Since the emergence of microsurgery in reconstructive surgery, free flaps have become a key tool in the management of patients with breast cancer. One such flap is the profunda artery perforator(PAP) flap. To date,the...Since the emergence of microsurgery in reconstructive surgery, free flaps have become a key tool in the management of patients with breast cancer. One such flap is the profunda artery perforator(PAP) flap. To date,there is no scientific consensus on whether voluminous free flaps remain dependent on their vascular pedicle throughout their lifespan. Therefore, the pedicle should always be carefully protected during revision surgery.In this article, we review the case of a middle-aged woman who suffered a pedicle transection needing reanastomosis during revision surgery six months after free-flap breast reconstruction. A 52-year-old woman who noticed a firm nodule in her right breast and armpit was referred to our department for surgical management. The Caucasian woman presented with no significant medical history or symptoms at the first consultation. Ultrasound-guided biopsy confirmed an invasive grade Ⅲ lobular carcinoma. Following staging,the patient underwent neoadjuvant chemotherapy before a right mastectomy with a complete homolateral axillary lymph node dissection and postoperative radiotherapy. One year after completing radiotherapy, free flap reconstruction with a PAP flap was performed, and six months later, revision surgery was required to enhance the volume of the reconstructed breast with a tissue expander and later an implant. Unfortunately,pedicle transection occurred during revision surgery, causing complete devascularization of the flap, which was confirmed by intraoperative Indocyanine Green imaging. The authors elected to perform salvage reanastomosis during the surgery. In keeping with the author’s 23-year experience with free flaps, the vascular pedicle should always be preserved in voluminous free flaps, as neovascularization alone may not ensure whole flap survival. The authors suggest always attempting re-anastomosis if vessels are compromised during revision surgery.展开更多
AIM:To discuss the impact of Lycium Barbarum Polysaccharide (LBP) and Danshensu purified from Traditional Chinese Medicine (TCM) on vascular endothelial growth factor (VEGF) of rabbits with retinal neovascularization....AIM:To discuss the impact of Lycium Barbarum Polysaccharide (LBP) and Danshensu purified from Traditional Chinese Medicine (TCM) on vascular endothelial growth factor (VEGF) of rabbits with retinal neovascularization. METHODS:Forty rabbits were divided into normal control group, model control group, LBP group and Danshensu group. Animals in the normal control group were fed in the normal oxygen environment. Animals in the other three groups were put into the environment with 70% oxygen for 5 days in order to build the model of oxygen-induced vascular proliferation retinopathy. And then different TCM extract was injected into the abdominal cavities of these annimals. After 7 days, the VEGF content of in the serum of rabbit was measured by double antibody sandwich method. RESULTS:Data analysis indicated that VEGF content was as follows:Danshensu group was lower than model control group (12.92 ±3.84ng/L vs 19.32 ±4.15ng/L, P < 0.05); LBP group and normal control group were lower than model control group (12.92±3.84ng/L, 9.26±1.61ng/L vs 19.32±4.15ng/L, P<0.01); total blood viscosity, plasma viscosity, cholesterol content, fibrinogen content and triacylglycerol content after peritoneal injection of LBP and Danshensu were obviously lower than before injection. CONCLUSION:TCM extract-LBP and Danshensu can prominently reduce the content of VEGF in the process of vascular proliferative retinopathy of rabbit; can prevent the occurrence of retinal microvascular disease by improving partial oxygen -deficient environment or affecting all kinds of new growth factor.展开更多
BACKGROUND: Studies have shown that abnormal innervation is an important factor impacting occurrence and development of pathological pain in endometriosis. OBJECTIVE: To observe uterine innervation of adenomyosis mice...BACKGROUND: Studies have shown that abnormal innervation is an important factor impacting occurrence and development of pathological pain in endometriosis. OBJECTIVE: To observe uterine innervation of adenomyosis mice and to analyze the cause of innervation changes due to nerve growth factor (NGF) expression, inflammation, and vascularization. DESIGN, TIME AND SETTING: This randomized, controlled, animal experiment was performed at the Research Institute of Obstetrics and Gynecology Hospital, and Central Laboratory of Zhong- shan Hospital, Fudan University from March to December 2008. MATERIALS: Tamoxifen was provided by Fudan Forward, China. Rabbit anti-mouse NGF was purchased from Santa Cruz Corporation, USA; rabbit anti-protein gene product 9.5 (PGP9.5) and rabbit antisubstance P (SP) were purchased from Chemicon, USA. METHODS: A total of 40 newborn ICR mice were randomly assigned to adenomyosis model and control groups, with 20 animals in each group. Mice in the adenomyosis model group were orally administrated 2.7 μmol/kg tamoxifen on days 2-5 after birth, while the controls were not treated. MAIN OUTCOME MEASURES: Both uteri from all mice were harvested at days 135-145 after birth. Expressions of polyclonal PGP9.5 and SP were immunohistochemically detected to demonstrate pan- and sensory nerve fibers. Microvessel density was quantified in the endometrium and myometrium using immunochemical staining for polyclonal rabbit anti-CD31, which stained vessels. Gene expression for NGF, high-affinity tyrosine kinase receptor (trkA), p75 neurotrophin receptor (p75NTR), bradykinin receptor-1 (BKR-1), and 2 (BKR-2), as well as substance P receptor (neurokinin1 receptor, NK1-R), were detected by reverse transcription-polymerase chain reaction. NGF-β protein expression was detected by Western blot analysis. RESULTS: More nerve fibers were stained with PGP9.5 in the endometrium and myometrium, and with SP in the endometrium, in adenomyosis mice compared with controls (P < 0.01 and P < 0.05). Microvessel density in the myometrium of adenomyosis mice was significantly greater than the controls (P < 0.01). In the uterus of adenomyosis mice, mRNA expression of NGF and its two receptors (trkA and p75 NTR), BKR-1, and NK1-R, as well as protein expression of NGF-β, were greater than the control mice (P < 0.01 or P < 0.05). CONCLUSION: Uterine innervation in the adenomyosis mice was increased compared with the controls. Moreover, NGF expression, inflammation, and vascularization, which have been shown to be impact factors of innervation, were abnormal in the uteri of adenomyosis mice.展开更多
Whether long non-coding RNA myocardial infarction-associated transcript is involved in oxygen-induced retinopathy remains poorly understood. To validate this hypothesis, we established a newborn mouse model of oxygen-...Whether long non-coding RNA myocardial infarction-associated transcript is involved in oxygen-induced retinopathy remains poorly understood. To validate this hypothesis, we established a newborn mouse model of oxygen-induced retinopathy by feeding in an oxygen concentration of 75 ± 2% from postnatal day 8 to postnatal day 12, followed by in normal air. On postnatal day 11, the mice were injected with the myocardial infarction-associated transcript siRNA plasmid via the vitreous cavity to knockdown long non-coding RNA myocardial infarction-associated transcript. Myocardial infarction-associated transcript siRNA transcription significantly inhibited myocardial infarctionassociated transcript mRNA expression, reduced the phosphatidylinosital-3-kinase, phosphorylated Akt and vascular endothelial growth factor immunopositivities, protein and mRNA expression, and alleviated the pathological damage to the retina of oxygen-induced retinopathy mouse models. These findings suggest that myocardial infarction-associated transcript is likely involved in the retinal neovascularization in retinopathy of prematurity and that inhibition of myocardial infarction-associated transcript can downregulate phosphatidylinosital-3-kinase, phosphorylated Akt and vascular endothelial growth factor expression levels and inhibit neovascularization. This study was approved by the Animal Ethics Committee of Shengjing Hospital of China Medical University, China(approval No. 2016 PS074 K) on February 25, 2016.展开更多
Vascularization is an important factor in nerve graft survival and function. The specific molecular regulations and patterns of angiogenesis following peripheral nerve injury are in a broad complex of pathways. This r...Vascularization is an important factor in nerve graft survival and function. The specific molecular regulations and patterns of angiogenesis following peripheral nerve injury are in a broad complex of pathways. This review aims to summarize current knowledge on the role of vascularization in nerve regeneration, including the key regulation molecules, and mechanisms and patterns of revascularization after nerve injury. Angiogenesis, the maturation of pre-existing vessels into new areas, is stimulated through angiogenic factors such as vascular endothelial growth factor and precedes the repair of damaged nerves. Vascular endothelial growth factor administration to nerves has demonstrated to increase revascularization after injury in basic science research. In the clinical setting, vascularized nerve grafts could be used in the reconstruction of large segmental peripheral nerve injuries. Vascularized nerve grafts are postulated to accelerate revascularization and enhance nerve regeneration by providing an optimal nutritional environment, especially in scarred beds, and decrease fibroblast infiltration. This could improve functional recovery after nerve grafting, however, conclusive evidence of the superiority of vascularized nerve grafts is lacking in human studies. A well-designed randomized controlled trial comparing vascularized nerve grafts to non-vascularized nerve grafts involving patients with similar injuries, nerve graft repair and follow-up times is necessary to demonstrate the efficacy of vascularized nerve grafts. Due to technical challenges, composite transfer of a nerve graft along with its adipose tissue has been proposed to provide a healthy tissue bed. Basic science research has shown that a vascularized fascial flap containing adipose tissue and a vascular bundle improves revascularization through excreted angiogenic factors, provided by the stem cells in the adipose tissue as well as by the blood supply and environmental support. While it was previously believed that revascularization occurred from both nerve ends, recent studies propose that revascularization occurs primarily from the proximal nerve coaptation. Fascial flaps or vascularized nerve grafts have limited applicability and future directions could lead towards off-the-shelf alternatives to autografting, such as biodegradable nerve scaffolds which include capillary-like networks to enable vascularization and avoid graft necrosis and ischemia.展开更多
