5-氨基酮戊酸光动力疗法对瘢痕成纤维细胞的作用研究

Impact of ALA-PDT on fibroblasts from keloid and its mechanism

摘要:

研究5- 氨基酮戊酸光动力疗法（ALA-PDT）对瘢痕成纤维细胞的作用及其机制，为临床运用ALA-PDT 治疗瘢痕疙瘩提供理论依据。方法取人瘢痕成纤维细胞原代培养，并将生长状态无明显差异的瘢痕成纤维细胞随机分为空白对照组、磷酸盐缓冲液（PBS）组以及6 个不同浓度的5- 氨基酮戊酸（ALA）培养液组。每组给予635 nm 发光二极管（LED）激光器照射。采用细胞计数试剂盒（CCK-8）法检测 ALA-PDT 对瘢痕成纤维细胞增殖的影响，并应用实时聚合酶链反应（real-time PCR）和Western blot 检测各组瘢痕成纤维细胞中胶原蛋白Ⅲ（Collagen Ⅲ）mRNA 水平和蛋白表达水平。结果与空白对照组及PBS组比较，CCK-8 结果表明，ALA-PDT 能抑制瘢痕成纤维细胞生长，其抑制作用随着ALA 浓度增加逐渐增强，当ALA 浓度为0.500 mmol/L时抑制作用最明显。RT-PCR 和Western blot结果表明，随着ALA 浓度增大，CollagenⅢ mRNA 水平和蛋白表达水平逐渐下降，当ALA 溶液浓度达到0.500 mmol/L 时下降至最低值。结论ALA-PDT 能有效抑制瘢痕成纤维细胞增殖，抑制作用在ALA 浓度为0.500 mmol/L 时最强。ALA-PDT 可能是通过抑制瘢痕成纤维细胞中CollagenⅢ的合成，进而抑制瘢痕疙瘩的形成。

Abstract:

To study the impact of 5-aminolevulinic acid based photodynamic therapy (ALA-PDT) on proliferation of fibroblasts from keloid and its mechanism for guiding the use of ALA-PDT treating keloid in clinic. Methods Fibroblasts from patients' keloid were primarily cultured, then those cells without significant difference in growth state were divided into control group, PBS group and 6 ALA groups. Each group received PDT of 635 nm light emitting diode (LED) laser irradiation. Cell counting kit-8 (CCK-8) was used to detect the suppressive rate of ALA-PDT on the proliferation of the fibroblasts. RT-PCR and Western blot were used to detect the mRNA and protein levels of Collagen III. Results Compared with the control and PBS groups, ALA-PDT inhibited the proliferation of fibroblasts and the suppressive rate was gradually enhanced with the increase of the ALA concentration. When the concentration of ALA was 0.500 mmol/L, the inhibition was the strongest. The levels of CollagenⅢ mRNA and protein were decreased with the increase of the ALA concentration, and the levels of CollagenⅢ mRNA and protein were the lowest when the concentration of ALA was 0.500 mmol/L. Conclusions ALA-PDT could inhibit the proliferation of fibroblasts from keloid and the inhibition would be the strongest at the ALA concentration of 0.500 mmol/L. The mechanism may be associated with the suppression of the synthesis of CollagenⅢ in scar fibroblasts.