State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060, PR China.
Biochem Biophys Res Commun. 2011 Jan 7;404(1):349-56. doi: 10.1016/j.bbrc.2010.11.122. Epub 2010 Dec 4.
To investigate the effect of mitochondria-targeted antioxidant peptide SS31 on prevention of high glucose-induced injury on human retinal endothelial cells (HRECs).
Cultured P3-P5 HRECs were divided into three groups: 5 mM glucose group, 30 mM glucose group and 30 mM glucose co-treated with 100 nM SS31 group. 24 and 48 h after treatment, Annexin V-FITC/PI staining was used to evaluate the survival of HRECs. Overproduction of ROS was assessed by MitoSOX staining under confocal microscope. Change of mitochondrial potential (ΔΨ(m)) of HRECs was measured by flow cytometry after JC-1 fluorescent probe staining. Release of cytochrome c was assessed by confocal microscopy and western blot. Expression of caspase-3 and thioredoxin-2 (Trx-2) were measured by western blot and real-time PCR.
Compared to the high glucose group, co-treatment with 100 nM SS31 significantly protected HRECs from high glucose-induced injury, reduced the production of ROS in mitochondria, stabilized ΔΨ(m), decreased the release of cytochrome c from mitochondria to cytoplasm, decreased the expression of caspase-3 and increased the expression of Trx-2 in high glucose-treated HRECs.
SS31 attenuates the high glucose-induced injuries on HRECs by stabilizing ΔΨ(m), decreasing ROS production, preventing the release of cytochrome c from mitochondria, decreasing the expression of caspase-3 and increasing the expression of Trx-2. Our study suggests that SS31 may be as a potential new treatment for diabetic retinopathy and other oxidative stress-related diseases.
研究线粒体靶向抗氧化肽 SS31 对高糖诱导的人视网膜内皮细胞（HRECs）损伤的预防作用。
将培养的 P3-P5 HRECs 分为三组：5 mM 葡萄糖组、30 mM 葡萄糖组和 30 mM 葡萄糖+100 nM SS31 组。处理 24 和 48 h 后，通过 Annexin V-FITC/PI 染色评估 HRECs 的存活率。通过共聚焦显微镜下 MitoSOX 染色评估 ROS 的过度产生。用 JC-1 荧光探针染色后通过流式细胞术测量 HRECs 的线粒体膜电位（ΔΨ(m)）变化。通过共聚焦显微镜和 Western blot 评估细胞色素 c 的释放。通过 Western blot 和实时 PCR 测量 caspase-3 和硫氧还蛋白-2（Trx-2）的表达。
与高糖组相比，100 nM SS31 共处理显著保护 HRECs 免受高糖诱导的损伤，减少线粒体中 ROS 的产生，稳定 ΔΨ(m)，减少细胞色素 c 从线粒体向细胞质的释放，减少高糖处理的 HRECs 中 caspase-3 的表达，增加 Trx-2 的表达。
SS31 通过稳定 ΔΨ(m)、减少 ROS 产生、防止细胞色素 c 从线粒体释放、减少 caspase-3 的表达和增加 Trx-2 的表达，减轻高糖诱导的 HRECs 损伤。我们的研究表明，SS31 可能是治疗糖尿病视网膜病变和其他氧化应激相关疾病的一种潜在新方法。
