核心岩藻糖基化修饰调控肺间质周细胞转分化的机制及干预

Pulmonary interstitial fibrosis is a common outcome of a variety of pulmonary interstitial disease. Myofibroblast is a main culprit for this occurrence. Recent studies have found that pericyte is one of major sources of myofibroblasts. The transition of pericyte is a complex process involving the activation of mutiple signal pathways such as PDGF, TGF-β pathways. Our previous findings sugggested that the key receptor proteins PDGFR, TGF-βR of core fucosylated modification (CF) after translation is an essential factor regulating the activity pathway. CF is virtually catalyzed by its specific -1,6 alpha fucosyltransferase (FUT8). Unfortunately, it remains barren in the field of pulmonary interstitial fibrosis. We previously found that CF plays an important role in the regulation lung pericyte transtion in vitro. But it is unclarified as to the pattern of CF activity, the underlying mechanism in the process of pericyte transition. In addition, whether it can be used as a therapeutic target for pulmonary interstitial fibrosis or not remains unknown. Very different from other studies, we plan to use glycomics technique to analyse the pattern of CF activity in the pulmonary interstitial fibrosis; The already established PDGFR-GFP transgenic mice and FUT8 gene knockout mice are planed to be used in the research of CF in the process of pericyte transition in the pulmonary interstitial fibrosis; The therapeutic comparisons between the suppression on CF of multiple signal pathways and the single inhibition of TGF-β pathway will be made to evaluate the efficacy of CF on pericyte transition and pulmonary interstitial fibrosis. Our findings may throw a light for the mechanism of pulmonary interstitial fibrosis and might provide a novel therapeutic strategy in clinic.

肺间质纤维化是多种肺间质疾病的共同结局，肌成纤维细胞是其进展的重要执行者。新近发现周细胞是肌成纤维细胞重要来源，周细胞转分化是PDGF、TGF-β等多信号通路活化的复杂过程。我们前期研究发现关键受体蛋白PDGFR、TGF-βR翻译后的核心岩藻糖基化（CF）是上述通路活化之必需和重要的调控者。CF由特异性α-1,6岩藻糖转移酶（FUT8）催化，在肺间质纤维化的研究尚空白。我们研究发现CF在体外培养肺周细胞转分化中起重要作用，但CF在周细胞转分化和肺间质纤维化中的变化规律、作用机制及是否可作为治疗靶点均尚未知晓。与以往蛋白质表达的多数研究不同，本研究从CF全新角度，利用糖生物组学技术揭示肺间质纤维化中CF的变化特征；利用基因修饰小鼠明确CF在肺间质纤维化的作用及机制；比较抑制CF降低多信号通路活性与给予TGF-βR抑制剂阻断单信号通路活性对小鼠肺间质纤维化影响的差异，探索干预肺纤维化的新思路。

肺间质肌成纤维细胞增多及微血管毁损是肺间质纤维化的核心事件。新近研究发现周细胞是肌成纤维细胞的主要来源。周细胞活化后会脱离内皮细胞，导致微血管毁损、组织缺血缺氧，加重肺间质纤维化。同时周细胞转化为肌成纤维细胞，导致细胞外基质沉积，形成肺间质纤维化。上述研究表明：周细胞活化是微血管毁损与肺间质肌成纤维细胞聚集的重要连接点，抑制周细胞活化有可能成为未来治疗肺间质纤维化的新策略。研究表明：转化生长因子β1（TGF-β1）、血小板源性生长因子（PDGF）、等多条“致肺间质纤维化信号通路”过度激活是周细胞活化发生的基础，抑制多条信号通路的过度激活可阻止肺脏周细胞活化。.我们课题组前期研究发现上述信号通路受体蛋白均为糖蛋白，存在蛋白质翻译后必需有糖基化修饰。核心岩藻糖基化（CF）是重要的蛋白质糖修饰，是由α-1,6 岩藻糖转移酶（FUT8）特异性地催化完成。我们课题组前期研究也发现： TGF-β1刺激体外培养的肺脏周细胞，导致其FUT8过度表达和CF修饰水平明显增加，沉默FUT8能明显抑制肺脏周细胞转分化。进一步在体内的研究显示，体内注射FUT8shRNA腺病毒、抑制FUT8催化的CF表达，可以减轻博来霉素诱导的周细胞活化；并发现其作用部分是通过抑制TGF-β、PDGF通路关键受体TGFβR、PDGFβR的CF修饰、进而降低TGF-β、PDGF信号通路活性来实现抑制周细胞活化。为了证实CF的干预效果，课题组率先开展了以CF作为多条信号通路阻断靶点与PDGFβR/ERK、TGFβR/Smad2/3单信号通路受体阻断剂的干预效果进行比较，试验结果显示抑制CF修饰，明显抑制PDGFβR/ERK、TGFβR/Smad2/3信号通路中下游磷酸化的表达，抑制周细胞转分化及肺间质纤维化的进展。