Proteomic study of retinal proteins associated with transcorneal electric stimulation in rats.
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ABSTRACT:
SUBMITTER: Kanamoto T
PROVIDER: S-EPMC4364380 | biostudies-literature | 2015
REPOSITORIES: biostudies-literature
Publications
Proteomic study of retinal proteins associated with transcorneal electric stimulation in rats.
Kanamoto Takashi T Souchelnytskyi Nazariy N Kurimoto Takuji T Ikeda Yasuhiro Y Sakaue Hiroaki H Munemasa Yasunari Y Kiuchi Yoshiaki Y
Journal of ophthalmology 20150304
Background. To investigate how transcorneal electric stimulation (TES) affects the retina, by identifying those proteins up- and downregulated by transcorneal electric stimulation (TES) in the retina of rats. Methods. Adult Wistar rats received TES on the left eyes at different electrical currents while the right eyes received no treatment and served as controls. After TES, the eye was enucleated and the retina was isolated. The retinas were analyzed by proteomics. Results. Proteomics showed tha ...[more]
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Project description:Purpose: To investigate the effect of transcorneal electrical stimulation (TES) on the retina of wildtype Brown Norway (BN) rats by gene expression profiling. Methods: TES was applied to BN adult wildtype rat retina in vivo for 1h (1ms biphasic pulses at 20Hz; current: 200 M-BM-5A). RNA was isolated, processed and used for microarray-based expression profiling 4hrs after initial TES. An expression profile was generated for genes differentially expressed at 4hrs after TES vs. sham stimulated animals using a fold change cutoff of 1.2. We validated the profile by real-time quantitative reverse transcription-polymerase chain reaction (qPCR). In addition, the application of TES was verified at the structural and functional level. Results: Transcriptome changes associated with TES vs. sham stimulated BN wildtype retina were identified. 490 genes were differentially expressed in TES and included well-known genes as well as a large number of novel genes. 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To ensure proper integrity all RNA used in our experiments showed single peaks for the 18S and 28S bands as determined by the Agilent 2100 Bioanalyzer using the RNA 6000 Nano LabChip Kit (both Agilent Technologies Inc., CA, USA) following the manufacturer's instructions. The purity and concentration of total RNA was determined by absorbance measurements at 260 and 280nm using a Nanodrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE, USA). All RNA that did not have a 260/280 ratio between 1.9 and 2.1 was discarded to ensure high quality of samples. Respective RNAs from stimulated and sham samples were used to generate double-stranded cDNA with the T7-oligo (dT) primer according to GeneChipM-BM-. (Affymetrix Inc., Santa Clara, CA, USA) two cycle target labeling protocol as described by the manufacturer. The processed cDNA was hybridized to Affymetrix GeneChipM-BM-. Rat Gene 1.0 ST arrays for 18M-bM-^@M-^S24h (Affymetrix Inc.). 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Project description:Objective:To explore the impact of weekly transcorneal electrical stimulation (TES) over a 6-month period as a treatment for retinitis pigmentosa (RP). Methods and analysis:A prospective open-label observational trial was carried out assessing weekly TES in participants with RP for a period of 6?months followed by observation for a further 6?months. Clinical examination and investigations were carried out at 3?monthly intervals for a total of 12 months. The primary outcome measure explored safety through a descriptive analysis of adverse effects with secondary outcome measures evaluating structural and functional efficacy. Results:Seven male and seven female participants with RP aged 18-80 years were recruited. TES was well tolerated with no serious adverse events reported. Two participants reported transient foreign body sensation and one participant had discomfort underneath the skin electrode. Following 6?months of TES, best-corrected visual acuity increased by 1.1±1.4 letters in the control arm and 0.93±1.4 letters in the treated arm. Central microperimetry threshold sensitivity rose by 0.02±0.5 decibels (dB) and 0.37±0.4?dB and Goldmann visual field volume by 0.16±0.09 steradians (sr) vs 0.22±0.12?sr for the control and treated eye, respectively. There was no statistical significance seen between eyes following the treatment or observation period. Conclusion:This small open-label clinical trial showed that TES was safe and well tolerated in patients with RP. Visual function measurements at 6 months demonstrated no significant difference between the control and treated eyes. The results justify a larger clinical trial over a longer period of time in order to identify any treatment effect.
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