Abstract
Purpose: :
The increased expression of two intermediate filaments (IF), vimentin and GFAP, has long been a hallmark of Müller cell (MC) reactivity in response to retinal insult, including retinal detachment (RD). Nestin is a relatively new addition to the family of 50+ proteins classified as IF. Its expression is developmentally regulated and is ultimately replaced by other IF proteins as CNS development proceeds. Here we sought to determine whether RD affects the expression of nestin in adult rat retinas and how its distribution correlates with GFAP and vimentin.
Methods: :
Experimental RD of 1, 3 or 7 days duration were created in the right eyes of Long Evans rats. Retinal sections were labeled for immunofluorescence with anti-GFAP, -vimentin, -nestin, and -BrdU (to detect dividing cells). Nestin also was detected by Western blots, and the amount of mRNA was determined by qPCR.
Results: :
Immunofluorescence shows that all MC strongly increase their expression of nestin, vimentin and GFAP at 3 and 7 days post-RD but the relative levels of the 3 proteins varies dramatically between MC. In addition, nestin is the predominant IF in those MC that extend into the subretinal space and is present in the leading edge of these rapidly growing processes. Nestin is also predominantly expressed in regions of BrdU labeled dividing MC. qPCR shows a greater than 2-fold increase in nestin mRNA at 1 day which is maintained at 3 and 7 days. Western blots show a single band at 220 kDa and the intensity of this band increases at 3 and 7 days.
Conclusions: :
RD induces an up-regulation of the IF nestin, a third IF cytoskeleton protein involved in the reactivity of MC after retinal injury. While the exact functional role for increased nestin expression is unknown, its differential expression with GFAP and vimentin, and the association with proliferating MC and those growing into the subretinal space suggest a significant role in glial scar formation.
Keywords: cytoskeleton • Muller cells • retinal detachment