Banca de QUALIFICAÇÃO: YASMIN RANA DE MIRANDA SILVA

Uma banca de QUALIFICAÇÃO de DOUTORADO foi cadastrada pelo programa.
STUDENT : YASMIN RANA DE MIRANDA SILVA
DATE: 07/06/2023
TIME: 14:00
LOCAL: São Bernardo do Campo
TITLE:

Telomerase inhibition in human neurospheres as a model for studying aging

PAGES: 87
BIG AREA: Ciências Biológicas
AREA: Fisiologia
SUBÁREA: Fisiologia de Órgãos e Sistemas
SPECIALTY: Neurofisiologia
SUMMARY:

The development of new drugs is a slow, expensive, and often inefficient process. Particularly for diseases related to the nervous system, this situation is even more complex, as human nervous tissue is difficult to access due to practical and ethical reasons. Many substances, even after being approved in preclinical tests, end up being unsuccessful in clinical trials, revealing difficulties in translational research. Limitations in the models used in preclinical tests may be at the core of this issue - thus, the use of in vitro models with human cells that possess sufficient complexity to accurately mimic the physiology of the human nervous system becomes extremely necessary. In this context, there is a type of in vitro model that is particularly interesting: the neurospheres. This model has a 3D conformation and is composed of different types of cells of interest such as NSCs, neurons, and astrocytes. The discovery of a way to induce cellular aging in neurospheres can greatly contribute to translational research, especially regarding neurodegenerative diseases. Since a significant marker of aging is telomere shortening, the present study aims to investigate whether the inhibition of telomerase, the enzyme responsible for maintaining telomere length in proliferative cells, can lead to telomere shortening in neural stem cells (NSCs) and neurospheres, promote aging-associated phenotypes, and consequently generate a 3D in vitro model to mimic aging in the central nervous system. NSCs and neurospheres were treated for a total of 28 days with 10 and 15µM of the telomerase inhibitor BIBR1532 and evaluated in different assays. The SA-β-gal assay showed a 3.5x increase in the marker in cells treated with 15µM compared to the control (from 0.31% to 1.18% in the SA-β-gal marked area/neurosphere area - p<0.0001). Regarding the immunofluorescence assays, there was a reduction in MAP2 for the 15µM treatment (p<0.01), a decrease in nestin quantity with the 10µM (p<0.0001) and 15µM (p<0.0001) treatments, and an increase in γH2Ax in the 10µM treatment (p<0.01). Telomere length was quantified by real-time PCR, and there was a reduction in telomere length of NSCs treated with 5, 10, 15, 20, and 25µM (T/S=1.1) compared to the control (T/S=1.4). The telomerase activity assay (TRAP) is still being standardized. Our results suggest that treatment with BIBR1532 decreases the telomere length of cells and produces markers associated with aging, such as increased SA-β-gal activity, decreased MAP2 quantity indicating a decrease in dendritic stability and plasticity, decreased nestin quantity suggesting a reduction in neural progenitor cells, and increased γH2Ax indicating increased DNA damage.

COMMITTEE MEMBERS:
Presidente - Interno ao Programa - 1994696 - SILVIA HONDA TAKADA
Membro Titular - Examinador(a) Externo à Instituição - LÍVIA CLEMENTE MOTTA TEIXEIRA - SantaCasa SP
Membro Titular - Examinador(a) Externo à Instituição - CRISTIANA LIBARDI MIRANDA FURTADO
Membro Suplente - Examinador(a) Interno ao Programa - 1872537 - MARCELA BERMUDEZ ECHEVERRY
Membro Suplente - Examinador(a) Interno ao Programa - 1887027 - FERNANDO AUGUSTO DE OLIVEIRA RIBEIRO