Objective

Keloid is a kind of disfiguring pathological scarring specific to human skin. For various cutaneous and internal tumours, 5-aminolevulinic acid-photodynamic therapy (5-ALA-PDT) is a curative choice, and we explored its mechanism on keloid fibroblast (KFB) proliferation and invasion via regulating forkhead box protein O6 (FoxO6) antioxidant stress.

Methods

Human keloid fibroblasts (HKFs) were cultured in vitro, treated with 5-ALA-PDT and simultaneously transfected with overexpression plasmid (oe-FoxO6). HKF proliferation, invasion, apoptosis and cell cycle were assessed by CCK-8/Transwell/TUNEL/flow cytometry assays. The protein levels of Bax, Bcl-2, FoxO6, Cyclin D1, Cyclin A1 and Cyclin B1 were determined by Western blot. SOD and CAT activities, and MDA and ROS levels were examined using the kits.

Results

Pretreatment of 5-ALA-PDT prominently inhibited the proliferation of HKFs, up-regulated Bax level, down-regulated Bcl-2 level, promoted HKF apoptosis, and notably inhibited HKF invasion. 5-ALA-PDT treatment decreased the expression of FoxO6 protein, promoted oxidative stress in HKFs, up-regulated ROS and MDA levels in HKFs and reduced SOD and CAT antioxidant enzyme activities. In addition, 5-ALA-PDT pretreatment increased the level of cell cycle-associated protein Cyclin D1, decreased the levels of Cyclin A1 and Cyclin B1, and accelerated cell cycle arrest in the G₀/G₁ phase of HKFs. Overexpression of FoxO6 partially annulled the promoting effects of 5-ALA-PDT on G₀/G₁ phase cell cycle arrest and oxidative stress in HKFs and enhanced cell proliferation and invasion.

Conclusion

5-ALA-PDT limited HKF proliferation and invasion by down-regulating the expression of FoxO6.

中文翻译：

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5-氨基乙酰丙酸-光动力疗法预处理通过介导叉头盒蛋白O6（FoxO6）抗氧化应激抑制瘢痕疙瘩成纤维细胞增殖和侵袭的机制

客观的

疤痕疙瘩是一种人类皮肤特有的毁容性病理性疤痕。对于各种皮肤和内部肿瘤，5-氨基乙酰丙酸光动力疗法（5-ALA-PDT）是一种治疗选择，我们通过调节叉头盒蛋白O6（FoxO6）抗氧化剂探讨其对瘢痕疙瘩成纤维细胞（KFB）增殖和侵袭的机制压力。

方法

体外培养人瘢痕疙瘩成纤维细胞（HKF），用5-ALA-PDT处理，同时转染过表达质粒（oe-FoxO6）。通过CCK-8/Transwell/TUNEL/流式细胞术测定HKF增殖、侵袭、凋亡和细胞周期。Western blot检测Bax、Bcl-2、FoxO6、Cyclin D1、Cyclin A1、Cyclin B1蛋白水平。使用该试剂盒检查 SOD 和 CAT 活性以及 MDA 和 ROS 水平。

结果

5-ALA-PDT预处理显着抑制HKF增殖，上调Bax水平，下调Bcl-2水平，促进HKF凋亡，并显着抑制HKF侵袭。5-ALA-PDT处理降低了FoxO6蛋白的表达，促进了HKF的氧化应激，上调了HKF中的ROS和MDA水平，降低了SOD和CAT抗氧化酶活性。此外，5-ALA-PDT预处理增加了细胞周期相关蛋白Cyclin D1的水平，降低了Cyclin A1和Cyclin B1的水平，并加速了HKFs细胞周期停滞在G ₀ /G _{1期。FoxO6的过表达部分抵消了5-ALA-PDT对HKFs中G 0} /G ₁期细胞周期阻滞和氧化应激的促进作用，并增强了细胞增殖和侵袭。

结论

5-ALA-PDT通过下调FoxO6的表达来限制HKF增殖和侵袭。