Data di Pubblicazione:
2019
Citazione:
PROTEOMIC ANALYSIS OF NEURONAL CELLS AND TISSUES / F. Grassi Scalvini ; tutor: G. Tedeschi ; coordinatore: A. Prinetti. DIPARTIMENTO DI MEDICINA VETERINARIA, 2019 Dec 05. 32. ciclo, Anno Accademico 2019. [10.13130/grassi-scalvini-francesca_phd2019-12-05].
Abstract:
Abstract
During my three years of PhD I had the opportunity to work with different biological samples analyzing
the content and type of proteins through the proteomic approach.
In the thesis I describe how a proteomic approach could be useful to analyze different aspect connected
to central nervous system. Indeed, the present work is divided in three different parts but the fil rouge is
represented by the same analysis technique, a shotgun label-free proteomic approach, for the identification and
quantification of expressed proteins, applied to different neuronal cells and tissues: 1) PC12 cells, a wellstudied
neuronal cell model due to the ability to easily differentiate into neuron-like cells, 2) Neuro2a cells,
another neuronal cellular model very well studied, and 3) the brain of Zebrafish. It is a poikilotherm and
eurytherm and therefore it has a wide thermal tolerance, from 6°C to 38°C, temperatures between 24 and 30°C
are more suitable for its development, growth and reproduction. Moreover, Danio rerio represents a good
animal model for different type of research because it has a generation time of 3-4 months, its maintenance is
cheaper than that for rats and mice and required little space.
Starting with the first project the proteomic approach is used to dissect at the proteome level similarities
and differences between the biochemically and mechanotransductively promoted neuronal differentiation of
PC12 cells growth on cluster-assembled zirconia surface with 15nm of roughness, polylysine coated glass in
the presence (NGF) or absence (PLL) of NGF.
This work lays a substantial cell biological foundation for the intelligent design of substrates for cell
culturing based on nanostructured surfaces produced by cluster assembling that mimic more closely
physiological 3D extracellular microenvironmental features. Our data suggest that the nanoscale information
provided by these surfaces could have a strong potential in favoring neurogenic processes by mechanotransductive
processes.
The results obtained on zirconia nanostructure can be the fundamental starting point to further
characterize the neuronal differentiation process in adequate primary and stem cell systems for regenerative
medicine approach.
The second part of this work has the aim to understand how the activation of the TrkA pathway is able
to trigger biochemical signaling, like ERK1/2, leading to cell differentiation when GM1 oligosaccharide,
II3Neu5Ac-Gg4 (OligoGM1), which interacts with NGF receptor TrkA, is administered to cultured murine
Neuro2a neuroblastoma cells.
The results of our work confirm and reinforce the idea that the molecular mechanisms underlying the
GM1 neurotrophic and neuroprotective effects depend on its oligosaccharide chain, suggesting the activation
of a positive signaling starting at plasma membrane level.
The third part of this thesis regards the determination of the effect of ambient temperature on the
molecular mechanism and the behavioural responses in Danio rerio. This project can be framed in a quite
interesting area of research, important because global warming occurring in our planet is, especially nowadays,
an urgent problem amplified by the anthropic action, the release of CO2 and other greenhouse gases. The huge
temperature increase causes climate changes that can deeply alter the habitat of the species, leading to
substantial environmental changes possibly impairing the prosecution of the species. We applied for the first
time a shotgun proteomic approach to analyze the effect of acclimatization on zebrafish brain proteome and to
correlate the results at the protein level with the behavioural tests.
As stated above, the shotgun proteomic approach adopted is a powerful analytical method for
Tipologia IRIS:
Tesi di dottorato
Elenco autori:
F. GRASSI SCALVINI
Link alla scheda completa: