The integrity of retinal ganglion cells is tightly associated with diabetic macular degeneration that leads to damage and death of retinal ganglion cells,affecting vision.The major clinical treatments for diabetic mac...The integrity of retinal ganglion cells is tightly associated with diabetic macular degeneration that leads to damage and death of retinal ganglion cells,affecting vision.The major clinical treatments for diabetic macular edema are anti-vascular endothelial growth factor drugs and laser photocoagulation.However,although the macular thickness can be normalized with each of these two therapies used alone,the vision does not improve in many patients.This might result from the incomplete recovery of retinal ganglion cell injury.Therefore,a prospective,non-randomized,controlled clinical trial was designed to investigate the effect of anti-vascular endothelial growth factor drugs combined with laser photocoagulation on the integrity of retinal ganglion cells in patients with diabetic macular edema and its relationship with vision recovery.In this trial,150 patients with diabetic macular edema will be equally divided into three groups according to therapeutic methods,followed by treatment with anti-vascular endothelial growth factor drugs,laser photocoagulation therapy,and their combination.All patients will be followed up for 12 months.The primary outcome measure is retinal ganglion cell-inner plexiform layer thickness at 12 months after treatment.The secondary outcome measures include retinal ganglion cell-inner plexiform layer thickness before and 1,3,6,and 9 months after treatment,retinal nerve fiber layer thickness,best-corrected visual acuity,macular area thickness,and choroidal thickness before and 1,3,6,9,and 12 months after treatment.Safety measure is the incidence of adverse events at 1,3,6,9,and 12 months after treatment.The study protocol hopes to validate the better efficacy and safety of the combined treatment in patients with diabetic macula compared with the other two monotherapies alone during the 12-month follow-up period.The trial is designed to focus on clarifying the time-effect relationship between imaging measures related to the integrity of retinal ganglion cells and best-corrected visual acuity.The trial protocol was approved by the Medical Ethics Committee of the Affiliated Hospital of Beihua University with approval No.(2023)(26)on April 25,2023,and was registered with the Chinese Clinical Trial Registry(registration number:ChiCTR2300072478,June 14,2023,protocol version:2.0).展开更多
Although bone marrow mesenchymal stem cells(BMSCs)might have therapeutic potency in ischemic stroke,the benefits are limited.The current study investigated the effects of BMSCs engineered to overexpress vascular endot...Although bone marrow mesenchymal stem cells(BMSCs)might have therapeutic potency in ischemic stroke,the benefits are limited.The current study investigated the effects of BMSCs engineered to overexpress vascular endothelial growth factor(VEGF)on behavioral defects in a rat model of transient cerebral ischemia,which was induced by middle cerebral artery occlusion.VEGF-BMSCs or control grafts were injected into the left striatum of the infarcted hemisphere 24 hours after stroke.We found that compared with the stroke-only group and the vehicle-and BMSCs-control groups,the VEGF-BMSCs treated animals displayed the largest benefits,as evidenced by attenuated behavioral defects and smaller infarct volume 7 days after stroke.Additionally,VEGF-BMSCs greatly inhibited destruction of the blood-brain barrier,increased the regeneration of blood vessels in the region of ischemic penumbra,and reducedneuronal degeneration surrounding the infarct core.Further mechanistic studies showed that among all transplant groups,VEGF-BMSCs transplantation induced the highest level of brain-derived neurotrophic factor.These results suggest that BMSCs transplantation with vascular endothelial growth factor has the potential to treat ischemic stroke with better results than are currently available.展开更多
Neovascularization and angiogenesis in the brain are important physiological processes for normal brain development and repair/regeneration following insults. Integrins are cell surface adhesion receptors mediating im...Neovascularization and angiogenesis in the brain are important physiological processes for normal brain development and repair/regeneration following insults. Integrins are cell surface adhesion receptors mediating important function of cells such as survival, growth and development during tissue organization, differentiation and organogenesis. In this study, we used an integrin-binding array platform to identify the important types of integrins and their binding peptides that facilitate adhesion, growth, development, and vascular-like network formation of rat primary brain microvascular endothelial cells. Brain microvascular endothelial cells were isolated from rat brain on post-natal day 7. Cells were cultured in a custom-designed integrin array system containing short synthetic peptides binding to 16 types of integrins commonly expressed on cells in vertebrates. After 7 days of culture, the brain microvascular endothelial cells were processed for immunostaining with markers for endothelial cells including von Willibrand factor and platelet endothelial cell adhesion molecule. 5-Bromo-2′-dexoyuridine was added to the culture at 48 hours prior to fixation to assess cell proliferation. Among 16 integrins tested, we found that α5β1, αvβ5 and αvβ8 greatly promoted proliferation of endothelial cells in culture. To investigate the effect of integrin-binding peptides in promoting neovascularization and angiogenesis, the binding peptides to the above three types of integrins were immobilized to our custom-designed hydrogel in three-dimensional(3 D) culture of brain microvascular endothelial cells with the addition of vascular endothelial growth factor. Following a 7-day 3 D culture, the culture was fixed and processed for double labeling of phalloidin with von Willibrand factor or platelet endothelial cell adhesion molecule and assessed under confocal microscopy. In the 3 D culture in hydrogels conjugated with the integrin-binding peptide, brain microvascular endothelial cells formed interconnected vascular-like network with clearly discernable lumens, which is reminiscent of brain microvascular network in vivo. With the novel integrin-binding array system, we identified the specific types of integrins on brain microvascular endothelial cells that mediate cell adhesion and growth followed by functionalizing a 3 D hydrogel culture system using the binding peptides that specifically bind to the identified integrins, leading to robust growth and lumenized microvascular-like network formation of brain microvascular endothelial cells in 3 D culture. This technology can be used for in vitro and in vivo vascularization of transplants or brain lesions to promote brain tissue regeneration following neurological insults.展开更多
Objective:This study aimed to investigate the potential of human umbilical cord mesenchymal stem cell(hucMSC)-derived exosomes(hucMSC-Exos)in inhibiting hypoxia-induced cell hyper proliferation and overexpression of v...Objective:This study aimed to investigate the potential of human umbilical cord mesenchymal stem cell(hucMSC)-derived exosomes(hucMSC-Exos)in inhibiting hypoxia-induced cell hyper proliferation and overexpression of vascular endothelial growth factor A(VEGF-A)in immature human fetal retinal microvascular endothelial cells(hfRMECs).Methods:Exosomes were isolated from hucMSCs using cryogenic ultracentrifugation and characterized through various techniques,including transmission electron microscopy,nanoparticle tracking analysis,bicinchoninic acid assays,and western blotting.The hfRMECs were identified using von Willebrand factor(vWF)co-staining and divided into four groups:a control group cultured under normoxic condition,a hypoxic model group,a hypoxic group treated with low-concentration hucMSC-Exos(75μg/mL)and a hypoxic group treated with high-concentration hucMSC-Exos(100μg/mL).Cell viability and proliferation were assessed using Cell Counting Kit-8(CCK-8)assay and EdU(5-ethynyl-2′-deoxyuridine)assay respectively.Expression levels of VEGF-A were evaluated using RT-PCR,western blotting and immunofluorescence.Results:Hypoxia significantly increased hfRMECs’viability and proliferation by upregulating VEGF-A levels.The administration of hucMSC-Exos effectively reversed this response,with the high-concentration group exhibiting greater efficacy compared to the lowconcentration group.Conclusion:In conclusion,hucMSC-Exos can dose-dependently inhibit hypoxia-induced hyperproliferation and VEGF-A overexpression in immature fetal retinal microvascular endothelial cells.展开更多
AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the...AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the retinal tissue in diabetic rats.METHODS:The expression of HDAC7 in HRMECs under high glucose and the retinal tissue from normal or diabetic rats were detected with immunohistochemistry and Western blot.LV-shHDAC7 HRMECs were used to study the effect of HDAC7 on cell activities.Cell count kit-8(CCK-8),5-ethynyl2’-deoxyuridine(EdU),flow cytometry,scratch test,Transwell test and tube formation assay were used to examine the ability of cell proliferation,migration,and angiogenesis.Finally,a preliminary exploration of its mechanism was performed by Western blot.RESULTS:The expression of HDAC7 was both upregulated in retinal tissues of diabetic rats and high glucosetreated HRMECs.Down-regulation of HDAC7 expression significantly reduced the ability of proliferation,migration,and tube formation,and reversed the high glucose-induced high expression of CDK1/Cyclin B1 and vascular endothelial growth factor in high glucose-treated HRMECs.CONCLUSION:High glucose can up-regulate the expression of HDAC7 in HRMECs.Down-regulation of HDAC7 can inhibit HRMECs activities.HDAC7 is proposed to be involved in pathogenesis of diabetic retinopathy and a therapeutic target.展开更多
Endogenous neural stem cells become "activated" after neuronal injury, but the activation sequence and fate of endogenous neural stem cells in focal cerebral ischemia model are little known. We evaluated the relatio...Endogenous neural stem cells become "activated" after neuronal injury, but the activation sequence and fate of endogenous neural stem cells in focal cerebral ischemia model are little known. We evaluated the relationships between neural stem cells and hypoxia-inducible factor-1α and vascular endothelial growth factor expression in a photothromobotic rat stroke model using immunohistochemistry and western blot analysis. We also evaluated the chronological changes of neural stem cells by 5-bromo-2′-deoxyuridine(BrdU) incorporation. Hypoxia-inducible factor-1α expression was initially increased from 1 hour after ischemic injury, followed by vascular endothelial growth factor expression. Hypoxia-inducible factor-1α immunoreactivity was detected in the ipsilateral cortical neurons of the infarct core and peri-infarct area. Vascular endothelial growth factor immunoreactivity was detected in bilateral cortex, but ipsilateral cortex staining intensity and numbers were greater than the contralateral cortex. Vascular endothelial growth factor immunoreactive cells were easily found along the peri-infarct area 12 hours after focal cerebral ischemia. The expression of nestin increased throughout the microvasculature in the ischemic core and the peri-infarct area in all experimental rats after 24 hours of ischemic injury. Nestin immunoreactivity increased in the subventricular zone during 12 hours to 3 days, and prominently increased in the ipsilateral cortex between 3–7 days. Nestin-labeled cells showed dual differentiation with microvessels near the infarct core and reactive astrocytes in the peri-infarct area. BrdU-labeled cells were increased gradually from day 1 in the ipsilateral subventricular zone and cortex, and numerous BrdU-labeled cells were observed in the peri-infarct area and non-lesioned cortex at 3 days. BrdU-labeled cells rather than neurons, were mainly co-labeled with nestin and GFAP. Early expressions of hypoxia-inducible factor-1α and vascular endothelial growth factor after ischemia made up the microenvironment to increase the neuronal plasticity of activated endogenous neural stem cells. Moreover, neural precursor cells after large-scale cortical injury could be recruited from the cortex nearby infarct core and subventricular zone.展开更多
Hypoxia and transforming growth factor-β1 (TGF-β1) increase vascular endothelial growth factor A (VEGFA) expression in a number of malignancies. This effect of hypoxia and TGF-β1 might be responsible for tumor ...Hypoxia and transforming growth factor-β1 (TGF-β1) increase vascular endothelial growth factor A (VEGFA) expression in a number of malignancies. This effect of hypoxia and TGF-β1 might be responsible for tumor progression and metastasis of advanced prostate cancer. In the present study, TGF-β1 was shown to induce VEGFA165 secretion from both normal cell lines (HPV7 and RWPE1) and prostate cancer cell lines (DU 145 and PC3). Conversely, hypoxia-stimulated VEGFA165 secretion was observed only in prostate cancer cell lines. Hypoxia induced TGF-β1 expression in PC3 prostate cancer cells, and the TGF-β1 type I receptor (ALK5) kinase inhibitor partially blocked hypoxia-mediated VEGFA16s secretion. This effect of hypoxia provides a novel mechanism to increase VEGFA expression in prostate cancer cells. Although autocrine signaling of VEGFA has been implicated in prostate cancer progression and metastasis, the associated mechanism is poorly characterized. VEGFA activity is mediated via VEGF receptor (VEGFR) 1 (Fit-l) and 2 (FIk-I/KDR). Whereas VEGFR-1 mRNA was detected in normal prostate epithelial cells, VEGFR-2 mRNA and VEGFR protein were expressed only in PC3 cells. VEGFA165 treatment induced phosphorylation of extracellular signal-regulated kinase 1/2 (ERKI/2) in PC3 cells but not in HPV7 cells, suggesting that the autocrine function of VEGFA may be uniquely associated with prostate cancer. Activation of VEGFR-2 by VEGFA165 was shown to enhance migration of PC3 cells. A similar effect was also observed with endogenous VEGFA induced by TGF-β1 and hypoxia. These findings illustrate that an autocrine loop of VEGFA via VEGFR-2 is critical for the tumorigenic effects of TGF-β1 and hypoxia on metastatic prostate cancers.展开更多
Objective: To investigate the effect of Fuzhenghuayu decoction on autocrine activation of hepatic stellate cell (HSC). Methods: The drug serum containing Fuzhenghuayu decoction was collected from normal rats, and cul-...Objective: To investigate the effect of Fuzhenghuayu decoction on autocrine activation of hepatic stellate cell (HSC). Methods: The drug serum containing Fuzhenghuayu decoction was collected from normal rats, and cul- tured with activated HSC in vitro. The conditioned medium from the drug serum treated HSC was added to primary cultured quiescent HSC. Cell prolifera- tion was assayed by tetrazolium colorimetric test, and the contents of type Ⅰ collagen and vascular endo- thelial growth factor (VEGF) in the supernatant were measured with ELISA. Results: The conditioned medium from activated HSC could stimulate the quiescent HSC proliferation and type Ⅰ collagen secretion. The drug serum inhibi- ted this stimulating action and VEGF secretion from the activated HSC. Conclusion: Fuzhenghuayu decoction acts effectively against the autocrine activation pathway of HSC. The mechanism may be associated with the inhibition of the secretion of VEGF by activated HSC.展开更多
Neural stem/progenitor cell (NSC) transplantation has been shown to effectively improve neurological function in rats with hypoxic-isch- emic brain damage. Vascular endothelial growth factor (VEGF) is a signaling ...Neural stem/progenitor cell (NSC) transplantation has been shown to effectively improve neurological function in rats with hypoxic-isch- emic brain damage. Vascular endothelial growth factor (VEGF) is a signaling protein that stimulates angiogenesis and improves neural regeneration. We hypothesized that transplantation of VEGF-transfected NSCs would alleviate hypoxic-ischemic brain damage in neo- natal rats. We produced and transfected a recombinant lentiviral vector containing the VEGF165gene into cultured NSCs. The transfected NSCs were transplanted into the left sensorimotor cortex of rats 3 days after hypoxic-ischemic brain damage. Compared with the NSCs group, VEGF mRNA and protein expression levels were increased in the transgene NSCs group, and learning and memory abilities were significantly improved at 30 days. Furthermore, histopathological changes were alleviated in these animals. Our findings indicate that transplantation of VEGF-transfected NSCs may facilitate the recovery of neurological function, and that its therapeutic effectiveness is better than that of unmodified NSCs.展开更多
