Alpha-synuclein fine-tunes neuronal response to pro-inflammatory cytokines.

Pro-inflammatory cytokines are emerging as neuroinflammatory mediators in Parkinson's disease (PD) due to their ability to act through neuronal cytokine receptors. Critical questions persist regarding the role of cytokines in neuronal dysfunction and their contribution to PD pathology. Specifically, the potential synergy of the hallmark PD protein alpha-synuclein (α-syn) with cytokines is of interest. We therefore investigated the direct impact of pro-inflammatory cytokines on neurons and hypothesized that α-syn pathology exacerbates cytokine-induced neuronal deficits in PD. iPSC-derived cortical neurons (CNs) from healthy controls and patients with α-syn gene locus duplication (SNCA dupl) were stimulated with IL-17A, TNF-α, IFN-γ, or a combination thereof. For rescue experiments, CNs were pre-treated with α-syn anti-oligomerisation compound NPT100-18A prior to IL-17A stimulation. Cytokine receptor expression, microtubule cytoskeleton, axonal transport and neuronal activity were assessed. SNCA dupl CNs displayed an increased IL-17A receptor expression and impaired IL-17A-mediated cytokine receptor regulation. Cytokines exacerbated the altered distribution of tubulin post-translational modifications in SNCA dupl neurites, with SNCA dupl-specific IL-17A effects. Tau pathology in SNCA dupl CNs was also aggravated by IL-17A and cytokine mix. Cytokines slowed down mitochondrial axonal transport, with IL-17A-mediated retrograde slowing in SNCA dupl only. The pre-treatment of SNCA dupl CNs with NPT100-18A prevented the IL-17A-induced functional impairments in axonal transport and neural activity. Our work elucidates the detrimental effects of pro-inflammatory cytokines, particularly IL-17A, on human neuronal structure and function in the context of α-syn pathology, suggesting that cytokine-mediated inflammation represents a second hit to neurons in PD which is amenable to disease modifying therapies that are currently in clinical trials.

A reversible state of hypometabolism in a human cellular model of sporadic Parkinson's disease.

Sporadic Parkinson's Disease (sPD) is a progressive neurodegenerative disorder caused by multiple genetic and environmental factors. Mitochondrial dysfunction is one contributing factor, but its role at different stages of disease progression is not fully understood. Here, we showed that neural precursor cells and dopaminergic neurons derived from induced pluripotent stem cells (hiPSCs) from sPD patients exhibited a hypometabolism. Further analysis based on transcriptomics, proteomics, and metabolomics identified the citric acid cycle, specifically the α-ketoglutarate dehydrogenase complex (OGDHC), as bottleneck in sPD metabolism. A follow-up study of the patients approximately 10 years after initial biopsy demonstrated a correlation between OGDHC activity in our cellular model and the disease progression. In addition, the alterations in cellular metabolism observed in our cellular model were restored by interfering with the enhanced SHH signal transduction in sPD. Thus, inhibiting overactive SHH signaling may have potential as neuroprotective therapy during early stages of sPD.

Scaffold-Based (Matrigel™) 3D Culture Technique of Glioblastoma Recovers a Patient-like Immunosuppressive Phenotype.

Conventional 2D cultures are commonly used in cancer research though they come with limitations such as the lack of microenvironment or reduced cell heterogeneity. In this study, we investigated in what respect a scaffold-based (Matrigel™) 3D culture technique can ameliorate the limitations of 2D cultures. NGS-based bulk and single-cell sequencing of matched pairs of 2D and 3D models showed an altered transcription of key immune regulatory genes in around 36% of 3D models, indicating the reoccurrence of an immune suppressive phenotype. Changes included the presentation of different HLA surface molecules as well as cellular stressors. We also investigated the 3D tumor organoids in a co-culture setting with tumor-infiltrating lymphocytes (TILs). Of note, lymphocyte-mediated cell killing appeared less effective in clearing 3D models than their 2D counterparts. IFN-γ release, as well as live cell staining and proliferation analysis, pointed toward an elevated resistance of 3D models. In conclusion, we found that the scaffold-based (Matrigel™) 3D culture technique affects the transcriptional profile in a subset of GBM models. Thus, these models allow for depicting clinically relevant aspects of tumor-immune interaction, with the potential to explore immunotherapeutic approaches in an easily accessible in vitro system.

