Relative proliferative rates of limbal and corneal epithelia. Implications of corneal epithelial migration, circadian rhythm, and suprabasally located DNA-synthesizing keratinocytes

An important element of the recently proposed limbal stem cell model is that corneal epithelial cells migrate centripetally. The driving force for this migration is unknown, although it has been suggested that limbal epithelium, proliferates at a higher rate than central corneal epithelium, thus creating a population pressure toward the central cornea. This hypothesis was tested by measuring the relative proliferative rates of limbal and central corneal epithelia using 3H-thymidine autoradiographic techniques. The results indicate that, in both the New Zealand white rabbit and SENCAR mouse, the labeling index (LI) of limbal epithelium is actually lower than that of central corneal epithelium. This difference in LI persists throughout the circadian rhythm cycle. These results suggest that population pressure per se cannot be responsible for the centripetal migration of corneal epithelium and raise the possibility that preferential desquamation of central corneal epithelium may "draw" peripheral cells toward the central cornea. In both epithelia, the LI peak precedes the mitotic index (MI) peak during circadian cycle by 4-6 hr. These data therefore are in close agreement with earlier results on several nonocular stratified epithelia but contradict an earlier suggestion that the LI and MI peaks of corneal epithelium coincide. Finally, although most of the 3H-thymidine incorporating cells in central cornea may appear to be suprabasally located, they are only partially displaced into the suprabasal compartment. In most cases, such cells are still connected with the basement membrane through a thin stalk of cytoplasm. Since corneal epithelium rests on an exceptionally flat and rigid substratum, an increase in cellular volume in DNA-synthesizing cells may not be tolerated well in an already crowded basal layer. This may explain why an unusually large proportion of DNA-synthesizing cells are expelled preferentially into either a "second tier basal layer" or into the suprabasal compartment.     

Limbal stem cells of the corneal epithelium

Stem cells have certain unique characteristics, which include longevity, high capacity of self-renewal with a long cell cycle time and a short S-phase duration, increased potential for error-free proliferation, and poor differentiation. The ocular surface is made up of two distinct types of epithelial cells, constituting the conjunctival and the corneal epithelia. Although anatomically continuous with each other at the corneoscleral limbus, the two cell phenotypes represent quite distinct subpopulations. Stem cells for the cornea reside at the corneoscleral limbus. The limbal palisades of Vogt and the interpalisade rete ridges are believed to be repositories of stem cells. The microenvironment of the limbus is considered to be important in maintaining the stemness of stem cells. Limbal stem cells also act as a "barrier" to conjunctival epithelial cells and normally prevent them from migrating on to the corneal surface. Under certain conditions, however, the limbal stem cells may be partially or totally depleted, resulting in varying degrees of stem cell deficiency with resulting abnormalities in the corneal surface. Such deficiency of limbal stem cells leads to "conjunctivalization" of the cornea with vascularization, appearance of goblet cells, and an irregular and unstable epithelium. This results in ocular discomfort and reduced vision. Partial stem cell deficiency can be managed by removing the abnormal epithelium and allowing the denuded cornea, especially the visual axis, to resurface with cells derived from the remaining intact limbal epithelium. In total stem cell deficiency, autologous limbus from the opposite normal eye or homologous limbus from living related or cadaveric donors can be transplanted on to the affected eye. With the latter option, systemic immunosuppression is required. Amniotic membrane transplantation is a useful adjunct to the above procedures in some instances.     

