Based on the physiological structure of the intestine, a Caco-2/EAhy926 tandem compound model was constructed in order to simulate the intestinal-vascular barrier. This model was applied in the study of transcytosis o...Based on the physiological structure of the intestine, a Caco-2/EAhy926 tandem compound model was constructed in order to simulate the intestinal-vascular barrier. This model was applied in the study of transcytosis of nanoparticles, and it was compared with the traditional intestinal cell model in the whole study. Briefly, Fe3O4 nanoparticles with a size about 30 nm were used as model nanoparticles, which remained steady during transcytosis. The nanoparticles hardly had cytotoxicity to Caco-2 cells and EAhy926 cells within the incubation concentrations. The cell tandem model was established by connecting upper Caco-2 monolayer and lower EAhy926 monolayer. Based on the FD4 permeability or TEER, all cell models remained integrity within certain period of culture time. The expression of Claudin-4 or VE Cadherin demonstrated the presence of tight junctions. The intact morphology of microfilament F-actin indicated the favorable intracellular connection. It was found that the two-layer cell tandem model created a bigger barrier for the transcytosis of FD4 than Caco-2 and EAhy926 monolayer models, and the translocation of Fe3O4 nanoparticles showed a similar pattern. Interestingly, we found that the main barrier of tandem model for nanoparticles was caused by the upper Caco-2 cell monolayer, while the lower layer of EAhy926 monolayer remained high permeability. Generally, the cell tandem compound model established here enabled us to evaluate the impact of both intestinal epithelial and endothelial layer on transcytosis, and it might provide a novel approach to study bio-nano interaction in the intestine.展开更多
目的:为探讨桂皮醛对糖尿病视网膜病变新生血管的作用及机制,观察了桂皮醛对血管内皮生长因子(VEGF)诱导EA. hy926细胞增殖、迁移、成管以及Janus激酶2/信号传导与转录激活因子3(JAK2/STAT3)通路的影响。方法:将EA.hy926细胞分成空白组...目的:为探讨桂皮醛对糖尿病视网膜病变新生血管的作用及机制,观察了桂皮醛对血管内皮生长因子(VEGF)诱导EA. hy926细胞增殖、迁移、成管以及Janus激酶2/信号传导与转录激活因子3(JAK2/STAT3)通路的影响。方法:将EA.hy926细胞分成空白组、模型组(7μg·L-1VEGF),VEGF+桂皮醛(60,90,120,150μmol·L-1)组,分别采用噻唑蓝(MTT)比色法和划痕实验检测桂皮醛对VEGF诱导EA. hy 926细胞增殖和迁移作用的影响;将EA. hy 926细胞分成空白组,模型组(7μg·L-1VEGF),VEGF+桂皮醛(90,150μmol·L-1)组,采用管腔形成实验检测桂皮醛对VEGF诱导EA. hy 926细胞成管作用的影响;将EA. hy 926细胞分成空白组,模型组(7μg·L-1VEGF),VEGF+AG490 (50μmol·L-1)组,VEGF+桂皮醛(90μmol·L-1)组,VEGF+桂皮醛(150μmol·L-1)组,VEGF+桂皮醛(150μmol·L-1)+AG490(50μmol·L-1)组,采用蛋白免疫印迹法(Western blot)检测桂皮醛对VEGF诱导EA. hy 926细胞JAK2/STAT3通路的影响。结果:与空白组比较,模型组能够显著地促进EA. hy 926细胞增殖和迁移(P <0. 01)。与模型组比较,桂皮醛(60,90,120,150μmol·L-1)组能显著抑制VEGF诱导EA. hy 926细胞的增殖和迁移(P <0. 01)。与空白组比较,VEGF对EA. hy 926细胞成管具有一定的促进作用,成管的节点数、交叉点数、网眼数和血管分支数均有增加,但无统计学差异。与模型组比较,桂皮醛(90,150μmol·L-1)组对成管的节点数、交叉点数和网眼数均有明显抑制作用(P <0. 05,P <0. 01)。与空白组比较,模型组p-JAK2,p-STAT3,STAT3蛋白表达明显升高(P <0. 05,P <0. 01)。与模型组比较,桂皮醛(150μmol·L-1)能够显著抑制VEGF引起的p-JAK2,p-STAT3,STAT3蛋白表达升高(P <0. 01),桂皮醛(90μmol·L-1)能够显著抑制VEGF引起的p-STAT3,STAT3蛋白表达升高(P <0. 05,P <0. 01)。结论:桂皮醛对VEGF诱导EA. hy 926细胞的增殖、迁移、成管具有明显的抑制作用,该作用与抑制JAK2/STAT3通路的激活有关。展开更多