AIM: To evaluate the effects of lentivirus-mediated pigment epithelium-derived factor(PEDF) gene transfer performed in treatment of rats with established choroidal neovascularization(CNV), and investigates the mechani...AIM: To evaluate the effects of lentivirus-mediated pigment epithelium-derived factor(PEDF) gene transfer performed in treatment of rats with established choroidal neovascularization(CNV), and investigates the mechanism by which PEDF inhibits CNV in rats.METHODS: Brown Norway(BN) rats(n =204) were induced by exposure to a laser, and then randomly assigned to 3 groups: no treatment; treatments with intravitreal injection of lentivirus-PEDF-green fluorescent protein(GFP) or lentivirus-control GFP(free fluorescent protein). Following induction and treatment,the CNV tissue was assessed for form, size and vessel leakage by fluorescein fundus angiography(FFA), optical coherence tomography(OCT), histopathology, and examination of choroidal flat mounts. VEGF, Flk-1, and PEDF expression were evaluated by real-time polymerase chain reaction(PCR) and Western blot.RESULTS: A stable laser-induced rat model of CNV was successfully established, and used to demonstrate lentivirus-mediated PEDG gene transfer by intravitreal injection. Expression of green fluorescence labelled PEDF was observed in the retina up to 28 d after injection. An intravitreal injection of lentivirus-PEDF-GFP at 7d led to a significant reduction in the size,thickness and area of CNV showed by FFA, OCT and choroidal flat mounts. PEDF was up-regulated while VEGF and Flk-1 were down-regulated in the lentivirus-PEDF-GFP group. The differences in VEGF and Flk-1 expression in the control and lentivirus-PEDF groups at 7, 14, 21 and 28 d after laser induction were all statistically significant. CONCLUSION: Lentivirus-mediated PEDF gene transfer is effective for use in treatment of laser-induced CNV, and PEDF exerts its therapeutic effects by inhibiting expression of VEGF and Flk-1.展开更多
AIM: To investigate the effects of the phosphatidylinositol 3-kinase(PI3 K) inhibitor LY294002 on retinal neovascularization(RNV) in the oxygen-induced retinopathy(OIR) mouse model and human umbilical vein endothelial...AIM: To investigate the effects of the phosphatidylinositol 3-kinase(PI3 K) inhibitor LY294002 on retinal neovascularization(RNV) in the oxygen-induced retinopathy(OIR) mouse model and human umbilical vein endothelial cells(HUVECs).METHODS: C57 BL/6 J mice were randomly divided into normoxia-control, OIR-control and LY294002 treatment groups. LY294002 or phosphate-buffered solution was intraperitoneally injected daily into mouse pups from P6 to P9 in LY294002 treatment group or OIR-control group. Morphological and pathological changes in RNV, as well as expression levels of PI3 K, serine-threonine kinase(AKT) and vascular endothelial growth factor(VEGF) were observed. HUVECs treating with LY294002 were exposed to hypoxia; the expression of PI3 K, AKT and VEGF were examined by Western blot and RT-PCR analyses.RESULTS: Compared with the OIR-control group, LY294002 significantly inhibit RNV. Adenosine diphosphatase(ADPase) staining and hematoxylin and eosin staining indicated that the clock hour scores of neovascularization and the nuclei of pre-retinal neovascular cells in the LY294002 treatment group were clearly less than those in the OIR-control group(1.41±0.52 vs 6.20±1.21; 10.50±1.58 vs 22.25±1.82, both P<0.05). Intravitreal injection of LY294002(in the LY294002 treatment group) markedly decreased PI3 K/AKT-VEGF expression compared with the OIR-control group by immunohistochemistry, Western blotting and RT-PCR(all P<0.05). In HUVECs treated with hypoxia, expression of PI3 K, AKT and VEGF were downregulated in the hypoxia-LY294002 group(all P<0.05).CONCLUSION: The PI3 K inhibitor LY294002 can inhibit RNV by downregulating PI3 K, AKT, and VEGF expression in vivo and in vitro. LY294002 may provide an effective method for preventing retinopathy of prematurity(ROP).展开更多
BACKGROUND A major problem in the healing of bone defects is insufficient or absent blood supply within the defect.To overcome this challenging problem,a plethora of approaches within bone tissue engineering have been...BACKGROUND A major problem in the healing of bone defects is insufficient or absent blood supply within the defect.To overcome this challenging problem,a plethora of approaches within bone tissue engineering have been developed recently.Bearing in mind that the interplay of various diffusible factors released by endothelial cells(ECs)and osteoblasts(OBs)have a pivotal role in bone growth and regeneration and that adjacent ECs and OBs also communicate directly through gap junctions,we set the focus on the simultaneous application of these cell types together with platelet-rich plasma(PRP)as a growth factor reservoir within ectopic bone tissue engineering constructs.AIM To vascularize and examine osteogenesis in bone tissue engineering constructs enriched with PRP and adipose-derived stem cells(ASCs)induced into ECs and OBs.METHODS ASCs isolated from adipose tissue,induced in vitro into ECs,OBs or just expanded were used for implant construction as followed:BPEO,endothelial and osteogenic differentiated ASCs with PRP and bone mineral matrix;BPUI,uninduced ASCs with PRP and bone mineral matrix;BC(control),only bone mineral matrix.At 1,2,4 and 8 wk after subcutaneous implantation in mice,implants were extracted and endothelial-related and bone-related gene expression were analyzed,while histological analyses were performed after 2 and 8 wk.RESULTS The percentage of vascularization was significantly higher in BC compared to BPUI and BPEO constructs 2 and 8 wk after implantation.BC had the lowest endothelial-related gene expression,weaker osteocalcin immunoexpression and Spp1 expression compared to BPUI and BPEO.Endothelial-related gene expression and osteocalcin immunoexpression were higher in BPUI compared to BC and BPEO.BPEO had a higher percentage of vascularization compared to BPUI and the highest CD31 immunoexpression among examined constructs.Except Vwf,endothelial-related gene expression in BPEO had a later onset and was upregulated and well-balanced during in vivo incubation that induced late onset of Spp1 expression and pronounced osteocalcin immunoexpression at 2 and 8 wk.Tissue regression was noticed in BPEO constructs after 8 wk.CONCLUSION Ectopically implanted BPEO constructs had a favorable impact on vascularization and osteogenesis,but tissue regression imposed the need for discovering a more optimal EC/OB ratio prior to considerations for clinical applications.展开更多
Choroidal neovascularization characterizes wet age-related macular degeneration.Choroidal neovascularization formation involves a primarily angiogenic process that is combined with both inflammation and proteolysis.A ...Choroidal neovascularization characterizes wet age-related macular degeneration.Choroidal neovascularization formation involves a primarily angiogenic process that is combined with both inflammation and proteolysis.A primary cause of choroidal neovascularization pathogenesis is alterations in pro-and anti-angiogenic factors derived from the retinal pigment epithelium,with vascular endothelium growth factor being mainly responsible for both clinical and experimental choroidal neovascularization.MicroRNAs(miRNAs)which are short,non-coding,endogenous RNA molecules have a major role in regulating various pathological processes,including inflammation and angiogenesis.A review of recent studies with the mouse laser-induced choroidal neovascularization model has shown alterations in miRNA expression in choroidal neovascularization tissues and could be potential therapeutic targets for wet age-related macular degeneration.Upregulation of miR-505(days 1 and 3 post-laser),miR-155(day 14)occurred in retina;miR-342-5p(days 3 and 7),miR-126-3p(day 14)in choroid;miR-23a,miR-24,miR-27a(day 7)in retina/choroid;miR-505(days 1 and 3)in retinal pigment epithelium/choroid;downregulation of miR-155(days 1 and 3),miR-29a,miR-29b,miR-29c(day 5),miR-93(day 14),miR-126(day 14)occurred in retinal pigment epithelium/choroid.Therapies using miRNA mimics or inhibitors were found to decrease choroidal neovascularization lesions.Choroidal neovascularization development was reduced by overexpression of miR-155,miR-188-5p,miR-(5,B,7),miR-126-3p,miR-342-5p,miR-93,miR-126,miR-195a-3p,miR-24,miR-21,miR-31,miR-150,and miR-184,or suppression of miR-505,miR-126-3p,miR-155,and miR-23/27.Further studies are warranted to determine miRNA expression in mouse laser-induced choroidal neovascularization models in order to validate and extend the reported findings.Important experimental variables need to be standardized;these include the strain and age of animals,gender,number and position of laser burns to the eye,laser parameters to induce choroidal neovascularization lesions including wavelength,power,spot size,and duration.展开更多