Histomorphometric profile of the corneal response to short-term reverse-geometry orthokeratology lens wear in primate corneas: a pilot study

PURPOSE: To investigate the histological changes in primate cornea induced by short-term overnight orthokeratology (OK). METHODS: Nine young adult primates were used. One animal served as negative control. The remaining 8 animals wore reverse-geometry OK lenses for periods of 4, 8, 16, and 24 hours on 1 eye with the other eye as control. Central and midperipheral corneal thickness, as well as ultrastructural changes in corneal epithelium, stroma and endothelium in response to OK lenses, were evaluated. RESULTS: OK significantly reduced the thickness of the central cornea in all treatment groups. The central corneal thinning was both stromal and epithelial in origin. Substantial midperipheral corneal thickening was seen in 16-hour and 24-hour lens-wear groups and this effect was both stromal and epithelial in origin as well. Histology evidence indicated the primary epithelial response in the central cornea was compression of cells that resulted in wing cells becoming shorter and basal cells being squatted rather than lost or migration of cell layers. These pronounced cell shape changes occurred without compromising the structural integrity of the desmosomes. The thickened corneal epithelium has normal cell layers. The squamous cells have larger surface sizes and are composed of oval instead of flattened nuclei. This implied delayed surface cell exfoliation at the thickened midperipheral epithelium. Physical presence of OK lens over the cornea did not influence the microstructures of microvilli and microplicae, endothelium, and collagen distribution. CONCLUSIONS: The primate cornea, particularly the corneal epithelium, responds rapidly to the application of reverse-geometry OK lenses with significant epithelial cell shape alterations with short-term OK lens wear. This finding suggests that the corneal epithelium is moldable in response to the physical forces generated by the OK lenses.     

Effects of low and hyper Dk rigid gas permeable contact lenses on Bcl-2 expression and apoptosis in the rabbit corneal epithelium.

PURPOSE: To study Bcl-2 expression and apoptotic cell shedding of the rabbit corneal epithelium during extended wear of low and hyper Dk rigid gas permeable (RGP) contact lenses. METHODS: Rabbits were fit with either a low or a hyper Dk RGP lens (Dk/Ltotal= 10 and 97). The rabbits wore the lenses for either 24 hours, 3 days, or 1 week at which point they were humanely sacrificed. Immunocytochemistry and western blot analyses were performed to detect Bcl-2 in the corneal epithelium; TUNEL assay (TdT-mediated dUTP nick-end labeling) was used to identify apoptotic epithelial cells. RESULTS: 1) Immunocytochemistry: In the normal cornea, antibodies to Bcl-2 uniformly stained nuclei of all epithelial cell layers. Occasional surface epithelial cells, however, showed no anti-Bcl-2 nuclear staining; concomitant TUNEL assay revealed that all TUNEL-labeled-surface cells were Bcl-2 negative. By contrast, RGP contact lens wear, regardless of test lens oxygen transmissibility or lens wearing interval, significantly decreased both the total number of Bcl-2 negative and TUNEL-labeled cells on the epithelial surface (P < 0.05). In addition, contact lens wear was associated with labeling of keratocytes with TUNEL assay in the anterior stroma. 2) Western blot analysis: Total epithelial layer Bcl-2 expression was markedly decreased in the low Dk lens test group but was similar to control values in the hyper Dk lens test group. CONCLUSION: Bcl-2 protein seems to play an important role in the regulation of apoptotic cell shedding in the normal rabbit corneal epithelium. The identical staining pattern was seen in previous studies of the normal human cornea. RGP contact lens wear, however, appears to block the changes in Bcl-2 protein prior to apoptotic surface cell shedding, suggesting a lens-related anti-apoptotic effect. Taken together, these findings may explain why contact lens wear reduces surface cell exfoliation as previously reported in human studies.     

Adherence of Pseudomonas aeruginosa to the rabbit corneal epithelium

Adherence of bacteria to the corneal epithelium is the first step in the pathogenesis of corneal infection. Keratitis caused by Pseudomonas aeruginosa usually occurs among the contact lens wearers. Adherence of Pseudomonas aeruginosa to rabbit corneal epithelium, damaged by one week of hard contact lens wear, was examined histologically. The cornea was excised for scanning electron microscopy at 5, 15, 30 and 60 minutes after inoculation of Pseudomonas aeruginosa (0.2 ml, 10(8)CFU/ml). Pseudomonas aeruginosa did not adhere to the intact corneal epithelium, but traumatized cornea provided a site for adherence. In rabbits in which the eyelid was opened by lid retractors, large numbers of organisms were observed adhering to the injured cornea mediated by ocular surface mucin. Thirty minutes after inoculation, the adherent bacteria began to penetrate the epithelial cells and surface mucin by the formation of pockets surrounding the organism.     

