Consensus Paper: Radiological Biomarkers of Cerebellar Diseases

Hereditary and sporadic cerebellar ataxias represent a vast and still growing group of diseases whose diagnosis and differentiation cannot only rely on clinical evaluation. Brain imaging including magnetic resonance (MR) and nuclear medicine techniques allows for characterization of structural and functional abnormalities underlying symptomatic ataxias. These methods thus constitute a potential source of radiological biomarkers, which could be used to identify these diseases and differentiate subgroups of them, and to assess their severity and their evolution. Such biomarkers mainly comprise qualitative and quantitative data obtained from MR including proton spectroscopy, diffusion imaging, tractography, voxel-based morphometry, functional imaging during task execution or in a resting state, and from SPETC and PET with several radiotracers. In the current article, we aim to illustrate briefly some applications of these neuroimaging tools to evaluation of cerebellar disorders such as inherited cerebellar ataxia, fetal developmental malformations, and immune-mediated cerebellar diseases and of neurodegenerative or early-developing diseases, such as dementia and autism in which cerebellar involvement is an emerging feature. Although these radiological biomarkers appear promising and helpful to better understand ataxia-related anatomical and physiological impairments, to date, very few of them have turned out to be specific for a given ataxia with atrophy of the cerebellar system being the main and the most usual alteration being observed. Consequently, much remains to be done to establish sensitivity, specificity, and reproducibility of available MR and nuclear medicine features as diagnostic, progression and surrogate biomarkers in clinical routine

Nerve diffusion tensor imaging

Diffusion tensor imaging (DTI) is a magnetic resonance imaging technique that in vivo visualises random translational movement of water molecules. DTI has inherent difficulties with low signal-to-noise ratio, sensitivity to patient motion, field inhomogeneities and fast T2 relaxation. It has been used in the central nervous system, although it has not been assessed in the peripheral nervous system. The aim of this thesis was to investigate if DTI in peripheral nerves was feasible, and if so, to investigate clinical implications. Study I showed that in healthy volunteers the peripheral nerves, the sciatic nerves, could be visualised in vivo using DTI and fiber tracking. Study II showed that sciatic nerves, including their division into the tibial and common fibular nerves, have a characteristic diffusion pattern with most impaired diffusion perpendicular to the nerve direction. This allowed nerves, excluding other tissues and artifacts, to be visualised using a novel approach called diffusion-direction- dependent imaging and with a simple unidirectional diffusion maximum-intensity projection approach. Study III showed that the olfactory bulbs (OBs) and olfactory tracts could be visualised in vivo using DTI and fiber tracking. In Study IV, Parkinson’s disease (PD) patients with impaired olfaction were evaluated with DTI of the OBs. A novel approach of DTI was used, taking advantage of the technique’s inherent directional information, for region of interest placement and diffusion measurements in the OBs. In the PD patient group diffusion was altered in the OBs, compared to healthy controls. This was hypothesised, since α-synuclein inclusions and Lewy neurites interfering with nerve structure have been detected in the OBs. However, the coefficient of variation between two identical DTI series was high, due to the small size of the OBs and their location in an area susceptible to artifacts, and the difference between the groups was statistically significant only for the first of two series. In Study V, patients of the Swedish ‘Huddinge Spinocerebellar ataxia (SCA) Family’ with peripheral neuropathy, were evaluated with DTI of a peripheral nerve. Diffusion alterations were found in peripheral nerves in SCA patients, compared to healthy controls, which was statistically significant. In conclusion, DTI in peripheral nerves is feasible and can be used to detect diffusion alterations in OBs in PD patients and in peripheral nerves in SCA patients with peripheral neuropathy

Oxidative stress in DNA repeat expansion disorders: a focus on NRF2 signaling involvement

DNA repeat expansion disorders are a group of neuromuscular and neurodegenerative diseases that arise from the inheritance of long tracts of nucleotide repetitions, located in the regulatory region, introns, or inside the coding sequence of a gene. Although loss of protein expression and/or the gain of function of its transcribed mRNA or translated product represent the major pathogenic effect of these pathologies, mitochondrial dysfunction and imbalance in redox homeostasis are reported as common features in these disorders, deeply affecting their severity and progression. In this review, we examine the role that the redox imbalance plays in the pathological mechanisms of DNA expansion disorders and the recent advances on antioxidant treatments, particularly focusing on the expression and the activity of the transcription factor NRF2, the main cellular regulator of the antioxidant response

