia and death approximately 15–20 years after onset (). HD is an autosomal dominant inheritable disease, caused by mutations in the huntingtin (HTT) gene, leading to an increased number of polyglutamine repeats in the encoded protein (). How mutant HTT protein causes neuronal dysfunction and neurodegeneration has not yet been understood in detail, and besides the existence of some symptomatic treatments, so far there is no causal therapy available for patients. Numerous laboratories showed that in a large number of HD mouse models BDNF or BDNF/TRKB signaling is strongly reduced due to a mutant htt-mediated mechanism (, ). Besides causing changes in vesicular transport of BDNF (), mutant HTT has been described to cause transcriptional downregulation of the BDNF gene through translocation of RE1 silencing transcription factor to the nucleus (). In addition to HD mouse models, a systematic and quantitative assessment of BDNF levels in human cerebral cortex samples, examined post mortem, confirmed that the production of this neurotrophin was impaired in the brains of HD patients (). As striatal medium spiny neurons (MSNs) depend on BDNF activity, a number of studies attempted striatal neuroprotection by providing exogenous BDNF delivered to the diseased rodent striatum either by adenoassociated viral transfer or by transplantation of diverse genetically modified cell types (e.g., fibroblasts) (, ). Altogether these studies showed enhanced neuroprotection, but no or only mild effects on long-term functional improvement in HD rodent models. On the other hand, cell transplantation as a promising therapeutic strategy, which aims to replace striatal neurons, has yielded some preliminary, but only modest and short-lived clinical benefits when using fetal neural cells (, ). Therefore, specifically embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) are considered to be an appropriate cell source, as ESCs can be differentiated in vitro into an MSN-like phenotype (, , , ). However, their long-term survival, long-term functional improvement, and safety in vivo still need to be proved.In the present study, we aimed to establish a combination therapy approach composed of cellular replacement by ESC-derived neural progenitors linked to BDNF supply. For this reason, we have generated BDNF-overexpressing mouse ESCs by knockin technology that display an enhanced neuronal and GABAergic differentiation in vitro (href="#bib23" rid="bib23" class=" bibr popnode">Leschik et al., 2013). Very recently, we were able to show that polysialylated neuronal cell adhesion molecule (PSA-NCAM)-positive progenitors derived from these ESCs lead to functional improvement when transplanted into mice with contusion spinal cord injury (href="#bib6" rid="bib6" class=" bibr popnode">Butenschön et al., 2016). With small modifications to the published protocol, which comprises magnetic-activated cell sorting (MACS) technology for purification, we tested in this study the efficiency and safety in three divergent HD mouse models. At present, a variety of different HD mouse models exist, chemically or genetically induced, which match with some aspects of HD. However, until now none of them perfectly recapitulates human neuropathological hallmarks as well as progressive cognitive and motor impairments. So far, genetic HD mouse models have been used only in a few cell transplantation studies. In most cases, cell transplantation was performed in toxin-lesioned mice, in which vast neurodegeneration occurs. This is clearly an advantage over genetic mouse models, which harbor less neurotoxicity. In contrast, genetic accuracy is missing in toxin-induced lesions and it is therefore questionable whether this represents an appropriate model system to test therapeutics for human pathology. For this reason, we decided to use besides the toxin-lesioned model with quinolinic acid (QA) the two widely used transgenic mouse lines R6/2 and N171-82Q, which differ in their extent of pathological features and degree of impairment (href="#bib28" rid="bib28" class=" bibr popnode">Ramaswamy et al., 2007). Typical behavioral assays for each HD mouse model were chosen based on previous publications. We also included the automated gait analysis system CatWalk as a very sensitive method for measuring subtle changes in motor behavior. Here, we show that the CatWalk assay is a valid method to address motor behavior in the QA-lesion and N171-82Q mouse models. Cell transplantation with purified BDNF-expressing neural progenitors revealed an improved motor function in QA-lesioned mice, whereas only subtle effects on motor behavior were detected in both transgenic mouse lines. Therapeutic effects of BDNF-expressing progenitors in the QA model could be attributed to enhanced neuronal and striatal in vivo differentiation compared with control cells. Furthermore, tumor formation was completely absent, in contrast to other studies (href="#bib1" rid="bib1" class=" bibr popnode">Aubry et al., 2008).