Proteomic analysis of regenerated rabbit lenses reveal crystallin expression characteristic of adult rabbits.

PURPOSE

To explore lens crystallin characteristics and morphology of rabbit regenerated lenses in comparison with wild type natural lenses by means of proteomic analysis and histological assay.

METHODS

The lens regeneration model of the New Zealand rabbit was established, and lens regeneration was observed by slit lamp examination and photography. A histological assay was evaluated under light microscopy and transmission electron microscopy (TEM). Protein samples of regenerated lenses were collected from experimental rabbit eyes 2, 4, and 16 weeks after surgery. Two-dimensional gel electrophoresis (2-DE) was performed. Image analyses was done using the ImageMaster 2D Elite 3.01 software package. The protein spots were trypsinized and identified by matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry.

RESULTS

Lens regeneration began in the periphery of the capsule bag about one to two weeks after the surgery and proceeded to regenerate toward the center. The regenerated lens appeared spherical in shape with a fairly translucent cortical structure and a nuclear opacity. Histological findings showed that the remnant lens epithelial cells differentiate at the lens capsule equator and new lens fibers form in a concentric pattern in a manner similar to that observed in natural lenses. However, TEM showed morphological changes in the epithelial cells of the regenerated lenses as compared with natural lenses. 2-D electrophoresis revealed that the patterns of protein spots from regenerated lenses (two weeks, four weeks, and 16 weeks) were analogous to those of 16-week-old natural lenses but were substantially different from those of two-week-old natural lenses, particularly when the two-week-old regenerated lenses were compared with the two-week-old natural lenses.

CONCLUSIONS

Proteomic analysis revealed that crystallin expression in regenerated rabbit lenses was analogous to that of natural lenses of adult rabbits but was different from that of very young rabbits (two weeks old), and TEM revealed the presence of morphological changes in the epithelial cells of regenerated lenses. These results suggest that the regrowth of lens materials in the lens capsule after endocapsular phacoemulsification might actually represent the regeneration of "mature" lens substances, which have led us to the conclusion that the regenerative process does not exactly mimic embryonic development.