Anti-vascular endothelial growth factor drugs combined with laser photocoagulation maintain retinal ganglion cell integrity in patients with diabetic macular edema: study protocol for a prospective, non-randomized, controlled clinical trial

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).

Vascular endothelial growth factor/connective tissue growth factor and proteomic analysis of aqueous humor after intravitreal conbercept for proliferative diabetes retinopathy

AIM:To investigate the role of connective tissue growth factor(CTGF)and vascular endothelial growth factor(VEGF)in the protein profile of the aqueous humor in patients with proliferative diabetic retinopathy(PDR)following intravitreal injection of conbercept.METHODS:This study included 72 PDR patients and 8 cataract patients as controls.PDR patients were divided into 3 groups according to the intervals of 3,5,and 7d between intravitreal conbercept(IVC,0.5 mg/0.05 mL)injection and pars plana vitrectomy(PPV)performed.Aqueous humor samples were collected before and after IVC and PPV for VEGF and CTGF levels detected with enzyme-linked immunosorbent assay(ELISA).The differential proteomics of 10 patients who underwent PPV surgery 5d after IVC and 8 normal controls was studied,Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis were performed on the data,and the protein interaction network of 23 differential proteins was studied.RESULTS:Post-IVC,VEGF levels decreased and CTGF levels increased significantly in aqueous humor,with the CTGF/VEGF ratio rising significantly at all intervals.Liquid chromatography tandem mass spectrometry(LC-MS/MS)identified differentially expressed proteins between preand post-IVC samples.GO and KEGG analyses revealed involvement in immune response,stress response,complement and coagulation cascades,ferroptosis,and PPAR signaling pathways.PPI analysis highlighted key proteins like APOA1,C3,and transferrin(TF).ELISA assay confirmed the differential expression of proteins such as HBA1,SERPINA1,COL1A1,and ACTB,with significant changes in the IVC groups.CONCLUSION:The study demonstrates that IVC effectively reduces VEGF levels while increasing CTGF levels,thereby modifying the CTGF/VEGF ratio,and IVC significantly alters the protein profile in the aqueous humor of patients with PDR.Proteomic analysis reveals that these changes are associated with critical biological pathways and protein interactions involved in immune response,stress response,and cellular metabolism.

Hypoxia-inducible factor 1alpha and vascular endothelial growth factor in Glioblastoma Multiforme:a systematic review going beyond pathologic implications

Glioblastoma multiforme(GBM)is an aggressive primary brain tumor characterized by extensive heterogeneity and vascular proliferation.Hypoxic conditions in the tissue microenvironment are considered a pivotal player leading tumor progression.Specifically,hypoxia is known to activate inducible factors,such as hypoxia-inducible factor 1alpha(HIF-1α),which in turn can stimulate tumor neo-angiogenesis through activation of various downward mediators,such as the vascular endothelial growth factor(VEGF).Here,we aimed to explore the role of HIF-1α/VEGF immunophenotypes alone and in combination with other prognostic markers or clinical and image analysis data,as potential biomarkers of GBM prognosis and treatment efficacy.We performed a systematic review(Medline/Embase,and Pubmed database search was completed by 16th of April 2024 by two independent teams;PRISMA 2020).We evaluated methods of immunoassays,cell viability,or animal or patient survival methods of the retrieved studies to assess unbiased data.We used inclusion criteria,such as the evaluation of GBM prognosis based on HIF-1α/VEGF expression,other biomarkers or clinical and imaging manifestations in GBM related to HIF-1α/VEGF expression,application of immunoassays for protein expression,and evaluation of the effectiveness of GBM therapeutic strategies based on HIF-1α/VEGF expression.We used exclusion criteria,such as data not reporting both HIF-1αand VEGF or prognosis.We included 50 studies investigating in total 1319 GBM human specimens,18 different cell lines or GBM-derived stem cells,and 6 different animal models,to identify the association of HIF-1α/VEGF immunophenotypes,and with other prognostic factors,clinical and macroscopic data in GBM prognosis and therapeutic approaches.We found that increased HIF-1α/VEGF expression in GBM correlates with oncogenic factors,such as miR-210-3p,Oct4,AKT,COX-2,PDGF-C,PLDO3,M2 polarization,or ALK,leading to unfavorable survival.Reduced HIF-1α/VEGF expression correlates with FIH-1,ADNP,or STAT1 upregulation,as well as with clinical manifestations,like epileptogenicity,and a favorable prognosis of GBM.Based on our data,HIF-1αor VEGF immunophenotypes may be a useful tool to clarify MRI-PET imaging data distinguishing between GBM tumor progression and pseudoprogression.Finally,HIF-1α/VEGF immunophenotypes can reflect GBM treatment efficacy,including combined first-line treatment with histone deacetylase inhibitors,thimerosal,or an active metabolite of irinotecan,as well as STAT3 inhibitors alone,and resulting in a favorable tumor prognosis and patient survival.These data were supported by a combination of variable methods used to evaluate HIF-1α/VEGF immunophenotypes.Data limitations may include the use of less sensitive detection methods in some cases.Overall,our data support HIF-1α/VEGF’s role as biomarkers of GBM prognosis and treatment efficacy.

