Abstract:
In fish it is well known that Müller cells proliferate, gain stem cell characteristics and generate new neurons upon injury. Proliferation is as essential step in retina regeneration and in mammals its tight regulation in Müller cells as well as age-dependent restriction of cell cycle re-entry has been proposed to limit retinal regeneration. Müller glia can be stimulated to proliferate by exogenous growth factors such as FGF2 in mammals, however the efficiency of neurogenesis remains highly limited. In this study MIO-M1 cells, which can be stimulated with FGF2 treatment to transdifferentiate into neurons, were used as the model system to better characterize FGF2 dependent MIO-M1 transdifferentiation process and to provide clues for the potential contribution of SIK2, as a negative regulator of FGF2 dependent Müller cell proliferation, to Müller cell reprogramming. Formation of nestin-positive neurospheres with self-renewal capacity confirmed transdifferentiation potential of MIO-M1 cells. Upregulation in Pax6 and Chx10 expression showed that MIO-M1 cells gain progenitor properties. Downregulation in vimentin expression with higher calretinin and Prox1 expression verified that MIO-M1 cells lose their glia characteristics and differentiate into neurons, particularly horizontal cells. Notch signaling, as in other Müller reprogramming events, might have dual roles; it promotes progenitor characteristics and amplification of progenitors in the early phase and involved in horizontal cell fate decision in the late phase. Our results indicate that ERK activity is required for increase in Müller population that will dedifferentiate into progenitors, but an unknown signaling pathway might regulate amplification of progenitor pool. Significant decrease in SIK2 protein level in early phase of the process is required for Müller glia to re-enter cell cycle. Overexpression of SIK2 blocked ERK activation, cell cycle entry and further transdifferentiation process. In the light of these findings, we suggest a novel role for SIK2 in Müller reprogramming through downregulation of ERK activity.
