Retinopathy of prematurity (ROP) is a vision-threatening disorder that leads to pathological growth of the retinal vasculature due to hypoxia. Here, we investigated the potential effects of alamandine, a novel heptapeptide in the renin-angiotensin system (RAS), on hypoxia-induced retinal neovascularization and its underlying mechanisms. In vivo, the C57BL/6J mice with oxygen-induced retinopathy (OIR) were injected intravitreally with alamandine (1.0 μmol/kg per eye). In vitro, human retinal microvascular endothelial cells (HRMECs) were utilized to investigate the effects of alamandine (10 μg/mL) on proliferation, apoptosis, migration, and tubular formation under vascular endothelial growth factor (VEGF) stimulation. Single-cell RNA sequencing (scRNA-seq) matrix data from the Gene Expression Omnibus (GEO) database and RAS-related genes from the Molecular Signatures Database (MSigDB) were sourced for subsequent analyses. By integrating scRNA-seq data across multiple species, we identified that RAS-associated endothelial cell populations were highly related to retinal neovascularization. The liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed a significant decrease in alamandine levels in both the serum and retina of OIR mice compared to those in the control group. Next, alamandine ameliorated hypoxia-induced retinal pathological neovascularization and physiologic revascularization in OIR mice. In vitro, alamandine effectively mitigated VEGF-induced proliferation, scratch wound healing, and tube formation of HRMECs primarily by inhibiting the hypoxia-inducible factor-1α (HIF-1α)/VEGF pathway. Further, coincubation with D-Pro⁷ (Mas-related G protein-coupled receptor D (MrgD) antagonist) hindered the beneficial impacts of alamandine on hypoxia-induced pathological angiogenesis both in vivo and in vitro. Our findings suggested that alamandine could mitigate retinal neovascularization by targeting the MrgD-mediated HIF-1α/VEGF pathway, providing a potential therapeutic agent for OIR prevention and treatment.
Animals ; Retinal Neovascularization/prevention & control* ; Mice, Inbred C57BL ; Vascular Endothelial Growth Factor A/metabolism* ; Humans ; Mice ; Hypoxia-Inducible Factor 1, alpha Subunit/metabolism* ; Oligopeptides/therapeutic use* ; Signal Transduction/drug effects* ; Cell Proliferation/drug effects* ; Endothelial Cells/drug effects* ; Retinopathy of Prematurity/drug therapy* ; Apoptosis/drug effects* ; Cell Movement/drug effects* ; Renin-Angiotensin System/drug effects* ; Cells, Cultured

OBJECTIVETo explore the effects of rat bone mesenchymal stem cell (BMSC) transplantation on retinal neovascularization, and to observe the changes of hypoxia-inducible factor-1 alpha (HIF-1α) and vascular endothelial growth factors (VEGF) in rats with oxygen-induced retinopathy (OIR).

METHODSSeventy-two seven-day-old Sprague-Dawley rats were randomly divided into three groups: normal control (CON), model (OIR) and BMSC transplantation. In the BMSC transplantation group, BMSCs were transplanted 5 days after oxygen conditioning. The phosphate buffered saline of the same volume was injected in the CON and OIR groups. The OIR model was prerpared according to the classic hyperoxygen method. At seven days after transplantation, retinal neovascularization was examined by retinal flat-mount staining and hematoxylin eosin (HE) staining. The expression of HIF-1α and VEGF proteins was examined by immunohistochemistry staining and Western blot analysis.

RESULTSThe retinal flat-mount staining results showed that the vessels were well organized in the CON group, but the vessels were irregularly organized, and lots of nonperfusion areas were observed in the OIR group. The large vessels were a bit circuitous, the retinal vessels were relatively organized, and less nonperfusion areas were noted in the BMSC transplantation group. The HE staining results showed that many neovessels and preretinal neovascular (pre-RNC) cells were observed on the internal limiting membrane in the OIR group. There were less pre-RNC cells in the BMSC transplantation group compared with the OIR group (P<0.01). The immunohistochemistry analysis showed that more HIF-1αand VEGFcells were observed in the OIR group compared with the CON group, and less HIF-1αand VEGFcells were observed in the BMSC transplantation group compared with OIR group (P<0.05). The Western blot analysis showed the expression of HIF-1α and VEGF proteins in the OIR group was significantly higher than that in the CON group. The expression of HIF-1α and VEGF proteins in the BMSC transplantation group was lower than that in the OIR group (P<0.01).

CONCLUSIONSBMSC transplantation therapy could alleviate retinal neovascularization in OIR rats, and its mechanisms might be associated with the inhibition of the expression of HIF-1α and VEGF proteins.