Magnetic resonance imaging of the substantia nigra in parkinson’s disease : neuromelanin, iron and diffusion tensor imaging

Tese de doutoramento, Medicina (Imagiologia), Universidade de Lisboa, Faculdade de Medicina, 2015In the last years, extensive developments in neuroimaging MR techniques have profoundly changed the study of Parkinson’s disease (PD), evolving from the role of excluding secondary parkinsonism to the emergence as a disease biomarker. MR advanced sequences in high field magnets opened the possibility to visualize in vivo the substantia nigra (SN) and to investigate specific PD pathological changes, enabling the development of high accuracy tools for disease diagnosis in early stages and for the comprehension of disease pathophysiology. Our work was centered on the application of new MR imaging techniques to study the SN in PD, early in the disease course, mainly focusing on untreated patients at the time of clinical diagnosis. The primary objectives were centered on the application of high field MR imaging sequences in two main areas: diagnosis of PD in early disease stages and differential diagnosis with Essential tremor (ET). The development and application of neuromelanin sensitive MR imaging in 3.0 Tesla allowed the detection of significant changes in the SN of PD patients, with high sensitivity and specificity for disease diagnosis, even in early disease stages (namely at the time of clinical diagnosis). These imaging findings reproduced in vivo the characteristic pathological changes of PD with greater alteration in the ventrolateral SN region and preservation of the dorsal segment. These results were obtained with several image evaluation methods: semi-automated area assessments, manual width measurements and simple visual inspection by Neuroradiologists, corroborating the reproducibility of the data and enabling wider applications of this image technique in the clinical practice. The MR correlation of neuromelanin with iron in the SN of PD patients allowed the in vivo investigation of the influence of local iron concentration in the SN on the signal of neuromelanin-sensitive sequences. A quantification T2-relaxometry study showed that the SN paramagnetic iron effects do not seem to influence significantly the neuromelanin MR signal reduction in PD patients. Several studies with diffusion tensor MR imaging (DTI) have allowed the detection of microstructural changes in the SN of PD patients in early disease changes, emerging as a possible disease biomarker. So, the reproducibility of DTI metrics in this specific brain area was particularly relevant for future applications of this MR technique. We conducted a reproducibility DTI study in PD patients that showed a good reproducibility of DTI metrics supporting the use of these measurements in further studies, namely longitudinal within-subject evaluation, and cross-sectional comparisons. The differential diagnosis of PD with ET is particularly relevant and there was the need of high accurate tools to aid the clinical assessment. The application of neuromelanin-sensitive MR techniques was able to discriminate ET from early stage tremor-dominant PD with high sensitivity and specificity values, in the same range as nuclear medicine techniques and may become a useful clinical tool in the evaluation of tremor disorders. Our research showed an important role of neuromelanin sensitive MR imaging for the diagnosis PD in early disease stages and its differential diagnosis with ET. A multi-modal MR approach with iron assessment and diffusion tensor imaging can further elucidate the SN disease changes and aid future research of disease pathophysiology.Nos últimos anos, o extenso desenvolvimento das técnicas de neuroimagem modificou profundamente a investigação da Doença de Parkinson (PD), evoluindo de um simples papel na exclusão de parkinsonismo secundário para a emergência de biomarcadores imagiológicos da doença. Sequências avançadas de RM em aparelhos de alto campo magnético abriram a possibilidade de visualizar in vivo a substantia nigra (SN) e a investigação de alterações patológicas específicas da PD, permitindo o desenvolvimento de ferramentas com elevada fiabilidade para o diagnóstico em fases precoces da evolução da doença e para a compreensão da sua fisiopatologia. A nossa investigação centrou-se na aplicação de novas técnicas de imagem RM para estudar a SN na PD, em fases precoces da doença, com enfoque especial em doentes não tratados na altura do diagnóstico clínico. Os objectivos principais centraram-se na aplicação de sequências de RM em alto campo em duas áreas major: diagnóstico da PD em fases precoces da doença e o diagnóstico diferencial com o Tremor essencial (ET). O desenvolvimento e aplicação da imagem RM sensível à neuromelanina em 3.0T permitiu a detecção de alterações significativas na SN de doentes com PD, com elevada sensibilidade e especificidade para o diagnóstico da doença, mesmo em fases precoces da sua evolução dela (nomeadamente na altura do diagnóstico clínico). Estes achados de imagem reproduziram in vivo as alterações patológicas características da PD, com uma maior alteração na região ventero-lateral da SN e preservação do segmento dorsal. Estes resultados foram obtidos com vários métodos de avaliação de imagem: avaliação semi-automática da área, medição manual da espessura e avaliação visual por neurorradiologistas, corroborando a reproductibilidade dos dados e permitindo uma aplicação abrangente desta técnica de imagem na prática clínica. A correlação por RM da neuromelanina com o ferro, na SN de doentes com PD, permitiu a investigação in vivo da influência da concentração local de ferro na SN com o sinal das sequências sensíveis à neuromelanina. Um estudo quantitativo de relaxometria T2* mostrou que os efeitos paramagnéticos do ferro não influenciam significativamente a redução de sinal RM da neuromelanina em doentes com PD. Vários estudos com tensores de difusão (DT) permitiram a detecção de alterações microestruturais na SN de doentes com PD em fases precoces de doença, emergindo como um possível biomarcador de doença. Assim, a reproductibilidade das métricas de DTI, nesta área específica do encéfalo, é particularmente relevante para aplicações futuras desta técnica de RM. Conduzimos um estudo de reproductibilidade de DTI em doentes com PD que demonstrou uma boa reproductibilidade das métricas de DTI, suportando a utilização destas medidas em estudos futuros, nomeadamente avaliações longitudinais “within-subject” e comparações “cross-sectional”. O diagnóstico diferencial da PD com ET é particularmente relevante e ferramentas fiáveis para auxiliar a avaliação clínica eram necessárias. A aplicação de técnicas de RM sensíveis à neuromelanina possibilitou a discriminação de ET de PD “tremor-dominant” em fases precoces com elevados valores de sensibilidade e especificidade, no mesmo espectro das técnicas de medicina nuclear e pode tornar-se uma ferramenta clínica útil para a avaliação do tremor. A nossa investigação demonstrou um importante papel das técnicas de RM sensíveis à neuromelanina para o diagnóstico de PD em fases precoces da doença e para o seu diagnóstico diferencial com ET. Uma abordagem multi-modal de RM com avaliação do ferro e DTI pode, adicionalmente, permitir estudar as alterações da SN e auxiliar a investigação futura da fisiopatologia da doença

