Human retinal pigment epithelial cell proliferation by the combined stimulation of hydroquinone and advanced glycation end-products via up-regulation of VEGF gene
| Autor: | Hiroki Tsujinaka, Maiko Takeda, Akiyo Yamauchi, Sumiyo Sakuramoto-Tsuchida, Asako Itaya-Hironaka, Nahoko Ogata, Takanori Fujimura, Hiroyo Ota, Shin Takasawa |
|---|---|
| Rok vydání: | 2015 |
| Předmět: |
RT-PCR
reverse transcription polymerase chain reaction SP1 specificity protein 1 medicine.medical_specialty Small interfering RNA WST-8 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2 4-disulfophenyl)-2H-tetrazolium monosodium salt medicine.medical_treatment Hydroquinone Cell Biophysics HQ hydroquinone IdU 5ʹ-Indo-2ʹ-deoxyuridine Biology TUNEL terminal deoxynucleotidyl transferase dUTP nick end labeling Biochemistry Paracrine signalling chemistry.chemical_compound SR scavenger receptor Advanced glycation endproduct(s) Internal medicine medicine AMD age-related macular degeneration Autocrine signalling Transcription factor Age-related macular degeneration Growth factor AGE advanced glycation endproduct ELISA enzyme-linked immunosorbent assay VEGF vascular endothelial growth factor Molecular biology eye diseases RPE retinal pigment epithelial Vascular endothelial growth factor Retinal pigment epithelial cells medicine.anatomical_structure Endocrinology chemistry siRNA small interfering RNA Apoptosis BSA bovine serum albumin RAGE receptor for advanced glycation endproduct FCS fetal calf serum sense organs Research Article |
| Zdroj: | Biochemistry and Biophysics Reports |
| ISSN: | 2405-5808 |
| DOI: | 10.1016/j.bbrep.2015.05.005 |
| Popis: | Although recent research showed that advanced glycation endproduct (AGE) and hydroquinone (HQ) are related to the pathogenesis of age-related macular degeneration (AMD), the mechanism how AGE and HQ induce or accelerate AMD remains elusive. In the present study, we examined the effects of AGE and HQ on changes of human retinal pigment epithelial (RPE) cell numbers and found that the viable cell numbers were markedly reduced by HQ by apoptosis and that AGE prevented the decreases of HQ-treated cell numbers by increased replicative DNA synthesis of RPE cells without changing apoptosis. Real-time RT-PCR revealed that vascular endothelial growth factor (VEGF)-A mRNA was increased by HQ treatment and the addition of HQ+AGE resulted in a further increment. The increase of VEGF secretion was confirmed by ELISA, and inhibition of VEGF signaling by chemical inhibitors and small interfering RNA decreased the HQ+AGE-induced increases in RPE cell numbers. The deletion analysis demonstrated that −102 to −43 region was essential for the VEGF-A promoter activation. Site-directed mutaions of specificity protein 1 (SP1) binding sequences in the VEGF-A promoter and RNA interference of SP1 revealed that SP1 is an essential transcription factor for VEGF-A expression. These results indicate that HQ induces RPE cell apoptosis, leading to dry AMD, and suggest that AGE stimulation in addition to HQ enhances VEGF-A transcription via the AGE-receptor for AGE pathway in HQ-damaged cells. As a result, the secreted VEGF acts as an autocrine/paracrine growth factor for RPE and/or adjacent vascular cells, causing wet AMD. Graphical abstract Highlights • HQ treatment induced RPE cell apoptosis. • AGE stimulated DNA replication of HQ-treated RPE cells via VEGF-A up-regulation. • The –102 to –43 region of VEGF-A is essential for promoter activation. • RNA interference showed SP1 was the key transcription factor for VEGF-A expression. • AGE-RAGE pathway leads to up-regulation of VEGF-A to induce wet-type AMD. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|