Animals ; Animals, Newborn ; Female ; Hypoxia-Inducible Factor 1, alpha Subunit ; analysis ; Male ; Mesenchymal Stem Cell Transplantation ; Rats ; Rats, Sprague-Dawley ; Retina ; chemistry ; Retinal Neovascularization ; prevention & control ; Retinopathy of Prematurity ; metabolism ; therapy ; Vascular Endothelial Growth Factor A ; analysis

BACKGROUNDRetinopathy of prematurity (ROP) has become one of the leading causes of visual loss in children. Vascular endothelial growth factor A (VEGF-A) is the principal stimulator of angiogenesis. VEGF was differentially spliced from exon 8 to exons 8a and 8b to form two families: the pro-angiogenic VEGFxxx family and the anti-angiogenic VEGFxxxb family. Previous research has shown variable effeteness of bevacizumab in inhibiting retinal neovascularization in ROP. This study aimed to investigate whether the effectiveness of this inhibition depends on the relative ratio of the two VEGF isoforms.

METHODSIntravitreal bevacizumab injection (IVB) was performed in the oxygen-induced-retinopathy (OIR) mice on postnatal day 12 (P12) (intravitreal phosphate buffered saline (PBS) injection as control). The Evans blue perfused retina were used to test the retinal neovascularization-leakage (NVL) area and non-perfusion (NP) area.

RESULTSThe retinal NVL and NP area in the IVB group were significantly smaller than the intravitreal PBS injection group (IVP group). On P17, the protein level of total VEGF isoforms was significantly inhibited compared to IVP group (P < 0.05) while VEGF(165)b isoform was slight reduced (P > 0.05). The switch from pro-angiogenic isoforms to anti-angiogenic isoforms after IVB could be found. The relative protein expression of VEGF(165)b isoform was significantly higher in IVB group than in IVP group (P < 0.05) on P17 which was correlated with the reduced ischemia-induced angiogenesis in OIR mice after IVB.

CONCLUSIONSThe anti-angiogenic effectiveness might depend on the relative high expression of VEGF(165)b after intravitreal bevacizumab injection. Anti-angiogenic therapy is a more effective therapy for ROP.

Angiogenesis Inhibitors ; administration & dosage ; Animals ; Animals, Newborn ; Antibodies, Monoclonal, Humanized ; administration & dosage ; Bevacizumab ; Disease Models, Animal ; Intravitreal Injections ; Mice ; Mice, Inbred C57BL ; Protein Isoforms ; analysis ; Retinal Neovascularization ; prevention & control ; Retinopathy of Prematurity ; drug therapy ; Vascular Endothelial Growth Factor A ; analysis

OBJECTIVETo study the inhibition effect of HIF-1α specific siRNA expression vector pSUPERH1-siHIF-1α on retinal neovascularization in a mouse model of retinopathy of prematurity (ROP).

METHODSThe mouse model of ROP was prepared by the method Smith described. Forty-eight ROP mice were randomly divided into two groups: an experimental group that was intravitreously injected with pSUPERH1-siHIF-1α and a control group that was injected with pSUPER retro vector. The levels of HIF-1α and vascular endothelia growth factor (VEGF) in the retina were examined by Western blot. The retinal neovascularization was evaluated by angiography using FITC Dextran and quantitated histologically.

RESULTSThe levels of HIF-1α and VEGF in the retina in the experimental group were reduced 90% and 65% respectively compared with those in the control group. Meanwhile, the number of retinal neovascular endothelial nucleus outbreaking the inner limit membrane in the experimental group was significantly reduced compared with that in the control group.

CONCLUSIONSThe development of retinal neovascularization of ROP can be markedly inhibited by RNA interference targeting HIF-1α.

Animals ; Disease Models, Animal ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; analysis ; antagonists & inhibitors ; genetics ; Infant, Newborn ; Mice ; Mice, Inbred C57BL ; RNA, Small Interfering ; genetics ; Retinal Neovascularization ; prevention & control ; Retinopathy of Prematurity ; therapy ; Vascular Endothelial Growth Factor A ; analysis