AIM:To determine the effects of intravitreal resveratrol(RSV)on murine laser-induced choroidal neovascularization(CNV).METHODS:The toxicity of RSV to choroidal endothelial cell(CEC)was measured using thiazolyl blue te...AIM:To determine the effects of intravitreal resveratrol(RSV)on murine laser-induced choroidal neovascularization(CNV).METHODS:The toxicity of RSV to choroidal endothelial cell(CEC)was measured using thiazolyl blue tetrazolium bromide(M一)assay.Effects of RSV on choroidal endothelial cell(CEC)migration were evaluated with a modified Boyden chamber assay,while tube formation was evaluated in a 2-D gel assay.CNV was induced by laser photocoagulation in mice.The effects of intravitreal injection of RSV on CNV development were evaluated by fluorescein angiography(FA),confocal analysis of isolectin B4 labeled choroidal flat mounts,and histologic examination of CNV membranes.Immunostaining was used to analyze the expression and phosphorylation of vascular endothelial growth factor receptor 2(VEGFR2).RESULTS:No significant cell toxicity was observed in CEC if the concentration of RSV was less than 200 pmol/L(P>0.05).RSV inhibited vascular endothelial growth factor(VEGF)-induced CEC migration(P<0.05)and tube formation(P<0.05)invitro.Furthermore,intravitrealinjectionof RSV significantly inhibited laser induced CNV formation in mice.The FA leakage,CNV volume and CNV area analysis revealed that there were 41%,45%,and 58%reduction in RSV-treated eyes(1.691±0.1032,178163±78623μm^3 and 6508±619.0μm^2,respectively)compared with those in control(2.724±0.08447,379676±98382μm3and16576±2646μm^2,respectively;P<0.05).Phospho-VEGFR2expression was much weaker in the sections of CNV lesions in RSV injected mice compared with that in control(P<0.05).CONCLUSION:Intravitreal injection of RSV exerts an inhibitory effect on CNV,which may through suppressing endothelial cell migration,tube formation and VEGFR2 phosphorylation.展开更多
AIM: To investigate the role of CCR7/p-ERK1/2/VEGF signaling in the mouse model of oxygen-induced retinopathy(OIR). METHODS: Neonatal C57BL/6J mice were evenly randomized into four groups: normoxia, OIR, OIR control(t...AIM: To investigate the role of CCR7/p-ERK1/2/VEGF signaling in the mouse model of oxygen-induced retinopathy(OIR). METHODS: Neonatal C57BL/6J mice were evenly randomized into four groups: normoxia, OIR, OIR control(treated with scramble si RNA), and OIR treated(treated with CCR7 siRNA). Normoxia group was not specially handled. Postnatal day 7(P7) mice in the OIR group were exposed to 75%±5% oxygen for 5d(P7-P12) and then maintained under normoxic conditions for 5d(P12-P17). Mice in the OIR control and OIR treated groups were given injections of scramble or CCR7 si RNA plasmid on P12 before returning to normoxic conditions for 5d(P12-P17). Retina samples were collected from all mice on P17, stained with adenosine diphosphatase(ADPase), and retinal neovascularization(RNV) was assessed. Retinas were also stained with hematoxylin and eosin(H&E) for RNV quantitation. The distribution and expression of CCR7, p-ERK1/2 and vascular endothelial growth factor(VEGF) were assessed via immunohistochemistry, Western blot, and quantitative real-time polymerase chain reaction(q RT-PCR). RESULTS: High oxygen promoted retinal neovascularization(P<0.05) and increased the number of endothelial nuclei in new vessels extending from the retina to the vitreous body; CCR7 promoted this process(P<0.05). CCR7 and VEGF m RNA were expressed at higher levels in the OIR and OIR control groups than in the normoxia and OIR treated groups. CCR7, p-ERK1/2, and VEGF protein were expressed in the retinas of mice in the OIR and OIR control groups. Intravitreal injection of CCR7 si RNA significantly reduced CCR7, p-ERK1/2, and VEGF expression in the OIR mouse model(all P<0.05). CCR7 significantly enhancedthe neovascularization and non-perfusion areas in the OIR group(P<0.05). CCR7 si RNA significantly reduced levels of p-ERK1/2 and VEGF as compared to OIR controls(P<0.05). CONCLUSION: These results suggest that CCR7/p-ERK 1/2/VEGF signaling plays an important role in OIR. CCR7 may be a potential target for the prevention and treatment of retinopathy of prematurity.展开更多
AIM: To investigate the signal transduction mechanism of matrix metalloproteinase-9(MMP-9) mediated-vascular endothelial growth factor(VEGF) expression and retinal neovascularization(RNV) in oxygen-induced retinopathy...AIM: To investigate the signal transduction mechanism of matrix metalloproteinase-9(MMP-9) mediated-vascular 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 C57BL/6J mice exposed to 75% ±2% oxygen for 5d.m RNA level and protein expression of MMP-9, TIMP-1and 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
Directed capillary ingrowth has long been considered synonymous with tumor vascularization.However,the vasculature of primary tumors and metastases is not necessarily formed by endothelial cell sprouting;instead,malig...Directed capillary ingrowth has long been considered synonymous with tumor vascularization.However,the vasculature of primary tumors and metastases is not necessarily formed by endothelial cell sprouting;instead,malignant tumors can acquire blood vessels via alternative vascularization mechanisms,such as intussusceptive microvascular growth,vessel co-option,and glomeruloid angiogenesis.Importantly,in response to anti-angiogenic therapies,malignant tumors can switch from one vascularization mechanism to another.In this article,we briefly review the biological features of these mechanisms and discuss on their significance in medical oncology.展开更多
AIM: To explore the process of retinal vascularization and risk factors for retinopathy of prematurity (ROP) treated with intravitreal ran ibizumab (IVR) as mono therapy.METHODS: Infants with type 1 ROP who received I...AIM: To explore the process of retinal vascularization and risk factors for retinopathy of prematurity (ROP) treated with intravitreal ran ibizumab (IVR) as mono therapy.METHODS: Infants with type 1 ROP who received IVR as primary treatment from August 2014 to October 2016 at Peking University People's HospitaPs Ophthalmology Department were included in the study. All eyes received 0.25 mg ranibizumab at initial treatment. Retinal vascularization was evaluated clinically. Potential risk factors were also recorded and examined.RESULTS: Retinal vascularization was completed in 126 eyes (62.7%), and retinal vascularization terminated in zone Ⅱ and zone Ⅲ with 16 eyes (7.9%) and 44 eyes (21.9%), respectively, after more than 1-year follow-up. In multivariate regression analysis, lower birth weight (BW), severity of ROP and repeated injections were found to be risk factors for peripheral avascular area (P<0.05).CONCLUSION: In our retrospective study, 29.8% of the ROP eyes treated with ranibizumab have peripheral avascular area at the last follow-up. Lighter BW and the severity of ROP are risk factors. Furthermore, repeated injections also increase the risk of retinal peripheral avascular area remaining in ROP patients.展开更多
AIM:To evaluate whether recombinant complement factor B(CFB)short hairpin RNA(sh RNA)reduces laserinduced choroidal neovascularization(CNV)in rats.METHODS:Laser-induced rat CNV model was established,and then the anima...AIM:To evaluate whether recombinant complement factor B(CFB)short hairpin RNA(sh RNA)reduces laserinduced choroidal neovascularization(CNV)in rats.METHODS:Laser-induced rat CNV model was established,and then the animals underwent fundus fluorescence angiography(FFA)and hematoxylin and eosin(HE)staining.On day 3 and 7 after photocoagulation,the expression of CFB and membrane attack complex(MAC)was detected by immunhischemistry.A recombinant CFBsh RNA plasmid was constructed.CFB and scrambled sh RNA plasmids were intravenous injected into rats via the tail vein on the day of laser treatment,respectively.On day 7,the incidence of CNV was determined by FFA,and the expression of CFB and vascular endothelial growth factor(VEGF)in retinal pigment epithelium(RPE)/choroidal tissues was detected by immunhischemistry,Western blot and/or semi-quantitative reverse transcription-polymerase chain reaction(RT-PCR)in CFB and scrambled sh RNA groups.The possible adverse effects of CFB-sh RNA injection were assessed by transmission electron microscopy and electroretinography.RESULTS:FFA and HE results indicated that a laserinduced rat CNV model was successfully established on day 7 after photocoagulation.The expression of CFB and MAC was extremely weak in normal retina and choroid,and increased on day 3 after photocoagulation.However,it started to reduce on day 7.CFB sh RNA plasmid was successfully constructed and induced CFB knockdown in the retinal and choroidal tissues.FFA showed CFB knockdown significantly inhibited incidence of CNV in rats.Moreover,CFB knockdown significantly inhibited the expression of VEGF in RPE/choroidal tissues.CFB sh RNA caused no obvious side effects in eyes.CONCLUSION:CFB knockdown significantly inhibits the formation and development of CNV in vivo through reducing the expression of VEGF,which is a potential therapy target.The alternative pathway of complement activation plays an important role in CNV formation.展开更多
Micro RNA-491-5 p(miR-491-5 p) plays an important role in regulating cell proliferation and migration;however,the effect of miR-491-5 p on neovascularization after traumatic brain injury remains poorly understood.In t...Micro RNA-491-5 p(miR-491-5 p) plays an important role in regulating cell proliferation and migration;however,the effect of miR-491-5 p on neovascularization after traumatic brain injury remains poorly understood.In this study,a controlled cortical injury model in C57 BL/6 mice and an oxygen-glucose deprivation model in microvascular endothelial cells derived from mouse brain were established to simulate traumatic brain injury in vivo and in vitro,respectively.In the in vivo model,quantitative real-time-polymerase chain reaction results showed that the expression of miR-491-5 p increased or decreased following the intracerebroventricular injection of an miR-491-5 p agomir or antagomir,respectively,and the expression of miR-491-5 p decreased slightly after traumatic brain injury.To detect the neuroprotective effects of miR-491-p,neurological severity scores,Morris water maze test,laser speckle techniques,and immunofluorescence staining were assessed,and the results revealed that miR-491-5 p downregulation alleviated neurological dysfunction,promoted the recovery of regional cerebral blood flow,increased the number of lectin-stained microvessels,and increased the survival of neurons after traumatic brain injury.During the in vitro experiments,the potential mechanism of miR-491-5 p on neovascularization was explored through quantitative real-time-polymerase chain reaction,which showed that miR-491-5 p expression increased or decreased in brain microvascular endothelial cells after transfection with an miR-491-5 p mimic or inhibitor,respectively.Dual-luciferase reporter and western blot assays verified that metallothionein-2 was a target gene for miR-491-5 p.Cell counting kit 8(CCK-8) assay,flow cytometry,and 2′,7′-dichlorofluorescein diacetate(DCFH-DA) assay results confirmed that the downregulation of miR-491-5 p increased brain microvascular endothelial cell viability,reduced cell apoptosis,and alleviated oxidative stress under oxygen-glucose deprivation conditions.Cell scratch assay,Transwell assay,tube formation assay,and western blot assay results demonstrated that miR-491-5 p downregulation promoted the migration,proliferation,and tube formation of brain microvascular endothelial cells through a metallothionein-2-dependent hypoxia-inducible factor-1α/vascular endothelial growth factor pathway.These findings confirmed that miR-491-5 p downregulation promotes neovascularization,restores cerebral blood flow,and improves the recovery of neurological function after traumatic brain injury.The mechanism may be mediated through a metallothionein-2-dependent hypoxia-inducible factor-1α/vascular endothelial growth factor signaling pathway and the alleviation of oxidative stress.All procedures were approved by Ethics Committee of the First Affiliated Hospital of Chongqing Medical University,China(approval No.2020-304) on June 22,2020.展开更多