Human Induced Pluripotent Stem Cell-Derived Pericytes as Scalable and Editable Source to Study Direct Lineage Reprogramming Into Induced Neurons.

Studying human somatic cell-to-neuron conversion using primary brain-derived cells as starting cell source is hampered by limitations and variations in human biopsy material. Thus, delineating the molecular variables that allow changing the identity of somatic cells, permit adoption of neuronal phenotypes, and foster maturation of induced neurons (iNs) is challenging. Based on our previous results that pericytes derived from the adult human cerebral cortex can be directly converted into iNs (Karow et al., 2018; Karow et al., 2012), we here introduce human induced pluripotent stem cell (hiPSC)-derived pericytes (hiPSC-pericytes) as a versatile and more uniform tool to study the pericyte-to-neuron conversion process. This strategy enables us to derive scalable cell numbers and allows for engineering of the starting cell population such as introducing reporter tools before differentiation into hiPSC-pericytes and subsequent iN conversion. Harvesting the potential of this approach, we established hiPSC-derived human-human neuronal cocultures that not only allow for independent manipulation of each coculture partner but also resulted in morphologically more mature iNs. In summary, we exploit hiPSC-based methods to facilitate the analysis of human somatic cell-to-neuron conversion.

Proteolytic Activity of the Paracaspase MALT1 Is Involved in Epithelial Restitution and Mucosal Healing.

The paracaspase MALT1 is a crucial regulator of immune responses in various cellular contexts. Recently, there is increasing evidence suggesting that MALT1 might represent a novel key player in mucosal inflammation. However, the molecular mechanisms underlying this process and the targeted cell population remain unclear. In this study, we investigate the role of MALT1 proteolytic activity in the context of mucosal inflammation. We demonstrate a significant enrichment of MALT1 gene and protein expression in colonic epithelial cells of UC patients, as well as in the context of experimental colitis. Mechanistically we demonstrate that MALT1 protease function inhibits ferroptosis, a form of iron-dependent cell death, upstream of NF-κB signaling, which can promote inflammation and tissue damage in IBD. We further show that MALT1 activity contributes to STAT3 signaling, which is essential for the regeneration of the intestinal epithelium after injury. In summary, our data strongly suggests that the protease function of MALT1 plays a critical role in the regulation of immune and inflammatory responses, as well as mucosal healing. Understanding the mechanisms by which MALT1 protease function regulates these processes may offer novel therapeutic targets for the treatment of IBD and other inflammatory diseases.

The Gut-Immune-Brain Axis: An Important Route for Neuropsychiatric Morbidity in Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) comprises Crohn's disease (CD) and ulcerative colitis (UC) and is associated with neuropsychiatric symptoms like anxiety and depression. Both conditions strongly worsen IBD disease burden. In the present review, we summarize the current understanding of the pathogenesis of depression and anxiety in IBD. We present a stepwise cascade along a gut-immune-brain axis initiated by evasion of chronic intestinal inflammation to pass the epithelial and vascular barrier in the gut and cause systemic inflammation. We then summarize different anatomical transmission routes of gut-derived peripheral inflammation into the central nervous system (CNS) and highlight the current knowledge on neuroinflammatory changes in the CNS of preclinical IBD mouse models with a focus on microglia, the brain-resident macrophages. Subsequently, we discuss how neuroinflammation in IBD can alter neuronal circuitry to trigger symptoms like depression and anxiety. Finally, the role of intestinal microbiota in the gut-immune-brain axis in IBD will be reviewed. A more comprehensive understanding of the interaction between the gastrointestinal tract, the immune system and the CNS accounting for the similarities and differences between UC and CD will pave the path for improved prediction and treatment of neuropsychiatric comorbidities in IBD and other inflammatory diseases.