Pathophysiological mechanisms in Parkinson`s Disease and Dystonia – converging aetiologies

In this thesis I used a range of experimental approaches including genetics, enzyme activity measurements, histology and imaging to explore converging pathophysiological mechanisms of Parkinson`s disease and dystonia, two conditions with frequent clinical overlap. First, based on a combined retro- and prospective cohort of patients, using a combination of lysosomal enzyme activity measurements in peripheral blood and brain samples, as well as a target gene approach, I provide first evidence of reduced levels of enzyme activity in Glucocerebrosidase and the presence of GBA mutations, indicating lysosomal abnormality, in a relevant proportion of patients with dystonia of previously unknown origin. Second, based on a retrospective cohort of patients, I detail that a relevant proportion of genetically confirmed mitochondrial disease patients present with a movement disorder phenotype - predominantly dystonia and parkinsonism. Analysing volumetric MRI data, I describe a patterned cerebellar atrophy in these particular patients. This also includes the first cases of isolated dystonia due to mitochondrial disease, adding the latter as a potential aetiology for dystonia of unknown origin. Third, I used a combination of post-GWAS population genetic approaches and tissue-based experiments to explore in how far the strong association between advancing age and Parkinson ́s disease is mediated via telomere length. Although the initial finding of an association between genetically determined telomere length and PD risk did not replicate in independent cohorts, I provide evidence that telomere length in human putamen physiologically shortens with advancing age and 3 is regulated differently than in other brain regions. This is unique in the human brain, implying a particular age-related vulnerability of the striatum, part of the nigro-striatal network, crucially involved in PD pathophysiology. I conclude by discussing the above findings in light of the current literature, expand on their relevance and possible direction of future experiments