球状脂联素对卵巢微血管内皮细胞增殖、迁移及管状结构形成的影响

发布时间：2018-01-21 23:17

本文关键词： 球状脂联素卵巢血管新生血管内皮细胞 AMPK　出处：《南方医科大学学报》2017年05期 　论文类型：期刊论文

【摘要】：目的分离并鉴定小鼠卵巢微血管内皮细胞,并利用卵巢微血管内皮细胞研究球状脂联素对小鼠卵巢血管新生的影响。方法采用Percoll密度梯度离心法分离纯化小鼠卵巢微血管内皮细胞,并采用免疫荧光检测细胞表面的卵泡刺激素受体、黄体生成素受体以及血管内皮细胞标记v WF;通过血管生成因子(VEGFA)处理检测卵巢微血管内皮细胞在Matrigel上的毛细管形成特性。重组的球状脂联素蛋白处理卵巢微血管内皮细胞,通过MTS检测细胞增殖;细胞划痕愈合法检测细胞迁移;Matrigel基质胶的血管形成,Western blot检测单磷酸腺苷活化蛋白激酶(AMPK)磷酸化激活。结果分离出的细胞表现为卵泡刺激素受体阴性,黄体生成素受体阳性,v WF阳性,且VEGFA能够促进血管生长,具有卵巢特异的血管内皮生长特性。高剂量浓度(1μg/m L和3μg/m L)的重组的脂联素球状结构域蛋白处理后卵巢微血管内皮细胞数量分别增加(158.72±14.50)%和(186.50±4.20)%,促进细胞增殖(P0.01);高剂量浓度(1μg/m L和3μg/m L)的重组的脂联素球状结构域蛋白处理后卵巢微血管划痕愈合率分别为(49.43±3.43)%(P0.05)和(69.67±1.2)%(P0.01);3μg/m L的脂联素处理卵巢微血管内皮细胞形成的毛细管状长度为对照组的7.63±0.66倍(P0.01)。3μg/m L的球状脂联素蛋白处理饥饿处理后的卵巢微血管内皮细胞培养15 min后p AMPK/AMPK、30 min后p AMPK/AMPK显著高于未加入蛋白处理的对照(P0.01)。Compound C抑制了球状脂联素促进的卵巢微血管内皮细胞的管状结构形成及AMPK磷酸化激活。结论成功分离了卵巢微血管内皮细胞,球状脂联素蛋白能够通过激活AMPK信号途径促进卵巢微血管内皮细胞的血管新生。
[Abstract]:Objective to isolate and identify mouse ovarian microvascular endothelial cells. The effects of globular adiponectin on ovarian angiogenesis in mice were studied by using ovarian microvascular endothelial cells. Methods Mouse ovarian microvascular endothelial cells were isolated and purified by Percoll density gradient centrifugation. The follicle stimulating hormone receptor, luteinizing hormone receptor and vascular endothelial cell labeled vWF were detected by immunofluorescence. The capillary formation characteristics of ovarian microvascular endothelial cells on Matrigel were detected by VEGF treatment. Recombinant spherical adiponectin was used to treat ovarian microvascular endothelial cells. Cell proliferation was detected by MTS. Cell migration was detected by scratch healing method. Angiogenesis of Matrigel matrix glue. The phosphorylation of adenosine monophosphate activated protein kinase (AMPK) was detected by Western blot. The results showed that the isolated cells were negative for follicle stimulating hormone receptor and positive for luteinizing hormone receptor. V WF was positive and VEGFA could promote blood vessel growth. Ovarian specific vascular endothelial growth characteristics. High dose concentration of 1 渭 g / mL and 3 渭 g / mL). The number of ovarian microvascular endothelial cells was increased by 158.72 卤14.50% and 186.50 卤4.20%, respectively. Promote cell proliferation (P0.01); The healing rate of ovarian microvascular scratches after treatment with 1 渭 g / mL and 3 渭 g / mL recombined adiponectin globular domain proteins was 49.43 卤3.43% (P < 0.05). P0.05) and 69.67 卤1.2% P0.01C; The capillary length of ovarian microvascular endothelial cells treated with 3 渭 g / mL adiponectin was 7.63 卤0.66 times longer than that of control group (P 0.01). 3 渭 g / mL of globular adiponectin treated with starvation treated ovarian microvascular endothelial cells cultured for 15 min, p AMPK/AMPK. After 30 min, p AMPK/AMPK was significantly higher than that of control without protein treatment (P0.01). Compound C inhibited tubular structure formation and AMPK phosphorylation activation of ovarian microvascular endothelial cells stimulated by globular adiponectin. Conclusion OVECs were isolated successfully. Spherical adiponectin can promote angiogenesis of ovarian microvascular endothelial cells by activating AMPK signaling pathway.
【作者单位】：暨南大学附属第一医院麻醉科;暨南大学再生医学教育部重点实验室;暨南大学生命科学技术学院发育与再生生物学系;
【基金】：国家自然科学基金(30900232) 中央高校基本科研业务费专项资金(21610202) 天河区科技计划项目(201404KW041) 广东省优秀博士学位论文作者资助项目(sybzzxm201033)~~
【分类号】：R614
【正文快照】： Effect of globular adiponectin on proliferation,migration and tube formation of ovarian mi-crovascular endothelial cellsCHEN Lei1,LU Xiaosheng2,3,LI Yalan1,MAO Zhoufei2,3,XIAO Luanjuan2,3,YU Yanhong2,3Department of Anesthesia,First Affiliated Hospital1,K

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