Background： The mechanisms of pathological retinal neovascularization （RNV） remain unknown. Several microRNAs were reported to be involved in the process of RNV. Oxygen-induced retinopathy （O1R） is a useful model...Background： The mechanisms of pathological retinal neovascularization （RNV） remain unknown. Several microRNAs were reported to be involved in the process of RNV. Oxygen-induced retinopathy （O1R） is a useful model to investigate RNV. Our present work explored the expression and the role of microRNA-128 （miR-218） in oxygen-induced RNV. Methods： OIR was used to establish RNV model. The expression level ofmiR-218 in the retina from OIR mice was assessed by quantitative real-time reverse transcriptase polymerase chain reaction. Fluorescein angiography was performed in retinae of OIR mice, and RNV was quantified by hematoxylin and eosin staining to evaluate the effect of pCDH-CMV-miR-218 intravitreal injection on RNV in OIR mice. Roundabout 1 （Robol） expression was detected by Western blotting in mouse retinal vascular endothelial cells expressing a high or low level of miR-218 and retinal tissues from OIR mice. Cell migration was evaluated by scratch wound assay. Results： In OIR mice, the expression level of miR-218 was significantly down-regulated （P = 0.006）. Retinal Robol expression was significantly increased at both mRNA and protein levels （P = 0.001, 0.008： respectively）, miR-218 intravitreal injection inhibited retinal angiogenesis in OIR mice, and the restoration of miR-218 in retina led to down-regulation of Robol. Conelusions： Our experiments showed that restoration ofmiR-218 inhibited retinal angiogenesis via targeting Robo 1. MiR-218 contributed to the inhibition of retinal angiogenesis and miR-218 might be a new therapeutic target for preventing RNV.展开更多
Retinopathy of prematurity（ROP） is a retinal vascular disorder frequently found in premature infants.Different therapeutic strategies have been developed to treat ROP.However,there are still many children with ROP s...Retinopathy of prematurity（ROP） is a retinal vascular disorder frequently found in premature infants.Different therapeutic strategies have been developed to treat ROP.However,there are still many children with ROP suffering by severe limitations in vision or even blindness.Recently,ROP has been suggested to be caused by abnormal development of the retinal vasculature,but not simply resulted by retinal neovascularization which takes about 4 to 6 wk after birth in premature infants.Thus,instead of focusing on how to reduce retinal neovascularization,understanding the pathological changes and mechanisms that occur prior to retinal neovascularization is meaningful,which may lead to identify novel target（s） for the development of novel strategy to promote the healthy growth of retinal blood vessels rather than passively waiting for the appearance of retinal neovascularization and removing it by force.In this review,we discussed recent studies about,1） the pathogenesis prior to retinal neovascularization in oxygen-induced retinopathy（OIR;a ROP in animal model） and in premature infants with ROP;2） the preclinical and clinical research on preventive treatment of early OIR and ROP.We will not only highlight the importance of the mechanisms and signalling pathways in regulating early stage of ROP but also will provide guidance for actively exploring novel mechanisms and discovering novel treatments for early phase OIR and ROP prior to retinal neovascularization in the future.展开更多
The Delta-like ligand 4/Notch signaling pathway was shown to participate in the process of retinal development and angiogenesis. However, the function of the Delta-like ligand 4/Notch signaling pathway in retinopathy ...The Delta-like ligand 4/Notch signaling pathway was shown to participate in the process of retinal development and angiogenesis. However, the function of the Delta-like ligand 4/Notch signaling pathway in retinopathy of prematurity requires further study. Retinopathy of prematurity was induced in 5-day-old Sprague-Dawley rats exposed to hyperoxia for 7 days, and then returned to room air. Reverse transcription-PCR and western blot revealed that Delta-like ligand 4 levels decreased at postnatal day 12 and increased at postnatal day 17 in retinopathy of prematurity rats. Flat-mounted adenosine diphosphatase stained retina and hematoxylin-eosin stained retinal tissue slices showed that the clock hour scores and the nuclei counts in retinopathy of prematurity rats were significantly different compared to normal control rats. After retinopathy of prematurity rats were intravitreally injected with Delta-like ligand 4 monoclonal antibody to inhibit the Delta-like ligand 4/Notch signaling pathway, there was a significant increase in the severity of retinal neovascularization （clock hours） in the intravitreally injected eyes. The nuclei count was highly correlated with the clock hour score. These results suggest that Delta-like ligand 4/Notch signaling plays an essential role in the process of physiological and pathological angiogenesis in the retina.展开更多
The mouse model of oxygen induced retinopathy is suitable for the study of various retinal neovascularization diseases,including retinopathy of prematurity.The maternally expressed gene 3(MEG3)has been demonstrated to...The mouse model of oxygen induced retinopathy is suitable for the study of various retinal neovascularization diseases,including retinopathy of prematurity.The maternally expressed gene 3(MEG3)has been demonstrated to have an inhibitory effect on diabetic retinopathy.In this study,we investigated the role of MEG3 overexpression in oxygen-induced retinopathy in mice.The results showed that MEG3 overexpression effectively inhibited the production of retinal neovascularization in oxygen-induced retinopathy mice.It acts by down-regulating the expression of phosphoinositide 3-kinase,serine/threonine kinase,and vascular endothelial growth factor and pro-inflammatory factors.MEG3 overexpression lentivirus has a future as a new method for the clinical treatment of retinopathy of prematurity.The animal experiments were approved by the Animal Ethics Committee of Shengjing Hospital of China Medical University,China(approval No.2016PS074K)on February 25,2016.展开更多
