Abstract
Purpose :
Vision is often impaired by middle age due to presbyopia, the age-related loss of near focal ability. Presbyopia is thought to be caused by changes to lens material properties, geometry, and size during growth. Lens growth is entirely subject to lens epithelial cell (LEC) proliferation after birth. Lens stretching is known to increase LEC proliferation rates in porcine models. This study employs a novel technique of generating accommodation-like forces on murine lenses through embedding a whole murine eye in a hydrogel cylinder and compressing the cylinder. RNA from paired compressed and sham lenses was quantified with cell-based ultralow input RNA-seq revealing alterations to RNA expression after compression.
Methods :
Fresh c57bl/6 murine eye globes were collected. These eyes were embedded in 20% agarose, with the optical axis positioned vertically. Hydrogel-eye cylinders compressed under static loading (sham: no compression). Zonular tension increased in eyes subject to static compression along the optic axis due to slight equatorial bulging as the corneosclera deforms. Paired eyes were incubated for 2 or 24-hours. After incubation, lenses were enucleated, cells isolated with flow cytometry, and RNA content measured utilizing a limiting-cell RNA-seq technique.
Results :
Over 5300 genes were identified across multiple lens samples. Lens pairs varied in the genetic expression of multiple genes after stretching. Differences in RNA content were noted between groups incubated for 2 hours versus 24 hours. Statically compressed lenses demonstrated increases to crystallin protein associated genes like Crybb1 and Cryaa. Actin genes including Acta2 and Actb also responded to lens stretching. The Apoe gene was shown to decrease in RNA expression after compression.
Conclusions :
Thousands of unique murine lens genes were identified with many shown to respond to stretching over time. Murine lens stretching shows great promise for uniting lens biomechanics with lens genetic research. This method will allow for further study of murine lens mechanobiology and help to shed light on how lens cells respond to an accommodation-like loading regimen. Furthering the understanding of genetic pathways governing lens mechanobiology may someday lead to preventative therapies for presbyopia.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.