This study was designed to determine the levels of early endothelial progenitor cells (EPCs), apelin, vascu- lar endothelial growth factor (VEGF) and stromal cell-derived growth factor-1 (SDF-1) after acute myoc...This study was designed to determine the levels of early endothelial progenitor cells (EPCs), apelin, vascu- lar endothelial growth factor (VEGF) and stromal cell-derived growth factor-1 (SDF-1) after acute myocardial infarction (AMI), and to investigate the relationships between these cytokines and early EPCs. Early EPCs, de- fined as CD133+, KDR+, and CD34~ cells, were quantified by flow cytometry. The levels of early EPCs and those cytokines in AMI patients were significantly different from those with coronary artery disease or controls (P 〈 0.05). Plasma apelin levels were inversely correlated with Gensini score and early EPCs (both P 〈 0.01). Early EPCs, VEGF and SDF-1 showed different patterns of changes in AMI patients during the first 24 h. The trend in the change of early EPCs was proportionally correlated with that of VEGF (P 〈 0.05). AMI patients exhibited in- creased early EPCs with remarkably decreased apelin levels and enhanced VEGF levels.展开更多
Neural stem cells are characterized by the ability to differentiate and stably express exogenous ge- nes. Vascular endothelial growth factor plays a role in protecting local blood vessels and neurons of newborn rats w...Neural stem cells are characterized by the ability to differentiate and stably express exogenous ge- nes. Vascular endothelial growth factor plays a role in protecting local blood vessels and neurons of newborn rats with hypoxic-ischemic encephalopathy. Transplantation of vascular endothelial growth factor-transfected neural stem cells may be neuroprotective in rats with cerebral palsy. In this study, 7-day-old Sprague-Dawley rats were divided into five groups: (1) sham operation (control), (2) cerebral palsy model alone or with (3) phosphate-buffered saline, (4) vascular en- dothelial growth factor 165 + neural stem cells, or (5) neural stem cells alone. The cerebral palsy model was established by ligating the left common carotid artery followed by exposure to hypox- ia. Phosphate-buffered saline, vascular endothelial growth factor + neural stem cells, and neural stem cells alone were administered into the sensorimotor cortex using the stereotaxic instrument and microsyringe. After transplantation, the radial-arm water maze test and holding test were performed. Immunohistochemistry for vascular endothelial growth factor and histology using hematoxylin-eosin were performed on cerebral cortex. Results revealed that the number of vas- cular endothelial growth factor-positive cells in cerebral palsy rats transplanted with vascular endothelial growth factor-transfected neural stem cells was increased, the time for finding water and the finding repetitions were reduced, the holding time was prolonged, and the degree of cell degeneration or necrosis was reduced. These findings indicate that the transplantation of vascu- lar endothelial growth factor-transfected neural stem cells alleviates brain damage and cognitive deficits, and is neuroprotective in neonatal rats with hypoxia ischemic-mediated cerebral palsy.展开更多
Summary: The effects of different concentrations of vascular endothelial cell growth factor (VEGF) on the fibrovascular ingrowth into rabbits hydroxyapatite orbital implant were investigated. Twelve New Zealand white ...Summary: The effects of different concentrations of vascular endothelial cell growth factor (VEGF) on the fibrovascular ingrowth into rabbits hydroxyapatite orbital implant were investigated. Twelve New Zealand white rabbits were divided into 3 groups and received hydroxyapatite orbital implant surgery in their right eyes. Before and after the operation, the implants were treated with 10 ng/ml VEGF, 100 ng/ml VEGF, or normal saline as control group. The animals received technetium bones scan at 2, 4, and 6 weeks postoperatively. The mean radioactivity counts within region of interest (ROI) of the surgery eye (R) and the non-surgery eye (L) in the same animal were tested, and the R/L ratios were calculated. The implants were harvested at 6th weeks and examined histopathologically. The results showed that at second week, there was no significant difference in mean R/L ratios between VEGF group and control group (F=2.83, P=0.111); At 4th week (F=7.728, P=0.011) and 6th week (F=7.831, P=0.011) postoperatively, the mean ratios in VEGF groups were significantly higher than that in control group. At 6th week postoperatively, the fibrovascularization rates in VEGF groups were higher than in control group significantly (F=8.711, P=0.008). It was suggested that VEGF could promote the fibrovascular ingrowth into hydroxyapatite orbital implant, thus might shorten the time required for complete vascularization of the HA orbital implant.展开更多
AIM:To investigate the effects of luteolin on apoptosis,the cell cycle,and the expression and secretion of vascular endothelial growth factor(VEGF)in human choroidal melanoma cells(C918 and OCM-1).METHODS:C918 and OCM...AIM:To investigate the effects of luteolin on apoptosis,the cell cycle,and the expression and secretion of vascular endothelial growth factor(VEGF)in human choroidal melanoma cells(C918 and OCM-1).METHODS:C918 and OCM-1 cells cultured in vitro were treated with various concentrations of luteolin(0,5,10,15μmol/L).Cell growth was observed with an inverted microscope,and cell cycle arrest was detected by propidium iodide(PI)staining using flow cytometry.Apoptosis was detected by Hoechst33342 staining,and apoptosis rate was determined by Annexin V-FITC/PI experiments using flow cytometry.The expression of apoptosis-related proteins Bcl-2,Bax and VEGF was analyzed using Western blots.The levels of VEGF secreted by the cells into the supernatant was analyzed using ELISA.RESULTS:After treating with 5 to 15μmol/L luteolin for 48 h,the fusion degree of C918 and OCM-1 cells decreased,and more floating apoptotic cells appeared.Luteolin treatment increased the G0-G1 phase ratio of the C918 and OCM-1 cells,blocked cell cycle progression,and increased the apoptosis rate of the C918 and OCM-1 cells.Western blot showed that luteolin decreased the expression of Bcl-2 and VEGF in the C918 and OCM-1 cells and increased the expression of Bax protein.The ELISA results showed that 10 to 15μmol/L luteolin decreased the cell secretion of VEGF.CONCLUSION:Luteolin may induce apoptosis by regulating the levels of apoptosis-related proteins in C918 and OCM-1 cells.Luteolin can induce cell cycle arrest,decrease the expression of VEGF.展开更多
AIM:To characterize the implications of vascular endothelial growth factor(VEGF)-A in stromal cells and colorectal cancer and the expression of VEGF-A splice variants.METHODS:VEGF-A expression in tumor and stromal cel...AIM:To characterize the implications of vascular endothelial growth factor(VEGF)-A in stromal cells and colorectal cancer and the expression of VEGF-A splice variants.METHODS:VEGF-A expression in tumor and stromal cells from 165 consecutive patients with colorectal cancer was examined by immunohistochemistry.The association between VEGF-A expression status and clinicopathological factors was investigated.Twenty freshfrozen samples were obtained for laser capture microdissection to analyze the splice variants of VEGF-A.RESULTS:VEGF-A was expressed in 53.9% and 42.4% of tumor and stromal cells,respectively.VEGF-A expression in tumor cells(t-VEGF-A) was associated with advanced clinical stage(stage 0,1/9;stage 1,2/16;stage 2,32/55;stage 3,38/66;stage 4,16/19,P < 0.0001).VEGF-A expression in stromal cells(s-VEGF-A) increased in the earlier clinical stage(stage 0,7/9;stage 1,6/16;stage 2,33/55;stage 3,22/66;stage 4,5/19;P = 0.004).Multivariate analyses for risk factors of recurrence showed that only s-VEGF-A expression was an independent risk factor for recurrence(relative risk 0.309,95% confidence interval 0.141-0.676,P = 0.0033).The five-year disease-free survival(DFS) rates of t-VEGF-A-positive and-negative cases were 51.4% and 62.9%,respectively.There was no significant difference in t-VEGF-A expression status.The five-year DFS rates of s-VEGF-A-positive and-negative cases were 73.8% and 39.9%,respectively.s-VEGFA-positive cases had significantly better survival than s-VEGF-A-negative cases(P = 0.0005).Splice variant analysis revealed that t-VEGF-A was mainly composed of VEGF165 and that s-VEGF-A included both VEGF165 and VEGF165b.In cases with no venous invasion(v0),the level of VEGF165b mRNA was significantly higher(v0 204.5 ± 122.7,v1 32.5 ± 36.7,v2 2.1 ± 1.7,P = 0.03).The microvessel density tended to be lower in cases with higher VEGF165b mRNA levels.CONCLUSION:s-VEGF-A appears be a good prognostic factor for colorectal cancer and includes VEGF165 and VEGF165b.展开更多