Aberrant NOVA1 function disrupts alternative splicing in early stages of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal disease characterized by aberrant alternative splicing (AS). Nuclear loss and cytoplasmic accumulation of the splicing factor TDP-43 in motor neurons (MN) are hallmarks of ALS at late stages of the disease. However, it is unknown if altered AS is present before TDP-43 pathology occurs. Here, we investigate altered AS and its origins in early stages of ALS using human induced pluripotent stem cell-derived motor neurons (MNs) from sporadic and familial ALS patients. We find high levels of the RNA-binding proteins NOVA1, NOVA2, and RBFOX2 in the insoluble protein fractions and observe that AS events in ALS-associated MNs are enriched for binding sites of these proteins. Our study points to an early disrupted function of NOVA1 that drives AS changes in a complex fashion, including events caused by a consistent loss of NOVA1 function. NOVA1 exhibits increased cytoplasmic protein levels in early stage MNs without TDP-43 pathology in ALS postmortem tissue. As nuclear TDP-43 protein level depletes, NOVA1 is reduced. Potential indications for a reduction of NOVA1 also came from mice over-expressing TDP-43 lacking its nuclear localization signal and iPSC-MN stressed with puromycin. This study highlights that additional RBP-RNA perturbations in ALS occur in parallel to TDP-43.

Efficient and Easy Conversion of Human iPSCs into Functional Induced Microglia-like Cells.

Current protocols converting human induced pluripotent stem cells (iPSCs) into induced microglia-like cells (iMGL) are either dependent on overexpression of transcription factors or require substantial experience in stem-cell technologies. Here, we developed an easy-to-use two-step protocol to convert iPSCs into functional iMGL via: (1) highly efficient differentiation of hematopoietic progenitor cells (HPCs) from iPSCs, and (2) optimized maturation of HPCs to iMGL. A sequential harvesting approach led to an increased HPC yield. The protocol implemented a freezing step, thus allowing HPC biobanking and flexible timing of differentiation into iMGL. Our iMGL responded adequately to the inflammatory stimuli LPS, and iMGL RNAseq analysis matched those of other frequently used protocols. Comparing three different coating modalities, we increased the iMGL yield by culturing on uncoated glass surfaces, thereby retaining differentiation efficiency and functional hallmarks of iMGL. In summary, we provide a high-quality, easy-to-use protocol, rendering generation and functional studies on iMGL an accessible lab resource.

Organoids in gastrointestinal diseases: from experimental models to clinical translation.

We are entering an era of medicine where increasingly sophisticated data will be obtained from patients to determine proper diagnosis, predict outcomes and direct therapies. We predict that the most valuable data will be produced by systems that are highly dynamic in both time and space. Three-dimensional (3D) organoids are poised to be such a highly valuable system for a variety of gastrointestinal (GI) diseases. In the lab, organoids have emerged as powerful systems to model molecular and cellular processes orchestrating natural and pathophysiological human tissue formation in remarkable detail. Preclinical studies have impressively demonstrated that these organs-in-a-dish can be used to model immunological, neoplastic, metabolic or infectious GI disorders by taking advantage of patient-derived material. Technological breakthroughs now allow to study cellular communication and molecular mechanisms of interorgan cross-talk in health and disease including communication along for example, the gut-brain axis or gut-liver axis. Despite considerable success in culturing classical 3D organoids from various parts of the GI tract, some challenges remain to develop these systems to best help patients. Novel platforms such as organ-on-a-chip, engineered biomimetic systems including engineered organoids, micromanufacturing, bioprinting and enhanced rigour and reproducibility will open improved avenues for tissue engineering, as well as regenerative and personalised medicine. This review will highlight some of the established methods and also some exciting novel perspectives on organoids in the fields of gastroenterology. At present, this field is poised to move forward and impact many currently intractable GI diseases in the form of novel diagnostics and therapeutics.

STAT1 coordinates intestinal epithelial cell death during gastrointestinal infection upstream of Caspase-8.

