Banca de DEFESA: LETÍCIA DIAS DOS SANTOS SILVA

Uma banca de DEFESA de MESTRADO foi cadastrada pelo programa.
STUDENT : LETÍCIA DIAS DOS SANTOS SILVA
DATE: 26/02/2025
TIME: 09:00
LOCAL: A definir
TITLE:

Expression of miR-7 and the enzyme PARP1 in SH-SY5Y dopaminergic cells exposed to the neurotoxin MPP+.

KEY WORDS:

PARP1; miR-7; microRNA; Parkinson’s disease; neurodegeneration.

PAGES: 100
BIG AREA: Ciências da Saúde
AREA: Medicina
SUMMARY:

The current study aimed to examine the expression of microRNA 7 (miR-7) and the enzyme poly(ADP-ribose) polymerase 1 (PARP1) in a model of dopaminergic cell injury using SHSY5Y cells exposed to MPP+. miR-7 plays a neuroprotective role in the underlying mechanisms of Parkinson's disease (PD) by reducing the accumulation of synuclein in Lewy bodies, as well as mitigating oxidative stress and apoptosis, among other factors. Moreover, miR-7 triggers the post-transcriptional silencing of the enzyme PARP1, thereby regulating its content. PARP1, in turn, has been implicated in the death of dopaminergic neurons in PD through the parthanatos pathway, which generates PAR polymers that interact with misfolded alpha-synuclein molecules, increasing the deposition of this protein in Lewy bodies. In this context, PARP1 inhibitors have proven effective in preventing the increase of PAR polymers and in attenuating motor deficits in animal models of experimental parkinsonism. Our group has already demonstrated that rotenone-induced parkinsonism results in a reduction of miR-7 in the striatum, accompanied by loss of dopaminergic neurons and motor impairment. The present study investigates whether this reduction of miR-7 is correlated with an increase in PARP1 expression and whether miR-7 supplementation through synthetic mimics (miR-7 mimics) could mitigate the effects of the neurotoxin MPP+, suggesting a neuroprotective effect. The cells were cultured in DMEM medium supplemented with fetal bovine serum (FBS), Glutamax, and an antibiotic/antimicrobial solution, and exposed to MPP+ for 24 hours. Cell viability was assessed through the MTT colorimetric assay, while miR-7 expression was determined by RT-qPCR. RNA was purified using commercial kits (RNeasy Plus Mini kit, Qiagen, and mirVana™, Invitrogen). For RT-qPCR of miRNAs, cDNA synthesis and qPCR were performed using the TaqMan™ MicroRNA Reverse Transcription and TaqMan™ Fast Advanced Master Mix kits. For PARP1 RT-qPCR, cDNA synthesis and qPCR were carried out with random primers (SuperScript II First-Strand Synthesis System, Invitrogen) and the SYBR Green method (Fast SYBR Green Master Mix, Applied Biosystems). Data were analyzed using the ∆∆Ct method, normalizing with endogenous genes and spike-ins. Exposure to MPP+ resulted in a significant, dose-dependent reduction in cell viability at concentrations of 0.5 mM, 1.0 mM, 2 mM, and 4 mM, with reductions of 87%, 81%, 59%, 34%, and 15%, respectively (P<0.05). Regarding miR-7, SH-SY5Y cells exposed to 1 mM MPP+ for 24 hours showed a reduction of miR-7 to 0.690 (1 mM) and 0.461 (2 mM). PARP1 expression was also analyzed using RT-qPCR, with GPB1 and GAPDH as calibrators. Our data indicate a decrease of 0.862 times in PARP1 expression in SH-SY5Y cells exposed to 1 mM, and a decrease of 0.790 times in cells exposed to 2 mM of MPP+ for 24 hours. In conclusion, the results demonstrate the reduction of gene expression of miR-7, a microRNA directly involved in controlling the most studied pathological protein in PD, alpha synuclein. This finding reinforces the role of miR-7 in the neuropathology of Parkinson's disease, considering that the decrease in miR-7 may result in the accumulation of the protein it regulates. Additionally, alterations in the expression of the enzyme PARP1 may lead to cellular changes that result in the death of dopaminergic cells. Thus, the present study highlights the potential role of two molecules involved in the underlying mechanisms of PD, contributing to a better understanding of this neurodegenerative disease and suggesting potential targets for innovative therapies aimed at slowing its progression. 

COMMITTEE MEMBERS:
Externa à Instituição - CLARA LUNA FREITAS MARINA - UnB
Externo ao Programa - 1156673 - FELIPE VON GLEHN SILVA - UnBExterno à Instituição - FERNANDO FRANCISCO BORGES RESENDE - UNICEPLAC
Presidente - 1127261 - RICARDO TITZE DE ALMEIDA