This study examined the effect of resveratrol on the secretion of vascular endothelial growth factor (VEGF) and subsequent proliferation of human leukemia U937 cells, and explored the mechanisms involved. Human leuk...This study examined the effect of resveratrol on the secretion of vascular endothelial growth factor (VEGF) and subsequent proliferation of human leukemia U937 cells, and explored the mechanisms involved. Human leukemia U937 cells were treated with resveratrol of different concentrations (12.5-200 μmol/L) for different time lengths (12-48 h). The proliferation of the U937 leukemic cells was determined by MTT assay. Apoptosis was observed by Annexin- V-FIFC/PI double staining and flow cytometry (FCM). Cells cycle was analyzed by PI staining and FCM. The content of VEGF was determined by ELISA. Human umbibical vein endothelial cells were examined for vasoformation in vitro after exposures to resveratrol of various concetrations. The results showed that resveratrol inhibited the proliferation of U937 leukemia cells in a dose- and time-dependent manner. Resveratrol induced apoptosis and S-phase cell cycle arrest in human leukemic U937 cells. Resveratrol inhibited the secretion of VEGF in U937 cells. Resveratrol inhibited the vasoformation of human vein endothelial cells in a dose-dependent manner. It was concluded that resveratrol could down-regulate the secretion of VEGE induce apoptosis and suppress the proliferation of U937 cells.展开更多
The effect of vascular endothelial growth factor (VEGF) overexpression on matrix metalloproteinase-2 (MMP-2) in nasopharyngeal carcinoma (NPC) cells in vitro and the possible mechanism involved were investigated...The effect of vascular endothelial growth factor (VEGF) overexpression on matrix metalloproteinase-2 (MMP-2) in nasopharyngeal carcinoma (NPC) cells in vitro and the possible mechanism involved were investigated, and the correlation between the expression of VEGF and MMP-2 in NPC evaluated. The NPC cells were transfected with PAd-trackVEGF165 plasmid. The expression levels of VEGF and MMP-2 mRNA and protein in NPC cells were detected by semi-quantitative RT-PCR and Western blot respectively. It was found that the expression of VEGF and MMP-2 mRNA and protein was significantly increased in NPC cells after transfection of VEGF 165. It was concluded that the expression of VEGF was correlated to the in vitro invasion of NPC cells, and the induction of MMP-2 by VEGF was a key process of NPC cell invasion.展开更多
To investigate the effect of meloxicam, a selected NSAIDs, on cell growth, expression of VEGF and angiopointin-2 (Ang-2) protein in HT-29 cell line, cultured HT-29 cells were treated with meloxicam of various concen...To investigate the effect of meloxicam, a selected NSAIDs, on cell growth, expression of VEGF and angiopointin-2 (Ang-2) protein in HT-29 cell line, cultured HT-29 cells were treated with meloxicam of various concentrations for various lengths of time. The proliferation of HT-29 was detected by cell counting kit-8 (CCK-8), the cell cycle was determined by flow cytometer and the levels of VEGF and Ang-2 protein in supernatants were examined by enzyme linked immunosorbent assay (ELISA). The mRNA expressions of VEGF and Ang-2 in cultured HT-29 were determined by real-time quantitative reverse-transcription polymerase chain reaction. Our results showed that treatment of meloxicam of different concentrations and for various lengths of time had a cytotoxicic effect on the cell proliferation of HT-29 cells in a concentration-dependant and time-dependant manner. Cell cycle analysis showed that the cells were mainly blocked in G0/G1 phase. The VEGF and Ang-2 protein levels in supernatants of the culture medium were decreased gradually in a concentration-dependent or time-dependent fashion. The mRNA expression of cox-2, VEGF and Ang-2 showed a gradual and concentration-dependent reduction. It is concluded that meloxicam can reduce the expression of VEGF and Ang-2 at the protein and mRNA level in colon carcinoma cell line.展开更多
AIM To test the hypothesis to block VEGFexpression of SMMC-7721 hepatoma cells mayinhibit tumor growth using the rat hepatomamodel.METHODS Amplifiy the 200 VEGF cDNAfragment and insert it into human U6 genecassette in...AIM To test the hypothesis to block VEGFexpression of SMMC-7721 hepatoma cells mayinhibit tumor growth using the rat hepatomamodel.METHODS Amplifiy the 200 VEGF cDNAfragment and insert it into human U6 genecassette in the reverse orientation transcribingsmall antisense RNA which could specificallyinteract with VEGF165, and VEGF121 mRNA.Construct the retroviral vector containing thisantisense VEGF U6 cassette and package thereplication-deficient recombinant retrovirus.SMMC-7721 cells were transduced with thesevirus and positive clones were selected withG418. PCR and Southern blot analysis wereperformed to determine if U6 cassette integratedinto the genomic DNA of positive clone.Transfected tumor cells were evaluated for RNAexpression by ribonuclease protection assays.The VEGF protein in the supernatant of parentaltumor cells and genetically modified tumor cellswas determined with ELISA. In vitro and in vivogrowth properties of antisense VEGF cell clonein nude mice were analyzed.RESULTS Restriction enzyme digestion andPCR sequencing verified that the antisense VEGFRNA retroviral vector was successfullyconstructed. After G418 selection, resistantSMMC-7721 cell clone was picked up. PCR andSouthern blot analysis suggested that U6cassette was integrated into the cell genomicDNA. Stable SMMC-7721 cell clone transducedwith U6 antisense RNA cassette could express200bp small antisense VEGF RNA and secretereduced levels of VEGF in culture condition.Production of VEGF by antisense transgeneexpressing cells was 65 ± 10 ng / L per 106 cells,420 ± 45 ng/L per 106 cells in sense group and 485± 30 ng/L per 106 cells in the negative control group, (P<0.05). The antisense-VEGF cell clone appeared phenotypically indistinguishable from SMMC-7721 cells and SMMC-7721 cells transfected sense VEGF. The growth rate of the antisense-VEGF cell clone was the same as the control cells. When S. C. was implanted into nude mice, growth of antisense-VEGF cell lines was greatly inhibited compared with control cells.CONCLUSION Expression of antisense VEGFRNA in SMMC-7721 cells could decrease thetumorigenicity, and antisense-VEGF genetherapy may be an adjuvant treatment forhepatoma.展开更多
AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endo...AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endothelial cells.METHODS: The KDR-TK fragment was extracted from pBluescript Ⅱ KDR-TK plasmid by enzymatic digestion with Xho I and Sal I. The enhanced green fluorescence protein (EGFP) carrier was extracted from pEGFP by the same procedure. The KDR-TK was inserted into the pEGFP carrier to construct pEGFP-KDR-TK. Using ultrasound irradiation and microbubble, pEGFP-KDR-TK was transferred into human umbilical vein endothelial cells (HUVECs). The transient infection rate was estimated by green fluorescent protein (GFP) expression. Transfected HUVECs, non-transfected HUVECs, and HepG2 cells were cultured in the presence of different concentrations of ganciclovir (GCV), and the killing efficacy of HSV-TK/GCV was analyzed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: The recombinant pEGFP-KDR-TK was successfully constructed by inserting the KDR-TK fragment into the pEGFP carrier. Transfected HUVECs showed cytoplasmic green fluorescence, and the transient transfection rate was about 20.3%. Pools of G418-resistant cells exhibited a higher sensitivity to theprodrug/GCV compared to non-transfected HUVECs or non-transfected HepG2 cells, respectively. CONCLUSION: KDR promoter and the suicide gene/prodrug system mediated by diagnostic ultrasound combined with microbubble can significantly kill HUVECs. Such therapy may present a novel and attractive approach to target gene therapy on tumor vessels.展开更多