AIM:To evaluate the potential efficacy and mechanisms of nintedanib in corneal neovascularization(NV)in rabbit models.METHODS:Corneal NV was induced using 1 mol/L Na OH.Rabbits(n=21)were randomized to 3 groups:Group 1...AIM:To evaluate the potential efficacy and mechanisms of nintedanib in corneal neovascularization(NV)in rabbit models.METHODS:Corneal NV was induced using 1 mol/L Na OH.Rabbits(n=21)were randomized to 3 groups:Group 1 were treated with 0.9%NaCl,Group 2 with Avastin(5 mg/mL),and Group 3 with nintedanib(1 mg/mL).All treatments star ted 1 d af ter alkaline burns and were topically performed 3 times a day for 2 wk.Photographs were taken on a slit lamp microscope on day 7 and 14.The NV area,the length of the vascularization and angiogenesis index(AI)were used to evaluate the corneal NV.On day 14,the immunohistochemical(IHC)studies of the cornea were examined.Western blot was performed to test the expression levels of vascular endothelial growth factor(VEGF),Akt,p-Akt,P38,p-P38,MMP-2 and MMP-9.RESULTS:The corneal NV area,vessel length and AI in Group 3 were significantly lower than Group 2,with both being lower than Group 1.IHC staining showed that VEGF was significantly overexpressed in the epithelium and stroma of cornea following alkaline burns.In contrast,the level of VEGF was significantly suppressed in both Group 2 and Group 3.Western blot results further confirmed that,compared with Group 1,Group 3 had significantly reduced expressions of VEGF,Akt,p-Akt,p-P38,MMP-2,and MMP-9 in corneal tissues.Trends of lower levels of MMP-2,AKT,and p-AKT in Group 3 than Group 2 were identified.CONCLUSION:Nintedanib and Avastin can effectively inhibit corneal NV,with P38 MAPK and AKT signaling pathways being possibly involved.Nintedanib seems more effective than Avastin and has the potential to be a novel therapy for preventing corneal NV.展开更多
AIM:To evaluate celastrol's effect on choroidal neovascularization(CNV).METHODS:In this study,neovascular formation in vitro(tube formation and aortic ring culture)and in vivo(laser induced neovascular in mice)was...AIM:To evaluate celastrol's effect on choroidal neovascularization(CNV).METHODS:In this study,neovascular formation in vitro(tube formation and aortic ring culture)and in vivo(laser induced neovascular in mice)was treated with celastrol to evaluate this natural compound's impact on CNV.Western blot was applied to explore the possible mechanism for it.For in vitro assay,triplicate for each group was repeated at least three times.For in vivo assay,each group contains 5 mice.RESULTS:Celastrol supressed tube formation and aortic ring sprout neovascularization.In vitro assay exhibited that celastrol inhibiting vascular endothelial growth factor(VEGF)-induced proliferation and migration of human umbilical vein endothelial cells and human choroidal endothelial cells,and by blocking VEGF signaling.Furthermore,intraperitoneal administration of celastrol significantly reduced the area of laser-induced CNV in an in vivo mouse model.By day 14,the area of CNV had decreased by 49.15%and 80.26%in the 0.1 mg/kg celastrol-treated group(n=5)and in the 0.5 mg/kg celastrol treated group(n=5),respectively,compared to the vehicle-treated group(n=5).CONCLUSION:Celastrol inhibits CNV by inhibiting VEGF-induced proliferation and migration of vascular endothelial cells,indicating that celastrol is a potent,natural therapeutic compound for the prevention of CNV.展开更多
AIM:To investigate whether ultrasound-targeted cationic microbubbles(CMBs)destruction could deliver endostatingreen fluorescent protein(GFP)plasmids efficiently to the human retinal endothelial cells(HRECs)and inhibit...AIM:To investigate whether ultrasound-targeted cationic microbubbles(CMBs)destruction could deliver endostatingreen fluorescent protein(GFP)plasmids efficiently to the human retinal endothelial cells(HRECs)and inhibit retinal neovascularization in mice.METHODS:CMBs were prepared and the presentation of GFP reporter was confirmed by flow cytometry and laser confocal microscopy.Experiments assessing HRECs migration and vascular formation were per formed to evaluate gene therapy’s efficiency in vitro.A mouse model of oxygen-induced retinopathy was employed and the expression of Bcl-xl,Bcl-2,vascular endothelial growth factor(VEGF)and endostatin in the retina of mice were determined by Western blotting and quantitative polymerase chain reaction(q PCR).The expression of endostatin-GFP in the retina was examined by laser confocal microscopy at 5,14,and 28 d after treatment.RESULTS:The gene expression of endostatin was the highest in the group of the CMBs.Besides,the inhibition and antiangiogenesis effect of the migration and development of HRECs were improved following treatment with CMBs compared with the other groups in vitro.In vivo,retinal neovascularization was significantly inhibited and the fluorescence intensity of endostatin-GFP in the mouse retina was importantly higher in the group of CMBs than that in other groups.CONCLUSION:The research illustrates ultrasoundtargeted CMBs destruction possessed distinct effect on the inhibition of the vascular formation and the development of retinal neovascularization both in vitro and in vivo.展开更多
BACKGROUND Diabetic retinopathy(DR)is a serious and potentially blinding complication of diabetes mellitus.Retinal neovascularization is one of the main pathological features of proliferative DR,and inhibiting retinal...BACKGROUND Diabetic retinopathy(DR)is a serious and potentially blinding complication of diabetes mellitus.Retinal neovascularization is one of the main pathological features of proliferative DR,and inhibiting retinal neovascularization is a research focus.AIM The aim was to evaluate the effect of intravitreal injection of recombinant human maspin on neovascularization in DR.METHODS An oxygen-induced retinopathy(OIR)mouse model was used to simulate neovascularization in DR.New born C57BL/6J mice were randomly divided to a normal control group,a maspin injection OIR group,and an OIR group.The mice in the maspin injection OIR group were injected with recombinant human maspin in the bilateral vitreous cavity on postnatal day P12,and those in the OIR group were injected with sterile phosphate buffered saline.The protein expression of vascular endothelial growth factor(VEGF)and hypoxia-inducible factor 1-alpha(HIF-1α)in the retina was measured by western blotting,and the mRNA expression of VEGF and HIF-1αwas measured by real-time polymerase chain reaction.The vascular cell nuclei that broke through the inner limiting membrane(ILM)were counted in haematoxylin-eosin stained retinal sections.RESULTS It was found that the number of vascular cell nuclei breaking through the ILM was 31.8±8.75 in the OIR group,which was significantly more than that in the normal control group(P<0.001).The number of vascular cell nuclei breaking through the ILM was 6.19±2.91 in the maspin injection OIR group,which was significantly less than that in OIR group(P<0.01).The relative protein and mRNA expression of VEGF and HIF-1αwas significantly lower in the retinas in the maspin injection OIR group than in those in the OIR group(P<0.01).CONCLUSION Maspin inhibited neovascularization in DR by modulating the HIF-1α/VEGF pathway,which provides a potential and effective strategy for the treatment of DR.展开更多
文摘Since the emergence of microsurgery in reconstructive surgery, free flaps have become a key tool in the management of patients with breast cancer. One such flap is the profunda artery perforator(PAP) flap. To date,there is no scientific consensus on whether voluminous free flaps remain dependent on their vascular pedicle throughout their lifespan. Therefore, the pedicle should always be carefully protected during revision surgery.In this article, we review the case of a middle-aged woman who suffered a pedicle transection needing reanastomosis during revision surgery six months after free-flap breast reconstruction. A 52-year-old woman who noticed a firm nodule in her right breast and armpit was referred to our department for surgical management. The Caucasian woman presented with no significant medical history or symptoms at the first consultation. Ultrasound-guided biopsy confirmed an invasive grade Ⅲ lobular carcinoma. Following staging,the patient underwent neoadjuvant chemotherapy before a right mastectomy with a complete homolateral axillary lymph node dissection and postoperative radiotherapy. One year after completing radiotherapy, free flap reconstruction with a PAP flap was performed, and six months later, revision surgery was required to enhance the volume of the reconstructed breast with a tissue expander and later an implant. Unfortunately,pedicle transection occurred during revision surgery, causing complete devascularization of the flap, which was confirmed by intraoperative Indocyanine Green imaging. The authors elected to perform salvage reanastomosis during the surgery. In keeping with the author’s 23-year experience with free flaps, the vascular pedicle should always be preserved in voluminous free flaps, as neovascularization alone may not ensure whole flap survival. The authors suggest always attempting re-anastomosis if vessels are compromised during revision surgery.