PURPOSE: Retinopathy of prematurity (ROP) is one of the leading causes of blindness, with retinal detachment occurring due to oxygen toxicity in preterm infants. Recently, advances in neonatal care have led to improved survival rates for preterm infants, and ROP has increased in incidence. In the present study, we aimed to determine whether or not resveratrol exhibits protective effects in an animal model of ROP and in primary retinal cell cultures of neonatal rat via nitric oxide (NO)-modulating actions using western blotting and real-time PCR with inducible nitric oxide synthase (iNOS), endothelial NOS (eNOS) and neuronal NOS (nNOS) antibodies and mRNAs. METHODS: In an in vivo oxygen-induced retinopathy (OIR) model, cyclic hyperoxia was induced with 80% O2 for one day and 21% O2 for one day from P1 to P14 in newborn Sprague-Dawley (SD) rats. Resveratrol was injected intravitreally for seven days and rats were sacrificed at P21. In vitro OIR primary retinal cell culture was performed using P0-2 SD rats. Hyperoxia injuries were induced through 100% O2 exposure for six hours. Western blotting and real-time PCR using iNOS, eNOS, nNOS antibodies and primers were performed in the rat model of ROP and the dispersed retinal cell culture. RESULTS: In both in vivo and in vitro OIR, the expression of iNOS antibody and mRNA was increased and of eNOS and nNOS were reduced in the resveratrol-treated group. CONCLUSIONS: In conclusion, resveratrol appeared to exert retinal protective effects via modulation of NO-mediated mechanism in in vivo and in vitro OIR models.
Analysis of Variance ; Animals ; Animals, Newborn ; Blotting, Western ; Disease Models, Animal ; Electrophoresis, Polyacrylamide Gel ; Humans ; Infant, Newborn ; Nitric Oxide Synthase/*metabolism ; Oxygen/toxicity ; RNA/metabolism ; RNA, Messenger/metabolism ; Rats ; Rats, Sprague-Dawley ; Retina/drug effects/pathology ; Retinopathy of Prematurity/*metabolism/pathology/*prevention & control ; Reverse Transcriptase Polymerase Chain Reaction ; Stilbenes/*pharmacology

OBJECTIVETo explore the etiology, incidence and methods to prevent and treat severe retinopathy of prematurity (ROP), which is rapidly becoming a threat to the vision of babies in areas of the world where increasing numbers of premature babies are surviving.

DATA SOURCESThe data used in this review were mainly from Medline and PubMed published in English. The search term was "retinopathy of prematurity and premature birth".

STUDY SELECTIONWe discuss the historical perspectives, prevalence and incidence, classification and treatment methods of ROP in premature babies.

RESULTSPeripheral retinal ablation for eyes with severe ROP can help prevent progression to blindness and several large clinical trials have shown the effectiveness of this treatment in high risk eyes. As a greater proportion of VLBW and ELBW babies survive, the population of babies at risk increases. In various regions of the world, different identification criteria are used to determine which babies are at risk of blindness in order to provide timely diagnostic examinations and treatment as needed. Methods for preventing ROP include better ante-natal and obstetric care leading to a reduction in the rate of prematurity, the use of ante-natal corticosteroids, and better neonatal care practices. Recent developments have indicated that management of oxygen supplementation is important for the prevention of severe ROP; however, there is not yet known what oxygen saturation target should be adopted. Sepsis increases severe ROP in very preterm infants. Genetic associations and a telemedicine approach may be explored to detect ROP. Treatment of anti-VEGF therapy are potentially useful in eyes with severe ROP, but long term effects are not yet known and such treatment should be used with great caution.

CONCLUSIONSROP is a potentially binding disease for premature babies which is becoming more prevalent with the development improving neonatal services in many countries in recent years. High priority should be placed on developing approaches to prevent ROP blindness by reducing preterm birth, improving care of premature babies in neonatal care units, and providing adequate ophthalmological services in those regions.

Humans ; Incidence ; Infant, Newborn ; Insulin-Like Growth Factor I ; physiology ; Prevalence ; Retinopathy of Prematurity ; classification ; epidemiology ; prevention & control ; Sepsis ; complications ; Vascular Endothelial Growth Factor A ; antagonists & inhibitors

OBJECTIVETo investigate the factors involved in the development of retinopathy of prematurity (ROP), and to provide the preliminary data for the evaluation of current criteria for ROP screening.

METHODPremature infants with birth body weight (BBW) ≤ 2000 g or gestational age (GA) ≤ 34 weeks in the two hospitals in Zhejiang between March 2005 and November 2008 were recruited and examined by indirect ophthalmoscopy. The records were analyzed.

RESULTOne thousand two hundred and twenty-five premature infants were included. Of them, 713 were male and 512 female. There were 179 twins and 21 triplets in the premature infants. The incidence of ROP was 10.8% (132 in 1225 patients). There were 12 cases (0.98%) to the point of pre threshold ROP. 4 cases (0.3%) developed threshold ROP. Only one case developed pre threshold ROP of low risk among 65 cases without history of oxygen treatment (1.5%). The percentage has significant difference compared to that of cases with history of oxygen (χ(2) = 5.115, P < 0.01).Between ROP and Non-ROP groups, there was significant difference in BBW(F = 26.39, P < 0.001), gestational age (F = 19.73, P < 0.001), but there was no significant difference in sex (χ(2) = 0.279, P > 0.05) or twins and triplets (χ(2) = 3.449, P > 0.05). The incidence of ROP among premature infants with BBW ≤ 1000 g was more than three times of that with BBW > 1000 g, and the incidence of ROP among premature infants with GA ≤ 28 weeks was about 2.5 times of that with GA > 28 weeks. Logistic regression analysis indicated that less BBW or shorter GA or undulation of blood oxygen concentration was a significant risk factor involved in the development of ROP (r = 0.57, P < 0.05). All ROP patients were cured.