Intestinal homeostasis and the maintenance of the intestinal epithelial barrier are essential components of host defense during gastrointestinal Salmonella Typhimurium infection. Both require a strict regulation of cell death. However, the molecular pathways regulating epithelial cell death have not been completely understood. Here, we elucidated the contribution of central mechanisms of regulated cell death and upstream regulatory components during gastrointestinal infection. Mice lacking Caspase-8 in the intestinal epithelium are highly sensitive towards bacterial induced enteritis and intestinal inflammation, resulting in an enhanced lethality of these mice. This phenotype was associated with an increased STAT1 activation during Salmonella infection. Cell death, barrier breakdown and systemic infection were abrogated by an additional deletion of STAT1 in Casp8ΔIEC mice. In the absence of epithelial STAT1, loss of epithelial cells was abolished which was accompanied by a reduced Caspase-8 activation. Mechanistically, we demonstrate that epithelial STAT1 acts upstream of Caspase-8-dependent as well as -independent cell death and thus might play a major role at the crossroad of several central cell death pathways in the intestinal epithelium. In summary, we uncovered that transcriptional control of STAT1 is an essential host response mechanism that is required for the maintenance of intestinal barrier function and host survival.

The Nuclear Receptor Nr4a1 Acts as a Microglia Rheostat and Serves as a Therapeutic Target in Autoimmune-Driven Central Nervous System Inflammation.

Microglia cells fulfill key homeostatic functions and essentially contribute to host defense within the CNS. Altered activation of microglia, in turn, has been implicated in neuroinflammatory and neurodegenerative diseases. In this study, we identify the nuclear receptor (NR) Nr4a1 as key rheostat controlling the activation threshold and polarization of microglia. In steady-state microglia, ubiquitous neuronal-derived stress signals such as ATP induced expression of this NR, which contributed to the maintenance of a resting and noninflammatory microglia phenotype. Global and microglia-specific deletion of Nr4a1 triggered the spontaneous and overwhelming activation of microglia and resulted in increased cytokine and NO production as well as in an accelerated and exacerbated form of experimental autoimmune encephalomyelitis. Ligand-induced activation of Nr4a1 accordingly ameliorated the course of this disease. Our current data thus identify Nr4a1 as regulator of microglia activation and potentially new target for the treatment of inflammatory CNS diseases such as multiple sclerosis.

Infiltrating T lymphocytes reduce myeloid phagocytosis activity in synucleinopathy model.

BACKGROUND: Synucleinopathies comprise a group of neurodegenerative diseases associated with abnormal accumulation of α-synuclein. One of the key factors that contribute to the progression of synucleinopathies is neuroinflammation. However, the role of lymphocytes in synucleinopathies like Parkinson's disease (PD) remains largely unclear. METHODS: To investigate how lymphocytes impact synucleinopathies, human wild-type α-synuclein (WTS) transgenic mice were crossed with mice lacking mature lymphocytes (Rag2(-/-)). In this in vivo model, we quantified α-synuclein aggregation in the substantia nigra (SN) and striatum and determined the numbers of innate and adaptive immune cells in the central nervous system (CNS). The activation state of resident and infiltrated CNS myeloid cells (M1 vs. M2) was further classified by gene and protein expression analyses. The impact of T and B lymphocytes on the phagocytic activity of microglia in the presence of α-synuclein aggregates was addressed in BV2 microglia in vitro. RESULTS: Compared to WTS(+) Rag2(+/+) mice, where T but not B lymphocytes infiltrated the CNS, decreased amounts of α-synuclein aggregates were found in WTS(+) Rag2(-/-) mice devoid of mature lymphocytes. The presence of T lymphocytes did not alter the number of Iba1(+) microglia but increased the frequency of the CD11b(+) CD45(hi) population in the CNS, indicative of an increased number of infiltrated macrophages. Moreover, the M1 phenotype was more prominent in WTS(+) Rag2(+/+) mice, whereas the M2 activation state was dominating in the absence of lymphocytes in WTS(+) Rag2(-/-) mice. In vitro, in the presence of T but not B lymphocytes, significantly less α-synuclein was phagocytosed by BV2 microglia, further supporting the prevalence of the M1 phenotype in the presence of T lymphocytes. CONCLUSIONS: Peripheral T lymphocytes strongly contribute to increased α-synuclein pathology via modulation of CNS myeloid cell function. In the presence of T lymphocytes, microglia phagocytosis of aggregated α-synuclein is reduced, which increases the severity of synucleinopathy.