Inhibiting retinal neovascularization is the optimal strategy for the treatment of retina-related diseases, but there is currently no effective treatment for retinal neovascularization. P-element-induced wimpy testis(...Inhibiting retinal neovascularization is the optimal strategy for the treatment of retina-related diseases, but there is currently no effective treatment for retinal neovascularization. P-element-induced wimpy testis(PIWI)-interacting RNA(piRNA) is a type of small non-coding RNA implicated in a variety of diseases. In this study, we found that the expression of piR-1245 and the interacting protein PIWIL2 were remarkably increased in human retinal endothelial cells cultured in a hypoxic environment, and cell apoptosis, migration, tube formation and proliferation were remarkably enhanced in these cells. Knocking down piR-1245 inhibited the above phenomena. After intervention by a p-JAK2 activator, piR-1245 decreased the expression of hypoxia inducible factor-1α and vascular endothelial growth factor through the JAK2/STAT3 pathway. For in vivo analysis, 7-day-old newborn mice were raised in 75 ± 2% hyperoxia for 5 days and then piR-1245 in the retina was knocked down. In these mice, the number of newly formed vessels in the retina was decreased, the expressions of inflammationrelated proteins were reduced, the number of apoptotic cells in the retina was decreased, the JAK2/STAT3 pathway was inhibited, and the expressions of hypoxia inducible factor-1α and vascular endothelial growth factor were decreased. Injection of the JAK2 inhibitor JAK2/TYK2-IN-1 into the vitreous cavity inhibited retinal neovascularization in mice and reduced expression of hypoxia inducible factor-1α and vascular endothelial growth factor. These findings suggest that piR-1245 activates the JAK2/STAT3 pathway, regulates the expression of hypoxia inducible factor-1α and vascular endothelial growth factor, and promotes retinal neovascularization. Therefore, piR-1245 may be a new therapeutic target for retinal neovascularization.展开更多
The content of somatostatin(SS) in hippocampus,striatum and frontal cortex tissues of rats exposed to 600 kpa hyperbaric oxygen was determined by means of radioimmunoassay. Initial time of convulsion, severity of conv...The content of somatostatin(SS) in hippocampus,striatum and frontal cortex tissues of rats exposed to 600 kpa hyperbaric oxygen was determined by means of radioimmunoassay. Initial time of convulsion, severity of convulsion and survival time of rats with convulsion exposed to 700 kPa hyperbaric oxygen after intraperitoneal injection of cysteamine (CSH) or intracerebroventricular injection of anti-somatostatin serum(ASS) were also observed. The results showed that the content of SS in hippocampus and striatum tissues increased remarkably when rats were at near-convulsion ; by the time the rats developed convulsion,it had a significant increase in all brain areas observed. Intraperitoneal injection of CSH or intracerebroventricular injection of ASS could delay initial time of convulsion (ITC),prolong survival time (ST) and reduce severity of convulsion (SOC). These results suggest that SS might play a role in oxygen-induced convulsion and be one of the endogenous agents which caused oxygeninduced convulsion.展开更多
This study sought to elucidate the correlation between cyclinD1 expression and the emergence of neovascularization in oxygen-induced retinopathy （OIR）. OIR was induced in Sprague-Dawley 7-day-old neonatal rats expos...This study sought to elucidate the correlation between cyclinD1 expression and the emergence of neovascularization in oxygen-induced retinopathy （OIR）. OIR was induced in Sprague-Dawley 7-day-old neonatal rats exposed to hyperoxia （75% O2） for 5 days, and then returned to room air. Adenosine diphosphatase staining showed that the neovascularization started to emerge at rat age of day 14, and reached a peak at day 17, then gradually decreased and resolved by day 26. Immunohistochemical and immunofluorescence staining revealed that cyclinD1 protein expression was seen in the OIR rats at the age of day 12, then gradually increased and returned to normal levels by day 26. These experimental findings demonstrated that the temporal pattern of cyclinD1 protein expression is consistent with the emergence of retinal neovascularization.展开更多
AIM: To explore the role of unc5b in retinal neovascularization in murine oxygen-induced retinopathy (OIR). METHODS: On postnatal 7 (P7), C57BL/6J mice were exposed to 75% +/- 2% oxygen for 5 days. On postnatal 12 (P1...AIM: To explore the role of unc5b in retinal neovascularization in murine oxygen-induced retinopathy (OIR). METHODS: On postnatal 7 (P7), C57BL/6J mice were exposed to 75% +/- 2% oxygen for 5 days. On postnatal 12 (P12), the mice were brought back to the room air (21% oxygen) to induce retinal neovascularization. Western blot analysis was performed to examine the temporal expression of unc5b in murine retinas. Double staining for unc5b and isolectin B4 were employed to determine the location of unc5b in murine retinas. The effect of unc5b on retinal neovascularization was evaluated by intravitreal injection of unc5b-FC in mice with OIR. Retinal neovascularization was measured by counting neovascular cell nuclei above the internal limiting membrane and by angiography of flat-mounted retinas perfused with fluorescein dextran. o RESULTS: Compared to age-matched normal mice, the expression of unc5b was significantly increased in retinas of OIR mice on P17 and P21. Unc5b was apparently expressed in retinal vessels of OIR while being negative in normal retinal vessels. Retinal neovascularization in eyes injected with unc5b-FC was significantly reduced. CONCLUSION: Unc5b-FC can effectively inhibit retinal neovascularization induced by OIR. It may serve as a powerful and novel therapy for ischemia-induced retinal disease.展开更多