基金National"Eleventh Five-year Plan"Science and Technology Support Project,China(No.2006BAI06A15-3)
文摘AIM:To discuss the impact of Lycium Barbarum Polysaccharide (LBP) and Danshensu purified from Traditional Chinese Medicine (TCM) on vascular endothelial growth factor (VEGF) of rabbits with retinal neovascularization. METHODS:Forty rabbits were divided into normal control group, model control group, LBP group and Danshensu group. Animals in the normal control group were fed in the normal oxygen environment. Animals in the other three groups were put into the environment with 70% oxygen for 5 days in order to build the model of oxygen-induced vascular proliferation retinopathy. And then different TCM extract was injected into the abdominal cavities of these annimals. After 7 days, the VEGF content of in the serum of rabbit was measured by double antibody sandwich method. RESULTS:Data analysis indicated that VEGF content was as follows:Danshensu group was lower than model control group (12.92 ±3.84ng/L vs 19.32 ±4.15ng/L, P < 0.05); LBP group and normal control group were lower than model control group (12.92±3.84ng/L, 9.26±1.61ng/L vs 19.32±4.15ng/L, P<0.01); total blood viscosity, plasma viscosity, cholesterol content, fibrinogen content and triacylglycerol content after peritoneal injection of LBP and Danshensu were obviously lower than before injection. CONCLUSION:TCM extract-LBP and Danshensu can prominently reduce the content of VEGF in the process of vascular proliferative retinopathy of rabbit; can prevent the occurrence of retinal microvascular disease by improving partial oxygen -deficient environment or affecting all kinds of new growth factor.
基金the Construction Program of Shanghai Medical Intensive Subject (Obstetrics and Gynaecology), No. 05-111-0165
文摘BACKGROUND: Studies have shown that abnormal innervation is an important factor impacting occurrence and development of pathological pain in endometriosis. OBJECTIVE: To observe uterine innervation of adenomyosis mice and to analyze the cause of innervation changes due to nerve growth factor (NGF) expression, inflammation, and vascularization. DESIGN, TIME AND SETTING: This randomized, controlled, animal experiment was performed at the Research Institute of Obstetrics and Gynecology Hospital, and Central Laboratory of Zhong- shan Hospital, Fudan University from March to December 2008. MATERIALS: Tamoxifen was provided by Fudan Forward, China. Rabbit anti-mouse NGF was purchased from Santa Cruz Corporation, USA; rabbit anti-protein gene product 9.5 (PGP9.5) and rabbit antisubstance P (SP) were purchased from Chemicon, USA. METHODS: A total of 40 newborn ICR mice were randomly assigned to adenomyosis model and control groups, with 20 animals in each group. Mice in the adenomyosis model group were orally administrated 2.7 μmol/kg tamoxifen on days 2-5 after birth, while the controls were not treated. MAIN OUTCOME MEASURES: Both uteri from all mice were harvested at days 135-145 after birth. Expressions of polyclonal PGP9.5 and SP were immunohistochemically detected to demonstrate pan- and sensory nerve fibers. Microvessel density was quantified in the endometrium and myometrium using immunochemical staining for polyclonal rabbit anti-CD31, which stained vessels. Gene expression for NGF, high-affinity tyrosine kinase receptor (trkA), p75 neurotrophin receptor (p75NTR), bradykinin receptor-1 (BKR-1), and 2 (BKR-2), as well as substance P receptor (neurokinin1 receptor, NK1-R), were detected by reverse transcription-polymerase chain reaction. NGF-β protein expression was detected by Western blot analysis. RESULTS: More nerve fibers were stained with PGP9.5 in the endometrium and myometrium, and with SP in the endometrium, in adenomyosis mice compared with controls (P < 0.01 and P < 0.05). Microvessel density in the myometrium of adenomyosis mice was significantly greater than the controls (P < 0.01). In the uterus of adenomyosis mice, mRNA expression of NGF and its two receptors (trkA and p75 NTR), BKR-1, and NK1-R, as well as protein expression of NGF-β, were greater than the control mice (P < 0.01 or P < 0.05). CONCLUSION: Uterine innervation in the adenomyosis mice was increased compared with the controls. Moreover, NGF expression, inflammation, and vascularization, which have been shown to be impact factors of innervation, were abnormal in the uteri of adenomyosis mice.
基金supported by the National Natural Science Foundation of China,No.81600747(to YD)the Start-Up Foundation for Doctors of Liaoning Province,China,No.201501020(to YD)。
文摘Whether long non-coding RNA myocardial infarction-associated transcript is involved in oxygen-induced retinopathy remains poorly understood. To validate this hypothesis, we established a newborn mouse model of oxygen-induced retinopathy by feeding in an oxygen concentration of 75 ± 2% from postnatal day 8 to postnatal day 12, followed by in normal air. On postnatal day 11, the mice were injected with the myocardial infarction-associated transcript siRNA plasmid via the vitreous cavity to knockdown long non-coding RNA myocardial infarction-associated transcript. Myocardial infarction-associated transcript siRNA transcription significantly inhibited myocardial infarctionassociated transcript mRNA expression, reduced the phosphatidylinosital-3-kinase, phosphorylated Akt and vascular endothelial growth factor immunopositivities, protein and mRNA expression, and alleviated the pathological damage to the retina of oxygen-induced retinopathy mouse models. These findings suggest that myocardial infarction-associated transcript is likely involved in the retinal neovascularization in retinopathy of prematurity and that inhibition of myocardial infarction-associated transcript can downregulate phosphatidylinosital-3-kinase, phosphorylated Akt and vascular endothelial growth factor expression levels and inhibit neovascularization. This study was approved by the Animal Ethics Committee of Shengjing Hospital of China Medical University, China(approval No. 2016 PS074 K) on February 25, 2016.
