Department of Plastic and Reconstructive Surgery, Xinhua Hospital, School of Medicine, Shanghai Jiao Tong University, People's Republic of China.
Department of Plastic and Reconstructive Surgery, Xinhua Hospital, School of Medicine, Shanghai Jiao Tong University, People's Republic of China.
Redox Biol. 2019 Jan;20:195-203. doi: 10.1016/j.redox.2018.10.011. Epub 2018 Oct 17.
Keloids exhibit cancer-like properties without spontaneous regression and usually recur post excision. Although photodynamic therapy (PDT) is a promising treatment, details of the mechanisms remain to be elucidated. In this study, we investigated mechanisms involved in 5-Aminolevulinic Acid (5-ALA)-based PDT against keloid. Found that 5-ALA-PDT induced superoxide anion-dependent autophagic cell death. Application of autophagy inhibitor 3-Methyladenine (3-MA) significantly prevented the effect that 5-ALA-PDT induced keloid-derived fibroblasts death, but Z-VAK-FMK (apoptotic inhibitor) did not. Interestingly, 5-ALA-PDT promoted the SIRT3 protein expression and the activity of mitochondrial superoxide dismutase 2 (SOD2), but SIRT1 protein expression level was decreased. SOD2 as a key enzyme can decrease mitochondrial ROS (mROS) level, Deacetylation of SOD2 by SIRT3 regulates SOD2 enzymatic activity has been identified. Then we explored SOD2 acetylation level with immunoprecipitation, found that 5-ALA-PDT significantly increased the acetylation levels of SOD2. In order to confirm deacetylation of SOD2 regulated by SIRT3, 3-TYP (SIRT3 inhibitor) was used. Found that inhibition of SIRT3 by 3-TYP significantly increased the level of SOD2 acetylation level compared with control group or 5-ALA-PDT group. To explore the connection of SIRT1 and SIRT3, cells were treated with EX527(SIRT1 inhibitor) or SRT1720 (SIRT1 activator), and EX527 increased SIRT3 protein level, however, SRT1720 displayed the opposite effect in the present or absence of 5-ALA-PDT. Moreover SIRT1-inhibited cells are more resistant to 5-ALA-PDT and showing decreased ROS accumulation. These results may demonstrate that 5-ALA-PDT induced SIRT1 protein level decreased, which promoted the effect of SIRT3 increased activity of SOD2 that can reduce mROS level, and then compromised 5-ALA-PDT induced autophagic cell death.
瘢痕疙瘩表现出类似于癌症的特性，不会自发消退，通常在切除后复发。光动力疗法 (PDT) 是一种很有前途的治疗方法，但具体的作用机制仍有待阐明。在这项研究中，我们研究了 5-氨基酮戊酸 (5-ALA) 为基础的 PDT 治疗瘢痕疙瘩的作用机制。结果发现，5-ALA-PDT 诱导活性氧依赖的自噬细胞死亡。应用自噬抑制剂 3-甲基腺嘌呤 (3-MA) 可显著预防 5-ALA-PDT 诱导的瘢痕疙瘩衍生成纤维细胞死亡，但 Z-VAK-FMK（凋亡抑制剂）没有。有趣的是，5-ALA-PDT 促进了 SIRT3 蛋白表达和线粒体超氧化物歧化酶 2 (SOD2) 的活性，但 SIRT1 蛋白表达水平降低。SOD2 作为一种关键酶，可降低线粒体 ROS (mROS) 水平，SIRT3 调节 SOD2 酶活性的去乙酰化已得到证实。然后我们通过免疫沉淀探索 SOD2 的乙酰化水平，发现 5-ALA-PDT 显著增加了 SOD2 的乙酰化水平。为了证实 SIRT3 调节的 SOD2 去乙酰化，我们使用了 3-TYP（SIRT3 抑制剂）。结果发现，与对照组或 5-ALA-PDT 组相比，用 3-TYP 抑制 SIRT3 显著增加了 SOD2 的乙酰化水平。为了探讨 SIRT1 和 SIRT3 之间的联系，我们用 EX527（SIRT1 抑制剂）或 SRT1720（SIRT1 激活剂）处理细胞，EX527 增加了 SIRT3 蛋白水平，而 SRT1720 在有或没有 5-ALA-PDT 的情况下表现出相反的效果。此外，SIRT1 抑制的细胞对 5-ALA-PDT 更有抵抗力，并且 ROS 积累减少。这些结果表明，5-ALA-PDT 诱导 SIRT1 蛋白水平降低，促进 SIRT3 活性增加，从而降低 mROS 水平，进而损害 5-ALA-PDT 诱导的自噬细胞死亡。
