δ-氨基酮戊酸光动力疗法对瘢痕疙瘩成纤维细胞增殖及胶原分泌的影响

摘要

Objective To investigate the effect of δ-aminolevulinic acid (ALA) photodynamic therapy (PDT) on the cell proliferation,apoptosis and collagen secretion in keloid fibroblasts and to provide the theoretical base for ALA-PDT treatment of keloids.Methods Fibroblasts from keloid patients were cultured to the third generation in vitro and incubated in 0,1,3,6,9 mmol / L of δ-aminolevulinic acid for 3 h in the darkness.Then they were exposed to 635 nm wavelength red light (30 J/cm2) and continued incubation 24 h after irradiation.CCK-8 assay was used to detect proliferation inhibition rate of fibroblasts.The content of hydroxyproline was measured by colorimetric method.The expression of p-Akt and programmed cell death 4 (PDCD4) were detected by Western blot.Results The inhibition rate of keloid fibroblasts were respectively0,(8.30 ± 1.01) ％,(29.48 ± 3.27) ％,(52.01 ± 5.34) ％,(79.99 ± 5.85) ％ with the presence of difference concentrations (0,1,3,6,9 mmol / L) of ALA.The content of hydroxyproline were respectively (9.540 0 ±0.352 42),(6.242 5 ± 0.224 85),(5.107 5 ± 0.534 88),(3.490 0 ±0.623 48),(2.945 0 ±0.514 10) μg/mg.The relative expression of p-Akt were respectively 1,0.75 ± 0.12,0.52 ± 0.14,0.41 ± 0.18,0.32 ± 0.09.The relative expression of PDCD4 were respectively 1,1.18 ± 0.19,1.51 ± 0.22,0.15 ±0.30,2.44 ± 0.22.The difference was statistically significant when compared the group of 1,3,6,9 mmol/L with 0 mmol/L(P ＜ 0.05).Conclusions In concentration within the range of 1-9 mmol/L,ALA could inhibit the proliferation of fibroblasts significantly,promote fibroblasts apoptosis and reduce the content of hydroxyproline in a dose-dependent manner,indicating that δ-aminolevulinic acid photodynamic therapy may be a potential treatment for keloid.%目的 探讨δ-氨基酮戊酸(ALA)光动力疗法(PDT)对瘢痕疙瘩成纤维细胞增殖、凋亡及胶原分泌的影响.方法 取体外培养的第3代瘢痕疙瘩成纤维细胞,给予0、1、3、6及9 mmol/L的ALA避光孵育3h,635 nm波长红光照射,能量密度为30 J/cm2,照射后继续孵育24 h,CCK-8法检测成纤维细胞增殖的抑制率;比色法检测羟脯氨酸含量;Western blot法检测丝氨酸/苏氨酸蛋白激酶Akt磷酸化激活形式p-Akt及程序性死亡因子4(PDCD4)的表达水平.结果 ALA在0、1、3、6及9 mmol/L的浓度下,瘢痕疙瘩成纤维细胞增殖的抑制率分别为0、(8.30±1.01)％、(29.48±3.27)％、(52.01±5.34)％、(79.99±5.85)％;羟脯氨酸含量分别是(9.540 0±0.352 42)、(6.242 5±0.224 85)、(5.107 5士0.534 88)、(3.4900±0.623 48)、(2.945 0±0.514 10) μg/mg;p-Akt的相对表达量分别为1、0.75±0.12、0.52±0.14、0.41±0.18、0.32±0.09;PDCD4的相对表达量分别为1、1.18±0.19、1.51±0.22、0.15±0.30、2.44±0.22.1、3、6及9 mmol/L组与0 mmol/L相比,差异均具有统计学意义(P＜0.05).结论ALA在1～9 mmol/L范围内,均可可显著抑制成纤维细胞的增殖,减少羟脯氨酸的含量,表明δ-氨基酮戊酸光动力疗法可能是瘢痕疙瘩的一种潜在的治疗方法.