基金supported by the National Institute of Neurological Disorders and Stroke of the National Institutes of Health under Award Number RO1 NS102360(to AYS)
文摘Vascularization is an important factor in nerve graft survival and function. The specific molecular regulations and patterns of angiogenesis following peripheral nerve injury are in a broad complex of pathways. This review aims to summarize current knowledge on the role of vascularization in nerve regeneration, including the key regulation molecules, and mechanisms and patterns of revascularization after nerve injury. Angiogenesis, the maturation of pre-existing vessels into new areas, is stimulated through angiogenic factors such as vascular endothelial growth factor and precedes the repair of damaged nerves. Vascular endothelial growth factor administration to nerves has demonstrated to increase revascularization after injury in basic science research. In the clinical setting, vascularized nerve grafts could be used in the reconstruction of large segmental peripheral nerve injuries. Vascularized nerve grafts are postulated to accelerate revascularization and enhance nerve regeneration by providing an optimal nutritional environment, especially in scarred beds, and decrease fibroblast infiltration. This could improve functional recovery after nerve grafting, however, conclusive evidence of the superiority of vascularized nerve grafts is lacking in human studies. A well-designed randomized controlled trial comparing vascularized nerve grafts to non-vascularized nerve grafts involving patients with similar injuries, nerve graft repair and follow-up times is necessary to demonstrate the efficacy of vascularized nerve grafts. Due to technical challenges, composite transfer of a nerve graft along with its adipose tissue has been proposed to provide a healthy tissue bed. Basic science research has shown that a vascularized fascial flap containing adipose tissue and a vascular bundle improves revascularization through excreted angiogenic factors, provided by the stem cells in the adipose tissue as well as by the blood supply and environmental support. While it was previously believed that revascularization occurred from both nerve ends, recent studies propose that revascularization occurs primarily from the proximal nerve coaptation. Fascial flaps or vascularized nerve grafts have limited applicability and future directions could lead towards off-the-shelf alternatives to autografting, such as biodegradable nerve scaffolds which include capillary-like networks to enable vascularization and avoid graft necrosis and ischemia.
基金Supported by the National Natural Science Foundation of China(No.81070735)
文摘AIM: To evaluate the effects of lentivirus-mediated pigment epithelium-derived factor(PEDF) gene transfer performed in treatment of rats with established choroidal neovascularization(CNV), and investigates the mechanism by which PEDF inhibits CNV in rats.METHODS: Brown Norway(BN) rats(n =204) were induced by exposure to a laser, and then randomly assigned to 3 groups: no treatment; treatments with intravitreal injection of lentivirus-PEDF-green fluorescent protein(GFP) or lentivirus-control GFP(free fluorescent protein). Following induction and treatment,the CNV tissue was assessed for form, size and vessel leakage by fluorescein fundus angiography(FFA), optical coherence tomography(OCT), histopathology, and examination of choroidal flat mounts. VEGF, Flk-1, and PEDF expression were evaluated by real-time polymerase chain reaction(PCR) and Western blot.RESULTS: A stable laser-induced rat model of CNV was successfully established, and used to demonstrate lentivirus-mediated PEDG gene transfer by intravitreal injection. Expression of green fluorescence labelled PEDF was observed in the retina up to 28 d after injection. An intravitreal injection of lentivirus-PEDF-GFP at 7d led to a significant reduction in the size,thickness and area of CNV showed by FFA, OCT and choroidal flat mounts. PEDF was up-regulated while VEGF and Flk-1 were down-regulated in the lentivirus-PEDF-GFP group. The differences in VEGF and Flk-1 expression in the control and lentivirus-PEDF groups at 7, 14, 21 and 28 d after laser induction were all statistically significant. CONCLUSION: Lentivirus-mediated PEDF gene transfer is effective for use in treatment of laser-induced CNV, and PEDF exerts its therapeutic effects by inhibiting expression of VEGF and Flk-1.
基金Supported by the National Natural Science Foundation of China (No.81600747)Startup Foundation for Docotors of Liaoning Province (No.201501020)
文摘AIM: To investigate the effects of the phosphatidylinositol 3-kinase(PI3 K) inhibitor LY294002 on retinal neovascularization(RNV) in the oxygen-induced retinopathy(OIR) mouse model and human umbilical vein endothelial cells(HUVECs).METHODS: C57 BL/6 J mice were randomly divided into normoxia-control, OIR-control and LY294002 treatment groups. LY294002 or phosphate-buffered solution was intraperitoneally injected daily into mouse pups from P6 to P9 in LY294002 treatment group or OIR-control group. Morphological and pathological changes in RNV, as well as expression levels of PI3 K, serine-threonine kinase(AKT) and vascular endothelial growth factor(VEGF) were observed. HUVECs treating with LY294002 were exposed to hypoxia; the expression of PI3 K, AKT and VEGF were examined by Western blot and RT-PCR analyses.RESULTS: Compared with the OIR-control group, LY294002 significantly inhibit RNV. Adenosine diphosphatase(ADPase) staining and hematoxylin and eosin staining indicated that the clock hour scores of neovascularization and the nuclei of pre-retinal neovascular cells in the LY294002 treatment group were clearly less than those in the OIR-control group(1.41±0.52 vs 6.20±1.21; 10.50±1.58 vs 22.25±1.82, both P<0.05). Intravitreal injection of LY294002(in the LY294002 treatment group) markedly decreased PI3 K/AKT-VEGF expression compared with the OIR-control group by immunohistochemistry, Western blotting and RT-PCR(all P<0.05). In HUVECs treated with hypoxia, expression of PI3 K, AKT and VEGF were downregulated in the hypoxia-LY294002 group(all P<0.05).CONCLUSION: The PI3 K inhibitor LY294002 can inhibit RNV by downregulating PI3 K, AKT, and VEGF expression in vivo and in vitro. LY294002 may provide an effective method for preventing retinopathy of prematurity(ROP).
基金Supported by Ministry of Education,Science and Technological Development of the Republic of Serbia,No.III 41017.
文摘BACKGROUND A major problem in the healing of bone defects is insufficient or absent blood supply within the defect.To overcome this challenging problem,a plethora of approaches within bone tissue engineering have been developed recently.Bearing in mind that the interplay of various diffusible factors released by endothelial cells(ECs)and osteoblasts(OBs)have a pivotal role in bone growth and regeneration and that adjacent ECs and OBs also communicate directly through gap junctions,we set the focus on the simultaneous application of these cell types together with platelet-rich plasma(PRP)as a growth factor reservoir within ectopic bone tissue engineering constructs.AIM To vascularize and examine osteogenesis in bone tissue engineering constructs enriched with PRP and adipose-derived stem cells(ASCs)induced into ECs and OBs.METHODS ASCs isolated from adipose tissue,induced in vitro into ECs,OBs or just expanded were used for implant construction as followed:BPEO,endothelial and osteogenic differentiated ASCs with PRP and bone mineral matrix;BPUI,uninduced ASCs with PRP and bone mineral matrix;BC(control),only bone mineral matrix.At 1,2,4 and 8 wk after subcutaneous implantation in mice,implants were extracted and endothelial-related and bone-related gene expression were analyzed,while histological analyses were performed after 2 and 8 wk.RESULTS The percentage of vascularization was significantly higher in BC compared to BPUI and BPEO constructs 2 and 8 wk after implantation.BC had the lowest endothelial-related gene expression,weaker osteocalcin immunoexpression and Spp1 expression compared to BPUI and BPEO.Endothelial-related gene expression and osteocalcin immunoexpression were higher in BPUI compared to BC and BPEO.BPEO had a higher percentage of vascularization compared to BPUI and the highest CD31 immunoexpression among examined constructs.Except Vwf,endothelial-related gene expression in BPEO had a later onset and was upregulated and well-balanced during in vivo incubation that induced late onset of Spp1 expression and pronounced osteocalcin immunoexpression at 2 and 8 wk.Tissue regression was noticed in BPEO constructs after 8 wk.CONCLUSION Ectopically implanted BPEO constructs had a favorable impact on vascularization and osteogenesis,but tissue regression imposed the need for discovering a more optimal EC/OB ratio prior to considerations for clinical applications.
文摘Choroidal neovascularization characterizes wet age-related macular degeneration.Choroidal neovascularization formation involves a primarily angiogenic process that is combined with both inflammation and proteolysis.A primary cause of choroidal neovascularization pathogenesis is alterations in pro-and anti-angiogenic factors derived from the retinal pigment epithelium,with vascular endothelium growth factor being mainly responsible for both clinical and experimental choroidal neovascularization.MicroRNAs(miRNAs)which are short,non-coding,endogenous RNA molecules have a major role in regulating various pathological processes,including inflammation and angiogenesis.A review of recent studies with the mouse laser-induced choroidal neovascularization model has shown alterations in miRNA expression in choroidal neovascularization tissues and could be potential therapeutic targets for wet age-related macular degeneration.Upregulation of miR-505(days 1 and 3 post-laser),miR-155(day 14)occurred in retina;miR-342-5p(days 3 and 7),miR-126-3p(day 14)in choroid;miR-23a,miR-24,miR-27a(day 7)in retina/choroid;miR-505(days 1 and 3)in retinal pigment epithelium/choroid;downregulation of miR-155(days 1 and 3),miR-29a,miR-29b,miR-29c(day 5),miR-93(day 14),miR-126(day 14)occurred in retinal pigment epithelium/choroid.Therapies using miRNA mimics or inhibitors were found to decrease choroidal neovascularization lesions.Choroidal neovascularization development was reduced by overexpression of miR-155,miR-188-5p,miR-(5,B,7),miR-126-3p,miR-342-5p,miR-93,miR-126,miR-195a-3p,miR-24,miR-21,miR-31,miR-150,and miR-184,or suppression of miR-505,miR-126-3p,miR-155,and miR-23/27.Further studies are warranted to determine miRNA expression in mouse laser-induced choroidal neovascularization models in order to validate and extend the reported findings.Important experimental variables need to be standardized;these include the strain and age of animals,gender,number and position of laser burns to the eye,laser parameters to induce choroidal neovascularization lesions including wavelength,power,spot size,and duration.