We constructed a lentiviral vector carrying vascular endothelial growth factor 165, which was used to transfect neural stem cells. The transfection rate was approximately 50%, as determined by flow cytometry. Vascular...We constructed a lentiviral vector carrying vascular endothelial growth factor 165, which was used to transfect neural stem cells. The transfection rate was approximately 50%, as determined by flow cytometry. Vascular endothelial growth factor protein was detected in neural stem cells and promoted proliferation.展开更多
基金supported by Science and Technology Research Project of Jilin Provincial Department of Education,No.JJKH20220072KJ(to XL)Science and Technology Development Program of Jilin Province,No.20200201495JC(to YL)。
文摘The integrity of retinal ganglion cells is tightly associated with diabetic macular degeneration that leads to damage and death of retinal ganglion cells,affecting vision.The major clinical treatments for diabetic macular edema are anti-vascular endothelial growth factor drugs and laser photocoagulation.However,although the macular thickness can be normalized with each of these two therapies used alone,the vision does not improve in many patients.This might result from the incomplete recovery of retinal ganglion cell injury.Therefore,a prospective,non-randomized,controlled clinical trial was designed to investigate the effect of anti-vascular endothelial growth factor drugs combined with laser photocoagulation on the integrity of retinal ganglion cells in patients with diabetic macular edema and its relationship with vision recovery.In this trial,150 patients with diabetic macular edema will be equally divided into three groups according to therapeutic methods,followed by treatment with anti-vascular endothelial growth factor drugs,laser photocoagulation therapy,and their combination.All patients will be followed up for 12 months.The primary outcome measure is retinal ganglion cell-inner plexiform layer thickness at 12 months after treatment.The secondary outcome measures include retinal ganglion cell-inner plexiform layer thickness before and 1,3,6,and 9 months after treatment,retinal nerve fiber layer thickness,best-corrected visual acuity,macular area thickness,and choroidal thickness before and 1,3,6,9,and 12 months after treatment.Safety measure is the incidence of adverse events at 1,3,6,9,and 12 months after treatment.The study protocol hopes to validate the better efficacy and safety of the combined treatment in patients with diabetic macula compared with the other two monotherapies alone during the 12-month follow-up period.The trial is designed to focus on clarifying the time-effect relationship between imaging measures related to the integrity of retinal ganglion cells and best-corrected visual acuity.The trial protocol was approved by the Medical Ethics Committee of the Affiliated Hospital of Beihua University with approval No.(2023)(26)on April 25,2023,and was registered with the Chinese Clinical Trial Registry(registration number:ChiCTR2300072478,June 14,2023,protocol version:2.0).
基金supported by Key Research and Development Plan of Xuzhou Science and Technology Bureau,No.KC21162(to XMZ)a grant from Jiangsu Key Laboratory of Brain Disease Bioinformationg,No.XZSYSKF2021018(to XMZ)+1 种基金Natural Science Fund for Colleges and Universities in Jiangsu Province,No.19KJB320024(to HNY)the Science and Technology Development Fund from Affiliated Hospital of Xuzhou Medical University,Nos.XYFM2021024(to XMZ),XYFM2021006(to DH).
文摘Although bone marrow mesenchymal stem cells(BMSCs)might have therapeutic potency in ischemic stroke,the benefits are limited.The current study investigated the effects of BMSCs engineered to overexpress vascular endothelial growth factor(VEGF)on behavioral defects in a rat model of transient cerebral ischemia,which was induced by middle cerebral artery occlusion.VEGF-BMSCs or control grafts were injected into the left striatum of the infarcted hemisphere 24 hours after stroke.We found that compared with the stroke-only group and the vehicle-and BMSCs-control groups,the VEGF-BMSCs treated animals displayed the largest benefits,as evidenced by attenuated behavioral defects and smaller infarct volume 7 days after stroke.Additionally,VEGF-BMSCs greatly inhibited destruction of the blood-brain barrier,increased the regeneration of blood vessels in the region of ischemic penumbra,and reducedneuronal degeneration surrounding the infarct core.Further mechanistic studies showed that among all transplant groups,VEGF-BMSCs transplantation induced the highest level of brain-derived neurotrophic factor.These results suggest that BMSCs transplantation with vascular endothelial growth factor has the potential to treat ischemic stroke with better results than are currently available.
基金supported by NIH grant RO1 NS093985 (to DS, NZ, XW) and RO1 NS101955 (to DS)the VCU Microscopy Facility,supported,in part,by funding from NIH-NCI Cancer Center Support Grant P30 CA016059。
文摘Neovascularization and angiogenesis in the brain are important physiological processes for normal brain development and repair/regeneration following insults. Integrins are cell surface adhesion receptors mediating important function of cells such as survival, growth and development during tissue organization, differentiation and organogenesis. In this study, we used an integrin-binding array platform to identify the important types of integrins and their binding peptides that facilitate adhesion, growth, development, and vascular-like network formation of rat primary brain microvascular endothelial cells. Brain microvascular endothelial cells were isolated from rat brain on post-natal day 7. Cells were cultured in a custom-designed integrin array system containing short synthetic peptides binding to 16 types of integrins commonly expressed on cells in vertebrates. After 7 days of culture, the brain microvascular endothelial cells were processed for immunostaining with markers for endothelial cells including von Willibrand factor and platelet endothelial cell adhesion molecule. 5-Bromo-2′-dexoyuridine was added to the culture at 48 hours prior to fixation to assess cell proliferation. Among 16 integrins tested, we found that α5β1, αvβ5 and αvβ8 greatly promoted proliferation of endothelial cells in culture. To investigate the effect of integrin-binding peptides in promoting neovascularization and angiogenesis, the binding peptides to the above three types of integrins were immobilized to our custom-designed hydrogel in three-dimensional(3 D) culture of brain microvascular endothelial cells with the addition of vascular endothelial growth factor. Following a 7-day 3 D culture, the culture was fixed and processed for double labeling of phalloidin with von Willibrand factor or platelet endothelial cell adhesion molecule and assessed under confocal microscopy. In the 3 D culture in hydrogels conjugated with the integrin-binding peptide, brain microvascular endothelial cells formed interconnected vascular-like network with clearly discernable lumens, which is reminiscent of brain microvascular network in vivo. With the novel integrin-binding array system, we identified the specific types of integrins on brain microvascular endothelial cells that mediate cell adhesion and growth followed by functionalizing a 3 D hydrogel culture system using the binding peptides that specifically bind to the identified integrins, leading to robust growth and lumenized microvascular-like network formation of brain microvascular endothelial cells in 3 D culture. This technology can be used for in vitro and in vivo vascularization of transplants or brain lesions to promote brain tissue regeneration following neurological insults.
基金supported by the following funds:1.Medical Scientific Research Foundation of Guangdong Province(A2022221)Natural Science Foundation of Guangdong Province(2019A1515011417)+2 种基金National Natural Science Foundation of China(81970806,82271094)Science and Technology Projects in Guangzhou(202201020030,202201020015)Guangdong High-Level Hospital Construction Fund(ynkt2021-zz16).
