Telomere Length Measurement in Different Ocular Structures: A Potential Implication in Corneal Endothelium Pathogenesis.

PURPOSE: Human chromosomes are protected at their end by a long portion of hexameric tandem repeats, the telomere. In somatic cells, telomere attrition caused by endogenous and exogenous oxidative stress as well as DNA replication can threaten genomic integrity and lead to the deterioration of tissue functions and an age-related physiological decline. The human eye is a complex organ in which cells of different ocular tissues are exposed to photo-oxidation, high mitochondrial metabolic activity, and/or replicative pressure. METHODS: We employed a highly sensitive quantitative PCR technique to determine relative telomere length in different human ocular structures. RESULTS: The longest telomeres in all ocular structures analyzed are found in neural retina, and the shortest are in the cornea. Within the retina, retinal pigment epithelium has four times shorter telomeres when compared to neural retina. However, no age-dependent telomere attrition in the retina and no difference between telomere lengths in the macular region and the rest of the retina have been found. In the cornea, stroma has the longer telomeres. In the corneal endothelium, we found a clear age-dependent telomere shortening. Since the endothelium is one of the most metabolically active ocular structure, this result suggests that endogenous oxidative stress from high mitochondrial activity is a major determinant of telomere loss in this structure. CONCLUSIONS: Taken together, our results imply that the aging process and telomere attrition in the different ocular structures are the result of multiple factors and could not be attributed to solely exogenous or endogenous oxidation or DNA replication.

Faster DNA Repair of Ultraviolet-Induced Cyclobutane Pyrimidine Dimers and Lower Sensitivity to Apoptosis in Human Corneal Epithelial Cells than in Epidermal Keratinocytes.

Absorption of UV rays by DNA generates the formation of mutagenic cyclobutane pyrimidine dimers (CPD) and pyrimidine (6-4) pyrimidone photoproducts (6-4PP). These damages are the major cause of skin cancer because in turn, they can lead to signature UV mutations. The eye is exposed to UV light, but the cornea is orders of magnitude less prone to UV-induced cancer. In an attempt to shed light on this paradox, we compared cells of the corneal epithelium and the epidermis for UVB-induced DNA damage frequency, repair and cell death sensitivity. We found similar CPD levels but a 4-time faster UVB-induced CPD, but not 6-4PP, repair and lower UV-induced apoptosis sensitivity in corneal epithelial cells than epidermal. We then investigated levels of DDB2, a UV-induced DNA damage recognition protein mostly impacting CPD repair, XPC, essential for the repair of both CPD and 6-4PP and p53 a protein upstream of the genotoxic stress response. We found more DDB2, XPC and p53 in corneal epithelial cells than in epidermal cells. According to our results analyzing the protein stability of DDB2 and XPC, the higher level of DDB2 and XPC in corneal epithelial cells is most likely due to an increased stability of the protein. Taken together, our results show that corneal epithelial cells have a better efficiency to repair UV-induced mutagenic CPD. On the other hand, they are less prone to UV-induced apoptosis, which could be related to the fact that since the repair is more efficient in the HCEC, the need to eliminate highly damaged cells by apoptosis is reduced.

Implication of ultraviolet light in the etiology of uveal melanoma: A review.

Uveal melanoma is the most frequent intraocular cancer and the second most common form of melanoma. It metastasizes in half of the patients and the prognostic is poor. Although ultraviolet (UV) radiation is a proven risk factor for skin melanoma, the role of UV light in the etiology of uveal melanoma is still contradictory. We have compared epidemiological and genetic evidences of the potential role of UV radiation in uveal melanoma with data on cutaneous melanoma. Even though frequently mutated genes in skin melanoma (e.g. BRAF) differ from those found in uveal melanoma (i.e. GNAQ, GNA11), their mutation pattern bears strong similarities. Furthermore, we provide new results showing that RAC1, a gene recently found harboring UV-hallmark mutation in skin melanoma, is also mutated in uveal melanoma. This article aims to review the work done in the last decades to understand the etiology of uveal melanoma and discuss new avenues, which shed some light on the potential role of UV exposure in uveal melanoma.

Wavelength-dependent ultraviolet induction of cyclobutane pyrimidine dimers in the human cornea.

