Abstract
Purpose:
Retinal progenitors unlike neural stem cells do not self-renew under normal culture conditions in vitro. Our recent study demonstrates that retinal progenitors can self-renew, but this property is non-cell autonomous, requiring interactions with other cell types. Here, we have examined the role of non-histone chromosomal high-mobility group protein A2; HMGA2 in the self-renewal.
Methods:
HMGA2 expression was determined by Q-PCR, immunohistochemistry and Western analysis in embryonic day 18 (E18) progenitors. Its involvement in self-renewal of E18 progenitors was determined by neurosphere assay in the presence of conditioned medium obtained from cultured endothelial cells (ECCM) and Hmga2 siRNA.
Results:
To determine the physiological relevance of HMGA2 in retina, we analyzed its temporal expression by Q-PCR, which revealed a temporal decrease in Hmga2 expression during retinal development in vivo. Analysis of spatial localization of HMGA2 by immunohistochemistry on E18 retinal sections showed a center to peripheral gradient of HMGA2 distribution, co-localized with BrdU positive progenitors. Involvement of HMGA2 in self-renewal of retinal progenitors was examined in ECCM generated neurospheres. A significant increase in Hmga2 transcript levels and Hmga2+ BrdU+ cells was observed in ECCM generated neurospheres compared to EGF controls. To determine whether Hmga2 regulatory axis is involved in ECCM mediated self - renewal, HMGA2 expression was attenuated using siRNA lentivirus in E18 retinal dissociates. HMGA2 knockdown led to decrease in the number of neurospheres and BrdU positive cells suggesting the loss of self-renewal. Data on HMGA2 axis involved in self-renewal of retinal progenitors will be provided.
Conclusions:
Retinal progenitors display a non-cell autonomous self-renewal property, which is likely to be regulated by a molecular axis, underpinned by HMGA2.
Keywords: 688 retina •
721 stem cells