AIM： To investigate the signal transduction mechanism of matrix metalloproteinase-9 （MMP-9） mediatedvascular endothelial growth factor （VEGF） expression and retinal neovascularization （RNV） in oxygen-induced re...AIM： To investigate the signal transduction mechanism of matrix metalloproteinase-9 （MMP-9） mediatedvascular endothelial growth factor （VEGF） expression and retinal neovascularization （RNV） in oxygen-induced retinopathy （OIR） model. METHODS： C57BL/6J mice were divided into four groups： control group, OIR group, OIR control group （phosphatebuffered saline by intravitreal injection） and treated group [tissue inhibitor of matrix metalloproteinase-1 （TIMP-1） by intravitreal injection]. OIR model was established in C57BIJ6J mice exposed to 75% ＋2% oxygen for 5d. mRNA level and protein expression of MMP-9, TIMP-1 and VEGF were measured by real-time polymerase chain reaction and Western blotting, and located by immunohistochemistry. RESULTS： Levels of MMP-9 and VEGF in retina were significantly increased in animals with OIR and OIR control group. Levels of TIMP -1 in retina was significantly reduced in animals with OIR and OIR control group. Furthermore, a significant correlation was found between MMP-9 and VEGF. Intravitreal injection of TIMP- 1 significantly reduced MMP-9 and VEGF expression of the OIR mouse model （all P〈0.05）. CONCLUSION： These results demonstrate that MMP- 9-mediated up-regulation of VEGF promotes RNV in retinopathy of prematurity （ROP）. TIMP-1 may be a potential target for the prevention and treatment of ROP.展开更多
In mild or moderate retinopathy of prematurity(ROP), retinal vessels undergo obliteration, proliferation, and regression. Despite complete regression of vessel abnormalities, a variety of visual impairments have been ...In mild or moderate retinopathy of prematurity(ROP), retinal vessels undergo obliteration, proliferation, and regression. Despite complete regression of vessel abnormalities, a variety of visual impairments have been reported. Rodent oxygen-induced retinopathy(OIR) is widely used as a model to study ROP. However, the long-term changes of OIR model remain unclear. The aim of this study is to examine long term changes of retinal vessel and visual function in a rodent OIR model resembling human mild or moderate ROP. In this study, after subjecting the animals to 80% oxygen(O_2) for 5–7 d, the retinal vessel density at postnatal day 12(P12) was approximately 30% lower than that in the age-matched control, but this difference was not significant between the groups. Vessel abnormalities, such as vessel tortuosity, neovascular tufts, and the number of vessels protruding into the vitreous, peaked between P17 and P20. Despite the regression of many abnormalities, vessel density in the OIR group was 36% and 32% lower than that in the control animals at 6 weeks and 4 months, respectively. The visual acuity and contrast sensitivity were impaired in the OIR group when measured at 2, 3 and 4 months. Therefore, the rodent OIR model exhibited long-lasting reduction in retinal vessel density and visual impairments, similar to those observed in mild or moderate human ROP. This study suggests that the rodent OIR model can be used to explore possible interventions for mild and moderate human ROP.展开更多
AIM: To identify disease-related miRNAs in retinas of mice with oxygen-induced retinopathy(OIR), and to explore their potential roles in retinal pathological neovascularization. METHODS: The retinal miRNA expression ...AIM: To identify disease-related miRNAs in retinas of mice with oxygen-induced retinopathy(OIR), and to explore their potential roles in retinal pathological neovascularization. METHODS: The retinal miRNA expression profile in mice with OIR and room air controls at postnatal day 17(P17) were determined through miRNA microarray analysis. Several miRNAs were significantly up-and down-regulated in retinas of mice with OIR compared to controls by quantitative real-time reverse transcription-polymerase chain reaction(qRT-PCR). Two databases including Targetscan7.1 and MirdbV5 were used to predict target genes that associated with those significantly altered mi RNAs in retinas of mice with OIR. Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway analyses were also conducted to identify possible biological functions of the target genes. RESULTS: In comparison with room air controls, 3 and 8 miRNAs were significantly up-and down-regulated, respectively, in retinas of mice with OIR. The qRT-PCR data confirmed that mmu-miR-350-3 p and mmu-miR-202-3 p were significantly up-regulated, while mmu-miR-711 and mmu-miR-30 c-1-3 p were significantly down-regulated in mice with OIR compared to controls. GO analysis demonstrated that the identified target genes were related to functions such as cellular macromolecule metabolic process. KEGG pathway analysis showed a group of pathways, such as Wnt signaling pathway, transcriptionalmisregulation in cancer, Mucin type O-glycan biosynthesis, and mitogen-activated protein kinase(MAPK) signaling pathway might be involved in pathological process of retinal neovascularization. CONCLUSION: Our findings suggest that the differentially expressed miRNAs in retinas of mice with OIR might provide potential therapeutic targets for treating retinal neovascularization.展开更多
