Motor Neuron Diseases (MND):
The Laboratory of Neuroscience is very busy in the study of the genetic and molecular bases of ALS as well as in the collection and characterization of cellular models of the disease obtained from the patient himself, as e.g. myoblast and fibroblast cells turned afterwards into induced pluripotent stem cells (iPSC). The Laboratory has available a large DNA bank as well as a serum and liquor collection from about 850 ALS cases with both sporadic and familiar disease, with or without known genetic mutations. Two Exome-sequencing projects carried out with the collaboration of Dr. John Landers of the University of Massachusetts Medical School, Worchester, USA, implied the sequencing of the entire exome, that is to say the whole exomic and therefore codifying portion of the human genome. DNA samples were obtained from a vast number of familiar forms without mutations in known genes and of sporadic forms with both living parents (TRIOS). The purpose was to identify new potential genes responsible for the disease. In 2012, through this approach of New Generation Sequencing, our group was able to identify a new gene, Profiline 1 (Wu et al, Nature, 2012). New candidate genes are actively studied. At the same time our Laboratory is focused on the identification of genetic risk factors for the sporadic forms through studies of the Genome-Wide Association (GWAs) of genetic polymorphisms (SNPs). For that purpose the SLAGEN Consortium has been created, having its location in our Institution. SLAGEN brings together 6 Centers referential for ALS in Italy and collected more than 3000 DNA samples of patients with the sporadic form of ALS. Two candidate loci have been recently defined (Fogh et al., 2014). The Laboratory together with the SLAGEN Consortium put also effort into the study of the genetic epidemiology of ALS in Italy, in collaboration with other International Groups with the target of defining the genetic factors of predisposition for ALS supported by different European projects.
Concerning the study of the molecular bases of ALS, ongoing studies aim to examine the role of the RNA- binding proteins TDP-43 and FUS in neural cells. TDP-43 and FUS are indeed mainly nuclear proteins and are ubiquitously expressed, but only in the ALS affected patients’ motor neurons they are subjected to a shift in sub-cellular localization forming potentially toxic protein aggregates in the cytoplasm. By defining the biological functions of these two proteins and of their molecular interactors, it is aimed to understand in how their malfunctioning can induce the motor neuronal selective degeneration. With this in mind the Laboratory of Neuroscience has recently contributed to prove that TDP-43, but not FUS, participates actively to the damage responses that start in the cells after stressful stimuli and producing “stress granules”, structures with a protective role able to stop temporarily the protein translation. Our experimental data show also that TDP-43 and FUS have different and barely superimposable roles in the regulation of the RNA metabolism in neuronal cells. There are different experimental approaches and cellular models that are used for that purpose, among which cellular models of the disease, which are directly obtained from patients (fibroblasts, myoblasts) also after the transformation into stem cells (iPSC).
Cognitive and Behavioral Disorders:
The Laboratory of Neuroscience collaborates in this clinical area supplying the genetic definition of some familiar forms of FTD, ALS/FTD and Cerebral Amyloid Angiopathy overlapping with Alzheimer’s Disease (definition of mutation for coding genes APP, ApoE, TTR, CST3, GSN). The study of the Cerebral Amyloid Angiopathy turned out to be of particular interest considering the vast amount of records collected through the influx of patients also through the Stroke Unit.
Parkinson’s Disease and Parkinsonisms:
The mutations of the gene PARK2, which codifies for the parkin protein, cause the most common form of juvenile Parkinsonism with autosomal recessive transmission (ARJP). A vast selection of the gene’s mutation associates with the disease, such as the substitution of single base pairs, little deletions or insertions of one or more base pairs and exomic rearrangements. In most of the cases the deletions or mutations in homozygosity or compound heterozygosity lead to the loss of expression of the parkin protein in the patients’ brain. These data sustain the idea of functional loss as a predominant pathogenetic mechanism in the PARK2-mediated Parkinsonism. The neuropathological analysis of brain of PARK2 mutation carrying patients has shown a selective loss of the dopaminergic neurons (DA) in the substantia nigra (SN), nevertheless, although the PARK2 gene has been identified more than 10 years ago, not much is known about the mechanism with which the mutant protein induces a progressive neuronal dysfunction and death. The excitotoxicity hypothesis has been inserted in Parkinson’s disease’s etiology, for the CNS’s neurons receive glutamatergic inputs from different areas of the brain, such as the cortex, pedunculopontine nucleus, thalamus and subthalamic nucleus. Concerning the ARJP, growing evidence suggests that parkin Wild-Type (WT) can modulate the glutamatergic receptors’ function and that the loss of parkin function in the brain of PARK2 patients makes the DA neurons more susceptible for the glutamate toxicity. Effectively, the silencing of WT-parkin sensitizes the mesencephalic neurons to kainate toxicity. On the contrary, the hyperexpression of WT-parkin protects the mesencephalic neurons from the kainate-mediated excitotoxicity. A more recent study has confirmed that parkin controls the function and stability of the excitatory glutamatergic synapses and it has also shown that the postsynaptic expression of mutant parkin is associated with a higher vulnerability to synaptic exitotoxicity. This evidence suggests a link between parkin and the function of the glutamatergic synapses, permitting the hypothesis that the loss of DA neurons in parkin-mediated ARJP could derive from a higher vulnerability to the glutamatergic stimulation. The research project undertaken in our Laboratory aims to identify molecular mechanisms through which parkin can modulate the glutamate receptors’ activity and to inspect whether those receptors could represent a new therapeutic target for ARJP, caused by mutations in PARK2.
The Huntington’s Disease is a hereditary neurologic disease that causes the progressive dysfunction and loss of neurons in different areas of the brain. The affection was reported for the first time by the physician George Huntington who described in 1872 the main clinical aspects of the disease: the appearance of behavioral disorders, the cognitive deficits and the movement alterations. In 1993 the cause for Huntington’s Disease was identified: the mutation of a gene called IT-15 localized on the short arm of chromosome 4. From that day on a lot has been understood about the molecular mechanisms that lead to the progressive loss of neuronal striatal cells, although there isn’t yet an efficient therapy for delaying the progress or preventing the onset of the disease in patients carrying the mutation. Nevertheless, today there are numerous prescriptions able to reduce significantly the motor and psychiatric symptoms, considerably improving the life standard of patients affected by MdH. The Laboratory of Neuroscience has recently dedicated its attention to define the molecular mechanisms of mitochondrial dysfunction in Huntington’s Disease. The further molecular mechanisms’ identification is the basis for development of new therapeutic strategies.
There are different research activities that are linked to stem cells, particularly for the treatment of neurodegenerative diseases that have been pursued in the Laboratory of Neuroscience for various years. The research’s first sector concerns the use of human mesenchymal stem cells for regenerative purposes in an animal model of Parkinson’s Disease. It has been verified that the transplanted cells have a protective effect and stimulate the endogenous neurogenesis. Furthermore, the transplanted cells have demonstrated a phenotypic change towards the glial phenotype expressing specific astrocyte markers, absent in the cell culture before the transplant. The transplanted cells’ traceability is of great importance for any project. For that aim different biomarkers have been used (IR colorants, magnetic nanoparticules) and the toxicity on different stem cell types (mesenchymal, from amniotic liquid, and chorionic villi) as well as the ease of assaying have been evaluated. A second sector of research concerns ALS with the aim of identifying disease biomarkers to define an accurate and early diagnosis. Furthermore, through the study of differentiated neuronal cells obtained from iPSC coming from affected patients’ fibroblasts, we intended to study and underline possible pathogenetic mechanisms at the basis of the disease’s onset, mainly in relation to the most common genetic forms.