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Vande Velde G.,Catholic University of Leuven | Rangarajan J.R.,Catholic University of Leuven | Toelen J.,Catholic University of Leuven | Dresselaers T.,Catholic University of Leuven | And 10 more authors.
Gene Therapy | Year: 2011

The development of in vivo imaging protocols to reliably track transplanted cells or to report on gene expression is critical for treatment monitoring in (pre)clinical cell and gene therapy protocols. Therefore, we evaluated the potential of lentiviral vectors (LVs) and adeno-associated viral vectors (AAVs) to express the magnetic resonance imaging (MRI) reporter gene ferritin in the rodent brain. First, we compared the induction of background MRI contrast for both vector systems in immune-deficient and immune-competent mice. LV injection resulted in hypointense (that is, dark) changes of T 2 /T 2 (spin-spin relaxation time)-weighted MRI contrast at the injection site, which can be partially explained by an inflammatory response against the vector injection. In contrast to LVs, AAV injection resulted in reduced background contrast. Moreover, AAV-mediated ferritin overexpression resulted in significantly enhanced contrast to background on T 2-weighted MRI. Although sensitivity associated with the ferritin reporter remains modest, AAVs seem to be the most promising vector system for in vivo MRI reporter gene imaging. © 2011 Macmillan Publishers Limited All rights reserved.

Van der Perren A.,Laboratory for Neurobiology and Gene Therapy | Casteels C.,Leuven University Hospital | Van Laere K.,Leuven University Hospital | Gijsbers R.,Laboratory for Molecular Virology and Gene Therapy | And 4 more authors.
Journal of Visualized Experiments | Year: 2016

In order to study the molecular pathways of Parkinson’s disease (PD) and to develop novel therapeutic strategies, scientific investigators rely on animal models. The identification of PD-associated genes has led to the development of genetic PD models. Most transgenic α-SYN mouse models develop gradual α-SYN pathology but fail to display clear dopaminergic cell loss and dopamine-dependent behavioral deficits. This hurdle was overcome by direct targeting of the substantia nigra with viral vectors overexpressing PD-associated genes. Local gene delivery using viral vectors provides an attractive way to express transgenes in the central nervous system. Specific brain regions can be targeted (e.g. The substantia nigra), expression can be induced in the adult setting and high expression levels can be achieved. Further, different vector systems based on various viruses can be used. The protocol outlines all crucial steps to perform a viral vector injection in the substantia nigra of the rat to develop a viral vector-based alpha-synuclein animal model for Parkinson's disease. © 2016 Journal of Visualized Experiments.

Van Camp N.,University of Antwerp | Vreys R.,University of Antwerp | Van Laere K.,University Hospital Leuven | Lauwers E.,Laboratory for Neurobiology and Gene Therapy | And 10 more authors.
Magnetic Resonance Materials in Physics, Biology and Medicine | Year: 2010

Object: In the present study, we aimed to evaluate the impact of neurodegeneration of the nigrostriatal tract in a rodent model of Parkinson's disease on the different MR contrasts (T2, T1, CBF and CBV) measured in the striatum. Material and methods: Animals were injected with 6-hydroxydopamine (6OHDA) in the substantia nigra resulting in massive loss of nigrostriatal neurons and hence dopamine depletion in the ipsilateral striatum. Using 7T MRI imaging, we have quantified T2, T1, CBF and CBV in the striata of 6OHDA and control rats. To validate the lesion size, behavioral testing, dopamine transporter μSPECT and tyrosine hydroxylase staining were performed. Results: No significant differences were demonstrated in the absolute MRI values between 6OHDA animals and controls; however, 6OHDA animals showed significant striatal asymmetry for all MRI parameters in contrast to controls. Conclusions: These PD-related asymmetry ratios might be the result of counteracting changes in both intact and affected striatum and allowed us to diagnose PD lesions. As lateralization is known to occur also in PD patients and might be expected in transgenic PD models as well, we propose that MR-derived asymmetry ratios in the striatum might be a useful tool for in vivo phenotyping of animal models of PD. © 2010 ESMRMB.

Fiesel F.C.,Hertie Institute for Clinical Brain Research | Voigt A.,RWTH Aachen | Weber S.S.,Hertie Institute for Clinical Brain Research | Van Den Haute C.,Laboratory for Neurobiology and Gene Therapy | And 15 more authors.
EMBO Journal | Year: 2010

TDP-43 is an RNA/DNA-binding protein implicated in transcriptional repression and mRNA processing. Inclusions of TDP-43 are hallmarks of frontotemporal dementia and amyotrophic lateral sclerosis. Besides aggregation of TDP-43, loss of nuclear localization is observed in disease. To identify relevant targets of TDP-43, we performed expression profiling. Thereby, histone deacetylase 6 (HDAC6) downregulation was discovered on TDP-43 silencing and confirmed at the mRNA and protein level in human embryonic kidney HEK293E and neuronal SH-SY5Y cells. This was accompanied by accumulation of the major HDAC6 substrate, acetyl-tubulin. HDAC6 levels were restored by re-expression of TDP-43, dependent on RNA binding and the C-terminal protein interaction domains. Moreover, TDP-43 bound specifically to HDAC6 mRNA arguing for a direct functional interaction. Importantly, in vivo validation in TDP-43 knockout Drosophila melanogaster confirmed the specific downregulation of HDAC6. HDAC6 is necessary for protein aggregate formation and degradation. Indeed, HDAC6dependent reduction of cellular aggregate formation and increased cytotoxicity of polyQ-expanded ataxin-3 were found in TDP-43 silenced cells. In conclusion, loss of functional TDP-43 causes HDAC6 downregulation and might thereby contribute to pathogenesis. © 2010 European Molecular Biology Organization.

Brito-Armas J.M.,University of La Laguna | Baekelandt V.,Laboratory for Neurobiology and Gene Therapy | Castro-Hernandez J.R.,University of La Laguna | Gonzalez-Hernandez T.,University of La Laguna | And 2 more authors.
Histology and Histopathology | Year: 2013

Two hallmarks of Parkinson's disease (PD) are dopaminergic cell loss and the presence of cytoplasmic inclusions (Lewy bodies). Different point mutations in alpha-synuclein, the main constituent of Lewy bodies, have been identified in familial PD. Alpha-synuclein also constitutes one of the main components of Lewy bodies in sporadic cases of PD. Moreover, oxidant stress and generation of free radicals from both mitochondrial impairment and dopamine metabolism are considered to play critical roles in PD etiopathogenesis. Melatonin, a known potent antioxidant secreted by the pineal gland, may protect against the effect of several Parkinsonogenic compounds that are associated with progressive impairment of mitochondrial function and increased oxidative damage. However, the neuroprotective effect of melatonin has never been tested in the newly available genetic models of PD based on the viral expression of mutated alpha-synuclein. Lentiviral vectors encoding A30P mutant human alphasynuclein (lenti-A30P) were stereotactically injected into the right substantia nigra of adult male Sprague-Dawley rats and neuroprotection was examined by administration of melatonin or vehicle from two days before nigral administration of lenti-A30P until eight weeks after injection. It was found that lenti-A30P induced a significant TH+ cell-loss both in the medial and lateral substantia nigra versus the contrallateral side injected with lenti-eGFP. However, melatonin administration showed a total neuroprotective effect in both regions of the substantia nigra. In conclusion, the data here show that melatonin is neuroprotective against mutant alpha-synuclein-induced injury in the substantia nigra.

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