AIM: To investigate whether the complement system is involved in a murine model of oxygen-induced retinopathy(OIR).METHODS: Forty C57BL/6J newborn mice were divided randomly into OIR group and control group. OIR was i...AIM: To investigate whether the complement system is involved in a murine model of oxygen-induced retinopathy(OIR).METHODS: Forty C57BL/6J newborn mice were divided randomly into OIR group and control group. OIR was induced by exposing mice to 75% ±2% oxygen from postnatal 7d(P7) to P12 and then recovered in room air.For the control group, the litters were raised in room air.At the postnatal 17d(P17), gene expressions of the complement components of the classical pathway(CP),the mannose-binding lectin(MBL) pathway and the alternative pathway(AP) in the retina were determined by quantitative real-time polymerase chain reaction(RT-PCR). Retinal protein expressions of the key components in the CP were examined by Western blotting.· RESULTS: Whole mounted retina in the OIR mice showed area of central hypoperfusion in both superficial and deep layers and neovascular tufts in the periphery.The expressions of C1 qb and C4 b genes in the OIR retina were significantly higher than those of the controls. The expression of retinal complement factor B(CFB) gene in OIR mice was significantly lower than those of the controls. However, the expressions of C3 and complement factor H(CFH) genes were higher. The protein synthesis of the key components involved in the CP(C1q, C4 and C3) were also significantly higher in OIR mouse retina. Although MBL-associated serine protease 1(MASP1) and MASP2 were detected in both the OIR and the control groups, the expressions were weak and the difference between the two groups was not significant.CONCLUSION: Our data suggest that the complement system CP is activated during the pathogenesis of murine model of OIR.展开更多
Background:Clinical trials have revealed that the antivascular endothelial growth factor(VEGF)therapies are effective in retinopathy of prematurity(ROP).But the low level of VEGF was necessary as a survival signal in ...Background:Clinical trials have revealed that the antivascular endothelial growth factor(VEGF)therapies are effective in retinopathy of prematurity(ROP).But the low level of VEGF was necessary as a survival signal in healthy conditions,and endogenous placental growth factor(PIGF)is redundant for development.The purpose of this study was to elucidate the PIGF expression under hypoxia as well as the infl uence of anti-VEGF therapy on PIGF.Methods:CoCl2-induced hypoxic human umbilical vein endothelial cells(HUVECs)were used for an in vitro study,and oxygen-induced retinopathy(OIR)mice models were used for an in vivo study.The expression patterns of PIGF under hypoxic conditions and the infl uence of anti-VEGF therapy on PIGF were evaluated by quantitative reverse transcription-polymerase chain reaction(RTPCR).The retinal avascular areas and neovascularization(NV)areas of anti-VEGF,anti-PIGF and combination treatments were calculated.Retina PIGF concentration was evaluated by ELISA after treatment.The vasoactive effects of exogenous PIGF on HUVECs were investigated by proliferation and migration studies.Results:PIGF mRNA expression was reduced by hypoxia in OIR mice,in HUVECs under hypoxia and anti-VEGF treatment.However,PIGF expression was reversed by anti-VEGF therapy in the OIR model and in HUVECs under hypoxia.Exogenous PIGF significantly inhibited HUVECs proliferation and migration under normal conditions,but it stimulated cell proliferation and migration under hypoxia.Anti-PIGF treatment was effective for neovascular tufts in OIR mice(P<0.05).Conclusion:The finding that PIGF expression is iatrogenically up-regulated by anti-VEGF therapy provides a consideration to combine it with anti-PIGF therapy.展开更多
AIM: To observe the effect of erythropoietin receptor antibody (EpoRA) on oxygen-induced retinal neovascularization. METHODS: C57BL / 63 mice, newly born 7 days, were exposed in high oxygen for 5 days and then placed ...AIM: To observe the effect of erythropoietin receptor antibody (EpoRA) on oxygen-induced retinal neovascularization. METHODS: C57BL / 63 mice, newly born 7 days, were exposed in high oxygen for 5 days and then placed in normal air for another 5 days, thus the animal models of retinal neovascularization were made. Experimental animals were allocated into 3 groups: normal, experimental and therapeutic. The normal group was fed in the normal environment. Into the vitreous cavity of mice in the therapeutic group were injected 2 mu L of EpoRA for 5 successive days. And the experimental group was injected the same amount of normal saline. Mice were sacrificed 17 days after birth and their eyeballs were removed for detection of malonaldehyde (MDA) content in the retina and by HE staining endothelial cells were counted the breaking through internal limiting membrane. RESULTS: In the experimental group, MDA content in the retina was 25.11 +/- 3.46 mu mol/g, which was obviously less than those in the normal group (5.34 +/- 1.79 mu mol/g, P<0.01) and those in the therapeutic group (12.04 +/- 1.91 mu mol/g). Pathological sections showed the nuclear number of the endothelial cells breaking through internal limiting membrane was 0.7 +/- 0.2 in normal group, and 46.2 +/- 6.5 in high oxygen induced experimental group. In the therapeutic group injected with EpoRA, it was lowered to 24.0 +/- 5.0(P < 0.01). CONCLUSION: EpoRA can effectively inhibit oxygen-induced neovascularization in retina of mouse by reducing oxidative damage.展开更多