文摘Objective:This study aimed to investigate the potential of human umbilical cord mesenchymal stem cell(hucMSC)-derived exosomes(hucMSC-Exos)in inhibiting hypoxia-induced cell hyper proliferation and overexpression of vascular endothelial growth factor A(VEGF-A)in immature human fetal retinal microvascular endothelial cells(hfRMECs).Methods:Exosomes were isolated from hucMSCs using cryogenic ultracentrifugation and characterized through various techniques,including transmission electron microscopy,nanoparticle tracking analysis,bicinchoninic acid assays,and western blotting.The hfRMECs were identified using von Willebrand factor(vWF)co-staining and divided into four groups:a control group cultured under normoxic condition,a hypoxic model group,a hypoxic group treated with low-concentration hucMSC-Exos(75μg/mL)and a hypoxic group treated with high-concentration hucMSC-Exos(100μg/mL).Cell viability and proliferation were assessed using Cell Counting Kit-8(CCK-8)assay and EdU(5-ethynyl-2′-deoxyuridine)assay respectively.Expression levels of VEGF-A were evaluated using RT-PCR,western blotting and immunofluorescence.Results:Hypoxia significantly increased hfRMECs’viability and proliferation by upregulating VEGF-A levels.The administration of hucMSC-Exos effectively reversed this response,with the high-concentration group exhibiting greater efficacy compared to the lowconcentration group.Conclusion:In conclusion,hucMSC-Exos can dose-dependently inhibit hypoxia-induced hyperproliferation and VEGF-A overexpression in immature fetal retinal microvascular endothelial cells.
基金Supported by the Shaanxi Province Traditional Chinese Medicine Project(No.SZY-KJCYC-2023-028)。
文摘AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the retinal tissue in diabetic rats.METHODS:The expression of HDAC7 in HRMECs under high glucose and the retinal tissue from normal or diabetic rats were detected with immunohistochemistry and Western blot.LV-shHDAC7 HRMECs were used to study the effect of HDAC7 on cell activities.Cell count kit-8(CCK-8),5-ethynyl2’-deoxyuridine(EdU),flow cytometry,scratch test,Transwell test and tube formation assay were used to examine the ability of cell proliferation,migration,and angiogenesis.Finally,a preliminary exploration of its mechanism was performed by Western blot.RESULTS:The expression of HDAC7 was both upregulated in retinal tissues of diabetic rats and high glucosetreated HRMECs.Down-regulation of HDAC7 expression significantly reduced the ability of proliferation,migration,and tube formation,and reversed the high glucose-induced high expression of CDK1/Cyclin B1 and vascular endothelial growth factor in high glucose-treated HRMECs.CONCLUSION:High glucose can up-regulate the expression of HDAC7 in HRMECs.Down-regulation of HDAC7 can inhibit HRMECs activities.HDAC7 is proposed to be involved in pathogenesis of diabetic retinopathy and a therapeutic target.
基金supported by the National Research Foundation of Korea Grant funded by the Korean Government,No.NRF-013-2011-1-E00045
文摘Endogenous neural stem cells become "activated" after neuronal injury, but the activation sequence and fate of endogenous neural stem cells in focal cerebral ischemia model are little known. We evaluated the relationships between neural stem cells and hypoxia-inducible factor-1α and vascular endothelial growth factor expression in a photothromobotic rat stroke model using immunohistochemistry and western blot analysis. We also evaluated the chronological changes of neural stem cells by 5-bromo-2′-deoxyuridine(BrdU) incorporation. Hypoxia-inducible factor-1α expression was initially increased from 1 hour after ischemic injury, followed by vascular endothelial growth factor expression. Hypoxia-inducible factor-1α immunoreactivity was detected in the ipsilateral cortical neurons of the infarct core and peri-infarct area. Vascular endothelial growth factor immunoreactivity was detected in bilateral cortex, but ipsilateral cortex staining intensity and numbers were greater than the contralateral cortex. Vascular endothelial growth factor immunoreactive cells were easily found along the peri-infarct area 12 hours after focal cerebral ischemia. The expression of nestin increased throughout the microvasculature in the ischemic core and the peri-infarct area in all experimental rats after 24 hours of ischemic injury. Nestin immunoreactivity increased in the subventricular zone during 12 hours to 3 days, and prominently increased in the ipsilateral cortex between 3–7 days. Nestin-labeled cells showed dual differentiation with microvessels near the infarct core and reactive astrocytes in the peri-infarct area. BrdU-labeled cells were increased gradually from day 1 in the ipsilateral subventricular zone and cortex, and numerous BrdU-labeled cells were observed in the peri-infarct area and non-lesioned cortex at 3 days. BrdU-labeled cells rather than neurons, were mainly co-labeled with nestin and GFAP. Early expressions of hypoxia-inducible factor-1α and vascular endothelial growth factor after ischemia made up the microenvironment to increase the neuronal plasticity of activated endogenous neural stem cells. Moreover, neural precursor cells after large-scale cortical injury could be recruited from the cortex nearby infarct core and subventricular zone.
文摘Hypoxia and transforming growth factor-β1 (TGF-β1) increase vascular endothelial growth factor A (VEGFA) expression in a number of malignancies. This effect of hypoxia and TGF-β1 might be responsible for tumor progression and metastasis of advanced prostate cancer. In the present study, TGF-β1 was shown to induce VEGFA165 secretion from both normal cell lines (HPV7 and RWPE1) and prostate cancer cell lines (DU 145 and PC3). Conversely, hypoxia-stimulated VEGFA165 secretion was observed only in prostate cancer cell lines. Hypoxia induced TGF-β1 expression in PC3 prostate cancer cells, and the TGF-β1 type I receptor (ALK5) kinase inhibitor partially blocked hypoxia-mediated VEGFA16s secretion. This effect of hypoxia provides a novel mechanism to increase VEGFA expression in prostate cancer cells. Although autocrine signaling of VEGFA has been implicated in prostate cancer progression and metastasis, the associated mechanism is poorly characterized. VEGFA activity is mediated via VEGF receptor (VEGFR) 1 (Fit-l) and 2 (FIk-I/KDR). Whereas VEGFR-1 mRNA was detected in normal prostate epithelial cells, VEGFR-2 mRNA and VEGFR protein were expressed only in PC3 cells. VEGFA165 treatment induced phosphorylation of extracellular signal-regulated kinase 1/2 (ERKI/2) in PC3 cells but not in HPV7 cells, suggesting that the autocrine function of VEGFA may be uniquely associated with prostate cancer. Activation of VEGFR-2 by VEGFA165 was shown to enhance migration of PC3 cells. A similar effect was also observed with endogenous VEGFA induced by TGF-β1 and hypoxia. These findings illustrate that an autocrine loop of VEGFA via VEGFR-2 is critical for the tumorigenic effects of TGF-β1 and hypoxia on metastatic prostate cancers.
文摘Objective: To investigate the effect of Fuzhenghuayu decoction on autocrine activation of hepatic stellate cell (HSC). Methods: The drug serum containing Fuzhenghuayu decoction was collected from normal rats, and cul- tured with activated HSC in vitro. The conditioned medium from the drug serum treated HSC was added to primary cultured quiescent HSC. Cell prolifera- tion was assayed by tetrazolium colorimetric test, and the contents of type Ⅰ collagen and vascular endo- thelial growth factor (VEGF) in the supernatant were measured with ELISA. Results: The conditioned medium from activated HSC could stimulate the quiescent HSC proliferation and type Ⅰ collagen secretion. The drug serum inhibi- ted this stimulating action and VEGF secretion from the activated HSC. Conclusion: Fuzhenghuayu decoction acts effectively against the autocrine activation pathway of HSC. The mechanism may be associated with the inhibition of the secretion of VEGF by activated HSC.
基金supported by the National Natural Science Foundation of China,No.81070523 and 81270728
文摘Neural stem/progenitor cell (NSC) transplantation has been shown to effectively improve neurological function in rats with hypoxic-isch- emic brain damage. Vascular endothelial growth factor (VEGF) is a signaling protein that stimulates angiogenesis and improves neural regeneration. We hypothesized that transplantation of VEGF-transfected NSCs would alleviate hypoxic-ischemic brain damage in neo- natal rats. We produced and transfected a recombinant lentiviral vector containing the VEGF165gene into cultured NSCs. The transfected NSCs were transplanted into the left sensorimotor cortex of rats 3 days after hypoxic-ischemic brain damage. Compared with the NSCs group, VEGF mRNA and protein expression levels were increased in the transgene NSCs group, and learning and memory abilities were significantly improved at 30 days. Furthermore, histopathological changes were alleviated in these animals. Our findings indicate that transplantation of VEGF-transfected NSCs may facilitate the recovery of neurological function, and that its therapeutic effectiveness is better than that of unmodified NSCs.
