CO2点阵激光介导5-FU乙醇脂质体靶向干预皮肤瘢痕增生的调控机制研究

This study of scar regulatory mechanisms uses a new method to deliver the anti-scarring drugs and to explore the new ways to study the regulatory mechanisms in scar tissue. The method involves the targeted intervention on scar with ethosomes containing the 5-FU mediated by CO2 fractional laser technique. The application of CO2 fractional laser-mediated 5-FU ethosomes is used in isolated human hypertrophic scar and rabbit ear scar tissue models for the first time. The objective of using this model is to observe its effects of penetration into scar tissue, to evaluate the drug release range as well as the pathological changes in the scar tissue in different time periods. And to detect the dynamic expression of regulatory proteins, cytokines etc associated with the hypertrophic scar tissue along with fibroblast’s apoptotic process. Clarifying whether the CO2 fractional laser itself promotes the collagen remodeling and/or the superimposed effect of the intervention associated with 5-FU carrying ethosomes. In all, the experimental data results show that the CO2 fractional laser effect to be promoting the drug penetration, improving the efficiency of the anti-scarring drug administration. It is very likely to provide a new avenue to reducing skin scar formation.

本实验研究CO2点阵激光介导乙醇脂质体携带抗瘢痕药物5-FU靶向干预皮肤瘢痕增生调控机制。首次将CO2点阵激光介导5-FU乙醇脂质体应用于人离体增生性瘢痕组织及在体兔耳增生性瘢痕模型，观测其对增生性瘢痕的渗透效果和药物释放范围的影响，以及作用不同时期瘢痕组织的病理变化，检测成纤维细胞凋亡及增生性瘢痕组织相关调控蛋白，细胞因子等分泌表达等动态影响，阐明CO2点阵激光本身对促进瘢痕组织胶原重塑作用及其介导载药乙醇脂质体后对皮肤瘢痕组织的干预的叠加效应和在不同给药时段皮肤瘢痕组织的修复状态, 为确定CO2点阵激光促进药物渗透瘢痕组织、提高抗瘢痕药物给药效率提供有效实验数据，为皮肤瘢痕组织修复研究提供新途径。

背景和目的：.观察CO2点阵激光对载5-氟尿嘧啶 （5-Fluorouracil, 5-FU）的乙醇脂质体凝胶（Ethosomal Gel, EG）在增生性瘢痕组织的促渗作用以及促渗作用的持续时间，观测CO2点阵激光联合载5-FU的EG作用于在体兔耳增生性瘢痕后的组织学及相关细胞因子的变化。.研究内容和结果：.借助离体透增生性瘢痕实验，评价CO2点阵激光的促渗作用：透瘢痕实验24h后，实验组5-FU的累积透过量为为107.61±13.27高于对照组20.73±3.77，P<0.05，实验组瘢痕组织滞留量 (µg/cm2)为24.42±4.37高于对照组12.45±1.64，P<0.05。罗丹明标记的载5-FU的ES在瘢痕组织的荧光强度24h后实验组为270.36±16.89高于对照组148.25±8.73，P<0.05。.借助在体兔耳增生性瘢痕实验，观测经CO2点阵激光照射后0h、6h、12h、24h、3天和7天时对罗丹明标记的载5-FU的ES的促渗作用：在激光照射后的0h和6h，通道开放率为100.00%；12h时通道开放率达90.59%；24h时通道开放率下降到15.58%；激光照射后3天和7天时，通道开放率为0%。.借助在体兔耳增生性瘢痕实验分别在3、7、10、14天行大体观测后处死并留取各组瘢痕样本进行病理切片和HE染色观察增生性瘢痕组织的相对厚度、胶原纤维排列等状态。同时留取样本抽提RNA并逆转录后采用实时定量PCR检测样本中Ⅰ型胶原、Ⅲ型胶原、TGF-β1、VEGF 和IL-6的表达格局。干预14天后，实验组中Ⅰ型胶原、Ⅲ型胶原以及TGF-β1的mRNA表达量均显著降低；VEGF的表达降低的程度在各组间为：联合治疗组＜激光治疗组＜脂质体治疗组＜对照组，其中联合治疗组和对照组之间有显著差异（p<0.05）；IL-6仅在激光治疗组中表达呈上升，与对照组相比有显著差异（p<0.05），而联合治疗组和脂质体治疗组中未见显著改变。.结论和科学意义：.CO2点阵激光可以迅速且显著地促进载5-FU的EG渗透增生性瘢痕，但其促渗效果在24h开始消失，故CO2点阵激光作用后24h内是促渗的关键时期。CO2点阵激光联合外用载5-FU的EG可以有效治疗增生性瘢痕，其机制可能与Ⅰ型胶原、Ⅲ型胶原、TGF-β1和VEGF表达受抑制有关，IL-6的调控机制尚待进一步研究。