AIM: To study the inhibitory effect of intravitreal captopril on oxygen-induced retinopathy (OIR) in mice. METHODS: Eighty postnatal day (P)7 C57BL/63 mice were randomly divided into treated group and control group wi...AIM: To study the inhibitory effect of intravitreal captopril on oxygen-induced retinopathy (OIR) in mice. METHODS: Eighty postnatal day (P)7 C57BL/63 mice were randomly divided into treated group and control group with forty mice in each group. The mice were exposed to 75% 2% oxygen for 5 days (P7-P11) and then returned to room air for 5 days (P12-P17) to induce retinal neovascularization (RNV). Beginning on P12, the mice in treated group received daily intravitreal injections of captopril(3.0mL/kg), while those in control group received daily intravitreal injections of phosphate-buffered saline (PBS) (3.0mL/kg) through P17. After anesthetized at P17, one eye was chosen randomly as experimental eye and were enucleated. RNV was examined by Adenosine diphosphate-ase (ADPase) stained retina flat-mounts and was quantitated histologically by counting the neovascular endothelial cell nuclei anterior to inner limiting membrane (ILM). The expressions of matrix metalloproteinase-2 (MMP-2) and vascular endothelial growth factor (VEGF) were measured by immunohistochemical method. RESULTS: Comparing with control group, more regular distributions, better branch and reduced density of RNV were observed in eyes of treated group. The number of neovascular cell nuclei was less in treated group than that in control group ( t =6.135, P <0.01). Stain of MMP-2 and VEGF was weaker in treated group than that in control group. CONCLUSION: The results indicate that captopril can significantly inhibit RNV in OIR mice.展开更多
基金This study was supported by the Natural Science Foundation of Tianjin
文摘Background： The mechanisms of pathological retinal neovascularization （RNV） remain unknown. Several microRNAs were reported to be involved in the process of RNV. Oxygen-induced retinopathy （O1R） is a useful model to investigate RNV. Our present work explored the expression and the role of microRNA-128 （miR-218） in oxygen-induced RNV. Methods： OIR was used to establish RNV model. The expression level ofmiR-218 in the retina from OIR mice was assessed by quantitative real-time reverse transcriptase polymerase chain reaction. Fluorescein angiography was performed in retinae of OIR mice, and RNV was quantified by hematoxylin and eosin staining to evaluate the effect of pCDH-CMV-miR-218 intravitreal injection on RNV in OIR mice. Roundabout 1 （Robol） expression was detected by Western blotting in mouse retinal vascular endothelial cells expressing a high or low level of miR-218 and retinal tissues from OIR mice. Cell migration was evaluated by scratch wound assay. Results： In OIR mice, the expression level of miR-218 was significantly down-regulated （P = 0.006）. Retinal Robol expression was significantly increased at both mRNA and protein levels （P = 0.001, 0.008： respectively）, miR-218 intravitreal injection inhibited retinal angiogenesis in OIR mice, and the restoration of miR-218 in retina led to down-regulation of Robol. Conelusions： Our experiments showed that restoration ofmiR-218 inhibited retinal angiogenesis via targeting Robo 1. MiR-218 contributed to the inhibition of retinal angiogenesis and miR-218 might be a new therapeutic target for preventing RNV.
基金Supported by the National Natural Science Foundation of China(No.81570873)the Science and Technology Department of Shaanxi Province(No.2015JM8481)
文摘Retinopathy of prematurity（ROP） is a retinal vascular disorder frequently found in premature infants.Different therapeutic strategies have been developed to treat ROP.However,there are still many children with ROP suffering by severe limitations in vision or even blindness.Recently,ROP has been suggested to be caused by abnormal development of the retinal vasculature,but not simply resulted by retinal neovascularization which takes about 4 to 6 wk after birth in premature infants.Thus,instead of focusing on how to reduce retinal neovascularization,understanding the pathological changes and mechanisms that occur prior to retinal neovascularization is meaningful,which may lead to identify novel target（s） for the development of novel strategy to promote the healthy growth of retinal blood vessels rather than passively waiting for the appearance of retinal neovascularization and removing it by force.In this review,we discussed recent studies about,1） the pathogenesis prior to retinal neovascularization in oxygen-induced retinopathy（OIR;a ROP in animal model） and in premature infants with ROP;2） the preclinical and clinical research on preventive treatment of early OIR and ROP.We will not only highlight the importance of the mechanisms and signalling pathways in regulating early stage of ROP but also will provide guidance for actively exploring novel mechanisms and discovering novel treatments for early phase OIR and ROP prior to retinal neovascularization in the future.
文摘The Delta-like ligand 4/Notch signaling pathway was shown to participate in the process of retinal development and angiogenesis. However, the function of the Delta-like ligand 4/Notch signaling pathway in retinopathy of prematurity requires further study. Retinopathy of prematurity was induced in 5-day-old Sprague-Dawley rats exposed to hyperoxia for 7 days, and then returned to room air. Reverse transcription-PCR and western blot revealed that Delta-like ligand 4 levels decreased at postnatal day 12 and increased at postnatal day 17 in retinopathy of prematurity rats. Flat-mounted adenosine diphosphatase stained retina and hematoxylin-eosin stained retinal tissue slices showed that the clock hour scores and the nuclei counts in retinopathy of prematurity rats were significantly different compared to normal control rats. After retinopathy of prematurity rats were intravitreally injected with Delta-like ligand 4 monoclonal antibody to inhibit the Delta-like ligand 4/Notch signaling pathway, there was a significant increase in the severity of retinal neovascularization （clock hours） in the intravitreally injected eyes. The nuclei count was highly correlated with the clock hour score. These results suggest that Delta-like ligand 4/Notch signaling plays an essential role in the process of physiological and pathological angiogenesis in the retina.
基金the National Natural Science Foundation of China,No.81600747(to YD)a grant from Liaoning Department of Education,No.QNZR2020010(to YD)a grant from 345 Talent Project of Shengjing Hospital(to YD).