基金supported by the program (No. CX10B_421Z to Jiaxin Ye) for Postgraduate Research Innovation in Universities of Jiangsu Provincethe grants (No. 81070195) and (No. 81000055) from Chinese National Science Fund of China (all to Biao Xu)grant (No.KF200938 to Lina Kang) from Jiangsu Province
文摘This study was designed to determine the levels of early endothelial progenitor cells (EPCs), apelin, vascu- lar endothelial growth factor (VEGF) and stromal cell-derived growth factor-1 (SDF-1) after acute myocardial infarction (AMI), and to investigate the relationships between these cytokines and early EPCs. Early EPCs, de- fined as CD133+, KDR+, and CD34~ cells, were quantified by flow cytometry. The levels of early EPCs and those cytokines in AMI patients were significantly different from those with coronary artery disease or controls (P 〈 0.05). Plasma apelin levels were inversely correlated with Gensini score and early EPCs (both P 〈 0.01). Early EPCs, VEGF and SDF-1 showed different patterns of changes in AMI patients during the first 24 h. The trend in the change of early EPCs was proportionally correlated with that of VEGF (P 〈 0.05). AMI patients exhibited in- creased early EPCs with remarkably decreased apelin levels and enhanced VEGF levels.
基金supported by grants from the National Natural Science Foundation of China,No.81070523,81270728
文摘Neural stem cells are characterized by the ability to differentiate and stably express exogenous ge- nes. Vascular endothelial growth factor plays a role in protecting local blood vessels and neurons of newborn rats with hypoxic-ischemic encephalopathy. Transplantation of vascular endothelial growth factor-transfected neural stem cells may be neuroprotective in rats with cerebral palsy. In this study, 7-day-old Sprague-Dawley rats were divided into five groups: (1) sham operation (control), (2) cerebral palsy model alone or with (3) phosphate-buffered saline, (4) vascular en- dothelial growth factor 165 + neural stem cells, or (5) neural stem cells alone. The cerebral palsy model was established by ligating the left common carotid artery followed by exposure to hypox- ia. Phosphate-buffered saline, vascular endothelial growth factor + neural stem cells, and neural stem cells alone were administered into the sensorimotor cortex using the stereotaxic instrument and microsyringe. After transplantation, the radial-arm water maze test and holding test were performed. Immunohistochemistry for vascular endothelial growth factor and histology using hematoxylin-eosin were performed on cerebral cortex. Results revealed that the number of vas- cular endothelial growth factor-positive cells in cerebral palsy rats transplanted with vascular endothelial growth factor-transfected neural stem cells was increased, the time for finding water and the finding repetitions were reduced, the holding time was prolonged, and the degree of cell degeneration or necrosis was reduced. These findings indicate that the transplantation of vascu- lar endothelial growth factor-transfected neural stem cells alleviates brain damage and cognitive deficits, and is neuroprotective in neonatal rats with hypoxia ischemic-mediated cerebral palsy.
文摘Summary: The effects of different concentrations of vascular endothelial cell growth factor (VEGF) on the fibrovascular ingrowth into rabbits hydroxyapatite orbital implant were investigated. Twelve New Zealand white rabbits were divided into 3 groups and received hydroxyapatite orbital implant surgery in their right eyes. Before and after the operation, the implants were treated with 10 ng/ml VEGF, 100 ng/ml VEGF, or normal saline as control group. The animals received technetium bones scan at 2, 4, and 6 weeks postoperatively. The mean radioactivity counts within region of interest (ROI) of the surgery eye (R) and the non-surgery eye (L) in the same animal were tested, and the R/L ratios were calculated. The implants were harvested at 6th weeks and examined histopathologically. The results showed that at second week, there was no significant difference in mean R/L ratios between VEGF group and control group (F=2.83, P=0.111); At 4th week (F=7.728, P=0.011) and 6th week (F=7.831, P=0.011) postoperatively, the mean ratios in VEGF groups were significantly higher than that in control group. At 6th week postoperatively, the fibrovascularization rates in VEGF groups were higher than in control group significantly (F=8.711, P=0.008). It was suggested that VEGF could promote the fibrovascular ingrowth into hydroxyapatite orbital implant, thus might shorten the time required for complete vascularization of the HA orbital implant.
文摘AIM:To investigate the effects of luteolin on apoptosis,the cell cycle,and the expression and secretion of vascular endothelial growth factor(VEGF)in human choroidal melanoma cells(C918 and OCM-1).METHODS:C918 and OCM-1 cells cultured in vitro were treated with various concentrations of luteolin(0,5,10,15μmol/L).Cell growth was observed with an inverted microscope,and cell cycle arrest was detected by propidium iodide(PI)staining using flow cytometry.Apoptosis was detected by Hoechst33342 staining,and apoptosis rate was determined by Annexin V-FITC/PI experiments using flow cytometry.The expression of apoptosis-related proteins Bcl-2,Bax and VEGF was analyzed using Western blots.The levels of VEGF secreted by the cells into the supernatant was analyzed using ELISA.RESULTS:After treating with 5 to 15μmol/L luteolin for 48 h,the fusion degree of C918 and OCM-1 cells decreased,and more floating apoptotic cells appeared.Luteolin treatment increased the G0-G1 phase ratio of the C918 and OCM-1 cells,blocked cell cycle progression,and increased the apoptosis rate of the C918 and OCM-1 cells.Western blot showed that luteolin decreased the expression of Bcl-2 and VEGF in the C918 and OCM-1 cells and increased the expression of Bax protein.The ELISA results showed that 10 to 15μmol/L luteolin decreased the cell secretion of VEGF.CONCLUSION:Luteolin may induce apoptosis by regulating the levels of apoptosis-related proteins in C918 and OCM-1 cells.Luteolin can induce cell cycle arrest,decrease the expression of VEGF.
文摘AIM:To characterize the implications of vascular endothelial growth factor(VEGF)-A in stromal cells and colorectal cancer and the expression of VEGF-A splice variants.METHODS:VEGF-A expression in tumor and stromal cells from 165 consecutive patients with colorectal cancer was examined by immunohistochemistry.The association between VEGF-A expression status and clinicopathological factors was investigated.Twenty freshfrozen samples were obtained for laser capture microdissection to analyze the splice variants of VEGF-A.RESULTS:VEGF-A was expressed in 53.9% and 42.4% of tumor and stromal cells,respectively.VEGF-A expression in tumor cells(t-VEGF-A) was associated with advanced clinical stage(stage 0,1/9;stage 1,2/16;stage 2,32/55;stage 3,38/66;stage 4,16/19,P < 0.0001).VEGF-A expression in stromal cells(s-VEGF-A) increased in the earlier clinical stage(stage 0,7/9;stage 1,6/16;stage 2,33/55;stage 3,22/66;stage 4,5/19;P = 0.004).Multivariate analyses for risk factors of recurrence showed that only s-VEGF-A expression was an independent risk factor for recurrence(relative risk 0.309,95% confidence interval 0.141-0.676,P = 0.0033).The five-year disease-free survival(DFS) rates of t-VEGF-A-positive and-negative cases were 51.4% and 62.9%,respectively.There was no significant difference in t-VEGF-A expression status.The five-year DFS rates of s-VEGF-A-positive and-negative cases were 73.8% and 39.9%,respectively.s-VEGFA-positive cases had significantly better survival than s-VEGF-A-negative cases(P = 0.0005).Splice variant analysis revealed that t-VEGF-A was mainly composed of VEGF165 and that s-VEGF-A included both VEGF165 and VEGF165b.In cases with no venous invasion(v0),the level of VEGF165b mRNA was significantly higher(v0 204.5 ± 122.7,v1 32.5 ± 36.7,v2 2.1 ± 1.7,P = 0.03).The microvessel density tended to be lower in cases with higher VEGF165b mRNA levels.CONCLUSION:s-VEGF-A appears be a good prognostic factor for colorectal cancer and includes VEGF165 and VEGF165b.
