Keratocytes

The cornea is the first structure of the eye to be in contact with incident light. It is composed of five distinct layers lying parallel to its surface the outer epithelium, which is continuous with the epithelial layers of the conjunctiva the epithelial basal lamina the keratocyte-containing stroma, which is a collagen structure arranged so that it is transparent Descemet s membrane and, finally, the endothelium adjacent to the aqueous humour. 

Microscopically, the cornea shows a rather simple and multilayered structure that can be divided into six layers the epithelium, basement membrane, Bowman s layer, stroma, Descemet s membrane, and endothelium. The corneal tissue consists of three different cell types epithelial cells, keratocytes (corneal fibroblasts), and endothelial cells. The outermost corneal surface is covered with the preocular tear film, which is functionally associated with the cornea. The epithelial surface must be kept moist and smooth, a role played by the tear film in conjunction with a spreading function of the eyelids during blinking motions. Furthermore, the tear film provides a protection against infectious agents that may gain access into the eye. 

In another approach, Parnigotto and coworkers reconstructed corneal structures in vitro by using corneal stroma containing keratocytes to which corneal epithelial cells from bovine primary cultures were overlaid [73], However, this particular corneal model did not contain an endothelial layer. This model was histochemically characterized and the toxicity of different surfactants was tested using MTT methods. This stroma-epithelium model has been reported to show a cornea-like morphology, where a multilayered epithelial barrier composed of basal cells (of a cuboidal shape) and superficial cells (of a flattened shape) is noted. Furthermore, the formation of a basement membrane equivalent and expression of the 64-kDa keratin were reported, indicating the presence of differentiated epithelial cells. The toxicity data for various surfactants obtained with this model correlate well with those seen by the Draize test [73], However, this corneal equivalent was not further validated or used as a model for permeation studies. 

Orwin EJ, Hubei A. In vitro culture characteristics of comeal epithelial, endothelial and keratocyte cells in a native collagen matrix. Tissue Eng 6 307-319 (2000). 

Nevertheless, the stroma might also have some ability to regenerate the cornea. That is what the Thill and his assistant study [7] seems to demonstrate with the highlighting of a new population of repairing stem cells located inside the adult human corneal stroma. These cells might evolve toward the monocyte-macrophage specie or toward the ti broblast specie (keratocytes). 

There is a parallel to draw between the Thill and assistant study and the study by Kubota and Fagerlhom [15] who have demonstrated that the importance of the initial corneal edema, resulting from a bum, is correlated to the importance of the sequelar cicatricial leukoma that causes the drop of vision. The stromal lacunae, fonned by the edema, will be colonized by the keratocytes. After the resorption of the edema and at the level of these lacunae, the keratocytes form a zone of cicatricial tissue, which is the origin of the leukoma. These keratocytes also produce an unorganized network of collagen fibrillae, thus causing the drop of transparency of the cornea. 

This study should be considered in relation with the Jester study [16] on rabbits. The latter suggests that the postnatal development of the comeal transparency of rabbits might be associated with the decline of the density of keratocytes, with their evolution toward resting state and with their excretion of two water soluble proteins class lAl dehydrogenase aldehyde and transketolase. 

At last, we must mention the essential role of vitamin C in the making by keratocytes of an unorganized network of even, uniform and parallel to each other collagen fibrillae [15]. And yet when ocular bums caused by basic aggressors occur, the ascorbic acid rate in aqueous humor remains low during the month following the bum [15]. 

The comeal stroma, made of collagen flbrillae interwoven according to a specific architecture (and between which are located some keratocytes), 450 pm thick it is the framework of the... 

The outer surface of the cornea is covered with a smooth layer of stratified corneal epithelium (Figure 3.4). The lower layer of cells is columnar in shape and rests on a basement membrane that sits on top of a thick limiting structure termed Bowman s membrane derived from the corneal stroma below. The corneal stroma is composed of parallel bundles of collagen fibrils termed lamellae and rows or layers of branching corneal fibroblasts termed keratocytes. The posterior of the cornea is covered with a low cuboidal epithelium with a wide basement membrane (Descemet s membrane) and rests on the posterior portion of the corneal stroma. The corneal epithelium is normally under tension due to the pressure that is present in the anterior chamber just behind the cornea. The intraocular pressure is between 10 and 20 mm of mercury and is enough to cause the cornea to contract about 5% when it is excised from the eye. Therefore this pressure must be transferred between epithelium via cell-cell junctions. 

FIGURE 39.1. The three cell layers of the cornea. Shown on the left is a frozen section of an adult rabbit cornea stained with hematoxylin and eosin. On the right is a frozen section of a 6 week old mouse cornea. The mouse cornea is not fully mature until 8 weeks after birth. The left half visualizes the cornea by phase contrast microscopy. The right half shows the nuclei by DAPI staining. The density of the cells in the epithelial layer, and the paucity of keratocytes in the stromal layer are clearly seen with the nuclear stain. 

The collagen fibrils occupy considerable space and thereby increase the path of diffusion. The net effect of impeding diffusion is to increase by several times the equivalent fluid layer thickness of the actual stroma. Nevertheless, the stroma is transparent to molecular species below approximately 500,000 Da. The stroma serves as the major ocular depot for topically applied hydrophilic drugs, and the keratocytes presumably provide a reservoir for lipophilic compounds as well. 

The stroma constitutes approximately 90% of the total corneal thickness and is primarily composed of collagen fibers, keratocytes, and glycosaminoglycans. The imiform arrangement of the collagen fibers is the major determinant of corneal transparency, in contrast to the opaque and less regularly arranged fibers of the sclera. Disruption of the stromal layer regularity results in loss of corneal transparency and potential scar formation. 

Krebs, F. Miller, S. Catalone, B. Welsh, P. Malamud, D. Howett, M. Wigdahl, B. Sodium dodecyl sulfate and C31G as microbicidal alternatives to N-9 comparative sensitivity of primary human vaginal keratocytes. Antimicrobial agents Chemotherapy 2000, 44, 1954-1960. 

The histopathological findings in the cornea have been described in such cases they include epithelial edema with bullous changes, marked loss of keratocytes, thickening of Descemet s membrane, and an attenuated disrupted cell layer (21). 

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