UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts
Abstract Background Pterygium 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...
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| Format: | Article |
| Language: | English |
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BMC
2018-12-01
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| Series: | BMC Ophthalmology |
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| Online Access: | http://link.springer.com/article/10.1186/s12886-018-0987-8 |
| _version_ | 1811326952858976256 |
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| author | Alexios J. Aletras Ioannis Trilivas Maria-Elpida Christopoulou Sotiria Drakouli Constantine D. Georgakopoulos Nikolaos Pharmakakis |
| author_facet | Alexios J. Aletras Ioannis Trilivas Maria-Elpida Christopoulou Sotiria Drakouli Constantine D. Georgakopoulos Nikolaos Pharmakakis |
| author_sort | Alexios J. Aletras |
| collection | DOAJ |
| description | Abstract Background Pterygium 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. Methods Normal 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. Results RT-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. Conclusion In 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. |
| first_indexed | 2024-04-13T14:57:41Z |
| format | Article |
| id | doaj.art-091c0e9306804b7891edf2babca5d524 |
| institution | Directory Open Access Journal |
| issn | 1471-2415 |
| language | English |
| last_indexed | 2024-04-13T14:57:41Z |
| publishDate | 2018-12-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Ophthalmology |
| spelling | doaj.art-091c0e9306804b7891edf2babca5d5242022-12-22T02:42:23ZengBMCBMC Ophthalmology1471-24152018-12-0118111010.1186/s12886-018-0987-8UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblastsAlexios J. Aletras0Ioannis Trilivas1Maria-Elpida Christopoulou2Sotiria Drakouli3Constantine D. Georgakopoulos4Nikolaos Pharmakakis5Laboratory of Biochemistry, Department of Chemistry, University of PatrasDepartment of Opthalmology, Medical School, University of PatrasLaboratory of Biochemistry, Department of Chemistry, University of PatrasLaboratory of Biochemistry, Department of Chemistry, University of PatrasDepartment of Opthalmology, Medical School, University of PatrasDepartment of Opthalmology, Medical School, University of PatrasAbstract Background Pterygium 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. Methods Normal 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. Results RT-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. Conclusion In 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.http://link.springer.com/article/10.1186/s12886-018-0987-8PterygiumFibroblastsProteasomeUVB irradiationNrf2-ARE pathwaySrc kinases |
| spellingShingle | Alexios J. Aletras Ioannis Trilivas Maria-Elpida Christopoulou Sotiria Drakouli Constantine D. Georgakopoulos Nikolaos Pharmakakis UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts BMC Ophthalmology Pterygium Fibroblasts Proteasome UVB irradiation Nrf2-ARE pathway Src kinases |
| title | UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts |
| title_full | UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts |
| title_fullStr | UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts |
| title_full_unstemmed | UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts |
| title_short | UVB-mediated down-regulation of proteasome in cultured human primary pterygium fibroblasts |
| title_sort | uvb mediated down regulation of proteasome in cultured human primary pterygium fibroblasts |
| topic | Pterygium Fibroblasts Proteasome UVB irradiation Nrf2-ARE pathway Src kinases |
| url | http://link.springer.com/article/10.1186/s12886-018-0987-8 |
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