Kdm6b and Pmepa1 as Targets of Bioelectrically and Behaviorally Induced Activin A Signaling.

The transforming growth factor-ß (TGF-ß) family member activin A exerts multiple neurotrophic and protective effects in the brain. Activin also modulates cognitive functions and affective behavior and is a presumed target of antidepressant therapy. Despite its important role in the injured and intact brain, the mechanisms underlying activin effects in the CNS are still largely unknown. Our goal was to identify the first target genes of activin signaling in the hippocampus in vivo. Electroconvulsive seizures, a rodent model of electroconvulsive therapy in humans, were applied to C57BL/6J mice to elicit a strong increase in activin A signaling. Chromatin immunoprecipitation experiments with hippocampal lysates subsequently revealed that binding of SMAD2/3, the intracellular effectors of activin signaling, was significantly enriched at the Pmepa1 gene, which encodes a negative feedback regulator of TGF-ß signaling in cancer cells, and at the Kdm6b gene, which encodes an epigenetic regulator promoting transcriptional plasticity. Underlining the significance of these findings, activin treatment also induced PMEPA1 and KDM6B expression in human forebrain neurons generated from embryonic stem cells suggesting interspecies conservation of activin effects in mammalian neurons. Importantly, physiological stimuli such as provided by environmental enrichment proved already sufficient to engender a rapid and significant induction of activin signaling concomitant with an upregulation of Pmepa1 and Kdm6b expression. Taken together, our study identified the first target genes of activin signaling in the brain. With the induction of Kdm6b expression, activin is likely to gain impact on a presumed epigenetic regulator of activity-dependent neuronal plasticity.

Gene dosage-dependent rescue of HSP neurite defects in SPG4 patients' neurons.

The hereditary spastic paraplegias (HSPs) are a heterogeneous group of motorneuron diseases characterized by progressive spasticity and paresis of the lower limbs. Mutations in Spastic Gait 4 (SPG4), encoding spastin, are the most frequent cause of HSP. To understand how mutations in SPG4 affect human neurons, we generated human induced pluripotent stem cells (hiPSCs) from fibroblasts of two patients carrying a c.1684C>T nonsense mutation and from two controls. These SPG4 and control hiPSCs were able to differentiate into neurons and glia at comparable efficiency. All known spastin isoforms were reduced in SPG4 neuronal cells. The complexity of SPG4 neurites was decreased, which was paralleled by an imbalance of axonal transport with less retrograde movement. Prominent neurite swellings with disrupted microtubules were present in SPG4 neurons at an ultrastructural level. While some of these swellings contain acetylated and detyrosinated tubulin, these tubulin modifications were unchanged in total cell lysates of SPG4 neurons. Upregulation of another microtubule-severing protein, p60 katanin, may partially compensate for microtubuli dynamics in SPG4 neurons. Overexpression of the M1 or M87 spastin isoforms restored neurite length, branching, numbers of primary neurites and reduced swellings in SPG4 neuronal cells. We conclude that neurite complexity and maintenance in HSP patient-derived neurons are critically sensitive to spastin gene dosage. Our data show that elevation of single spastin isoform levels is sufficient to restore neurite complexity and reduce neurite swellings in patient cells. Furthermore, our human model offers an ideal platform for pharmacological screenings with the goal to restore physiological spastin levels in SPG4 patients.

Role of α-synuclein in adult neurogenesis and neuronal maturation in the dentate gyrus.

