CARATTERIZZAZIONE DELL¿ORGANELLO DI NEUROMELANINA DELLA SUBSTANTIA NIGRA UMANA CON APPROCCIO PROTEOMICO

Substantia nigra is the most affected brain region in Parkinson's disease. Substantia nigra is characterized by strong pigmentation derived by the presence of neuromelanin (NM), a particular melanic pigment that accumulates during aging in neurons in typical membrane-bounded organelles with high levels of lipids (dolichols). The occurrence of an elevated number of NM organelles in the most vulnerable brain region during Parkinson’s disease is of special interest. It is not clear if NM organelles may interfere with the vulnerability of neurons during aging. Nevertheless, many evidences support the correlation between aging and neurodegeneration, and the accumulation of intracellular indigestible materials (damaged proteins, inclusion bodies and lipofuscins). The accumulation of indigestible materials inside the cell may increase cellular stress and neurodegeneration. In this study, samples of NM organelles (whole and without membranes) and NM pigment were isolated from human post-mortem tissue (substantia nigra) of healthy subjects. Samples were analysed in duplicate to obtaining the proteomic profile of the NM organelle using MudPIT proteomic technology. A total of 1142 proteins were identified, of which 48 were observed with high confidence as representative of samples. Comparing results from different samples, some proteins were assigned to different regions of the NM organelle (NM pigment, membrane, soluble portion). From the proteomic profile emerges the lysosomal nature of the NM organelle: among the 48 representative proteins, more than 20 are lysosomal proteins. In addition, other groups of proteins were identified: proteins involved in protein folding (alpha-crystallin) and degradation (ubiquitin), proteins important to iron homeostasis (ferritin L), antioxidant proteins and proteins involved in intracellular aggregates formation. Furthermore, typical biomarkers of lysosomal storage disease are observed (mitochondrial subunit c, saposins). On the other hand, any enzyme potentially involved in the melanogenic process and any lipids biosynthetic enzyme were identified. This study demonstrates that the NM organelle has the features of both a lysosomal/autophagic organelle and an aged and damaged lysosome accumulating indigestible material. These data are in agreement with recent hypothesis suggesting that the NM organelle formation originate by sequestration of NM and other potentially toxic molecules as damaged proteins, damaged lipids and metals. Our data suggest that this protective mechanism, during aging, may induce accumulation of indigestible material inside the NM organelles and, as consequence, a decreased degradative function of the cell. In summary, e suggest that NM organelle could confer protection to neurons and, at the same time, could increase cellular vulnerability in aging conditions.