含化学促渗剂驻极体5-FU贴剂体外经皮转运和抑制瘢痕生长的动物实验研究

发布时间：2018-08-12 14:31

【摘要】：当人体皮肤损伤至真皮层,组织修复过程中往往会形成增生性瘢痕。增生性瘢痕颜色暗红,高于正常皮肤表面,易造成疼痛和瘙痒,有时还会造成关节部位移动受限。瘢痕的发病机制复杂,导致临床上治疗难度大,复发率高。5-FU作为一种抗肿瘤药,已证明对增生性瘢痕有确切疗效。5-FU常采用病灶内注射的给药方式,此种给药方式一次性剂量大,常引起红斑、色素沉着等副作用,而且重复注射导致患者顺应性差。因此改变5-FU的给药方式对增生性瘢痕的治疗具有重要意义。经皮给药系统具有给药方便,无侵入性,药物释放可控等优点,在针对某些疾病的治疗时,此种给药方式要优于口服和局部注射。本文拟将物理促渗因子驻极体和化学促渗剂相结合,将5-FU制备成经皮给药贴剂作用于大鼠创面,以期达到良好的抑制瘢痕生长的效果。本文主要内容包括:(一)含化学促渗剂5-FU贴剂处方筛选及驻极体对贴剂性质的影响本文以尤特奇E100为基质,柠檬酸丁三酯为增塑剂,水溶性氮酮为化学促渗剂,将5-FU制备成压敏胶贴剂。通过单因素考察发现E100、增塑剂和化学促渗剂对贴剂黏性均有显著影响,之后以初黏力、持黏力和体外释药量为指标,进行正交设计试验,最终确定0.25gE100,55%m(增塑剂)/m(E100)和3%浓度的氮酮为最优处方。最优处方制备的贴剂皮肤刺激性,处方稳定性和皮肤追随性均符合药典要求。本文还考察了驻极体对贴剂黏性及体外释药量的影响,结果发现较高电位的驻极体通过降低贴剂持黏力,并与5-FU产生静电作用力而促进了贴剂中药物的体外释放。(二)含化学促渗剂驻极体5-FU贴剂体外经大鼠背部皮肤和体外经大鼠瘢痕皮肤转运规律研究将不同电位不同极性含化学促渗剂驻极体5-FU贴剂(以下简称驻极体贴剂),分别作用于离体大鼠背部皮肤和瘢痕皮肤,利用改良的Franz扩散池和高效液相色谱仪检测驻极体贴剂体外经正常皮肤和经瘢痕皮肤的转运规律。结果显示:(1)驻极体贴剂经两类皮肤的累积释药规律和皮肤中药物滞留规律相似。(2)较高电位驻极体贴剂的累积透皮量和皮肤中的药物滞留量也较高,且相同电位负极性驻极体贴剂的累积透皮量和皮肤中药物滞留量均高于正极性驻极体贴剂。(3)正常皮肤的累积透皮量高于瘢痕皮肤,而皮肤组织中的药物含量却低于瘢痕皮肤。(4)较高表面电位的驻极体贴剂增加了皮肤组织中的药量含量,为后期驻极体贴剂的在体动物实验奠定了基础。(三)含化学促渗剂驻极体5-FU贴剂的动物实验研究首先制造深及筋膜层的大鼠创面,于急性炎症期过后,分别贴敷驻极体空白贴剂和驻极体药物贴剂,将皮肤样品于术后4周取样,分别做HE和Masson染色,研究驻极体和驻极体贴剂对瘢痕的治疗效果。结果显示:(1)自然愈合组的表皮和真皮层明显增生,胶原纤维沉积显著,排列紊乱,符合增生性瘢痕的特征。(2)驻极体空白贴剂治疗组形态学结构与自然愈合组相似,未表现出显著抑制瘢痕生长的效果。(3)含化学促渗剂5-FU贴剂治疗组一定程度上抑制了瘢痕的生长。相较化学促渗组,驻极体贴剂治疗组更大程度的抑制了瘢痕的增生。(四)含化学促渗剂驻极体5-FU贴剂抑制增生性瘢痕生长的机制研究本文利用免疫组织化学方法,以平均光密度值评估增生性瘢痕组织中Ⅰ、Ⅲ型胶原、TGF-β1和HSP47的阳性表达结果,进一步探讨驻极体贴剂抑制增生性瘢痕生长的机制。结果发现:(1)Ⅰ、Ⅲ型胶原、TGF-β1和HSP47在正常皮肤中均低表达,而增生性瘢痕组织中Ⅰ、Ⅲ型胶原、TGF-β1和HSP47阳性表达显著上升。进一步证明我们成功构建了大鼠瘢痕模型。(2)与自然愈合组相比,驻极体治疗组降低了Ⅲ型胶原的平均光密度值,对其它指标无显著影响。(3)含化学促渗剂5-FU贴剂治疗组、驻极体贴剂治疗组均降低了Ⅰ、Ⅲ型胶原、TGF-β1和HSP47的平均光密度值,而驻极体贴剂治疗组对各指标表达的抑制程度更显著。综上所述,驻极体贴剂在体外透皮试验中显著增加了皮肤中的药物含量,在动物实验研究中驻极体贴剂通过抑制Ⅰ、Ⅲ型胶原、TGF-β1和HSP47的表达显著抑制了大鼠瘢痕的生长,因此含化学促渗剂驻极体5-FU贴剂可以为临床增生性瘢痕的治疗提供一种新的思路和方法。
[Abstract]:Hypertrophic scars often form in the process of tissue repair when human skin is damaged to the dermis. Hypertrophic scars are dark red in color, higher than the normal skin surface, which can easily cause pain and itching, and sometimes restrict the movement of the joint. The pathogenesis of scars is complex, leading to difficult clinical treatment and high recurrence rate. 5-FU as an anti-inflammatory agent. Tumor drugs have been proved to have a definite therapeutic effect on hypertrophic scars. 5-FU is often administered by intralesional injection. This method of administration has a large dose at one time, which often causes side effects such as erythema and pigmentation, and the patient's compliance is poor after repeated injection. Therefore, it is important to change the administration of 5-FU in the treatment of hypertrophic scars. The transdermal delivery system has the advantages of convenient administration, non-invasiveness and controlled drug release. It is superior to oral and local injection in the treatment of some diseases. In this paper, the electret of physical osmotic enhancer and chemical osmotic enhancer are combined to prepare 5-FU transdermal patch to act on the wound of rats in order to achieve good results. The main contents of this paper are as follows: (1) Formula selection of 5-FU patch containing chemical penetration enhancer and the effect of electret on the properties of patch. In this paper, the pressure-sensitive patch was prepared from 5-FU patch based on EUTEC E100, butyl triester citrate and water-soluble azone. Plasticizers and chemical penetration enhancers had significant effects on the adhesive properties of patches. Then orthogonal design tests were carried out to determine the optimal formulation of 0.25 g E100, 55% m (plasticizer) / m (E100) and 3% azone with the indexes of initial adhesive strength, adhesive strength and drug release in vitro. The results showed that electrets with higher potential promoted the release of patches in vitro by reducing the adhesion of patches and producing electrostatic force with 5-FU. In vitro transport of electret 5-FU patches containing different potentials and different polarities of chemical penetration enhancers (hereinafter referred to as electret patches) on rat back skin and scar skin were studied. The electret patches were tested by modified Franz diffusion cell and high performance liquid chromatography (HPLC). The results showed that: (1) The cumulative release of electret patches through the two types of skin was similar to the retention of skin medicines. (2) The cumulative transdermal amount of electret patches with higher potential and the drug retention in the skin were also higher, and the cumulative transdermal amount of electret patches with the same potential and negative polarity and the retention of skin medicines were also higher. (3) The cumulative transdermal volume of normal skin was higher than that of scar skin, but the drug content in skin was lower than that of scar skin. (4) Electret patches with higher surface potential increased the drug content in skin tissue, which laid a foundation for the in vivo animal experiment of electret patches. (3) Inclusion In order to study the therapeutic effect of electret and electret patches on scars, the skin samples were collected 4 weeks after operation and stained with HE and Mason respectively. The results showed that: (1) The epidermis and dermis of the natural healing group were markedly hyperplasia, collagen fibers were deposited and arranged disorderly, which accorded with the characteristics of hypertrophic scars. (2) The morphological structure of the electret blank patch group was similar to that of the natural healing group, but the effect of inhibiting scar growth was not obvious. (3) The treatment group with 5-FU patch containing chemical osmotic enhancer had certain effect. Compared with the chemical osmosis group, the electret patch group inhibited the growth of hypertrophic scar to a greater extent. (4) The mechanism of the inhibitory effect of electret 5-FU patch containing chemical osmosis enhancer on the growth of hypertrophic scar. The results of positive expression of type I collagen, TGF-beta 1 and HSP47 showed that: (1) Collagen I, type III, TGF-beta 1 and HSP47 were low expressed in normal skin, while the positive expression of type I, type III collagen, TGF-beta 1 and HSP47 in hypertrophic scar tissue was significantly increased. (2) Compared with the natural healing group, the electret group decreased the average optical density of collagen type III, but had no significant effect on other indexes. (3) The electret patch group decreased the average optical density of collagen type I, III, TGF-beta 1 and HSP47, while the electret group decreased the average optical density of collagen type III, TGF-beta 1 and HSP47. In conclusion, electret patch significantly increased the drug content in the skin in vitro transdermal test. In animal experiments, electret patch significantly inhibited the growth of rat scar by inhibiting the expression of collagen I, III, TGF-beta 1 and HSP47, thus containing chemical stimulation. Infiltration electret 5-FU patch can provide a new idea and method for the treatment of hypertrophic scars.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R622

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