GSK-3ß-induced Tau pathology drives hippocampal neuronal cell death in Huntington's disease: involvement of astrocyte-neuron interactions.

Glycogen synthase kinase-3ß (GSK-3ß) has emerged as a critical factor in several pathways involved in hippocampal neuronal maintenance and function. In Huntington's disease (HD), there are early hippocampal deficits both in patients and transgenic mouse models, which prompted us to investigate whether disease-specific changes in GSK-3ß expression may underlie these abnormalities. Thirty-three postmortem hippocampal samples from HD patients (neuropathological grades 2-4) and age- and sex-matched normal control cases were analyzed using real-time quantitative reverse transcription PCRs (qPCRs) and immunohistochemistry. In vitro and in vivo studies looking at hippocampal pathology and GSK-3ß were also undertaken in transgenic R6/2 and wild-type mice. We identified a disease and stage-dependent upregulation of GSK-3ß mRNA and protein levels in the HD hippocampus, with the active isoform pGSK-3ß-Tyr(216) being strongly expressed in dentate gyrus (DG) neurons and astrocytes at a time when phosphorylation of Tau at the AT8 epitope was also present in these same neurons. This upregulation of pGSK-3ß-Tyr(216) was also found in the R6/2 hippocampus in vivo and linked to the increased vulnerability of primary hippocampal neurons in vitro. In addition, the increased expression of GSK-3ß in the astrocytes of R6/2 mice appeared to be the main driver of Tau phosphorylation and caspase3 activation-induced neuronal death, at least in part via an exacerbated production of major proinflammatory mediators. This stage-dependent overactivation of GSK-3ß in HD-affected hippocampal neurons and astrocytes therefore points to GSK-3ß as being a critical factor in the pathological development of this condition. As such, therapeutic targeting of this pathway may help ameliorate neuronal dysfunction in HD.

Cystamine/cysteamine rescues the dopaminergic system and shows neurorestorative properties in an animal model of Parkinson's disease.

The neuroprotective properties of cystamine identified in pre-clinical studies have fast-tracked this compound to clinical trials in Huntington's disease, showing tolerability and benefits on motor symptoms. We tested whether cystamine could have such properties in a Parkinson's disease murine model and now provide evidence that it can not only prevent the neurodegenerative process but also can reverse motor impairments created by a 6-hydroxydopamine lesion 3 weeks post-surgery. Importantly, we report that cystamine has neurorestorative properties 5 weeks post-lesion as seen on the number of nigral dopaminergic neurons which is comparable with treatments of cysteamine, the reduced form of cystamine used in the clinic, as well as rasagiline, increasingly prescribed in early parkinsonism. All three compounds induced neurite arborization of the remaining dopaminergic cells which was further confirmed in ex vivo dopaminergic explants derived from Pitx3-GFP mice. The disease-modifying effects displayed by cystamine/cysteamine would encourage clinical testing.

Single-cell suspension methodology favors survival and vascularization of fetal striatal grafts in the YAC128 mouse model of Huntington's disease.

Cell replacement therapies have yielded variable and short-lived benefits in Huntington's disease (HD) patients. This suboptimal outcome is likely due to the fact that graft survival is compromised long term because grafts are subjected to a host's microglial inflammatory response, to a lack of adequate trophic support, and possibly to cortical excitotoxicity. However, graft demise may also relate to more straightforward issues such as cell preparation methodology (solid grafts vs. cell suspension). Indeed, we recently reported that solid grafts are poorly revascularized in HD patients transplanted 9 and 12 years previously. To evaluate whether methodological issues relating to cell preparation may have an impact on graft viability, we implanted green fluorescent protein (GFP(+)) single-cell suspensions of fetal striatal neuronal cells into the striatum of YAC128 HD mice. Postmortem evaluation yielded comparable graft survival in YAC128 mice and their wild-type littermates (noncarrier) at 1 and 3 months posttransplantation. Additionally, the degrees of graft revascularization in the YAC128 and noncarrier mice were similar, with both capillaries and large-caliber vessels observable within the grafted tissue. Furthermore, GFP(+) cells interacted well with host blood vessels, indicating integration of the donor cells within the recipient brain. These observations, combined with our recent report of poor revascularization of solid grafts in the HD-transplanted patients, suggest that the success of cell transplantation can be improved by optimizing methodological aspects relating to cell preparation.

