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Analisys of autophagy during protein aggregation containing alpha-synuclein and hyperphosphorylated tau in cultured cells from hipoccampus, substantia nigra and locus COREULEUS

Grant number: 13/07942-1
Support type:Scholarships in Brazil - Scientific Initiation
Effective date (Start): June 01, 2013
Effective date (End): May 31, 2014
Field of knowledge:Biological Sciences - Morphology - Cytology and Cell Biology
Principal Investigator:Merari de Fátima Ramires Ferrari
Grantee:Vinícius de Andrade Torrecillas
Home Institution: Instituto de Biociências (IB). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Changes in cell structure and biochemistry, formation of senile plaques and neurofibrillary tangles are common features of the brain in degenerative process. These neurofibrillary tangles are consisted essentially of hyperphosphorylated tau protein. The presence of these protein aggregates leads to intracellular damage culminating in cell death. Failure in the process of cellular degradation could favor the aggregation of intracellular and extracellular proteins and culminate in neurodegeneration. For this reason, we intend to analyze the expression of proteins involved in cellular autophagy, the formation of autophagolysosomes and the colocalization of autophagosomes with hyperphosphorylated tau and alpha-synuclein, in order to study the relation between autophagy and protein aggregation. For this analysis, specific antibodies will be used to detect proteins of the autophagy pathway such as Rab7, p62, Ulk1 and Beclin-1, in addition to antibodies for the labeling of hyperphosphorylated tau and alpha-synuclein. The analysis of autophagolysosomes will be performed using fluorescent probes which mark acidic organelles with different colors depending upon the pH range. This study will be performed on primary cell cultures of the hippocampus, substantia nigra and locus coeruleus subjected to rotenone exposure in concentrations from 0.1 to 0.5 nM, considering that 0.5nM is able to induce aggregates containing hyperphosphorylated tau protein and alpha-synuclein in the cells.