Abstract
Purpose :
We examined the inner retina and ILM structures of the Rs1-/Y rat exon-1-del model of X-linked retinoschisis (XLRS). We seek to understand whether surface structural alterations may help explain the ready access of AAV viral vector to deeper retinal layers after intravitreal administration.
Methods :
Protocols were overseen by the UCD IACUC. Rats were sacrificed, eyes collected (N=3), fixed by freeze substitution method, paraffin-embedded, and sectioned (5um thick). Histologic changes of retinal structure were assessed by paraffin sections and flatmounts. Samples were labeled by H&E and immunohistochemistry using antibodies for recoverin, glutamine synthetase, chx10, prox1, GFAP and vimentin, with secondary fluorescent antibodies. Images were collected by confocal microscopy.
Results :
INL cellularity appears increased in Rs1-KO at P7 compared to WT. In WT retina Chx10 labels all cells in the INL (principally bipolar cells) up to the OPL border, while the INL of Rs1-KO shows an extra band of unlabeled cells 1-2 thick bordering the OPL. ILM continuity is disrupted as early as P30, presumably compromising its barrier role. Glutamine synthetase labeling Müller cell feet shows the most significant continuity breaks of the ILM. Vimentin staining of ILM intermediate filaments gives sparse, irregular, and clumped retina surface morphology on flat mount compared to WT. As expected, in Rs1-KO after schisis collapse, GFAP activation reaches to deeper inner retinal layers than in WT.
Conclusions :
This Rs1−/Y rat model displays a developmental difference at the early age prior to P12 schisis cavity formation, as there are more INL cells in the Rs1-KO compared to WT. The presence of these cells suggests possible delay in cell differentiation or degradation. Disruption of the ILM structures by P30 may contribute to AAV entry from the vitreous.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.