Abstract
BackgroundPterygium is a condition characterized by epithelial overgrowth of the cornea, inflammatory cell infiltration and an abnormal extracellular matrix accumulation. Chronic UV exposure is considered as a pathogenic factor of this disease. Proteasome is an intracellular multi-subunit protease complex that degrades intracellular proteins. Among proteasome subunits the β5 (PSMB5), bearing chymotrypsin-like activity. It is considered as the main proteasome subunit and its expression is mediated by Nrf2-ARE pathway in many cell types. This study investigates the expression of PSMB5 in pterygium and the effect of UVB irradiation on its expression and activity in pterygium fibroblasts.MethodsNormal conjunctival and pterygium specimens were obtained from the bulbar conjunctiva of patients undergoing cataract surgery and from patients with pterygium undergoing surgical removal of primary tissue, respectively. Fibroblasts were isolated upon treatment of specimens with clostridium collagenase. The expression of PSMB5 and Nrf2 in tissues and cells was ascertained by RT-PCR analysis and western blotting. Cell survival was measured by the MTT method and the proteasome chymotrypsin-like activity was determined by fluorometry.ResultsRT-PCR analysis showed that the expression of PSMB5 was significantly lower in pterygium than in normal conjunctiva. The expression of PSMB5 was mediated by the Nrf2/ARE pathway as indicated by using the Nrf2 activator Oltipraz. The expression of PSMB5 and Nrf2 by pterygium fibroblasts was suppressed in a dose dependent manner following UVB radiation of 0–50 mJ/cm2 doses. The expression of PSMB5, but not of Nrf2, remained at almost the control levels, when UVB exposure was performed after pre-incubation of cells with the src kinases inhibitor PP2. UVB irradiation had very low deleterious effect on fibroblasts survival, while it did not affect the proteasome chymotrypsin-like activity.ConclusionIn pterygium fibroblasts, UVB exposure leads to down-regulation of Nrf2/ARE-mediated PSMB5 gene expression, in which src kinases may be implicated. This effect may be partially responsible for the lower expression of PSMB5 detected in pterygium as compared to normal conjunctiva.
Highlights
Pterygium is a condition characterized by epithelial overgrowth of the cornea, inflammatory cell infiltration and an abnormal extracellular matrix accumulation
The expression of proteasome subunits the β5 (PSMB5) in pterygium tissues was statistically significantly lower than normal conjunctiva. This result was not attributed to differences in the age of patients with pterygium and normal conjunctiva since there were no significant differences between the age distribution and mean age of two groups of donors
Effects of Ultraviolet B (UVB) irradiation on PSMB5 expression and activity in fibroblasts Since the UVB exposure is considered as a main pathogenetic factor in pterygium disease, we examined whether the UVB irradiation has an effect on the expression of proteasome in pterygium fibroblasts, since fibroblasts produce many components of extracellular matrix and are implicated in pterygium formation
Summary
Pterygium is a condition characterized by epithelial overgrowth of the cornea, inflammatory cell infiltration and an abnormal extracellular matrix accumulation. Pterygium is an eye disorder featuring hyperplasia of the cornea epithelium, usually bilateral, overgrowth of stromal fibroblasts and blood vessels, significant neovascularization, inflammatory cell infiltration and abnormal extracellular matrix accumulation, with elastin and collagen being the main components. High expression of hypoxia inducible factor-1α (HIF-1α) and heat shock proteins (Hsp), such as Hsp, Hsp and Hsp in pterygium tissues has been previously reported [14, 15]. They may contribute in the cellular defense mechanisms against stressful conditions
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