CONCLUSIONLess BBW, shorter GA and undulation of blood oxygen concentration are the important risk factors for the development of ROP. Premature infants with BBW ≤ 1000 g or GA ≤ 28 weeks, who had oxygen history, should be given very special attention in the ROP screening. The current criteria for ROP screening should be narrowed. In general, the ROP screening has lowered the incidence of blindness among children by investigating and treating ROP timely.

China ; epidemiology ; Female ; Humans ; Incidence ; Infant, Newborn ; Infant, Premature ; Male ; Neonatal Screening ; Retinopathy of Prematurity ; diagnosis ; epidemiology ; prevention & control ; Risk Factors

OBJECTIVETo analyze the risk factors of retinopathy of prematurity (ROP) and provide evidence for the rational establishment of screening standard.

METHODSThe clinical data of 1675 preterm infants at gestational age < or = 36 weeks or birth weight < or = 2500 g who were admitted to the neonatal intensive care unit and had been screened in our hospital from July 2006 to May 2008 were analyzed retrospectively by univariate analysis and Logistic regression analysis. Gender, birth count, gestational age, birth weight, oxygen therapy, and mother's conditions were recorded.

RESULTSROP was detected in 195 (11.6%) of 1675 infants, of whom 35 infants (2.1%) had type 1 or threshold ROP. The lower the birth weight, the smaller the gestational age and the longer the time of oxygen therapy were, the higher the incidence of ROP was. For the infants whose birth weight was < or = 1200 g, 1201 - 1500 g, 1501 - 2000 g, 2001 - 2500 g, the incidence of ROP was 73.2%, 30.4%, 8.0%, and 1.1%; for those at gestational age < or = 30 weeks, 30(+1)-32 weeks, 32(+1)-34 weeks, 34(+1)-36 weeks, the incidence of ROP was 67.6%, 16.9%, 3.9%, and 1.0%; for the infants underwent oxygen therapy for 0 d, -3 d, -5 d, -8 d, > 8 d, the incidence of ROP was 1.5%, 3.3%, 9.6%, 23.2% and 38.8%;in the infants who inhaled oxygen at concentrations of 0.40, -0.60, -0.80 and > 0.80, the incidence of ROP was 11.8%, 18.1%, 26.8%, and 52.6%, respectively. Logistic regression analysis indicated that low birth weight, small gestational age, asphyxia, apnea, oxygen therapy were the high risk factors of ROP (the odds ratio was 0.957, 1.052, 1.186, 5.314, and 1.881).

CONCLUSIONSLow birth weight, small gestational age, asphyxia, apnea, and oxygen therapy were the high risk factors of ROP. It is recommended that all preterm infants with high risk factors should be screened.

Birth Weight ; Female ; Gestational Age ; Humans ; Infant, Newborn ; Infant, Small for Gestational Age ; Male ; Neonatal Screening ; Retinopathy of Prematurity ; epidemiology ; prevention & control ; Retrospective Studies ; Risk Factors

The effect of the trace elements on retinopathy of prematurity (ROP) were studied. Thirty preterm infants who had potential high risk factors of ROP were selected as observation group and 18 normal infants as control groups. By using atom spectrophotometer, the contents of serum trace elements (Mg, Cu, Zn, Mn, Se) were measured and analyzed statistically. The contents of serum Zn, Cu and Se in observation group were 0.75+/-0.22, 0.41+/-0.20 and (134.07+/-71.57)x10(-3) mg/L respectively, and 0.55+/-0.12, 0.65+/-0.194 and (202.92+/-44.71)x10(-3) mg/L in control group respectively (P<0.01). The contents of Cu and Se were obviously lower and that of Zn higher in observation group than those in control group. The same results were obtained between the infants with ROP and controls (P<0.01). However, there was no significant difference in the contents of serum Mg and Mn between two groups (P>0.05). It was concluded that the contents of serum Cu and Se in preterm infants who had high risk factors of ROP were obviously lower than in the controls. The contents of serum Cu and Se in the ROP infants were also much lower while contents of Zn much higher. Attention should be paid to the detection of the trace elements in preterm infants in order to prevent the deficiencies of Cu and Se. Only in this way can we prevent the deficiencies of Cu and Se, so as to decrease the ROP risk factors and prevent the disease.
Copper/blood ; Infant, Premature/*blood ; Retinopathy of Prematurity/*blood ; Retinopathy of Prematurity/*prevention & control ; Risk Factors ; Selenium/blood ; Spectrophotometry, Atomic ; Trace Elements/*blood ; Zinc/blood