miR-206对细胞外基质的影响在婴幼儿血管瘤自发消退中的作用

Infantile hemangioma(IH) is the most common benign tumor in infant, with spontaneous regression as its typical characteristic. The basement membrane of capillary wall in proliferative phase of IH is thicker than normal ones. Extracellular matrix(ECM) and fat replaced the tumor organization gradually. Studies confirm that non-code RNAs, which is a key modulated component in ECM deposition, played an important role in IH process. The level of MMPs in IH tumor and urine is higher in proliferative phase than involuting and involuted phase. These studies indicates that the metabolic disorders of ECM adjusted by MMPs took part in the process of IH spontaneous regression process. Our previous studies show that miR-206 played a part in IH process, and MMPs might be the target of miR-206. Based on this, we proposed that ECM disorder medieated by miR-206 and MMPs could be play important role in IH spontaneous involuted process. To verify the hypothesis, we isolated infantile hemangioma derived endothelial cells (HemECs) from IH tumor in proliferative phase, and use dual luciferase report system and transwell system to detect the effect of miR-206 on MMPs and ECM metabolism. Animal model would be duplicated to test the effect of miR-206 on IH process. In conclusion, the study could provide theoretical basis of IH spontaneous regression and new therapeutic directions.

婴幼儿血管瘤(IH)是婴幼儿最常见软组织肿瘤，有自发消退病程特征，机制不明。增生期IH毛细血管壁基底膜增厚，自发消退过程细胞外基质成分和脂肪逐步替代瘤体组织。研究证实在细胞外基质沉淀中有关键调控作用的非编码RNAs在IH病程中发挥有重要作用，而增生期IH瘤体组织和尿液中MMPs水平明显高于消退期和消退完成期。提示MMPs调控的细胞外基质代谢异常参与了IH自发消退病程发展过程。我们前期发现miR-206在婴幼儿血管瘤的病程发展中有重要调控作用，而MMPs很可能是miR-206的靶向基因。据此我们提出：miR-206介导MMPs调控下的细胞外基质代谢异常，可能促进了婴幼儿血管瘤的自发消退过程。为验证假设拟采用自增生期IH瘤体组织中提取的内皮细胞为研究对象，通过双荧光素酶报告基因等手段检测miR-206介导MMPs对ECM的影响，并通过动物实验验证对IH病程发展的影响，为IH自发消退提供理论依据

婴幼儿血管瘤是婴幼儿最常见软组织良性肿瘤，病程特异，1岁内呈现肿瘤样增生特性，之后逐渐稳定并消退为纤维脂肪组织，瘤体具有自发消退特性，但机制不明，可能和细胞凋亡、纤维化过程等相关。婴幼儿血管瘤标本内可见基底膜明显增厚，病程发展中细胞成分减少，细胞外基质逐渐增多，和病程发展相关，但相关机制研究未有相关报道。本项目组成员在前期研究中发现miR-206在婴幼儿血管瘤组织中表达异常并可可能参与细胞外基质代谢过程。在本课题研究中我们主要的研究内容包括：通过qRT-PCR确定miR-206在婴幼儿血管瘤不同病程时期中的表达；组织块结合磁珠分选法分离培养婴幼儿血管瘤内皮细胞（HemECs）；通过表达miR-206研究HemECs表达谱芯片改变，结合miRNA靶蛋白数据库和文献分析筛选在细胞外基质代谢方面的miR-206靶蛋白，并通过双荧光素酶报告基因进行验证；研究通过筛选获得的靶蛋白对HemECs的功能影响，及对ECM代谢相关蛋白的影响，并通过动物实验进行体内水平的验证。通过本课题的研究，我们确定了miR-206在婴幼儿血管瘤增殖期的低表达状态，对HemECs的增殖、迁移侵袭和凋亡具有明显抑制作用，miR-206具有靶向DNMT3A蛋白的作用，miR-206对ECM代谢相关蛋白MMP2、MMP9、COL2A1和Aggrecan的调控作用，能够被DNMT3A的过表达所阻断，在婴幼儿血管瘤动物模型中，miR-206对细胞外基质的调控作用明显抑制了婴幼儿血管瘤病程的发展，体内实验结果与体外实验结果一致。通过本课题的实验研究，证实了miR-206参与的ECM的代谢过程，对婴幼儿血管瘤自发消退病程发展机制具有重要调控作用，丰富了婴幼儿血管瘤的病程机制，为治疗婴幼儿血管瘤提供了理论依据。