富勒醇对血管生成的作用机制研究

发布时间：2018-12-11 14:11

【摘要】：研究背景：肿瘤血管生成与肿瘤生长、转移密切相关。抗血管生成治疗成为多种实体肿瘤治疗中的有效方案。富勒醇C60(OH)22是一种低细胞毒性纳米材料，体内研究显示其具有抗肿瘤作用，机制包括免疫调节和抗肿瘤血管生成等，但它抗血管生成的作用机制尚不明确。既往研究显示C60衍生物可以调控HDAC，而HDAC抑制剂通过下调HIF-1α、VEGF的表达从而抑制肿瘤血管生成。本课题进一步研究C60(OH)22是否通过调控HDAC而影响血管生成。方法：通过检测细胞活性（MTT法）来反映C60(OH)22对永生化人脐静脉血管内皮细胞（HUVEC）增殖的影响。通过细胞侵袭实验（Transwell法）和网格形成实验来检测C60(OH)22对体外HUVEC血管生成的影响。通过检测C60(OH)22处理后细胞内活性氧（ROS）以及HDAC1、HDAC2、HIF-1α、VEGF、RECK的表达和MMP2、MMP9酶活性的变化，并进一步通过干扰HDAC1或HDAC2的表达后检测体外HUVEC介导的血管生成、细胞内ROS以及HIF-1α、VEGF、、RECK表达和MMP2、MMP9酶活性的变化，来研究C60(OH)22对血管生成的作用机制。通过裸鼠的Matrigel plug模型，进一步研究C60(OH)22对体内血管生成及对HDAC1、HDAC2、HIF-1α、VEGF、MMP2、MMP9、RECK表达水平的影响。结果： C60(OH)22能够抑制血管内皮细胞侵袭转移和网格形成，但不影响细胞活性。C60(OH)22能够降低细胞内ROS以及HDAC1、HDAC2、HIF-1α、VEGF的表达，并抑制MMP2、MMP9酶活性，但增加了RECK的表达。siRNA干扰HDAC1或HDAC2的表达，抑制了血管内皮细胞侵袭和网格形成，降低了细胞内ROS水平以及HIF-1α、VEGF的表达，并抑制了MMP2、MMP9酶活性，但增加了RECK的表达。体内实验显示C60(OH)22处理后Matrigel plug中血管数明显减少，，HDAC1、HDAC2、HIF-1α、VEGF、MMP2、MMP9的表达水平降低，RECK的表达水平增加。结论：C60(OH)22通过抑制HDAC1和HDAC2的表达从而抑制人脐静脉血管内皮细胞的血管生成。
[Abstract]:Background: tumor angiogenesis is closely related to tumor growth and metastasis. Anti-angiogenic therapy has become an effective regimen in the treatment of various solid tumors. Fullerenol C60 (OH) 22 is a low cytotoxic nano-material. In vivo studies show that C60 (OH) 22 has anti-tumor effects, including immunomodulation and anti-tumor angiogenesis, but its mechanism of anti-angiogenesis is unclear. Previous studies have shown that C60 derivatives can regulate HDAC, and HDAC inhibitors inhibit tumor angiogenesis by down-regulating the expression of HIF-1 伪 and VEGF. In this study, we further studied whether C 60 (OH) 22 affects angiogenesis by regulating HDAC. Methods: the effect of C60 (OH) 22 on (HUVEC) proliferation of human umbilical vein endothelial cells was measured by MTT assay. The effect of C60 (OH) 22 on HUVEC angiogenesis in vitro was detected by cell invasion assay (Transwell) and grid formation assay. After treatment with C60 (OH) 22, the expression of reactive oxygen (ROS), HDAC1,HDAC2,HIF-1 伪, VEGF,RECK and the activity of MMP2,MMP9 were detected. Furthermore, HUVEC mediated angiogenesis in vitro was detected by interfering with the expression of HDAC1 or HDAC2. The changes of ROS, HIF-1 伪, VEGF,,RECK expression and MMP2,MMP9 enzyme activity were used to study the mechanism of C60 (OH) 22 on angiogenesis. The effects of C60 (OH) 22 on angiogenesis and the expression of HDAC1,HDAC2,HIF-1 伪 and VEGF,MMP2,MMP9,RECK in nude mice were further studied by using Matrigel plug model of nude mice. Results: C60 (OH) 22 inhibited the invasion, metastasis and mesh formation of vascular endothelial cells, but did not affect cell activity. C60 (OH) 22 decreased the expression of ROS and HDAC1,HDAC2,HIF-1 伪, VEGF, and inhibited the activity of MMP2,MMP9 enzyme. SiRNA interfered with the expression of HDAC1 or HDAC2, inhibited the invasion and mesh formation of vascular endothelial cells, decreased the expression of ROS, HIF-1 伪 and VEGF, and inhibited the activity of MMP2,MMP9, but increased the expression of RECK. In vivo experiments showed that the number of blood vessels decreased, the expression of HDAC1,HDAC2,HIF-1 伪 and VEGF,MMP2,MMP9 decreased, and the expression of RECK increased after C60 (OH) 22 treatment. Conclusion: C60 (OH) 22 inhibits angiogenesis of human umbilical vein endothelial cells by inhibiting the expression of HDAC1 and HDAC2.
【学位授予单位】：中国人民解放军医学院
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R96

【相似文献】