Abstract
Purpose: :
We recently showed that p38 mitogen-activated protein kinase (MAPK) signaling becomes activated in an inducible rat model of ocular hypertension and contributes to the progression of retinal ganglion cell degeneration. As age is a significant risk factor for glaucoma, we sought to differentiate the effects of increasing age and ocular pressure on p38 MAPK activation using aged naïve mice and an inbred mouse model in which intraocular pressure increases with age.
Methods: :
We induced unilateral ocular pressure elevation in 3 month C57BL/6J (C57) mice via polystyrene microbead injection into the anterior chamber; the opposing eye received an equal volume saline injection. We also used the DBA/2J (DBA) mouse model of pigmentary glaucoma aged 3 to 10 months to represent various stages of progression. Naïve C57 mice aged 3 to 11 months were used as controls. Forty-eight hours prior to sacrifice and paraformaldehyde perfusion, mice received intravitreal injection of fluorescent cholera toxin beta subunit to label retinal ganglion cells. Dissected retinas were either lysed for protein extraction and subsequent western blotting or cross-sectioned for immunohistochemistry. Furthermore, retinal layer-specific staining intensities of activated p38 MAPK were quantified and compared across micrographs using a custom-developed algorithm.
Results: :
Microbead occlusion induced an average 28.3% elevation in C57 ocular pressure over a 4 week period. This elevation resulted in increased p38 MAPK immunolabeling throughout the retina, with significant increases in the outer plexiform layer (3.38 fold), inner nuclear layer (3.51 fold), inner plexiform layer (2.68 fold), and the ganglion cell and nerve fiber layers (which was highest at 9.98 fold). This expression pattern was similar in DBA mice as 10 month old mice, with an elevated intraocular pressure, had increased activated p38 MAPK over 3 month mice when compared by immuolabeling in retinas, again with the largest increases seen in the ganglion cell layer. Concordantly, whole retina western blots also showed nearly a 2-fold increase in activated p38 MAPK in 10 month over 3 month DBA. We did not find detectable differences in activated p38 MAPK expression across naïve C57 ages.
Conclusions: :
These results support the hypothesis that ocular pressure has a greater role than advanced age on p38 MAPK activation in the retina of relevant glaucoma models. Furthermore, these data implicate that specific cell-signaling responses may be regulated by particular stress signals. While aging may not directly activate p38 MAPK, it may be relevant in lowering the threshold for signal pathway activation and subsequent injury.
Keywords: signal transduction • intraocular pressure • retinal degenerations: cell biology