Dissertation
Dissertation > Medicine, health > Ophthalmology > Retina and optic nerve diseases > Retinal disease

The Effect of Blue Light on α1D Subunit Protein Expression and Secretion Function of Human Retinal Pigment Epithelium Cells in Vitro

Author LiHaiHui
Tutor CaiShanJun
School Zunyi Medical College,
Course Ophthalmology
Keywords Photodamage Pigment epithelium Calcium α1D subunit VEGF
CLC R774.1
Type Master's thesis
Year 2011
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Objective To investigate the cause of human retinal pigment epithelial blue (retinal pigment epithelium, RPE) cells intracellular calcium concentration; L-type calcium channel subunit α1D changes in gene expression; RPE secretion of VEGF, bFGF and PEDF change its relationship with L- neuroendocrine-type calcium channel subtypes (α1D) subunit of the relationship between changes in protein expression. And dihydropyridine compounds (Dihydropyridines, DHPs) nifedipine, (-) Bay K8644 can intervene. Methods fourth generation of cultured human RPE cells were randomly divided into four groups, A group: control group; B Group: Lighting group; C Group: Lighting nifedipine group; D Group: Lighting (-) Bay K8644 group, with the 2000 ± 5001ux Blu-ray irradiation in vitro cultured human RPE cells 6h, 24h termination of culture, enzyme-linked immunosorbent assay cells secrete vascular endothelial growth factor (vascular endothelial cell growth factor, VEGF) basic fibroblast growth factor (basic fibroblast growth factor, bFGF), pigment epithelium-derived factor (pigment epithelium-derived factor, PEDF); protein immunoblot (Western blot) to detect L-type calcium channel α1D subunit protein; reverse transcription polymerase chain reaction (RT-RCR ) and quantitative PCR analysis of L-type calcium channel subunits α1DmRNA neuroendocrine expression; using laser confocal microscopy of intracellular calcium concentration. Results (1) RPE cell function changes: VEGF: light group and light (-) Bay K8644 set higher, the difference was statistically significant (P = 0.000,0.000); illumination (-) Bay K8644 high concentrations of VEGF in the light group (P = 0.000); illumination light nifedipine group than in the control group and the group, the difference was statistically significant (P = 0.000,0.001). bFGF: light group, light (-) Bay K8644 set higher, the difference was statistically significant (P = 0.019,0.000); illumination nifedipine bFGF concentrations below the light group, the difference was statistically significant (P = 0.000); illumination (-) BayK8644 group bFGF concentrations higher than the light group (P = 0.006) illumination nifedipine bFGF compared with the control group showed no significant difference (P = 0.078). PEDF: light group, light nifedipine, light (-) Bay K8644 compared with the control group, the difference was not statistically significant (P = 0.318,0.411,0.860); illumination nifedipine group, light (-) Bay K8644 Groups and light group, the difference was not statistically significant (P = 0.856,0.243); illumination (-) Bay K8644 nifedipine group and lighting group, the difference was not statistically significant (P = 0.321) (2) RPE cells L - type calcium channel α1D subunit protein expression changes: light group, light (-) Bay K8644 group α1D subunit / βactinIOD values ??were higher, the difference was statistically significant (P = 0.000,0.000); illumination (- ) Bay K8644 group than light nifedipine group, the difference was statistically significant (P = 0.000). Polyacrylamide gel group and display illumination light (-) Bay K8644 group α1D subunit protein expression was higher with light nifedipine group. (3) RPE cells in each group of L-type calcium channel α1D subunit protein expression in RPE cells secrete VEGF levels with a positive correlation function is Y = 0.7377X 0.4392, (Y is the concentration of VEGF, X is α1D subunit protein expression volume), the correlation coefficient r was statistically significant. (R = 0.674, F = 8.333, P = 0.016), RPE cells in each group of L-type calcium channel α1D subunit protein expression in RPE cells with bFGF concentration correlation function Y = 0.0739X 0.2466, (Y as bFGF concentration, X is α1D subunit protein expression), no statistically significant correlation coefficient (r) (r = 0.537, F = 4.061, P = 0.072). (4) changes in intracellular calcium ion concentration: light group, (-) BayK8644 group, light (-) BayK8644 group RPE intracellular calcium fluorescence intensity higher than that without light group, the difference was statistically significant (P = 0.000,0.000, 0.000), nifedipine group than in the absence of light group, the difference was statistically significant (P = 0.000), with no light illumination nifedipine group compared with the light nifedipine nifedipine group, the difference was statistical significance (P = 0.000,0.000), (-) BayK8644 group and light (-) BayK8644 group, the difference was not statistically significant (P = 0.155). (5) α1D subunit mRNA expression: light group, light (-) Bay K8644 light nifedipine group and α1D subunit mRNA were higher, the difference was statistically significant (P = 0.023,0.006,0.010) ; illumination (-) Bay K8644 group than light group and illumination nifedipine group, the difference was statistically significant (P = 0.032,0.039); illumination and illumination nifedipine group was not statistically significant (P = 0.459) . Conclusions (1) Blu-ray irradiation causes human RPE cells L-type calcium channel α1D subunit mRNA and protein expression increased, VEGF, bFGF increased secretion, α1D subunit protein expression and secretion of VEGF positive correlation between the amount; (2) calcium channel blocker nifedipine can make RPE cells α1D subunit protein expression decreased, VEGF, bFGF secretion decreases and the channel agonist (-) Bay K8644 makes RPE cells α1D subunit protein expression, VEGF, bFGF secretion increased. (3) calcium ion concentration in blue light led to cultured human RPE cell damage process.

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