文摘The mouse model of oxygen induced retinopathy is suitable for the study of various retinal neovascularization diseases,including retinopathy of prematurity.The maternally expressed gene 3(MEG3)has been demonstrated to have an inhibitory effect on diabetic retinopathy.In this study,we investigated the role of MEG3 overexpression in oxygen-induced retinopathy in mice.The results showed that MEG3 overexpression effectively inhibited the production of retinal neovascularization in oxygen-induced retinopathy mice.It acts by down-regulating the expression of phosphoinositide 3-kinase,serine/threonine kinase,and vascular endothelial growth factor and pro-inflammatory factors.MEG3 overexpression lentivirus has a future as a new method for the clinical treatment of retinopathy of prematurity.The animal experiments were approved by the Animal Ethics Committee of Shengjing Hospital of China Medical University,China(approval No.2016PS074K)on February 25,2016.
基金supported by the National Natural Science Foundation of China,No. 81570866 (to XLC)。
文摘Inhibiting retinal neovascularization is the optimal strategy for the treatment of retina-related diseases, but there is currently no effective treatment for retinal neovascularization. P-element-induced wimpy testis(PIWI)-interacting RNA(piRNA) is a type of small non-coding RNA implicated in a variety of diseases. In this study, we found that the expression of piR-1245 and the interacting protein PIWIL2 were remarkably increased in human retinal endothelial cells cultured in a hypoxic environment, and cell apoptosis, migration, tube formation and proliferation were remarkably enhanced in these cells. Knocking down piR-1245 inhibited the above phenomena. After intervention by a p-JAK2 activator, piR-1245 decreased the expression of hypoxia inducible factor-1α and vascular endothelial growth factor through the JAK2/STAT3 pathway. For in vivo analysis, 7-day-old newborn mice were raised in 75 ± 2% hyperoxia for 5 days and then piR-1245 in the retina was knocked down. In these mice, the number of newly formed vessels in the retina was decreased, the expressions of inflammationrelated proteins were reduced, the number of apoptotic cells in the retina was decreased, the JAK2/STAT3 pathway was inhibited, and the expressions of hypoxia inducible factor-1α and vascular endothelial growth factor were decreased. Injection of the JAK2 inhibitor JAK2/TYK2-IN-1 into the vitreous cavity inhibited retinal neovascularization in mice and reduced expression of hypoxia inducible factor-1α and vascular endothelial growth factor. These findings suggest that piR-1245 activates the JAK2/STAT3 pathway, regulates the expression of hypoxia inducible factor-1α and vascular endothelial growth factor, and promotes retinal neovascularization. Therefore, piR-1245 may be a new therapeutic target for retinal neovascularization.
文摘The content of somatostatin(SS) in hippocampus,striatum and frontal cortex tissues of rats exposed to 600 kpa hyperbaric oxygen was determined by means of radioimmunoassay. Initial time of convulsion, severity of convulsion and survival time of rats with convulsion exposed to 700 kPa hyperbaric oxygen after intraperitoneal injection of cysteamine (CSH) or intracerebroventricular injection of anti-somatostatin serum(ASS) were also observed. The results showed that the content of SS in hippocampus and striatum tissues increased remarkably when rats were at near-convulsion ; by the time the rats developed convulsion,it had a significant increase in all brain areas observed. Intraperitoneal injection of CSH or intracerebroventricular injection of ASS could delay initial time of convulsion (ITC),prolong survival time (ST) and reduce severity of convulsion (SOC). These results suggest that SS might play a role in oxygen-induced convulsion and be one of the endogenous agents which caused oxygeninduced convulsion.
文摘This study sought to elucidate the correlation between cyclinD1 expression and the emergence of neovascularization in oxygen-induced retinopathy （OIR）. OIR was induced in Sprague-Dawley 7-day-old neonatal rats exposed to hyperoxia （75% O2） for 5 days, and then returned to room air. Adenosine diphosphatase staining showed that the neovascularization started to emerge at rat age of day 14, and reached a peak at day 17, then gradually decreased and resolved by day 26. Immunohistochemical and immunofluorescence staining revealed that cyclinD1 protein expression was seen in the OIR rats at the age of day 12, then gradually increased and returned to normal levels by day 26. These experimental findings demonstrated that the temporal pattern of cyclinD1 protein expression is consistent with the emergence of retinal neovascularization.
基金National Natural Science Foundation of China (No.30872822)
文摘AIM: To explore the role of unc5b in retinal neovascularization in murine oxygen-induced retinopathy (OIR). METHODS: On postnatal 7 (P7), C57BL/6J mice were exposed to 75% +/- 2% oxygen for 5 days. On postnatal 12 (P12), the mice were brought back to the room air (21% oxygen) to induce retinal neovascularization. Western blot analysis was performed to examine the temporal expression of unc5b in murine retinas. Double staining for unc5b and isolectin B4 were employed to determine the location of unc5b in murine retinas. The effect of unc5b on retinal neovascularization was evaluated by intravitreal injection of unc5b-FC in mice with OIR. Retinal neovascularization was measured by counting neovascular cell nuclei above the internal limiting membrane and by angiography of flat-mounted retinas perfused with fluorescein dextran. o RESULTS: Compared to age-matched normal mice, the expression of unc5b was significantly increased in retinas of OIR mice on P17 and P21. Unc5b was apparently expressed in retinal vessels of OIR while being negative in normal retinal vessels. Retinal neovascularization in eyes injected with unc5b-FC was significantly reduced. CONCLUSION: Unc5b-FC can effectively inhibit retinal neovascularization induced by OIR. It may serve as a powerful and novel therapy for ischemia-induced retinal disease.