Spherical indentations of human and rabbit corneal epithelium following extended contact lens wear.

PURPOSE: Mucin balls appear to cause spherical indentations in the corneal epithelium during silicone hydrogel extended contact lens wear. The purpose of this report is to describe and quantify these spherical indentations, as examined in the human cornea by in vivo confocal microscopy and by in vitro immunocytochemistry in the rabbit cornea. METHODS: Confocal images of full-thickness corneal epithelium were taken from three human patients participating in a 1-year extended contact lens-wear trial. Diameter and depth of the indentations were determined and measured. Two rabbit corneas showing identical indentations were stained with propidium iodide (nuclear stain) and Ki-67 (proliferation marker) and were examined using a laser scanning confocal microscope. RESULTS: The diameter of the spherical indentations is largest on the epithelial surface, ranging from 33.9 to 78.8 microm. Indentations form spherical sections whose depth variably extends into the corneal epithelium, reaching as far as the basal lamina. The rabbit model showed no epithelial nuclei within the indentation. Furthermore, stromal cells localized immediately beneath the indentations were positive for Ki-67 (proliferation). DISCUSSION: Spherical indentations of the corneal epithelium induced by mucin balls appear to be gaps or holes that can extend deep into the corneal epithelium. Indentations may potentially open a pathway for infectious microorganisms to penetrate the cornea. Surprisingly, stromal cells immediately beneath the holes were stimulated to proliferate, and there seemed to be an increase in localized cell density.     

The conjunctiva in corneal epithelial wound healing

BACKGROUND/AIMS—During the healing of corneal epithelial wounds with limbal involvement, conjunctival epithelium often migrates across the denuded limbus to cover the corneal surface. It is believed that, over a period of time, conjunctival epithelium covering the cornea assumes characteristics of corneal epithelium by a process referred to as conjunctival transdifferentiation. The purpose of this study was to examine, clinically, the fate of conjunctival epithelial cells covering the cornea and to assess the healing of corneal epithelial wounds when the conjunctival epithelium was removed or actively prevented from crossing the limbus and extending onto the cornea.
METHODS—10 patients with conjunctivalisation of the cornea were followed for an average of 7.5 months. Five patients in this group had their conjunctival epithelium removed from the corneal surface and allowed to heal from the remaining intact corneal epithelium. In another four patients with corneal epithelial defects, the conjunctival epithelium was actively prevented from crossing the limbus by mechanically scraping it off.
RESULTS—The area of cornea covered by conjunctival epithelium appeared thin, irregular, attracted new vessels and was prone to recurrent erosions. Conjunctivalisation of the visual axis affected vision. Removal of conjunctival epithelium from the cornea allowed cells of corneal epithelial phenotype to cover the denuded area with alleviation of symptoms and improvement of vision. It was also established that migration of conjunctival epithelium onto corneal surface could be anticipated by close monitoring of the healing of corneal epithelial wounds, and prevented by scraping off conjunctival epithelium before it reached the limbus.
CONCLUSION—This study shows that there is little clinical evidence to support the concept that conjunctival transdifferentiation per se, occurs in humans. "Replacement" of conjunctival epithelium by corneal epithelial cells may be an important mechanism by which conjunctival "transdifferentiation" may occur. In patients with partial stem cell deficiency this approach can be a useful and effective alternative to partial limbal transplantation, as is currently practised.

Keywords: corneal epithelium; conjunctiva; stem cells; transdifferentiation     

Expression of K12 keratin in alkali-burned rabbit corneas.

