Corneal crystallins and the development of cellular transparency

Abstract

Past studies have established that the cornea like the lens abundantly expresses a few water-soluble enzyme/proteins in a taxon specific fashion. Based on these similarities it has been proposed that the lens and the cornea form a structural unit, the 'refracton', that has co-evolved through gene sharing to maximize light transmission and refraction to the retina. Thus far, the analogy between corneal crystallins and lens crystallins has been limited to similarities in the abundant expression, with few reports concerning their structural function. This review covers recent studies that establish a clear relationship between expression of corneal crystallins and light scattering from corneal stromal cells, i.e. keratocytes, that support a structural role for corneal crystallins in the development of transparency similar to that of lens crystallins that would be consistent with the 'refracton' hypothesis.

Fig. 1

Normal rabbit cornea as shown by histology (A and B) and in vivo confocal microscopy (C and D). (A) Cross-section stained with hematoxylin & eosin showing anterior stratified corneal epithelium overlying corneal stroma with keratocyte nuclei (arrowheads) and posterior corneal endothelium. (B) Coronal section through the corneal stroma stained with gold-chloride showing keratocyte cell bodies and nuclei (arrow). (C) 3-D reconstruction of confocal images through a living rabbit cornea showing surface epithelial cells (Epi), underlying stroma and posterior corneal endothelium (Endo). (D) 2-D in vivo confocal image taken from the 3-D data set through the corneal stroma showing light scattering from the keratocyte nuclei. Bar = 100 μm.

Fig. 2

Coomassie-blue stained SDS-PAGE of water-soluble protein extracts keratocytes isolated fresh from rabbit, mouse and bovine. Note that TKT and ALDH is abundantly expressed in all three species, representing over 55% of the total water-soluble protein in bovine keratocytes. Bar = 100 μm.

Fig. 3

In vivo confocal images of light scattering from rabbit corneas 2 months after excimer laser surface ablation treated with vehicle (A and B) or 0.02% mitomycin C (C and D) to block myofibroblast differentiation. (A and C) X-Z slice through a 3-dimensional data set from the cornea. (B and D) X-Y optical plane from the 3-dimensional data set taken just below the corneal epithelium. Note the marked haze (A, arrow) below the epithelium and the cellular light scattering (B) in the vehicle treated eye. Bar = 100 μm.

Fig. 4

In vivo confocal microscopy of a patient following intrastromal implantation of a corneal lens. (A) X-Z projection through the 3-dimensional data set showing the anterior placement of the intracorneal lens (arrow) and the associated increased light scattering at the posterior margin of the lens. (B) X-Y plane taken from the 3-dimensional data set just posterior to the intracorneal lens showing marked light scattering from corneal keratocytes. (C) X-Y plane taken from the 3-dimensional data set further posterior within the normal stroma showing light scattering limited to keratocyte nuclei. Bar = 100 μm.

Fig. 5

Clinical photographs (A and C) and 3-D reconstructions (B and D) of postnatal rabbit eyes at 4 and 20 days after birth. Note prominent light scattering through out the cornea from the eyes at 4 days compared to the keratocyte nuclear scattering seen at 20 days. Bar = 100 μm.

Fig. 6

Reflectance confocal micrographic of corneal keratocytes treated with control serum-free media (A) and TGFβ1 (1 ng/ml) to induce myofibroblast differentiation (B). Note that myofibroblasts appear much larger and spread out then keratocytes and appear to scatter more light. Bar = 100 μm.