基金Supported by National Natural Science Foundation of China (No.81371045)Science and Technology Project of Shenyang City, China (No.F13-220-9-37)
文摘AIM： To investigate the signal transduction mechanism of matrix metalloproteinase-9 （MMP-9） mediatedvascular endothelial growth factor （VEGF） expression and retinal neovascularization （RNV） in oxygen-induced retinopathy （OIR） model. METHODS： C57BL/6J mice were divided into four groups： control group, OIR group, OIR control group （phosphatebuffered saline by intravitreal injection） and treated group [tissue inhibitor of matrix metalloproteinase-1 （TIMP-1） by intravitreal injection]. OIR model was established in C57BIJ6J mice exposed to 75% ＋2% oxygen for 5d. mRNA level and protein expression of MMP-9, TIMP-1 and VEGF were measured by real-time polymerase chain reaction and Western blotting, and located by immunohistochemistry. RESULTS： Levels of MMP-9 and VEGF in retina were significantly increased in animals with OIR and OIR control group. Levels of TIMP -1 in retina was significantly reduced in animals with OIR and OIR control group. Furthermore, a significant correlation was found between MMP-9 and VEGF. Intravitreal injection of TIMP- 1 significantly reduced MMP-9 and VEGF expression of the OIR mouse model （all P〈0.05）. CONCLUSION： These results demonstrate that MMP- 9-mediated up-regulation of VEGF promotes RNV in retinopathy of prematurity （ROP）. TIMP-1 may be a potential target for the prevention and treatment of ROP.
基金supported by the National Natural Science Foundation of China Key Project(31030036)to SHa National Natural Science Foundation of China Project(81570863)Science and Technology Supporting Program by Department of Science and Technology,Sichuan Province(2016SZ0024)to FL
文摘In mild or moderate retinopathy of prematurity(ROP), retinal vessels undergo obliteration, proliferation, and regression. Despite complete regression of vessel abnormalities, a variety of visual impairments have been reported. Rodent oxygen-induced retinopathy(OIR) is widely used as a model to study ROP. However, the long-term changes of OIR model remain unclear. The aim of this study is to examine long term changes of retinal vessel and visual function in a rodent OIR model resembling human mild or moderate ROP. In this study, after subjecting the animals to 80% oxygen(O_2) for 5–7 d, the retinal vessel density at postnatal day 12(P12) was approximately 30% lower than that in the age-matched control, but this difference was not significant between the groups. Vessel abnormalities, such as vessel tortuosity, neovascular tufts, and the number of vessels protruding into the vitreous, peaked between P17 and P20. Despite the regression of many abnormalities, vessel density in the OIR group was 36% and 32% lower than that in the control animals at 6 weeks and 4 months, respectively. The visual acuity and contrast sensitivity were impaired in the OIR group when measured at 2, 3 and 4 months. Therefore, the rodent OIR model exhibited long-lasting reduction in retinal vessel density and visual impairments, similar to those observed in mild or moderate human ROP. This study suggests that the rodent OIR model can be used to explore possible interventions for mild and moderate human ROP.
基金Supported by National Natural Science Foundation of China (No.81700837 No.81800855)+2 种基金Natural Science Foundation of Hunan Province (No.2017JJ3452 No.2018JJ3765)Department of Science and Technology, Hunan (No.2015TP2007)
文摘AIM: To identify disease-related miRNAs in retinas of mice with oxygen-induced retinopathy(OIR), and to explore their potential roles in retinal pathological neovascularization. METHODS: The retinal miRNA expression profile in mice with OIR and room air controls at postnatal day 17(P17) were determined through miRNA microarray analysis. Several miRNAs were significantly up-and down-regulated in retinas of mice with OIR compared to controls by quantitative real-time reverse transcription-polymerase chain reaction(qRT-PCR). Two databases including Targetscan7.1 and MirdbV5 were used to predict target genes that associated with those significantly altered mi RNAs in retinas of mice with OIR. Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway analyses were also conducted to identify possible biological functions of the target genes. RESULTS: In comparison with room air controls, 3 and 8 miRNAs were significantly up-and down-regulated, respectively, in retinas of mice with OIR. The qRT-PCR data confirmed that mmu-miR-350-3 p and mmu-miR-202-3 p were significantly up-regulated, while mmu-miR-711 and mmu-miR-30 c-1-3 p were significantly down-regulated in mice with OIR compared to controls. GO analysis demonstrated that the identified target genes were related to functions such as cellular macromolecule metabolic process. KEGG pathway analysis showed a group of pathways, such as Wnt signaling pathway, transcriptionalmisregulation in cancer, Mucin type O-glycan biosynthesis, and mitogen-activated protein kinase(MAPK) signaling pathway might be involved in pathological process of retinal neovascularization. CONCLUSION: Our findings suggest that the differentially expressed miRNAs in retinas of mice with OIR might provide potential therapeutic targets for treating retinal neovascularization.
基金Supported partially by National Natural Science Foundation of China(No.81271033,81470621)
文摘AIM: To investigate whether the complement system is involved in a murine model of oxygen-induced retinopathy(OIR).METHODS: Forty C57BL/6J newborn mice were divided randomly into OIR group and control group. OIR was induced by exposing mice to 75% ±2% oxygen from postnatal 7d(P7) to P12 and then recovered in room air.For the control group, the litters were raised in room air.At the postnatal 17d(P17), gene expressions of the complement components of the classical pathway(CP),the mannose-binding lectin(MBL) pathway and the alternative pathway(AP) in the retina were determined by quantitative real-time polymerase chain reaction(RT-PCR). Retinal protein expressions of the key components in the CP were examined by Western blotting.· RESULTS: Whole mounted retina in the OIR mice showed area of central hypoperfusion in both superficial and deep layers and neovascular tufts in the periphery.The expressions of C1 qb and C4 b genes in the OIR retina were significantly higher than those of the controls. The expression of retinal complement factor B(CFB) gene in OIR mice was significantly lower than those of the controls. However, the expressions of C3 and complement factor H(CFH) genes were higher. The protein synthesis of the key components involved in the CP(C1q, C4 and C3) were also significantly higher in OIR mouse retina. Although MBL-associated serine protease 1(MASP1) and MASP2 were detected in both the OIR and the control groups, the expressions were weak and the difference between the two groups was not significant.CONCLUSION: Our data suggest that the complement system CP is activated during the pathogenesis of murine model of OIR.
