Contributor:

Łukomska, Barbara (Promotor) ; Stanaszek, Luiza Róża (Promotor pomocniczy)

Publisher:

Instytut Medycyny Doświadczalnej i Klinicznej im.M. Mossakowskiego PAN

Place of publishing:

Warszawa

Degree name:

doktor

Level of degree:

2

Degree discipline :

nauki medyczne

Abstract:

I aimed to investigate the influence of neuregulin-1 (NRG-1) on the functional properties of glial-restricted progenitor cells (GRPs). The hypothesis I sought to verify was that overexpressing neuregulin-1 in GRPs would alter their functional characteristics, potentially enhancing their application potential in the therapy of demyelination-related diseases. The study focused on GRPs isolated from mouse fetal nervous tissue (referred to as mGRPs). To induce overexpression of neuregulin-1 type I (NRG-1 type I), genetic modifications were performed on mGRPs through the transduction of cells using lentiviral vectors encoding NRG-1 type I. The research involved analyzing the expression levels of NRG-1 in the modified mGRPs, examining their phenotype, and investigating the functional properties of mGRPs-NRG-1 in vitro. Evaluation of mGRPs transduced with the pLenti-GIII-CMV-NRG-1 type I-mCherry vector demonstrated that these cells exhibit varying levels of NRG-1 type I transcript synthesis in subsequent transduction attempts, and their continued culture leads to spontaneous detachment of cells from the surface of the culture vessel and subsequent cell death. The unfavorable results obtained from transducing mGRPs with the pLenti-GIII-CMV-NRG-1 type I-mCherry vector prompted me to use the HIV-SFFV-NRG-1 type I-IRES-mRFP vector for the next phase of the study. This vector encodes NRG-1 type I sequence and the mRFP fluorescent reporter protein, separated by the IRES (Internal Ribosome Entry Site) sequence. This arrangement allows for the coexpression of both sequences under the transcriptional control of a shared promoter, while their translation occurs independently. In the next stage of the study, I attempted to transduce mGRPs with lentiviral activating particles designed to overexpress the endogenous NRG-1 gene (NRG-1-LAPs). However, simultaneous transduction of mGRPs with the three lentiviral vectors comprising the NRG-1-LAPs system proved ineffective. Therefore, the attempt to induce endogenous NRG-1 overexpression in mGRPs through transduction with lentiviral activating particles was unsuccessful. In light of the setbacks associated with genetic modifications aimed at overexpressing NRG-1 in mGRPs, I decided to conduct additional research based on the stimulation of mGRPs with exogenous recombinant mouse neuregulin-1 peptide (rmNRG-1) through its supplementation in the standard culture medium. Supplementation of mGRPs with rmNRG-1 peptide had a diverse impact on the phenotype of the cells in vitro. Based on the conducted research, the following conclusions can be drawn: - overexpression of NRG-1 type I, achieved by transducing mGRPs with lentiviral vectors, resulted in a decrease in the growth rate of these cells and promoted their differentiation towards oligodendrocytes; however, I did not observe a significant impact of NRG-1 type I overexpression on the myelination properties of mGRPs - my preliminary investigations into the stimulation of mGRPs using exogenous recombinant neuregulin-1 peptide indicate an enhanced proliferation rate and reduced migratory capacity of supplemented mGRPs, accompanied by a diminished capability of these cells to differentiate into mature oligodendrocytes - the findings presented in my doctoral thesis highlight that the influence of neuregulin-1 on the functional properties of mGRPs is contingent upon the experimental setup.

Detailed Resource Type:

PhD Dissertations

Format:

pdf

Resource Identifier:

oai:rcin.org.pl:240042

Source:

IMDiK PAN, sygn. ZS 427

Language:

pol

Language of abstract:

eng

Rights:

Creative Commons Attribution BY 4.0 license

Terms of use:

Copyright-protected material. [CC BY 4.0] May be used within the scope specified in Creative Commons Attribution BY 4.0 license, full text available at: ; -

Digitizing institution:

Mossakowski Medical Research Institute PAS

Original in:

Library of the Mossakowski Medical Research Institute PAS

Projects co-financed by:

Operational Program Digital Poland, 2014-2020, Measure 2.3: Digital accessibility and usefulness of public sector information; funds from the European Regional Development Fund and national co-financing from the state budget.

Access:

Open