基金Supported by the National Natural Science Foundation of China(No.81703134,No.81770952)Henan Province Nature Science Foundation of China(No.162300410296)Hunan Province Nature Science Foundation of China(No.2018JJ3772)。
文摘AIM:To determine the effects of intravitreal resveratrol(RSV)on murine laser-induced choroidal neovascularization(CNV).METHODS:The toxicity of RSV to choroidal endothelial cell(CEC)was measured using thiazolyl blue tetrazolium bromide(M一)assay.Effects of RSV on choroidal endothelial cell(CEC)migration were evaluated with a modified Boyden chamber assay,while tube formation was evaluated in a 2-D gel assay.CNV was induced by laser photocoagulation in mice.The effects of intravitreal injection of RSV on CNV development were evaluated by fluorescein angiography(FA),confocal analysis of isolectin B4 labeled choroidal flat mounts,and histologic examination of CNV membranes.Immunostaining was used to analyze the expression and phosphorylation of vascular endothelial growth factor receptor 2(VEGFR2).RESULTS:No significant cell toxicity was observed in CEC if the concentration of RSV was less than 200 pmol/L(P>0.05).RSV inhibited vascular endothelial growth factor(VEGF)-induced CEC migration(P<0.05)and tube formation(P<0.05)invitro.Furthermore,intravitrealinjectionof RSV significantly inhibited laser induced CNV formation in mice.The FA leakage,CNV volume and CNV area analysis revealed that there were 41%,45%,and 58%reduction in RSV-treated eyes(1.691±0.1032,178163±78623μm^3 and 6508±619.0μm^2,respectively)compared with those in control(2.724±0.08447,379676±98382μm3and16576±2646μm^2,respectively;P<0.05).Phospho-VEGFR2expression was much weaker in the sections of CNV lesions in RSV injected mice compared with that in control(P<0.05).CONCLUSION:Intravitreal injection of RSV exerts an inhibitory effect on CNV,which may through suppressing endothelial cell migration,tube formation and VEGFR2 phosphorylation.
基金Supported by the Science and Technology Planning Foundation of Liaoning Province(No.2010225034)
文摘AIM: To investigate the role of CCR7/p-ERK1/2/VEGF signaling in the mouse model of oxygen-induced retinopathy(OIR). METHODS: Neonatal C57BL/6J mice were evenly randomized into four groups: normoxia, OIR, OIR control(treated with scramble si RNA), and OIR treated(treated with CCR7 siRNA). Normoxia group was not specially handled. Postnatal day 7(P7) mice in the OIR group were exposed to 75%±5% oxygen for 5d(P7-P12) and then maintained under normoxic conditions for 5d(P12-P17). Mice in the OIR control and OIR treated groups were given injections of scramble or CCR7 si RNA plasmid on P12 before returning to normoxic conditions for 5d(P12-P17). Retina samples were collected from all mice on P17, stained with adenosine diphosphatase(ADPase), and retinal neovascularization(RNV) was assessed. Retinas were also stained with hematoxylin and eosin(H&E) for RNV quantitation. The distribution and expression of CCR7, p-ERK1/2 and vascular endothelial growth factor(VEGF) were assessed via immunohistochemistry, Western blot, and quantitative real-time polymerase chain reaction(q RT-PCR). RESULTS: High oxygen promoted retinal neovascularization(P<0.05) and increased the number of endothelial nuclei in new vessels extending from the retina to the vitreous body; CCR7 promoted this process(P<0.05). CCR7 and VEGF m RNA were expressed at higher levels in the OIR and OIR control groups than in the normoxia and OIR treated groups. CCR7, p-ERK1/2, and VEGF protein were expressed in the retinas of mice in the OIR and OIR control groups. Intravitreal injection of CCR7 si RNA significantly reduced CCR7, p-ERK1/2, and VEGF expression in the OIR mouse model(all P<0.05). CCR7 significantly enhancedthe neovascularization and non-perfusion areas in the OIR group(P<0.05). CCR7 si RNA significantly reduced levels of p-ERK1/2 and VEGF as compared to OIR controls(P<0.05). CONCLUSION: These results suggest that CCR7/p-ERK 1/2/VEGF signaling plays an important role in OIR. CCR7 may be a potential target for the prevention and treatment of retinopathy of prematurity.
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) mediated-vascular 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 C57BL/6J mice exposed to 75% ±2% oxygen for 5d.m RNA level and protein expression of MMP-9, TIMP-1and 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 Grants from KTIA AIK 12-1-2013-0041Semmelweis University Start-Up Grant 40148-11658+3 种基金OTKA K109626,OTKA SNN1 14490,OTKA K100931,OTKA PD109201,EUREKA_HU_12-1-2012-0057TAMOP 4.2.4.A/1-11-1-2012-0001ONB Jubilaumsfondsprojekt Nr.14043 and Nr.14574the Vienna Fund for Innovative Interdisciplinary Cancer Research
文摘Directed capillary ingrowth has long been considered synonymous with tumor vascularization.However,the vasculature of primary tumors and metastases is not necessarily formed by endothelial cell sprouting;instead,malignant tumors can acquire blood vessels via alternative vascularization mechanisms,such as intussusceptive microvascular growth,vessel co-option,and glomeruloid angiogenesis.Importantly,in response to anti-angiogenic therapies,malignant tumors can switch from one vascularization mechanism to another.In this article,we briefly review the biological features of these mechanisms and discuss on their significance in medical oncology.
基金Supported by Peking University People’s Hospital Research and Development Funds(No.RDY2017-17)
文摘AIM: To explore the process of retinal vascularization and risk factors for retinopathy of prematurity (ROP) treated with intravitreal ran ibizumab (IVR) as mono therapy.METHODS: Infants with type 1 ROP who received IVR as primary treatment from August 2014 to October 2016 at Peking University People's HospitaPs Ophthalmology Department were included in the study. All eyes received 0.25 mg ranibizumab at initial treatment. Retinal vascularization was evaluated clinically. Potential risk factors were also recorded and examined.RESULTS: Retinal vascularization was completed in 126 eyes (62.7%), and retinal vascularization terminated in zone Ⅱ and zone Ⅲ with 16 eyes (7.9%) and 44 eyes (21.9%), respectively, after more than 1-year follow-up. In multivariate regression analysis, lower birth weight (BW), severity of ROP and repeated injections were found to be risk factors for peripheral avascular area (P<0.05).CONCLUSION: In our retrospective study, 29.8% of the ROP eyes treated with ranibizumab have peripheral avascular area at the last follow-up. Lighter BW and the severity of ROP are risk factors. Furthermore, repeated injections also increase the risk of retinal peripheral avascular area remaining in ROP patients.
文摘AIM:To evaluate whether recombinant complement factor B(CFB)short hairpin RNA(sh RNA)reduces laserinduced choroidal neovascularization(CNV)in rats.METHODS:Laser-induced rat CNV model was established,and then the animals underwent fundus fluorescence angiography(FFA)and hematoxylin and eosin(HE)staining.On day 3 and 7 after photocoagulation,the expression of CFB and membrane attack complex(MAC)was detected by immunhischemistry.A recombinant CFBsh RNA plasmid was constructed.CFB and scrambled sh RNA plasmids were intravenous injected into rats via the tail vein on the day of laser treatment,respectively.On day 7,the incidence of CNV was determined by FFA,and the expression of CFB and vascular endothelial growth factor(VEGF)in retinal pigment epithelium(RPE)/choroidal tissues was detected by immunhischemistry,Western blot and/or semi-quantitative reverse transcription-polymerase chain reaction(RT-PCR)in CFB and scrambled sh RNA groups.The possible adverse effects of CFB-sh RNA injection were assessed by transmission electron microscopy and electroretinography.RESULTS:FFA and HE results indicated that a laserinduced rat CNV model was successfully established on day 7 after photocoagulation.The expression of CFB and MAC was extremely weak in normal retina and choroid,and increased on day 3 after photocoagulation.However,it started to reduce on day 7.CFB sh RNA plasmid was successfully constructed and induced CFB knockdown in the retinal and choroidal tissues.FFA showed CFB knockdown significantly inhibited incidence of CNV in rats.Moreover,CFB knockdown significantly inhibited the expression of VEGF in RPE/choroidal tissues.CFB sh RNA caused no obvious side effects in eyes.CONCLUSION:CFB knockdown significantly inhibits the formation and development of CNV in vivo through reducing the expression of VEGF,which is a potential therapy target.The alternative pathway of complement activation plays an important role in CNV formation.