α-Synuclein has been reported to be important in modulating brain plasticity and to be a key protein in neurodegenerative diseases, including Lewy body dementia (LBD). We investigated how α-synuclein levels modulate adult neurogenesis and the development of dendritic arborization and spines in the dentate gyrus, in which new neurons are constantly added. In the human hippocampus, levels of endogenous α-synuclein were increased in LBD, and the numbers of SOX2-positive cells were decreased. We investigated whether newly generated neurons were modulated by endogenous α-synuclein, and we found increased adult neurogenesis in α/ß-synuclein knock-out mice. In contrast, overexpression of human wild-type α-synuclein (WTS) decreased the survival and dendritic development of newborn neurons. Endogenous α-synuclein expression levels increased the negative impact of WTS on dendrite development, suggesting a toxic effect of increasing amounts of α-synuclein. To attempt a rescue of the dendritic phenotype, we administered rolipram to activate the cAMP response element-binding protein pathway, which led to a partial rescue of neurite development. The current work provides novel insights into the role of α-synuclein in adult hippocampal neurogenesis.

α-Synuclein induces alterations in adult neurogenesis in Parkinson disease models via p53-mediated repression of Notch1.

Parkinson disease is characterized by the loss of dopaminergic neurons mainly in the substantia nigra. Accumulation of α-synuclein and cell loss has been also reported in many other brain regions including the hippocampus, where it might impair adult neurogenesis, contributing to nonmotor symptoms. However, the molecular mechanisms of these alterations are still unknown. In this report we show that α-synuclein-accumulating adult rat hippocampus neural progenitors present aberrant neuronal differentiation, with reduction of Notch1 expression and downstream signaling targets. We characterized a Notch1 proximal promoter that contains p53 canonical response elements. In vivo binding of p53 represses the transcription of Notch1 in neurons. Moreover, we demonstrated that α-synuclein directly binds to the DNA at Notch1 promoter vicinity and also interacts with p53 protein, facilitating or increasing Notch1 signaling repression, which interferes with maturation and survival of neural progenitors cells. This study provides a molecular basis for α-synuclein-mediated disruption of adult neurogenesis in Parkinson disease.

Dopamine receptor activation promotes adult neurogenesis in an acute Parkinson model.

Cell proliferation of neural progenitors in the subventricular zone (SVZ) of Parkinson disease (PD) patients and animal models is decreased. It was previously demonstrated that the neurotransmitter dopamine modulates cell proliferation in the embryonic brain. The aim of the present study was to analyze whether oral treatment with the dopamine receptor agonist pramipexole (PPX) modulates adult neurogenesis in the SVZ/olfactory bulb system in a dopaminergic lesion model. 6-Hydroxydopamine (6-OHDA) lesioned adult rats received either PPX (1.0 mg/kg) or PBS orally twice daily and bromodeoxyuridine (BrdU, a cell proliferation marker) for 10 days and were perfused immediately after treatment or 4 weeks after PPX withdrawal. Stereological analysis revealed a significant augmentation in SVZ proliferation by PPX. Consecutively, enhanced neuronal differentiation and more new neurons were present in the olfactory bulb 4 weeks after PPX withdrawal. In addition, dopaminergic neurogenesis was increased in the olfactory bulb after PPX treatment. Motor activity as assessed by using an open field paradigm was permanently increased even after long term PPX withdrawal. In addition, we demonstrate that D2 and D3 receptors are present on adult rat SVZ-derived neural progenitors in vitro, and PPX specifically increased mRNA levels of epidermal growth factor receptor (EGF-R) and paired box gene 6 (Pax6). Oral PPX treatment selectively increases adult neurogenesis in the SVZ-olfactory bulb system by increasing proliferation and cell survival of newly generated neurons. Analyzing the neurogenic fate decisions mediated by D2/D3 signaling pathways may lead to new avenues to induce neural repair in the adult brain.

A Nurr1/CoREST pathway in microglia and astrocytes protects dopaminergic neurons from inflammation-induced death.