Exposition to ultraviolet (UV) light is involved in the initiation and the progression of skin cancer. The genotoxicity of UV light is mainly attributed to the induction of cyclobutane pyrimidine dimers (CPDs), the most abundant DNA damage generated by all UV types (UVA, B and C). The human cornea is also exposed to the harmful UV radiations, but no UV-related neoplasm has been reported in this ocular structure. The probability that a specific DNA damage leads to a mutation and eventually to cellular transformation is influenced by its formation frequency. To shed light on the genotoxic effect of sunlight in the human eye, we have analyzed CPD induction in the cornea and the iris following irradiation of ex vivo human eyes with UVA, B or C. The extent of CPD induction was used to establish the penetrance of the different UV types in the human cornea. We show that UVB- and UVC-induced CPDs are concentrated in the corneal epithelium and do not penetrate deeply beyond this corneal layer. On the other hand, UVA wavelengths penetrate deeper and induce CPDs in the entire cornea and in the first layers of the iris. Taken together, our results are undoubtedly an important step towards better understanding the consequences of UV exposure to the human eye.

The 3895-bp mitochondrial DNA deletion in the human eye: a potential involvement in corneal ageing and macular degeneration.

In human skin, the 3895-bp deletion of mitochondrial DNA (mtDNA(3895)) is catalysed by ultraviolet (UV) light through the generation of reactive oxygen species. Given its function in vision, the human eye is exposed to oxidising UV and blue light in its anterior (cornea, iris) and posterior (retina) structures. In this study, we employed a highly sensitive quantitative PCR technique to determine mtDNA(3895) occurrence in human eye. Our analysis shows that the mtDNA(3895) is concentrated in both the cornea and the retina. Within the cornea, the highest mtDNA(3895) level is found in the stroma, the cellular layer conferring transparency and rigidity to the human cornea. Moreover, mtDNA(3895) accumulates with age in the stroma, suggesting a role of this deletion in corneal ageing. Within the retina, mtDNA(3895) is concentrated in the macular region of both the neural retina and the retinal pigment epithelium, supporting the hypothesis that this deletion is implicated in retinal pathologies such as age-related macular degenerescence. Taken together, our results imply that UV and blue light catalyse mtDNA(3895) induction in the human eye.

Mitochondrial DNA common deletion in the human eye: a relation with corneal aging.

The most frequent mitochondrial DNA (mtDNA) mutation is a 4977 bp deletion known as the common deletion (mtDNA(CD4977)). mtDNA(CD4977) is related to skin photo-aging and to chronological aging of cells with high-energy demands such as neurons and muscle cells. The human eye contains both sun-exposed (cornea, iris) and high-energy demand structures (retina). In this study, we employed a highly sensitive quantitative PCR technique to determine mtDNA(CD4977) occurrence in different structures of the human eye. We found that the cornea, the most anterior structure of the eye, contains the highest amount of mtDNA(CD4977) (2.6%, 0.25% and 0.06% for the cornea, iris and retina, respectively). Within the cornea, mtDNA(CD4977) is almost exclusively found in the stroma, the cellular layer conferring transparency and rigidity to the human cornea (8.59%, 0.13% and 0.05% in the stroma, endothelium and epithelium, respectively). Moreover, we show that mtDNA(CD4977) accumulates with age in the corneal stroma. Taken together, our results suggest that mtDNA(CD4977) is related to photo-aging rather than chronological aging in the human eye. Similar to the involvement of mtDNA(CD4977) in skin photo-aging phenotypes, we believe that the clinical manifestations of corneal aging, including clouding and stiffening, are associated with the accumulation of mtDNA(CD4977) in the corneal stroma.

Ultraviolet light-induced cyclobutane pyrimidine dimers in rabbit eyes.

Sunlight exposure of the eye leads to pathologies including photokeratitis, cortical cataracts, pterygium, actinic conjunctivitis and age-related macular degeneration. It is well established that exposure to ultraviolet (UV) radiations leads to DNA damage, mainly cyclobutane pyrimidine dimers (CPDs). CPD formation is the principal factor involved in skin cancer. However, the exact mechanism by which sunlight induces ocular pathologies is not well understood. To shed light on this issue, we quantified the CPD formation onto DNA of rabbit ocular cells following UVB exposure. We found that CPDs were induced only in the structures of the ocular anterior chamber (cornea, iris and lens) and were more concentrated in the corneal epithelium. Residual UVB that pass through the cornea are completely absorbed by the anterior layers of the iris. CPDs were also detected in the central portion of the lens that is not protected by the iris (pupil). By determining the UV-induced DNA damage formation in eyes, we showed that anterior ocular structures are a reliable physical barrier that protects the subjacent structures from the toxic effects of UV. Although the corneal epithelium is the structure where most of the CPDs were detected, no cancer is related to solar exposure.