基金supported by the National Natural Science Foundation of China,Nos.82071397 (to XCS),82071332 (to ZDG)the Youth Fund of the National Natural Science Foundation of China,No.81801230 (to JJZ)the Excellent Scientific Research Talents Fund of the First Affiliated Hospital of Chongqing Medical University,China (to JJZ)。
文摘Micro RNA-491-5 p(miR-491-5 p) plays an important role in regulating cell proliferation and migration;however,the effect of miR-491-5 p on neovascularization after traumatic brain injury remains poorly understood.In this study,a controlled cortical injury model in C57 BL/6 mice and an oxygen-glucose deprivation model in microvascular endothelial cells derived from mouse brain were established to simulate traumatic brain injury in vivo and in vitro,respectively.In the in vivo model,quantitative real-time-polymerase chain reaction results showed that the expression of miR-491-5 p increased or decreased following the intracerebroventricular injection of an miR-491-5 p agomir or antagomir,respectively,and the expression of miR-491-5 p decreased slightly after traumatic brain injury.To detect the neuroprotective effects of miR-491-p,neurological severity scores,Morris water maze test,laser speckle techniques,and immunofluorescence staining were assessed,and the results revealed that miR-491-5 p downregulation alleviated neurological dysfunction,promoted the recovery of regional cerebral blood flow,increased the number of lectin-stained microvessels,and increased the survival of neurons after traumatic brain injury.During the in vitro experiments,the potential mechanism of miR-491-5 p on neovascularization was explored through quantitative real-time-polymerase chain reaction,which showed that miR-491-5 p expression increased or decreased in brain microvascular endothelial cells after transfection with an miR-491-5 p mimic or inhibitor,respectively.Dual-luciferase reporter and western blot assays verified that metallothionein-2 was a target gene for miR-491-5 p.Cell counting kit 8(CCK-8) assay,flow cytometry,and 2′,7′-dichlorofluorescein diacetate(DCFH-DA) assay results confirmed that the downregulation of miR-491-5 p increased brain microvascular endothelial cell viability,reduced cell apoptosis,and alleviated oxidative stress under oxygen-glucose deprivation conditions.Cell scratch assay,Transwell assay,tube formation assay,and western blot assay results demonstrated that miR-491-5 p downregulation promoted the migration,proliferation,and tube formation of brain microvascular endothelial cells through a metallothionein-2-dependent hypoxia-inducible factor-1α/vascular endothelial growth factor pathway.These findings confirmed that miR-491-5 p downregulation promotes neovascularization,restores cerebral blood flow,and improves the recovery of neurological function after traumatic brain injury.The mechanism may be mediated through a metallothionein-2-dependent hypoxia-inducible factor-1α/vascular endothelial growth factor signaling pathway and the alleviation of oxidative stress.All procedures were approved by Ethics Committee of the First Affiliated Hospital of Chongqing Medical University,China(approval No.2020-304) on June 22,2020.
基金Supported by the Natural Science Foundation of Tianjin City(No.2150000045)。
文摘AIM:To evaluate the potential efficacy and mechanisms of nintedanib in corneal neovascularization(NV)in rabbit models.METHODS:Corneal NV was induced using 1 mol/L Na OH.Rabbits(n=21)were randomized to 3 groups:Group 1 were treated with 0.9%NaCl,Group 2 with Avastin(5 mg/mL),and Group 3 with nintedanib(1 mg/mL).All treatments star ted 1 d af ter alkaline burns and were topically performed 3 times a day for 2 wk.Photographs were taken on a slit lamp microscope on day 7 and 14.The NV area,the length of the vascularization and angiogenesis index(AI)were used to evaluate the corneal NV.On day 14,the immunohistochemical(IHC)studies of the cornea were examined.Western blot was performed to test the expression levels of vascular endothelial growth factor(VEGF),Akt,p-Akt,P38,p-P38,MMP-2 and MMP-9.RESULTS:The corneal NV area,vessel length and AI in Group 3 were significantly lower than Group 2,with both being lower than Group 1.IHC staining showed that VEGF was significantly overexpressed in the epithelium and stroma of cornea following alkaline burns.In contrast,the level of VEGF was significantly suppressed in both Group 2 and Group 3.Western blot results further confirmed that,compared with Group 1,Group 3 had significantly reduced expressions of VEGF,Akt,p-Akt,p-P38,MMP-2,and MMP-9 in corneal tissues.Trends of lower levels of MMP-2,AKT,and p-AKT in Group 3 than Group 2 were identified.CONCLUSION:Nintedanib and Avastin can effectively inhibit corneal NV,with P38 MAPK and AKT signaling pathways being possibly involved.Nintedanib seems more effective than Avastin and has the potential to be a novel therapy for preventing corneal NV.
基金Supported by National Natural Science Foundation of China(No.81570826)。
文摘AIM:To evaluate celastrol's effect on choroidal neovascularization(CNV).METHODS:In this study,neovascular formation in vitro(tube formation and aortic ring culture)and in vivo(laser induced neovascular in mice)was treated with celastrol to evaluate this natural compound's impact on CNV.Western blot was applied to explore the possible mechanism for it.For in vitro assay,triplicate for each group was repeated at least three times.For in vivo assay,each group contains 5 mice.RESULTS:Celastrol supressed tube formation and aortic ring sprout neovascularization.In vitro assay exhibited that celastrol inhibiting vascular endothelial growth factor(VEGF)-induced proliferation and migration of human umbilical vein endothelial cells and human choroidal endothelial cells,and by blocking VEGF signaling.Furthermore,intraperitoneal administration of celastrol significantly reduced the area of laser-induced CNV in an in vivo mouse model.By day 14,the area of CNV had decreased by 49.15%and 80.26%in the 0.1 mg/kg celastrol-treated group(n=5)and in the 0.5 mg/kg celastrol treated group(n=5),respectively,compared to the vehicle-treated group(n=5).CONCLUSION:Celastrol inhibits CNV by inhibiting VEGF-induced proliferation and migration of vascular endothelial cells,indicating that celastrol is a potent,natural therapeutic compound for the prevention of CNV.
基金Supported by the National Natural Science Foundation for Youth of China(No.81901765)。
文摘AIM:To investigate whether ultrasound-targeted cationic microbubbles(CMBs)destruction could deliver endostatingreen fluorescent protein(GFP)plasmids efficiently to the human retinal endothelial cells(HRECs)and inhibit retinal neovascularization in mice.METHODS:CMBs were prepared and the presentation of GFP reporter was confirmed by flow cytometry and laser confocal microscopy.Experiments assessing HRECs migration and vascular formation were per formed to evaluate gene therapy’s efficiency in vitro.A mouse model of oxygen-induced retinopathy was employed and the expression of Bcl-xl,Bcl-2,vascular endothelial growth factor(VEGF)and endostatin in the retina of mice were determined by Western blotting and quantitative polymerase chain reaction(q PCR).The expression of endostatin-GFP in the retina was examined by laser confocal microscopy at 5,14,and 28 d after treatment.RESULTS:The gene expression of endostatin was the highest in the group of the CMBs.Besides,the inhibition and antiangiogenesis effect of the migration and development of HRECs were improved following treatment with CMBs compared with the other groups in vitro.In vivo,retinal neovascularization was significantly inhibited and the fluorescence intensity of endostatin-GFP in the mouse retina was importantly higher in the group of CMBs than that in other groups.CONCLUSION:The research illustrates ultrasoundtargeted CMBs destruction possessed distinct effect on the inhibition of the vascular formation and the development of retinal neovascularization both in vitro and in vivo.
基金Liaoning Province Natural Science Foundation,No.2020-BS-277.
文摘BACKGROUND Diabetic retinopathy(DR)is a serious and potentially blinding complication of diabetes mellitus.Retinal neovascularization is one of the main pathological features of proliferative DR,and inhibiting retinal neovascularization is a research focus.AIM The aim was to evaluate the effect of intravitreal injection of recombinant human maspin on neovascularization in DR.METHODS An oxygen-induced retinopathy(OIR)mouse model was used to simulate neovascularization in DR.New born C57BL/6J mice were randomly divided to a normal control group,a maspin injection OIR group,and an OIR group.The mice in the maspin injection OIR group were injected with recombinant human maspin in the bilateral vitreous cavity on postnatal day P12,and those in the OIR group were injected with sterile phosphate buffered saline.The protein expression of vascular endothelial growth factor(VEGF)and hypoxia-inducible factor 1-alpha(HIF-1α)in the retina was measured by western blotting,and the mRNA expression of VEGF and HIF-1αwas measured by real-time polymerase chain reaction.The vascular cell nuclei that broke through the inner limiting membrane(ILM)were counted in haematoxylin-eosin stained retinal sections.RESULTS It was found that the number of vascular cell nuclei breaking through the ILM was 31.8±8.75 in the OIR group,which was significantly more than that in the normal control group(P<0.001).The number of vascular cell nuclei breaking through the ILM was 6.19±2.91 in the maspin injection OIR group,which was significantly less than that in OIR group(P<0.01).The relative protein and mRNA expression of VEGF and HIF-1αwas significantly lower in the retinas in the maspin injection OIR group than in those in the OIR group(P<0.01).CONCLUSION Maspin inhibited neovascularization in DR by modulating the HIF-1α/VEGF pathway,which provides a potential and effective strategy for the treatment of DR.