基金The National Basic Research Program of China(973 program,Grant No.2015CB932100)the National Basic Research Program of China(Grant No.2015CB932100)+1 种基金National Natural Science Foundation of China(Grant No.81690264)the Innovation Team of the Ministry of Education(Grant No.BMU20110263)
文摘Based on the physiological structure of the intestine, a Caco-2/EAhy926 tandem compound model was constructed in order to simulate the intestinal-vascular barrier. This model was applied in the study of transcytosis of nanoparticles, and it was compared with the traditional intestinal cell model in the whole study. Briefly, Fe3O4 nanoparticles with a size about 30 nm were used as model nanoparticles, which remained steady during transcytosis. The nanoparticles hardly had cytotoxicity to Caco-2 cells and EAhy926 cells within the incubation concentrations. The cell tandem model was established by connecting upper Caco-2 monolayer and lower EAhy926 monolayer. Based on the FD4 permeability or TEER, all cell models remained integrity within certain period of culture time. The expression of Claudin-4 or VE Cadherin demonstrated the presence of tight junctions. The intact morphology of microfilament F-actin indicated the favorable intracellular connection. It was found that the two-layer cell tandem model created a bigger barrier for the transcytosis of FD4 than Caco-2 and EAhy926 monolayer models, and the translocation of Fe3O4 nanoparticles showed a similar pattern. Interestingly, we found that the main barrier of tandem model for nanoparticles was caused by the upper Caco-2 cell monolayer, while the lower layer of EAhy926 monolayer remained high permeability. Generally, the cell tandem compound model established here enabled us to evaluate the impact of both intestinal epithelial and endothelial layer on transcytosis, and it might provide a novel approach to study bio-nano interaction in the intestine.
文摘目的:为探讨桂皮醛对糖尿病视网膜病变新生血管的作用及机制,观察了桂皮醛对血管内皮生长因子(VEGF)诱导EA. hy926细胞增殖、迁移、成管以及Janus激酶2/信号传导与转录激活因子3(JAK2/STAT3)通路的影响。方法:将EA.hy926细胞分成空白组、模型组(7μg·L-1VEGF),VEGF+桂皮醛(60,90,120,150μmol·L-1)组,分别采用噻唑蓝(MTT)比色法和划痕实验检测桂皮醛对VEGF诱导EA. hy 926细胞增殖和迁移作用的影响;将EA. hy 926细胞分成空白组,模型组(7μg·L-1VEGF),VEGF+桂皮醛(90,150μmol·L-1)组,采用管腔形成实验检测桂皮醛对VEGF诱导EA. hy 926细胞成管作用的影响;将EA. hy 926细胞分成空白组,模型组(7μg·L-1VEGF),VEGF+AG490 (50μmol·L-1)组,VEGF+桂皮醛(90μmol·L-1)组,VEGF+桂皮醛(150μmol·L-1)组,VEGF+桂皮醛(150μmol·L-1)+AG490(50μmol·L-1)组,采用蛋白免疫印迹法(Western blot)检测桂皮醛对VEGF诱导EA. hy 926细胞JAK2/STAT3通路的影响。结果:与空白组比较,模型组能够显著地促进EA. hy 926细胞增殖和迁移(P <0. 01)。与模型组比较,桂皮醛(60,90,120,150μmol·L-1)组能显著抑制VEGF诱导EA. hy 926细胞的增殖和迁移(P <0. 01)。与空白组比较,VEGF对EA. hy 926细胞成管具有一定的促进作用,成管的节点数、交叉点数、网眼数和血管分支数均有增加,但无统计学差异。与模型组比较,桂皮醛(90,150μmol·L-1)组对成管的节点数、交叉点数和网眼数均有明显抑制作用(P <0. 05,P <0. 01)。与空白组比较,模型组p-JAK2,p-STAT3,STAT3蛋白表达明显升高(P <0. 05,P <0. 01)。与模型组比较,桂皮醛(150μmol·L-1)能够显著抑制VEGF引起的p-JAK2,p-STAT3,STAT3蛋白表达升高(P <0. 01),桂皮醛(90μmol·L-1)能够显著抑制VEGF引起的p-STAT3,STAT3蛋白表达升高(P <0. 05,P <0. 01)。结论:桂皮醛对VEGF诱导EA. hy 926细胞的增殖、迁移、成管具有明显的抑制作用,该作用与抑制JAK2/STAT3通路的激活有关。
