To date the expression of Id4 in the adult SVZ was fully characterized. We found that Id4 was expressed across the SVZ lineage. However, we found that stem cell were showing the highest levels of Id4 expression. Immuno-godd labelling followed by electron microscopy analysis confirms this expression. Moreover, BrdU incorporation paradigm to detect slow diving cells showed that a high percentage of these cells were Id4 positive suggesting that a big proportion of Id4 population correspond to quiescent stem cells. Additionally, we found Id4 expression in striatal astrocytes and ependymal cells, which are quiescent cells in the adult brain.
We then characterized the phenotype of the adult Id4-/- mouse. We observed a defect in SVZ morphology and a decreased in neurogenesis resulting in a decrease number of newborn neurons that reached the olfactory bulb, suggesting that Id4 may be playing an important role in the stem cell regulation. Complementary, electron microscopy analysis and wholemount stainings showed enlarged ventricles and defects in ependymal cell layer. These additional results open a new lead to investigate the potential role of Id4 in ependymal cell specification or maturation from radial glia cells.
In order to elucidate the Id4 function in the adult SVZ we acquired the conditional Id4 mouse line and cross it to a Tamoxifen inducible GlastCreERT2 mouse, which allowed us to delete specifically Id4 in stem cells. In vitro data showed an increase in proliferation of neurospheres cultured from adult SVZ, increasing the number of stem cells and the size of the spheres. Moreover, deletion of Id4 after differentiation showed an increase of division rate. Taken together this data suggest a role of Id4 in the maintenance of stem cell quiescence in the adult SVZ. To confirm these observations we analysed SVZ recombined cells in adult mice. Five days after Id4 deletion we observed an increase in the number of proliferating cells in the adult SVZ. We confirmed that this increase in the progenitors population occurred in detriment of stem cell pool observed as a result of Id4 deletion. However, we couldn’t detect an increase in the number of new born neurons but in oligodendrocytes in the whiter matter. Additionally, transcriptomic analysis showed deregulation of proliferative and cell adhesion pathways between others.
Finally, we studied the role of Id4 under pathological conditions. We developed tMCAO (transient Middle Cerebral Occlusion) model as stroke model. Immunofluorescence analysis of the SVZ in the tMCAO animal model at different time points idicated that SVZ proliferation upon stroke was preceded by Id4 downregulation and potentially it could be implicated in SVZ activation. However, further investigations need to be developed to fully understand the implication of Id4 in this process.
The results of this project have been disseminated in national meetings such as Journée de Biologie et Medicine in 2015 organized by University Pierre et Marie Curie (UPMC) and international congresses such as the ISSCR 2016 in San Francisco and 2017 in Basel. Additionally, the data was presented at the Doc and Postdoc Workshop organized by the ICM in 2015 and in the ICM Days in 2016. I also participated in the Marie Curie Alumni Association General Assembly in 2016 in Venice and the Chercheur en herbe program.
Moreover, in the team we have obtained the transcriptomic signature of Id4-overexpressing tumours. Strikingly, this signature overlaps with the signature of SVZ quiescent cells reported by other authors, suggesting that Id4 promote quiescence in gliomas. Taken together these data will be presented as a scientific article that will be submitted shortly in a peer-review journal.