基金supported by the National Basic Research Program of China(973 Program,2011CB510200).
文摘Background:Clinical trials have revealed that the antivascular endothelial growth factor(VEGF)therapies are effective in retinopathy of prematurity(ROP).But the low level of VEGF was necessary as a survival signal in healthy conditions,and endogenous placental growth factor(PIGF)is redundant for development.The purpose of this study was to elucidate the PIGF expression under hypoxia as well as the infl uence of anti-VEGF therapy on PIGF.Methods:CoCl2-induced hypoxic human umbilical vein endothelial cells(HUVECs)were used for an in vitro study,and oxygen-induced retinopathy(OIR)mice models were used for an in vivo study.The expression patterns of PIGF under hypoxic conditions and the infl uence of anti-VEGF therapy on PIGF were evaluated by quantitative reverse transcription-polymerase chain reaction(RTPCR).The retinal avascular areas and neovascularization(NV)areas of anti-VEGF,anti-PIGF and combination treatments were calculated.Retina PIGF concentration was evaluated by ELISA after treatment.The vasoactive effects of exogenous PIGF on HUVECs were investigated by proliferation and migration studies.Results:PIGF mRNA expression was reduced by hypoxia in OIR mice,in HUVECs under hypoxia and anti-VEGF treatment.However,PIGF expression was reversed by anti-VEGF therapy in the OIR model and in HUVECs under hypoxia.Exogenous PIGF significantly inhibited HUVECs proliferation and migration under normal conditions,but it stimulated cell proliferation and migration under hypoxia.Anti-PIGF treatment was effective for neovascular tufts in OIR mice(P<0.05).Conclusion:The finding that PIGF expression is iatrogenically up-regulated by anti-VEGF therapy provides a consideration to combine it with anti-PIGF therapy.
基金National Science Foundation for Youths of China (No.30801268,30800375)the Foundation for Disaster Medicine(2008),Second Military Medical University, China
文摘AIM: To observe the effect of erythropoietin receptor antibody (EpoRA) on oxygen-induced retinal neovascularization. METHODS: C57BL / 63 mice, newly born 7 days, were exposed in high oxygen for 5 days and then placed in normal air for another 5 days, thus the animal models of retinal neovascularization were made. Experimental animals were allocated into 3 groups: normal, experimental and therapeutic. The normal group was fed in the normal environment. Into the vitreous cavity of mice in the therapeutic group were injected 2 mu L of EpoRA for 5 successive days. And the experimental group was injected the same amount of normal saline. Mice were sacrificed 17 days after birth and their eyeballs were removed for detection of malonaldehyde (MDA) content in the retina and by HE staining endothelial cells were counted the breaking through internal limiting membrane. RESULTS: In the experimental group, MDA content in the retina was 25.11 +/- 3.46 mu mol/g, which was obviously less than those in the normal group (5.34 +/- 1.79 mu mol/g, P<0.01) and those in the therapeutic group (12.04 +/- 1.91 mu mol/g). Pathological sections showed the nuclear number of the endothelial cells breaking through internal limiting membrane was 0.7 +/- 0.2 in normal group, and 46.2 +/- 6.5 in high oxygen induced experimental group. In the therapeutic group injected with EpoRA, it was lowered to 24.0 +/- 5.0(P < 0.01). CONCLUSION: EpoRA can effectively inhibit oxygen-induced neovascularization in retina of mouse by reducing oxidative damage.
基金Natural Science Foundation of Liaoning Province, China (No.20052089)Science and Technology Project of Liaoning Province, China (No.2010225034)
文摘AIM: To study the inhibitory effect of intravitreal captopril on oxygen-induced retinopathy (OIR) in mice. METHODS: Eighty postnatal day (P)7 C57BL/63 mice were randomly divided into treated group and control group with forty mice in each group. The mice were exposed to 75% 2% oxygen for 5 days (P7-P11) and then returned to room air for 5 days (P12-P17) to induce retinal neovascularization (RNV). Beginning on P12, the mice in treated group received daily intravitreal injections of captopril(3.0mL/kg), while those in control group received daily intravitreal injections of phosphate-buffered saline (PBS) (3.0mL/kg) through P17. After anesthetized at P17, one eye was chosen randomly as experimental eye and were enucleated. RNV was examined by Adenosine diphosphate-ase (ADPase) stained retina flat-mounts and was quantitated histologically by counting the neovascular endothelial cell nuclei anterior to inner limiting membrane (ILM). The expressions of matrix metalloproteinase-2 (MMP-2) and vascular endothelial growth factor (VEGF) were measured by immunohistochemical method. RESULTS: Comparing with control group, more regular distributions, better branch and reduced density of RNV were observed in eyes of treated group. The number of neovascular cell nuclei was less in treated group than that in control group ( t =6.135, P <0.01). Stain of MMP-2 and VEGF was weaker in treated group than that in control group. CONCLUSION: The results indicate that captopril can significantly inhibit RNV in OIR mice.
