Rabbits are widely used as an animal model for the prevention of secondary cataract after surgery not only because the secondary cataract in the rabbit eye reflects the condition in the human eye, but also because of the short time required for secondary cataract formation in this animal model.25
Additional rationales for selecting the rabbit as animal model in the present study include the rabbit's large eyes and low cost. As (1) an intracapsular sustained release system for H-7 was not available, (2) long-term multiple topical administrations of a high dose of H-7 are drug-consumptive and thus very expensive and might be toxic to the cornea, and (3) frequent intracameral injections of the drug were intolerable for the cornea, H-7 was delivered into the rabbit eye by twice-weekly intravitreal injections in this study. Intravitreal injection is commonly used in studies of posterior segment diseases. A previous study indicated that H-7 in the vitreous could effectively diffuse into the AC.26
As intravitreal injection did not directly mechanically invade the AC, it only induced mild inflammation in the anterior segment as evidenced in the BSS-treated eyes.
PCO is generally caused by proliferation and EMT of residual equatorial LECs that have migrated from the periphery of the capsular bag to the center of the posterior capsule and thus can directly affect postoperative vision. In the present study, no significant difference in PCO was observed between the H-7–treated eye and the BSS-treated eye. As inflammation stimulates wound healing that shares many features with PCO formation,27,28
it may also stimulate migration and proliferation of residual LECs to the central area of the posterior capsule, which could attenuate H-7's inhibitory effects. Additionally, capsular deposition of the observed aqueous humor protein (AC flare) and cells may also contribute to the early PCO formation in the rabbit eye. Therefore, the slightly denser PCO in the 1,200
μM H-7–treated eye than in the BSS-treated eye may be not due to any direct effect of H-7 on the capsule, but rather due to the drug-induced intraocular inflammation (see below). A preliminary study has shown that H-7 substantially inhibits PCO formation in cultured human lens capsules,29
supporting this speculation.
SR is the major part of secondary cataract following surgery in the rabbit eye. Although SR usually does not affect the postoperative vision, the SR-induced adhesions of the lens capsule might affect the function of future accommodating IOLs.4,5
Additionally, SR is a direct precursor to PCO30,31
and, eventually, will extend to the central area of the posterior capsule. Therefore, prevention of SR is as important as that of PCO, especially for future accommodating IOL implantation. To prevent the possible inhibitory effect of IOLs on LEC migration from complicating the experimental results, IOLs were not implanted following phacoemulsification in this study. As residual lens cortex and equatorial LECs are responsible for SR formation,30,31
SR's size and mass represent the speed of the regeneration/proliferation of these residual lens materials within the capsular bag. SRA/PSRA and WW are indications of the SR size and mass or density. The smaller values of SRA, PSRA, and WW in the 1,200
μM H-7–treated eye compared with the BSS-treated eye indicate that 1,200
μM intravitreal H-7 significantly inhibits SR formation in the rabbit.
In previous studies, 300
μM H-7 maximally inhibits the actomyosin-driven contractility and perturbs the dynamics of the actin cytoskeleton and associated proteins in various cultured cells.14–16
μM intravitreal H-7 has shown a similar effect on the pupil's response to pilocarpine as 300
μM intracameral H-7 in monkeys,26
the peak concentration of H-7 in the AC following each treatment of the 1,200
μM intravitreal H-7 could be ~300
μM in the rabbit eye. Additionally, the effective PCO-inhibition concentration of H-7 in the cultured human lens capsule is also 300
Taken together, this suggests that H-7's inhibitory effects on the actomyosin-driven contractility and the actin cytoskeleton and associated proteins may be involved in the mechanism of the drug-induced SR inhibition. It is thought that differentiation of residual equatorial LECs into lens fiber cells plays an important role in SR formation32
and that cytoskeletal remodeling, an essential component of both fiber cell elongation and migration, is likely regulated by MLCK and Rho kinase.33
Additionally, transforming growth factor β is widely implicated in secondary cataract formation after surgery4
and serine threonine kinases, including MLCK, Rho kinase, and protein kinases C and A, play important roles in transforming growth factor β signaling.19,34,35
As a broad-spectrum serine-threonine kinase inhibitor, H-7 may reduce SR formation by its multiple inhibitory effects on those protein kinases. Further studies are needed to clarify this issue.
In the present study, the observed AC flare and cells in the H-7–treated eye represent responses of the eye to all the interventions including the lens surgery, the intravitreal injection, and the H-7 treatment. Therefore, the response to H-7 alone may be relatively slight. As H-7 might separate cell–cell junctions in the ciliary epithelium,14–16
the increased AC flare in the H-7–treated eye compared with the BSS-treated eye could be primarily due to the drug-induced breakdown of the blood-aqueous barrier (BAB). The high dose of H-7 required for the intravitreal injection in the proof-of-concept study may be the cause of the BAB breakdown and can be avoided in future studies. Long-term sustained release of a low dose of H-7 or an analog directly into the capsular bag may not only enhance the drug's effect on secondary cataract, but also reduce the drug-induced inflammation and BAB breakdown. Additionally, as the human eye has a much stronger BAB than the rabbit eye,36,37
the H-7–induced BAB breakdown and the consequent increase in aqueous humor protein may not occur in the human to the same degree as in the rabbit. Indeed, no AC flare or cells have been seen following H-7 treatments, including intravitreal injection to a final intravitreal H-7 concentration of 1,200
μM, in the eyes of nonhuman primates.26
Our previous studies of H-7's effects on the cornea, ciliary body, and retina in monkeys have shown that 300
μM H-7 in the AC or the vitreous achieved by intracameral or intravitreal administration did not affect the structures of the cornea and ciliary body or the retinal vascular permeability or electrophysiology.38,39
These findings suggest that the effective secondary cataract-inhibition concentration of H-7 will not produce adverse effects on adjacent ocular tissues. Although clinically apparent corneal edema occurred after surgery in some eyes in the study, it appeared in both the treated and control eyes and disappeared in all eyes within 3–5 weeks, indicating that the corneal edema was due to the surgery rather than to the drug treatment. Further studies of H-7 (or analogs such as specific MLCK or rho kinase inhibitors) or agents affecting the actomyosin system by different mechanisms, coated on IOLs or other intracapsular devices, in the live nonhuman primate eye may clarify whether these cytoskeleton-modulating compounds are viable candidates as secondary cataract inhibitors.