碱性成纤维细胞生长因子促压疮愈合中氧化应激和微循环改善调控的机制研究

Pressure ulcer (PU) is a kind of chronic and refractory wound,which is a commom and serious complication in clinical practice. The mechanism of schemic injury and the effect of growth factors in the development of UP become a focus on wound healing. Whereas the regulating effect of oxidative stress、melioration of the microcirculation and growth factors on pressure ulcer healing was little reported . This study was conducted to explore the mechanism on wound healing of PU by cell culture in vitro, animal model and clinical study, with control by basic fibroblast growth factor (bFGF). Research is concerned with: First, it was observed the proliferation, GAPDH, MDH和LDH performance and analyzed the effect of bFGF in the primary culture of fibroblast and muscle cells. Second, to test themolecular mechanism on oxidatie stress microcirculation change through the animal model on early and late stage. Meanwhile, it is analysis on regulatory effect of bFGF. Finally, with the clinical study, wound is observed and assessed by Near Infrared System on local skin blood perfusion, regional oxyhemoglobin saturation. At the same time, on the ulcer of skin, immunohistochemical method is take to analysis process of wound healing and the effects of bFGF protection. Combining the methods of different level, the purpose of this study is to clarify mechanism of cellular function which response of oxidative stress and microcirculation changes on pressure ulcer, to explore the mechanism of bFGF and its regulation effects. It will clarify the pathogenesis of presssure ulcer development and perfect the strategy on pressure ulcer treatment.

压疮为慢性难愈性创面，是临床上常见的严重并发症。缺血性损伤机制及生长因子在压疮发生、发展中的作用已成为关注焦点。然而，关于压疮损伤中氧化应激、微血管改善及生长因子调控作用研究报道尚少。本项以外源性生长因子（bFGF）为干预，分别在体外原代细胞培养、压疮动物模型及临床压疮探讨压疮难愈机制及其bFGF的调控作用：①通过原代培养成纤维细胞和肌细胞观察压疮细胞在氧化应激环境对成细胞增殖情况及其胞浆与核内所含氧化应激相关酶的变化趋势及bFGF调控作用；②通过压疮早期、晚期动物模型验证压力造成氧化应激和微血管障碍的分子机制及相关性，观察bFGF保护的作用靶点；③通过对压疮创面观察、早期压疮应用近红外光检测组织血流灌注、血氧饱和度等变化，晚期创面用免疫组化法分析等来进一步分析难愈压疮机制及bFGF对压疮的促修复意义。旨在阐明微循环障碍和氧化应激反应对压疮损伤及调控的机制和生长因子类药物的治疗策略。

压疮又称为压力性损伤，是临床长期卧床患者常见并发症，是临床护理工作中十分棘手的问题。缺血性损伤机制及生长因子在压疮发生、发展过程中具有重要作用。而压疮损伤中关于氧化应激、微循环改善及生长因子调控的报道较少。本研究将外源性生长因子（bFGF）做为干预，以压疮动物模型、细胞模型及临床压疮为载体，分析压疮形成、发展过程中氧化应激、血管生成和细胞凋亡等相关细胞因子变化，探讨氧化应激和微循环障碍在压疮发生发展中的作用及外源性bFGF保护作用，旨在明确压疮形成机制，寻找干预靶点，同时为生长因子临床应用提供科学理论依据。. 本项目主要内容包括：.1）.构建大鼠骨骼肌氧化应激模型，观察Bcl-2及PCNA等蛋白表达，探讨bFGF对骨骼肌细胞凋亡及增殖作用。结果显示：大鼠成肌细胞活力随氧化应激损伤程度加重而下降，外源性bFGF可通过下调氧化应激水平，减轻肌细胞凋亡，减轻组织损伤。.2）.构建压疮DTI动物模型，观察内质网应激及细胞凋亡在DTI形成中的作用及外源性bFGF对肌组织修复作用，结果显示：内质网应激诱导的细胞凋亡参与压疮深部组织损伤，其机制可能与chop，caspase12介导细胞凋亡有关；外源性bFGF可通过提高成肌素表达，激活肌肉生长相关信号通路，促进大鼠压疮DTI肌肉修复；.3）.通过检测ICU患者骶尾部组织血氧情况，观察Braden量表及近红外线监测仪对早期压疮的监测作用；收集临床急慢性创面、正常组织，检测生长因子及胶原等变化，探讨晚期压疮难愈机制。结果显示：随住院时间延长，ICU患者骶尾部组织rSO2持续下降，使用近红外光谱法结合Braden量表评估有利于临床早期压疮监测；晚期压疮创面中心组织bFGF及VEGF蛋白明显低于急性创面组织，胶原减少或消失，可能是压疮难愈的重要因素。.4）构建晚期压疮动物模型，验证炎症因子及胶原在晚期压疮发展中的作用。结果显示：大鼠压疮局部皮肤组织中TNF-α表达增高介导的炎症反应引起MMP-2高表达和胶原减少可能是压疮形成的重要原因之一；本项目从动物实验、细胞实验、临床压疮三个层面分别探讨压疮形成与发展的可能机制及外源性bFGF保护作用，为临床压疮防治奠定基础。