The healing of alkali-injured corneas is characterized by the persistence of polymorphonuclear leukocytes (PMN) in tissues and recurrent corneal epithelial defects. It has been suggested that the proteolytic enzymes secreted by PMN may account in part for the recurrent epithelial defects in the alkali-burned corneas. Cytoplasmic keratins, which form intracellular intermediate filaments, participate in the formation of hemidesmosomes and play a key role in the focal adhesion of epithelial cells to the basement membranes. The K3/K12 keratin pair is a major constituent of differentiated and stratified corneal epithelium. We have recently cloned the cDNA encoding the rabbit K12 keratin. In the present study we examined the expression of K12 keratin during the healing of alkali-burned rabbit corneas by slot-blot and in situ hybridization. Our results indicate that in normal cornea K12 keratin is equally expressed in all cell layers of stratified corneal epithelium and suprabasal layers of limbal epithelium, but not in bulbar conjunctival and other epithelia, i.e., lens, iris, and retinal pigment epithelium. The basal cells of the detached regenerating epithelium of the injured cornea express a very low level of K12 keratin. These observations are consistent with the notion that defective expression of K3/K12 keratins may play a role in the abnormal attachment of the regenerating epithelium to the basement membrane.     

角膜缘干细胞的研究

角膜缘干细胞是位于角膜缘基底上皮层底的一类特殊类型的上皮细胞,随着细胞培养技术的发展,角膜缘干细胞体外培养后移植用于治疗由于角膜缘干细胞缺乏或者功能不全引起的眼表疾病成为研究的热点。本文就其解剖学定位、生物学特性、组织工程化角膜的基础性研究及其临床应用做一综述。     

Reconstruction of the anterior chamber in posttraumatic epithelial ingrowths and epithelial anterior chamber cysts

The pathology of penetrating injuries of the cornea and corneal-scleral area with the ingrowth of epithelium and formation of anterior chamber cysts is characterized by polymorphism and can be defined by two main kinds: 1) plane ingrowth of epithelium along the posterior surface of the cornea involving the iris and the lens, and partially or totally obliterating the anterior chamber; 2) formation of cysts with a locked cavity and clear content. In 42 patients with epithelial ingrowths as a result of penetrating injuries of the cornea and corneal-scleral area, the following kinds of interventions were used: 1) transplantation of posterior layers of the cornea with dissection of epithelial growth; 2) dissection of anterior chamber cysts; 3) cystectomy; 4) dissection of the cite of epithelial ingrowth by means of corneal transplantation with removal of the cyst or reconstruction of the anterior chamber. Immediate and remote results of surgical treatment, complications in various forms of the mentioned pathology are described.     

钠尿肽受体A（NPR-A）在小鼠角膜和晶状体发育过程中的表达

目的　检测钠尿肽受体A（natriuretic peptide receptor A,NPR-A）在不同鼠龄小鼠角膜和晶状体内的表达,探讨其在小鼠眼发育过程中的作用。方法　选用C57BL/6转基因小鼠,收集从 E14到P90小鼠眼球标本120只,采用40 g·L^-1多聚甲醛灌注固定后,石蜡包埋切片,采用免疫组织化学方法对NPR-A在角膜和晶状体中的表达进行免疫荧光检测。结果　在E16,NPR-A高表达于角膜上皮细胞中,并持续至成年;其在角膜基质层和内皮细胞的表达也始于E16,而在P14之后,NPR-A表达随鼠龄的增加逐渐减弱。此外,NPR-A在P0小鼠晶状体上皮细胞膜内被检测到,并持续高表达至成年。在E16,由晶状体后壁上皮细胞生成的初级纤维开始高表达NPR-A,但随着鼠龄的增长和纤维结构的改变而逐渐减弱,直到P90消失。结论　NPR-A可能参与了角膜和晶状体生长和发育,并且对维持角膜上皮细胞的增生和修复以及晶状体的通透性具有重要作用。     

