Evaluation of Spatially Targeted Scleral Stiffening on Neuroprotection in a Rat Model of Glaucoma.

Purpose: Scleral stiffening may protect against glaucomatous retinal ganglion cell (RGC) loss or dysfunction associated with ocular hypertension. Here, we assess the potential neuroprotective effects of two treatments designed to stiffen either the entire posterior sclera or only the sclera adjacent to the peripapillary sclera in an experimental model of glaucoma. Methods: Rat sclerae were stiffened in vivo using either genipin (crosslinking the entire posterior sclera) or a regionally selective photosensitizer, methylene blue (stiffening only the juxtaperipapillary region surrounding the optic nerve). Ocular hypertension was induced using magnetic microbeads delivered to the anterior chamber. Morphological and functional outcomes, including optic nerve axon count and appearance, retinal thickness measured by optical coherence tomography, optomotor response, and electroretinography traces, were assessed. Results: Both local (juxtaperipapillary) and global (whole posterior) scleral stiffening treatments were successful at increasing scleral stiffness, but neither provided demonstrable neuroprotection in hypertensive eyes as assessed by RGC axon counts and appearance, optomotor response, or electroretinography. There was a weak indication that scleral crosslinking protected against retinal thinning as assessed by optical coherence tomography. Conclusions: Scleral stiffening was not demonstrated to be neuroprotective in ocular hypertensive rats. We hypothesize that the absence of benefit may in part be due to RGC loss associated with the scleral stiffening agents themselves (mild in the case of genipin, and moderate in the case of methylene blue), negating any potential benefit of scleral stiffening. Translational Relevance: The development of scleral stiffening as a neuroprotective treatment will require the identification of better tolerated stiffening protocols and further preclinical testing.

Sustained scleral stiffening in rats after a single genipin treatment.

Scleral stiffening has been proposed as a therapy for glaucoma and myopia. Previous in vivo studies have evaluated the efficacy of scleral stiffening after multiple treatments with a natural collagen crosslinker, genipin. However, multiple injections limit clinical translatability. Here, we examined whether scleral stiffening was maintained after four weeks following a single genipin treatment. Eyes from brown Norway rats were treated in vivo with a single 15 mM genipin retrobulbar injection, sham retrobulbar injection, or were left naive. Eyes were enucleated either 1 day or four weeks post-injection and underwent whole globe inflation testing. We assessed first principal Lagrange strain of the posterior sclera using digital image correlation as a proxy for scleral stiffness. Four weeks post-injection, genipin treatment resulted in a 58% reduction in scleral strain as compared to controls (p = 0.005). We conclude that a single in vivo injection of genipin effectively stiffened rat sclera for at least four weeks which motivates further functional studies and possible clinical translation of genipin-induced scleral stiffening.