The fate of cell grafts for the treatment of Huntington's disease: the post-mortem evidence.

The hope that cell transplantation therapies will provide an ideal treatment option for neurodegenerative diseases has been considerably revived with the remarkable advancements in genetic engineering towards active cell fate determination in vitro. However, for disorders such as Huntington's disease (HD), the challenges that we face are still enormous. This autosomal dominant genetic disorder leads, in part, to massive neuronal loss and severe brain atrophy which, despite the cell type used, cannot be easily repaired. And before large clinical trials are even considered, we must take a critical look at the outcomes of the pilot studies already available, not only from a clinical perspective but also by a careful assessment of what we can learn from the autopsies of HD patients who have undergone transplantation. In this review, we summarize and discuss the seven transplantation pilot trials that were initiated worldwide in HD patients more than a decade ago, with a particular emphasis on the post-mortem analyses of nine unique cases. Moreover, we describe a series of factors, both technical and related to patient selection, that we deem important to predict the outcome of cell grafts in HD therapy.

An in vitro perspective on the molecular mechanisms underlying mutant huntingtin protein toxicity.

Huntington's disease (HD) is a devastating neurodegenerative disorder whose main hallmark is brain atrophy. However, several peripheral organs are considerably affected and their symptoms may, in fact, manifest before those resulting from brain pathology. HD is of genetic origin and caused by a mutation in the huntingtin gene. The mutated protein has detrimental effects on cell survival, but whether the mutation leads to a gain of toxic function or a loss of function of the altered protein is still highly controversial. Most currently used in vitro models have been designed, to a large extent, to investigate the effects of the aggregation process in neuronal-like cells. However, as the pathology involves several other organs, new in vitro models are critically needed to take into account the deleterious effects of mutant huntingtin in peripheral tissues, and thus to identify new targets that could lead to more effective clinical interventions in the early course of the disease. This review aims to present current in vitro models of HD pathology and to discuss the knowledge that has been gained from these studies as well as the new in vitro tools that have been developed, which should reflect the more global view that we now have of the disease.

Hax1 lacks BH modules and is peripherally associated to heavy membranes: implications for Omi/HtrA2 and PARL activity in the regulation of mitochondrial stress and apoptosis.

Hax1 has an important role in immunodeficiency syndromes and apoptosis. A recent report (Chao et al., Nature, 2008) proposed that the Bcl-2-family-related protein, Hax1, suppresses apoptosis in lymphocytes and neurons through a mechanism that involves its association to the inner mitochondrial membrane rhomboid protease PARL, to proteolytically activate the serine protease Omi/HtrA2 and eliminate active Bax. This model implies that the control of cell-type sensitivity to pro-apoptotic stimuli is governed by the PARL/Hax1 complex in the mitochondria intermembrane space and, more generally, that Bcl-2-family-related proteins can control mitochondrial outer-membrane permeabilization from inside the mitochondrion. Further, it defines a novel, anti-apoptotic Opa1-independent pathway for PARL. In this study, we present evidence that, in vivo, the activity of Hax1 cannot be mechanistically coupled to PARL because the two proteins are confined in distinct cellular compartments and their interaction in vitro is an artifact. We also show by sequence analysis and secondary structure prediction that Hax1 is extremely unlikely to be a Bcl-2-family-related protein because it lacks Bcl-2 homology modules. These results indicate a different function and mechanism of Hax1 in apoptosis and re-opens the question of whether mammalian PARL, in addition to apoptosis, regulates mitochondrial stress response through Omi/HtrA2 processing.