Functions of aquaporins in the eye

The aquaporins (AQPs) are integral membrane proteins whose main function is to transport water across cell membranes in response to osmotic gradients. At the ocular surface, AQP1 is expressed in corneal endothelium, AQP3 and AQP5 in corneal epithelium, and AQP3 in conjunctival epithelium. AQPs are also expressed in lens fiber cells (AQP0), lens epithelium (AQP1), ciliary epithelium (AQP1, AQP4) and retinal Müller cells (AQP4). Mutations in AQP0 produce congenital cataracts in humans. Analysis of knockout mice lacking individual AQPs suggests their involvement in maintenance of corneal and lens transparency, corneal epithelial repair, intraocular pressure (IOP) regulation, retinal signal transduction and retinal swelling following injury. The mouse phenotype findings implicate AQPs as potential drug targets for therapy of elevated IOP and ocular disorders involving the cornea, lens and retina. However, much research remains in defining cell-level mechanisms for the ocular AQP functions, in establishing the relevance to human eye disease of conclusions from knockout mice, and in developing AQP-modulating drugs.     

Modeling glucose distribution in the cornea

The central cornea obtains its glucose by diffusion through the cornea from the aqueous humor to the epithelium. The diffusion of glucose in the cornea is analogous to the flow of current in an electrical resistance network. The cellular consumption of glucose can be compared to shunting a portion of the charge to electrical ground. An electrical analog model of the cornea was developed to predict the availability of glucose to the epithelium and the distribution of glucose in the stroma. The glucose constant concentration lines in the normal stroma are parallel to the corneal surface and have decreasing values from 880 to 580 micrograms/ml. The effects on epithelial glucose concentration by implanting an intracorneal lens (ICL) of varying diameter, depth, permeability and thickness can be modeled. Glucose permeability through the intracorneal lens has the most significant effect on glucose availability. The ICL profile i.e. power, can also be an important fact in determining glucose availability. A minus power design requires a thin central lens zone with a thick peripheral zone. The design results in relatively more glucose flux through the optical zone of the lens and thus improves central epithelial glucose availability.     

Vertical movement of epithelial basal cells toward the corneal surface during use of extended-wear contact lenses

PURPOSE: To study the effects of extended contact lens wear (EW) on the movement of basal epithelial cells toward the corneal surface. METHODS: Rabbits (n = 32) were injected with 5-bromo-2-deoxyuridine (BrdU) to label a group of proliferating basal epithelial cells, and, 24 hours later, one randomly chosen eye was fitted with a low- or medium-oxygen-transmissible (Dk/t) rigid gas permeable (RGP) contact lens, while the other eye served as the control (n = 28). Four rabbits were not fitted with any contact lens. Rabbits were euthanatized at different time points and the corneal epithelium was immunocytochemically stained for BrdU and/or Ki-67 and counterstained with propidium iodide or Syto 59. Corneal flatmount tissues were examined three dimensionally under a laser confocal microscope and the location of each BrdU-labeled cell in the corneal epithelium (basal or suprabasal) was determined. RESULTS: Four days after injection of BrdU, both low- (P < 0.001) and medium-Dk/t RGP (P < 0.001) lens groups showed significantly more BrdU-labeled cells in the basal cell layer than in the control eyes. Six days after injection of BrdU, a small percentage of BrdU-labeled cells (<0.5%) were Ki-67 positive. CONCLUSIONS: Within 6 days, the majority (80%) of BrdU-labeled basal cells became terminally differentiated and rarely divided secondarily in the central epithelium. Short-term use of low- and medium-Dk/t RGP EW contact lenses slows the normal movement of basal epithelial cells toward the surface in the central cornea. This is consistent with known EW-lens-induced decreases in corneal epithelial basal cell proliferation and surface cell exfoliation. Overall, the data suggest that EW lenses significantly inhibit the normal homeostatic turnover rate of the corneal epithelium.     