Nurr1, an orphan nuclear receptor, plays an essential role in the generation and maintenance of dopaminergic neurons in the brain. Rare mutations in Nurr1 are associated with familial Parkinson's disease, but the underlying basis for this relationship has not been established. Here, we demonstrate that Nurr1 unexpectedly functions to inhibit expression of pro-inflammatory neurotoxic mediators in both microglia and astrocytes. Reduced Nurr1 expression results in exaggerated inflammatory responses in microglia that are further amplified by astrocytes, leading to the production of factors that cause death of tyrosine hydroxylase-expressing neurons. Nurr1 exerts anti-inflammatory effects by docking to NF-kappaB-p65 on target inflammatory gene promoters in a signal-dependent manner. Subsequently, Nurr1 recruits the CoREST corepressor complex, resulting in clearance of NF-kappaB-p65 and transcriptional repression. These studies suggest that Nurr1 protects against loss of dopaminergic neurons in Parkinson's disease in part by limiting the production of neurotoxic mediators by microglia and astrocytes.

Cellular repair strategies in Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disorder after Alzheimer's disease, affecting 0.7% of the elderly population (defined as over 65 years of age). PD is clinically characterized by resting tremor, muscular rigidity, hypokinesia and postural instability. These motor symptoms result largely from the deficiency or dysfunction of dopaminergic neurons in the substantia nigra. Histopathological analysis reveals depletion of dopaminergic neurons as well as eosinophilic intracytoplasmic inclusions (Lewy bodies) in surviving neurons of the substantia nigra and other brain regions. The molecular pathogenesis is linked to protein misfolding by compromised alpha-synuclein and/or related proteins (synucleinopathy). Therefore, successful therapy of motor symptoms aims for the restoration of dopaminergic neurotransmission. Pharmacological drug treatment is usually effective only at an early stage of the disease but cannot halt progressive neuronal degeneration. With recent developments in stem cell technology, cell repair or replacement approaches came into focus. Here, we review new therapeutic strategies resulting from the innate propensity of the adult brain to generate new neurons, either by pharmacological stimulation of endogenous adult stem cell population or exogenous cell transplantation modalities.

Long-term course and mutational spectrum of spatacsin-linked spastic paraplegia.

OBJECTIVE: Hereditary spastic paraplegias (HSPs) comprise a heterogeneous group of neurodegenerative disorders resulting in progressive spasticity of the lower limbs. One form of autosomal recessive hereditary spastic paraplegia (ARHSP) with thin corpus callosum (TCC) was linked to chromosomal region 15q13-21 (SPG11) and associated with mutations in the spatacsin gene. We assessed the long-term course and the mutational spectrum of spatacsin-associated ARHSP with TCC. METHODS: Neurological examination, cerebral magnetic resonance imaging (MRI), 18fluorodeoxyglucose positron emission tomography (PET), nerve biopsy, linkage and mutation analysis are presented. RESULTS: Spastic paraplegia in patients with spatacsin mutations (n = 20) developed during the second decade of life. The Spastic Paraplegia Rating Scale (SPRS) showed severely compromised walking between the second and third decades of life (mean SPRS score, >30). Impaired cognitive function was associated with severe atrophy of the frontoparietal cortex, TCC, and bilateral periventricular white matter lesions. Progressive cortical and thalamic hypometabolism in the 18fluorodeoxyglucose PET was observed. Sural nerve biopsy showed a loss of unmyelinated nerve fibers and accumulation of intraaxonal pleomorphic membranous material. Mutational analysis of spatacsin demonstrated six novel and one previously reported frameshift mutation and two novel nonsense mutations. Furthermore, we report the first two splice mutations to be associated with SPG11. INTERPRETATION: We demonstrate that not only frameshift and nonsense mutations but also splice mutations result in SPG11. Mutations are distributed throughout the spatacsin gene and emerge as major cause for ARHSP with TCC associated with severe motor and cognitive impairment. The clinical phenotype and the ultrastructural analysis suggest a disturbed axonal transport of long projecting neurons.