血管内皮细胞是构成新生血管管腔的重要物质基础之一,其增殖是血管新生的第一步,从这一角度来说,内皮细胞增殖对缺血性心血管疾病的治疗有积极意义。DLL4（delta like ligand 4）作为一种重要的血管生长发育调节因子,其与邻近细胞相应受...血管内皮细胞是构成新生血管管腔的重要物质基础之一,其增殖是血管新生的第一步,从这一角度来说,内皮细胞增殖对缺血性心血管疾病的治疗有积极意义。DLL4（delta like ligand 4）作为一种重要的血管生长发育调节因子,其与邻近细胞相应受体Notch（Notchl或Notch4）相互作用启动细胞内的信号传递,从而调节血管的生成和分支过程。展开更多
Background Delta-like 4 （DLL4） is an endothelium specific Notch ligand and has been shown to function as a regulating factor during physiological and pathological angiogenesis. It has been reported that the DLL4-Not...Background Delta-like 4 （DLL4） is an endothelium specific Notch ligand and has been shown to function as a regulating factor during physiological and pathological angiogenesis. It has been reported that the DLL4-Notch signaling pathway is regulated by hypoxia and may prevent excessive angiogenesis through the inhibition of angiogenic branching and by triggering vessel maturation. Choroidal neovascularization （CNV） is a pathological form of angiogenesis in which hypoxia is thought to play an important role. This study was aimed to evaluate the role of DLL4 in the development of CNV. Methods We utilized chemical hypoxia induced by 200 pmol/L CoOl2 to observe the expression of DLL4 in choroid-retinal endothelial cells （RF/6A cells）, which are the primary cells involved in CNV. After transfection of a DLL4 small interfering RNA （siRNA）, mRNA and protein expression of DLL4 and key downstream genes, including HES1 and HEY1, in hypoxic RF/6A cells were investigated by RT-PCR, real-time PCR, and Western blotting analysis. Three controls were used： one without transfection, one with transfection reagent, and one with scrambled negative control siRNA. The effects of the DLL4 siRNA on the biological function of hypoxic RF/6A cells during angiogenesis, including cell proliferation, migration and tube formation, were investigated. Results The results showed that hypoxic conditions led to upregulation of DLL4 expression in RF/6A cells in vitro. After transfection, siRNA-duplexl targeting DLL4 depleted the DLL4 mRNA levels by as much as 91.4% compared with the scrambled siRNA control, and DLL4 protein expression was similarly effected. There was no significant difference in DLL4 expression among the blank control, transfection reagent control, and scrambled siRNA groups. In addition, after transfection of hypoxic RF/6A cells with the DLL4 siRNA-duplexl, the mRNA levels of HES1 and HEY1, which function downstream of DLL4-Notch signaling, were lowered by 75.1% and 86.3%, respectively, compared with the scrambled siRNA control. Furthermore, knockdown of DLL4 expression significantly promoted the proliferation of hypoxic RF/6A cells and led to their arrest in the S phase of the cell cycle. Migration and tube formation of hypoxic RF/6A cells were significantly induced by the DLL4 siRNA, with the number of migrated cells increased by 1.6-fold and total tube length increased by 82.3%, compared with the scrambled siRNA （P 〈0.05）. Conclusions DLL4 functions as a negative regulator of angiogenic branching and sprouting. Based on our results, DLL4 signaling appears to play an essential role in the biological behavior of choroid vascular endothelial cells under hypoxia. Therefore, DLL4 may represent a novel target for CNV therapy in the future.展开更多
文摘血管内皮细胞是构成新生血管管腔的重要物质基础之一,其增殖是血管新生的第一步,从这一角度来说,内皮细胞增殖对缺血性心血管疾病的治疗有积极意义。DLL4（delta like ligand 4）作为一种重要的血管生长发育调节因子,其与邻近细胞相应受体Notch（Notchl或Notch4）相互作用启动细胞内的信号传递,从而调节血管的生成和分支过程。
基金This research was supported by grants fi-om the National Natural Science Foundation of China （No. 30901640）, New Teacher Foundation of China Science and Technology Development Center of Education Ministry （No. 20090142120068）, Natural Science Foundation of Hubei Provincial Government （No. 2008CDB155） and Chenguang Program of Wuhan Science and Technology Bureau （No. 200950431170）.
文摘Background Delta-like 4 （DLL4） is an endothelium specific Notch ligand and has been shown to function as a regulating factor during physiological and pathological angiogenesis. It has been reported that the DLL4-Notch signaling pathway is regulated by hypoxia and may prevent excessive angiogenesis through the inhibition of angiogenic branching and by triggering vessel maturation. Choroidal neovascularization （CNV） is a pathological form of angiogenesis in which hypoxia is thought to play an important role. This study was aimed to evaluate the role of DLL4 in the development of CNV. Methods We utilized chemical hypoxia induced by 200 pmol/L CoOl2 to observe the expression of DLL4 in choroid-retinal endothelial cells （RF/6A cells）, which are the primary cells involved in CNV. After transfection of a DLL4 small interfering RNA （siRNA）, mRNA and protein expression of DLL4 and key downstream genes, including HES1 and HEY1, in hypoxic RF/6A cells were investigated by RT-PCR, real-time PCR, and Western blotting analysis. Three controls were used： one without transfection, one with transfection reagent, and one with scrambled negative control siRNA. The effects of the DLL4 siRNA on the biological function of hypoxic RF/6A cells during angiogenesis, including cell proliferation, migration and tube formation, were investigated. Results The results showed that hypoxic conditions led to upregulation of DLL4 expression in RF/6A cells in vitro. After transfection, siRNA-duplexl targeting DLL4 depleted the DLL4 mRNA levels by as much as 91.4% compared with the scrambled siRNA control, and DLL4 protein expression was similarly effected. There was no significant difference in DLL4 expression among the blank control, transfection reagent control, and scrambled siRNA groups. In addition, after transfection of hypoxic RF/6A cells with the DLL4 siRNA-duplexl, the mRNA levels of HES1 and HEY1, which function downstream of DLL4-Notch signaling, were lowered by 75.1% and 86.3%, respectively, compared with the scrambled siRNA control. Furthermore, knockdown of DLL4 expression significantly promoted the proliferation of hypoxic RF/6A cells and led to their arrest in the S phase of the cell cycle. Migration and tube formation of hypoxic RF/6A cells were significantly induced by the DLL4 siRNA, with the number of migrated cells increased by 1.6-fold and total tube length increased by 82.3%, compared with the scrambled siRNA （P 〈0.05）. Conclusions DLL4 functions as a negative regulator of angiogenic branching and sprouting. Based on our results, DLL4 signaling appears to play an essential role in the biological behavior of choroid vascular endothelial cells under hypoxia. Therefore, DLL4 may represent a novel target for CNV therapy in the future.
