Department of Burn and Plastic Surgery, Chengdu Second People's Hospital, 610021 Chengdu, Sichuan, China.
State Key Laboratory of Trauma, Burn and Combined Injury, Institute of Burn Research, Southwest Hospital, Third Military Medical University (Army Medical University), 400038 Chongqing, China.
Discov Med. 2024 Jul;36(186):1430-1440. doi: 10.24976/Discov.Med.202436186.133.
Keloid, a fibroproliferative disorder, significantly impacts patients' quality of life, yet effective therapies remain elusive. This study explored the role of silent information regulator 6 (SIRT6) in modulating the proliferation, invasion, and collagen synthesis of keloid fibroblasts.
Keloid and normal skin specimens were collected, and fibroblasts were isolated from the keloid tissue. SIRT6 recombinant adenovirus (Ad) was constructed to infect keloid fibroblasts to overexpress SIRT6. This study entails three groups: Control group, adenovirus-Negative Control (Ad-NC) group, and Ad-SIRT6 group. SIRT6 protein and mRNA levels were measured via Western blotting and Quantitative reverse transcription polymerase chain reaction (qRT-PCR), respectively. Cell viability was determined using 5-ethynyl-2'-deoxyuridine (EdU) assay. Flow cytometry was exploited to measure cell apoptosis. To investigate cell migration, wound healing assay and Transwell assay were employed. Western blotting was also utilized to study the expression levels of apoptotic proteins, collagen deposition-related proteins, and Mitogen-Activated Protein Kinases (MAPK)/extracellular regulated protein kinases (ERK) pathway-related proteins.
Compared to the control and Ad-NC groups, the Ad-SIRT6 group exhibited significantly elevated SIRT6 level; diminished cell proliferation, migration and invasion; reduced protein levels of α-smooth muscle actin (α-SMA), collagen I, collagen III, phospho SMAD Family Member 3 (p-Smad3), transforming growth factor-β 1 (TGF-β1), and MAPK/ERK pathway proteins (phospho extracellular signal-regulated protein kinase 1/2 (p-ERK1/2), phospho MAP kinase-ERK kinase (p-MEK) and phospho-c-Raf (p-c-Raf)). Treatment with epidermal growth factor (EGF), an MAPK/ERK pathway agonists, reversed the inhibitory effect of SIRT6 on cell activity and inhibited apoptosis in keloid fibroblasts.
SIRT6 overexpression in keloid fibroblasts attenuates proliferation, invasion, and collagen synthesis, while fostering apoptosis, likely through the suppression of MAPK/ERK pathway activity. This suggests a potential therapeutic target for keloid treatment.
瘢痕疙瘩是一种纤维增生性疾病，严重影响患者的生活质量，但有效的治疗方法仍难以捉摸。本研究探讨了沉默信息调节因子 6（SIRT6）在调节瘢痕疙瘩成纤维细胞增殖、侵袭和胶原合成中的作用。
收集瘢痕疙瘩和正常皮肤标本，从瘢痕疙瘩组织中分离出成纤维细胞。构建 SIRT6 重组腺病毒（Ad）感染瘢痕疙瘩成纤维细胞以过表达 SIRT6。本研究分为三组：对照组、腺病毒阴性对照（Ad-NC）组和 Ad-SIRT6 组。通过 Western blot 和定量逆转录聚合酶链反应（qRT-PCR）分别测量 SIRT6 蛋白和 mRNA 水平。使用 5-乙炔基-2'-脱氧尿苷（EdU）测定法测定细胞活力。通过流式细胞术测量细胞凋亡。为了研究细胞迁移，采用划痕愈合实验和 Transwell 实验。还利用 Western blot 研究凋亡蛋白、胶原沉积相关蛋白和丝裂原激活蛋白激酶（MAPK）/细胞外调节蛋白激酶（ERK）通路相关蛋白的表达水平。
与对照组和 Ad-NC 组相比，Ad-SIRT6 组 SIRT6 水平显著升高；细胞增殖、迁移和侵袭减少；α-平滑肌肌动蛋白（α-SMA）、胶原 I、胶原 III、磷酸 SMAD 家族成员 3（p-Smad3）、转化生长因子-β 1（TGF-β1）和 MAPK/ERK 通路蛋白（磷酸化细胞外信号调节蛋白激酶 1/2（p-ERK1/2）、磷酸化 MAP 激酶-ERK 激酶（p-MEK）和磷酸化 c-Raf（p-c-Raf））的蛋白水平降低。表皮生长因子（EGF）是 MAPK/ERK 通路激动剂，用 EGF 处理可逆转 SIRT6 对细胞活性的抑制作用，并抑制瘢痕疙瘩成纤维细胞凋亡。
瘢痕疙瘩成纤维细胞中 SIRT6 的过表达可减弱增殖、侵袭和胶原合成，同时促进细胞凋亡，可能通过抑制 MAPK/ERK 通路活性。这表明 SIRT6 可能成为瘢痕疙瘩治疗的潜在靶点。
