Inhibition of proliferation, migration and tube formation of choroidal microvascular endothelial cells by targeting HIF-1α with short hairpin RNA-expressing plasmid DNA in human RPE cells in a coculture system

Zhao, Wei; Wang, Yusheng; Hui, Yan-Nian; Zhang, Peng; Li, Xia; Dou, Guo‐Rui

doi:10.1007/s00417-008-0858-8

Cited by 36 publications

(25 citation statements)

References 33 publications

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“…HUVECs were isolated, cultured, and characterized as previously described. 19 For the hypoxia treatment, cells were cultured in a modular incubator chamber (Billups-Rothenberg, Del Mar, Calif), which was flushed with 1% O 2 , 5%CO 2 , and 94% N 2 ; and the cells were harvested after 3 hours, 6 hours, 9 hours, and 12 hours.…”

Section: Cell Culture and Hypoxia Modelmentioning

confidence: 99%

“…19 For each assay model, MCF-7 cells were transfected with the control plasmid, V14RhoA, or V14RhoA and were treated with nutlin-3 (Sigma-Aldrich, St. Louis, Mo). In the proliferation, migration, stress-fiber, and tubeformation assays, MCF-7 cells were plated in Costar Transwells (Costar/Corning, Corning, NY) with 0.40 lm poresize inserts, which were placed into wells in which HUVECs were plated.…”

Section: Coculture Assaysmentioning

confidence: 99%

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Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor‐dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions

Xue

Cui

et al. 2012

Cancer

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BACKGROUND: Tumor neovascularization (TNV) is a common pathologic basis for malignant growth and metastasis. However, the mechanism of TNV pathogenesis is not fully understood. Ras homolog gene family, member A (RhoA), a Rho guanosine triphosphatase (GTPase) family member, may be involved in a hypoxia-induced vascular endothelial growth factor (VEGF) pathway that regulates TNV angiogenesis through an unclear mechanism. METHODS: The regulation of RhoA on p53, the p53 binding protein homolog murine double minute 2 (MDM2), and VEGF was analyzed in hypoxic MCF-7 cells using Western blot analysis, real-time polymerase chain reaction (PCR) analysis, coimmunoprecipitation, and immunofluorescence staining assays. Changes in proliferation, invasion, migration, stress fiber formation, and tube formation were detected in an MCF-7 human umbilical vein endothelial cell (HUVEC) coculture system. Correlations of RhoA expression with MDM2, wild-type p53 (wt-p53), and VEGF expression in breast cancer tissues and relations between RhoA and breast cancer clinical features were analyzed by immunohistochemistry. RESULTS: Activated RhoA down-regulated p53 protein, which increased VEGF expression in hypoxic MCF-7 cells; whereas p53 messenger RNA levels were not altered. In addition, the ubiquitin-mediated degradation of p53 was enhanced by active RhoA. RhoA and MDM2 colocalized in the cytoplasm of hypoxic MCF-7 cells and interacted with each other physically. Furthermore, nutlin-3, a specific MDM2 inhibitor, was capable of reducing activated RhoA-induced p53 protein stability and attenuating VEGF accumulation. In an MCF-7-HUVEC coculture system, nutlin-3 effectively inhibited HUVEC proliferation, invasion, migration, stress fiber formation, and tube formation mediated by activated RhoA under hypoxic conditions. Data from 129 clinical breast cancer specimens with wt-p53 revealed that high RhoA expression was correlated with high MDM2 expression, low wt-p53 expression, and high VEGF expression. CONCLUSIONS: The current data suggested that activated RhoA promotes VEGF expression and hypoxia-induced angiogenesis through the up-regulation of MDM2 to decrease p53 stability. Cancer 2012;118:4105-16.

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Section: Cell Culture and Hypoxia Modelmentioning

confidence: 99%

Section: Coculture Assaysmentioning

confidence: 99%

Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor‐dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions

Xue

Cui

et al. 2012

Cancer

View full text Add to dashboard Cite

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“…25,26 Knocking down of HIF-1a gene results in the down regulation of VEGF. 27 Moreover, RPE cells senescence and hypoxia may decrease expression of angiogenesis inhibitors such as pigmentepithelium-derived factor (PEDF), further shifting the balance to a pro-angiogenic state in the aging eye. Thus, much attention has been drawn to this master modulator, because it may not only downregulate the VEGF expression, but may also adjust some other uncertain factors to achieve a synergistic effect.…”

Section: Introductionmentioning

confidence: 99%

Inhibitory efficacy of hypoxia-inducible factor 1α short hairpin RNA plasmid DNA-loaded poly (D, L-lactide-co-glycolide) nanoparticles on choroidal neovascularization in a laser-induced rat model

Zhang

et al. 2009

Gene Ther

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The aim of this study was to evaluate the possibility of poly (D, L-lactide-co-glycolide) nanoparticle (NPs) as a gene vector for functional plasmid DNA (pDNA) and to investigate its inhibitory efficacy on experimental choroidal neovascularization (CNV). We developed intravitreal administered, hypoxia-inducible factor 1a (HIF-1a) short hairpin RNA and green fluorescent protein (GFP) co-expressed pDNA-loaded NPs (pshHIF-1a NPs). CNV was induced by laser photocoagulation in 112 rats. The rats were then randomly assigned to be injected intravitreally with phosphate-buffered saline (PBS), blank NPs, naked pDNA, control pDNA NPs and pshHIF-1a NPs, respectively, and non-injection group was set as the control. Immunofluorescence staining, fluorescein fundus angiography and histologic analysis were performed to evaluate the inhibitory efficacy on CNV. The results showed that the expression of GFP preferentially localized in the retinal pigment epithelium cell layer and lasted for 4 weeks. The fluorescein leakage areas of CNV were significantly larger in the PBS, blank NPs, control pDNA NPs, non-injection group and naked pDNA group than in pshHIF-1a NPs group (Po0.01). The mean thickness of the CNV lesions in the intravitreally pshHIF1a NPs-treated group was significantly smaller than other groups (Po0.01). No signs of functional or ultrastructural destruction in retina were detected. Therefore, pshHIF-1a NPs may act as a novel therapeutic option to transfer specific pDNA and inhibit the formation of experimental CNV.

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“…[31][32][33][34][35][36] Although RNAi and many other small molecule inhibitors have been identified, their clinical translation is slow. Hence, interest is directed toward identifying new implications of existing drugs rather than discovering new molecules.…”

Section: Discussionmentioning

confidence: 99%

Ritonavir inhibits HIF-1α-mediated VEGF expression in retinal pigment epithelial cells in vitro

Vadlapatla

Vadlapudi

Pal

et al. 2013

Eye

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Inhibition of proliferation, migration and tube formation of choroidal microvascular endothelial cells by targeting HIF-1α with short hairpin RNA-expressing plasmid DNA in human RPE cells in a coculture system

Abstract: RNAi of HIF-1alpha in RPE cells can inhibit angiogenesis in vitro and provide a possible strategy for treatment of choroidal neovascularization diseases by targeting HIF-1alpha.

Cited by 36 publications

References 33 publications

Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor‐dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions

Ras homolog gene family, member A promotes p53 degradation and vascular endothelial growth factor‐dependent angiogenesis through an interaction with murine double minute 2 under hypoxic conditions

Inhibitory efficacy of hypoxia-inducible factor 1α short hairpin RNA plasmid DNA-loaded poly (D, L-lactide-co-glycolide) nanoparticles on choroidal neovascularization in a laser-induced rat model

Ritonavir inhibits HIF-1α-mediated VEGF expression in retinal pigment epithelial cells in vitro

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