基金This project was supported a grant from the National Natural Science Foundation of China (No 30572440)
文摘This study examined the effect of resveratrol on the secretion of vascular endothelial growth factor (VEGF) and subsequent proliferation of human leukemia U937 cells, and explored the mechanisms involved. Human leukemia U937 cells were treated with resveratrol of different concentrations (12.5-200 μmol/L) for different time lengths (12-48 h). The proliferation of the U937 leukemic cells was determined by MTT assay. Apoptosis was observed by Annexin- V-FIFC/PI double staining and flow cytometry (FCM). Cells cycle was analyzed by PI staining and FCM. The content of VEGF was determined by ELISA. Human umbibical vein endothelial cells were examined for vasoformation in vitro after exposures to resveratrol of various concetrations. The results showed that resveratrol inhibited the proliferation of U937 leukemia cells in a dose- and time-dependent manner. Resveratrol induced apoptosis and S-phase cell cycle arrest in human leukemic U937 cells. Resveratrol inhibited the secretion of VEGF in U937 cells. Resveratrol inhibited the vasoformation of human vein endothelial cells in a dose-dependent manner. It was concluded that resveratrol could down-regulate the secretion of VEGE induce apoptosis and suppress the proliferation of U937 cells.
基金This project was supported by grants from National Excellent Young Scientists Foundation of China (No.39925035) the Major Clinical Project of Ministry of Health (No.22012332).
文摘The effect of vascular endothelial growth factor (VEGF) overexpression on matrix metalloproteinase-2 (MMP-2) in nasopharyngeal carcinoma (NPC) cells in vitro and the possible mechanism involved were investigated, and the correlation between the expression of VEGF and MMP-2 in NPC evaluated. The NPC cells were transfected with PAd-trackVEGF165 plasmid. The expression levels of VEGF and MMP-2 mRNA and protein in NPC cells were detected by semi-quantitative RT-PCR and Western blot respectively. It was found that the expression of VEGF and MMP-2 mRNA and protein was significantly increased in NPC cells after transfection of VEGF 165. It was concluded that the expression of VEGF was correlated to the in vitro invasion of NPC cells, and the induction of MMP-2 by VEGF was a key process of NPC cell invasion.
基金This project was supported by a grant from R&D program of Hubei Provincial government (No 2005AA304B09)
文摘To investigate the effect of meloxicam, a selected NSAIDs, on cell growth, expression of VEGF and angiopointin-2 (Ang-2) protein in HT-29 cell line, cultured HT-29 cells were treated with meloxicam of various concentrations for various lengths of time. The proliferation of HT-29 was detected by cell counting kit-8 (CCK-8), the cell cycle was determined by flow cytometer and the levels of VEGF and Ang-2 protein in supernatants were examined by enzyme linked immunosorbent assay (ELISA). The mRNA expressions of VEGF and Ang-2 in cultured HT-29 were determined by real-time quantitative reverse-transcription polymerase chain reaction. Our results showed that treatment of meloxicam of different concentrations and for various lengths of time had a cytotoxicic effect on the cell proliferation of HT-29 cells in a concentration-dependant and time-dependant manner. Cell cycle analysis showed that the cells were mainly blocked in G0/G1 phase. The VEGF and Ang-2 protein levels in supernatants of the culture medium were decreased gradually in a concentration-dependent or time-dependent fashion. The mRNA expression of cox-2, VEGF and Ang-2 showed a gradual and concentration-dependent reduction. It is concluded that meloxicam can reduce the expression of VEGF and Ang-2 at the protein and mRNA level in colon carcinoma cell line.
基金Project supported by National Natural Science Foundation of China,No.863 Z2001-04
文摘AIM To test the hypothesis to block VEGFexpression of SMMC-7721 hepatoma cells mayinhibit tumor growth using the rat hepatomamodel.METHODS Amplifiy the 200 VEGF cDNAfragment and insert it into human U6 genecassette in the reverse orientation transcribingsmall antisense RNA which could specificallyinteract with VEGF165, and VEGF121 mRNA.Construct the retroviral vector containing thisantisense VEGF U6 cassette and package thereplication-deficient recombinant retrovirus.SMMC-7721 cells were transduced with thesevirus and positive clones were selected withG418. PCR and Southern blot analysis wereperformed to determine if U6 cassette integratedinto the genomic DNA of positive clone.Transfected tumor cells were evaluated for RNAexpression by ribonuclease protection assays.The VEGF protein in the supernatant of parentaltumor cells and genetically modified tumor cellswas determined with ELISA. In vitro and in vivogrowth properties of antisense VEGF cell clonein nude mice were analyzed.RESULTS Restriction enzyme digestion andPCR sequencing verified that the antisense VEGFRNA retroviral vector was successfullyconstructed. After G418 selection, resistantSMMC-7721 cell clone was picked up. PCR andSouthern blot analysis suggested that U6cassette was integrated into the cell genomicDNA. Stable SMMC-7721 cell clone transducedwith U6 antisense RNA cassette could express200bp small antisense VEGF RNA and secretereduced levels of VEGF in culture condition.Production of VEGF by antisense transgeneexpressing cells was 65 ± 10 ng / L per 106 cells,420 ± 45 ng/L per 106 cells in sense group and 485± 30 ng/L per 106 cells in the negative control group, (P<0.05). The antisense-VEGF cell clone appeared phenotypically indistinguishable from SMMC-7721 cells and SMMC-7721 cells transfected sense VEGF. The growth rate of the antisense-VEGF cell clone was the same as the control cells. When S. C. was implanted into nude mice, growth of antisense-VEGF cell lines was greatly inhibited compared with control cells.CONCLUSION Expression of antisense VEGFRNA in SMMC-7721 cells could decrease thetumorigenicity, and antisense-VEGF genetherapy may be an adjuvant treatment forhepatoma.
基金New Century Distinguished Scholar Supporting Program of Ministry of Education (80000-3171404) The National Natural Science Foundation of China, No. 30300082, No. 30470467
文摘AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endothelial cells.METHODS: The KDR-TK fragment was extracted from pBluescript Ⅱ KDR-TK plasmid by enzymatic digestion with Xho I and Sal I. The enhanced green fluorescence protein (EGFP) carrier was extracted from pEGFP by the same procedure. The KDR-TK was inserted into the pEGFP carrier to construct pEGFP-KDR-TK. Using ultrasound irradiation and microbubble, pEGFP-KDR-TK was transferred into human umbilical vein endothelial cells (HUVECs). The transient infection rate was estimated by green fluorescent protein (GFP) expression. Transfected HUVECs, non-transfected HUVECs, and HepG2 cells were cultured in the presence of different concentrations of ganciclovir (GCV), and the killing efficacy of HSV-TK/GCV was analyzed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: The recombinant pEGFP-KDR-TK was successfully constructed by inserting the KDR-TK fragment into the pEGFP carrier. Transfected HUVECs showed cytoplasmic green fluorescence, and the transient transfection rate was about 20.3%. Pools of G418-resistant cells exhibited a higher sensitivity to theprodrug/GCV compared to non-transfected HUVECs or non-transfected HepG2 cells, respectively. CONCLUSION: KDR promoter and the suicide gene/prodrug system mediated by diagnostic ultrasound combined with microbubble can significantly kill HUVECs. Such therapy may present a novel and attractive approach to target gene therapy on tumor vessels.
基金the National Natural Science Foundation of China, No. 30772341, 81070523
文摘We constructed a lentiviral vector carrying vascular endothelial growth factor 165, which was used to transfect neural stem cells. The transfection rate was approximately 50%, as determined by flow cytometry. Vascular endothelial growth factor protein was detected in neural stem cells and promoted proliferation.