长期配戴软性角膜接触镜者角膜上皮的病理改变

目的研究软性角膜接触镜对角膜上皮组织的基底细胞数目、上皮厚度、基底细胞形态、增殖细胞核抗原（PCNA）等的影响。方法前瞻性病例对照研究。选择配戴软性角膜接触镜超过3年的患者33例（66眼,角膜接触镜组）和从未配戴软性角膜接触镜患者33例（66眼,对照组）,对所选患者,在去瓣准分子激光上皮下角膜磨镶术（Flap—freeEpi．LASIK）手术中,用Epi-K角膜上皮刀取下角膜上皮瓣。2组中各取30例（60眼）采用超声进行离体角膜上皮瓣厚度的测量和采用显微镜进行基底细胞计数,各取3例（6眼）进行石蜡切片及免疫组化染色,观察角膜上皮瓣基底细胞形态的变化和进行PCNA的测定。数据采用独立样本t检验进行分析。结果角膜接触镜组角膜上皮层厚度为（55．33±4．56）μm;对照组角膜上皮层厚度为（57．19±3．82）μm,二者之间差异有统计学意义（t=-2．422,P〈0．05）。角膜接触镜组基底上皮细胞数目减少,为（3872．6±153．2）eells／mm^2;对照组基底上皮细胞数目为（3989．2±289．6）cells／mm^2,二者之间差异有统计学意义（t=-2．757,P〈0．01）。角膜接触镜组较对照组角膜基底细胞排列疏松,细胞形态欠规整,基底膜较厚,相对粗糙。角膜接触镜组角膜上皮基底细胞可发现PCNA阳性细胞,而对照组无阳性细胞。结论长期配戴软性角膜接触镜可导致角膜上皮损伤：使角膜上皮层变薄;单位面积内的的角膜上皮基底细胞数量减少;角膜上皮的组织结构发生病理学改变,细胞形态发生改变。细胞之间的连接和基底膜遭到破坏,PCNA阳性细胞存在。     

Effect of eyelid closure and overnight contact lens wear on viability of surface epithelial cells in rabbit cornea.

PURPOSE: To determine the effects of open, closed eye, and overnight contact lens wear on homeostatic epithelial surface cell death in the rabbit cornea. METHODS: One eye of each rabbit was either closed by eyelid suture or fitted with one of the following test contact lenses: (1) low Dk/t rigid gas permeable (RGP) lens, (2) hyper Dk/t RGP lens, (3) hyper Dk/t soft lens. The other eye served as a control. After 24 hours, whole corneas were carefully excised and immediately stained with a calcein-acetoxymethyl ester-ethidium homodimer viability assay to quantify the number of nonviable surface epithelial cells. In addition, exfoliated corneal epithelial cells were collected with an eye irrigation chamber to determine cell viability. RESULTS: In the normal cornea, open-eye conditions showed significantly more nonviable surface cells in the central cornea than in the periphery (p < 0.05). Overnight wear of all test lenses and eyelid closure induced significant decreases in the number of nonviable cells on the central corneal surface compared with controls (p < 0.05). All exfoliated corneal epithelial cells collected by eye irrigation were nonviable. CONCLUSION: In the rabbit model, overnight contact lens wear significantly downregulated spontaneous epithelial surface cell death independent of lens rigidity or material oxygen transmissibility. These effects were similar to eyelid closure without lens wear. Taken together, these results suggest that eyelid closure and the physical presence of the contact lens may protect against the shear stress forces exerted by eyelid blinking, which are believed to cause central surface cell death and subsequent exfoliation.     

The contribution of blood flow by the anterior ciliary arteries to the anterior segment in the primate eye

The rate and mode of corneal wound healing in severely diabetic rats were studied by light microscopy and scanning electron microscopy. Diabetes mellitus was induced in 52 rats by alloxan injection, and 52 nondiabetic rats were used as controls. After 3 weeks, a nonpenetrating razor-blade wound was made in the central cornea of both eyes in 48 diabetic and 48 normal rats. The incision passed through the epithelium and into the stroma. The effects of diabetes on the unwounded cornea were observed by comparison with corneas from eight unwounded rats (four diabetic and four normal). Whole corneas from wounded diabetic and normal rats were studied at 0, 1, 3, 6, 12 and 24 hr and at 2–7 days after wounding. The rate and mode of healing were not found to differ between diabetics and normals. The surfaces of corneal wounds in both groups appeared to be completely healed and indistinguishable from the surrounding unwounded epithelium after 24 hr. The epithelial cells involved in the initial healing process were derived primarily from the layer of wing cells which progressed across the wound close to the connective-tissue base. Only in the final stages of healing, after the wound had been filled by the deeper epithelial cells, did superficial epithelial cells migrate. There appeared to be more exfoliating superficial epithelial cells over the entire cornea in diabetic rats than in normals. Because the healing of central corneal incisions occurs initially and primarily by sliding of the deeper epithelial cells, and because the diabetic condition appears to be associated with increased exfoliation of surface cells, the healing of central incisions may be less affected by diabetes than the healing of defects of the whole corneal surface, where the superficial epithelial cells have been reported to be the main migratory cells in the initial healing process and where healing in diabetics is delayed.     

Actin filament localization in developing and pathologic human corneas

Actin is associated with motility, cell morphology, and cell-substrate adhesion. The molecular probe NBD phallacidin, which reacts with filamentous actin, was used to study the distribution of actin filaments in the corneal and conjunctival epithelium, stroma, and endothelium. Frozen sections of human fetal eyes from 8 weeks to 40 weeks of gestation were reacted with NBD phallacidin. Pathologic tissues included keratoplasty specimens from patients with hereditary posterior polymorphous corneal dystrophy (PPMD) and surgically excised tissues removed for treatment of epithelial down-growth. Normal human cornea was used as a control. Immunofluorescent staining disclosed actin filament distribution in corneal epithelium as early as 9-10 weeks of gestation. Staining increased with maturation until term. Adult human corneal epithelium showed more pronounced staining of the surface layers. Stromal staining was more extensive in earlier stages of gestation and decreased in later stages of gestation, after 20-21 weeks. In pathologic corneas with posterior polymorphous dystrophy, there was localization of actin, as well as keratin, in the abnormal epithelial-like layers lining the posterior cornea. In epithelial downgrowth, actin and keratin were demonstrated in multilayered squamous epithelium on the anterior iris surface. Actin appears to be involved in migration of corneal epithelial and endothelial cells.     

Effects of increasing Dk with rigid contact lens extended wear on rabbit corneal epithelium using confocal microscopy.

The effects of 24-h wear of various Dk-rigid gas-permeable (RGP) contact lenses on the rabbit corneal epithelium were studied by in vivo tandem scanning confocal microscopy (TSCM), and confirmed by scanning electron microscopy (SEM). Lenses used were polymethylmethacrylate (PMMA) (Dk/L = 0), RGP experimental A lens (siloxanylmethacrylate-fluoromethacrylate-methylmethacrylate , 33), experimental B (siloxanylmethacrylate-fluoromethacrylate, 56), and experimental C (siloxanylstyrene-fluoromethacrylate copolymer, 64 x 10(-9)) (cm/s) (ml O2/ml mm Hg) with 0.15-mm thickness (Dk/L measured by polarograph including boundary layer effect). After 24-h PMMA lens wear, TSCM showed no superficial epithelial cells but only exposed, underlying wing cells. The cornea with experimental A showed partial superficial epithelial desquamation. With experimental B wear, slight superficial epithelial cell swelling and desquamation were observed on the surface of the cornea. No changes were observed for the eye with experimental C and control. The observed severity of desquamation of superficial epithelial cells was dependent on the oxygen transmissibility (Dk/L) of RGP lenses worn. All in vivo findings were confirmed by SEM observations. Based on the results of this study, we conclude that (a) although Dk/L = 56 lens B shows no residual overnight corneal swelling, surface damage is still produced; (b) Dk/L = 64 lens C is best for epithelium showing the same corneal images as control; and (c) TSCM is a good way to evaluate the contact lens safety and efficacy in vivo at the cellular level noninvasively.