Expression and significance of pigment epithelium-derived factor and vascular endothelial growth factor in colorectal adenoma and cancer

BACKGROUND The incidence and mortality of colorectal cancer(CRC)are among the highest in the world,and its occurrence and development are closely related to tumor neovascularization.When the balance between pigment epithelium-derived factors(PEDF)that inhibit angiogenesis and vascular endothelial growth factors(VEGF)that stimulate angiogenesis is broken,angiogenesis is out of control,resulting in tumor development.Therefore,it is very necessary to find more therapeutic targets for CRC for early intervention and later treatment.AIM To investigate the expression and significance of PEDF,VEGF,and CD31-stained microvessel density values(CD31-MVD)in normal colorectal mucosa,adenoma,and CRC.METHODS In this case-control study,we collected archived wax blocks of specimens from the Digestive Endoscopy Center and the General Surgery Department of Chengdu Second People's Hospital from April 2022 to October 2022.Fifty cases of specimen wax blocks were selected as normal intestinal mucosa confirmed by electronic colonoscopy and concurrent biopsy(normal control group),50 cases of specimen wax blocks were selected as colorectal adenoma confirmed by electronic colonoscopy and pathological biopsy(adenoma group),and 50 cases of specimen wax blocks were selected as CRC confirmed by postoperative pathological biopsy after inpatient operation of general surgery(CRC group).An immunohistochemical staining experiment was carried out to detect PEDF and VEGF expression in three groups of specimens,analyze their differences,study the relationship between the two and clinicopathological factors in CRC group,record CD31-MVD in the three groups,and analyze the correlation of PEDF,VEGF,and CD31-MVD in the colorectal adenoma group and the CRC group.The F test or adjusted F test is used to analyze measurement data statistically.Kruskal-Wallis rank sum test was used between groups for ranked data.The chi-square test,adjusted chi-square test,or Fisher's exact test were used to compare the rates between groups.All differences between groups were compared using the Bonferroni method for multiple comparisons.Spearman correlation analysis was used to test the correlation of the data.The test level(α)was 0.05,and a two-sided P<0.05 was considered statistically significant.RESULTS The positive expression rate and expression intensity of PEDF were gradually decreased in the normal control group,adenoma group,and CRC group(100%vs 78%vs 50%,χ^(2)=34.430,P<0.001;++~++vs+~++vs-~+,H=94.059,P<0.001),while VEGF increased gradually(0%vs 68%vs 96%,χ^(2)=98.35,P<0.001;-vs-~+vs++~+++,H=107.734,P<0.001).In the CRC group,the positive expression rate of PEDF decreased with the increase of differen-tiation degree,invasion depth,lymph node metastasis,distant metastasis,and TNM stage(χ^(2)=20.513,4.160,5.128,6.349,5.128,P<0.05);the high expression rate of VEGF was the opposite(χ^(2)=10.317,13.134,17.643,21.844,17.643,P<0.05).In the colorectal adenoma group,the expression intensity of PEDF correlated negatively with CD31-MVD(r=-0.601,P<0.001),whereas VEGF was not significantly different(r=0.258,P=0.07).In the CRC group,the expression intensity of PEDF correlated negatively with the expression intensity of CD31-MVD and VEGF(r=-0.297,P<0.05;r=-0.548,P<0.05),while VEGF expression intensity was positively related to CD31-MVD(r=0.421,P=0.002).CONCLUSION It is possible that PEDF can be used as a new treatment and prevention target for CRC by upregulating the expression of PEDF while inhibiting the expression of VEGF.

Transforming growth factor-β1 and vascular endothelial growth factor levels in senile acute myeloid leukemia and correlation with prognosis

BACKGROUND Acute myeloid leukemia(AML)is a disease in which immature hematopoietic cells accumulate in the bone marrow and continuously expand,inhibiting hematopoiesis.The treatment and prognosis of this disease have always been unsatisfactory.AIM To investigate the correlation between vascular endothelial growth factor(VEGF)and transforming growth factor-β1(TGFβ1)expression and prognosis in older adults with AML.METHODS This study enrolled 80 patients with AML(AML group),including 36 with complete response(AML-CR),23 with partial response(AML-PR),and 21 with no response(AML-NR).The expression levels of VEGF and TGFβ1 were detected by reverse transcription polymerase chain reaction in bone marrow mononuclear cells isolated from 56 healthy controls.Kaplan-Meier analysis was performed to assess overall survival(OS)and progression-or disease-free survival(DFS).Prognostic risk factors were analyzed using a Cox proportional hazards model.RESULTS The AML group showed a VEGF level of 2.68±0.16.VEGF expression was lower in patients with AML-CR than those with AML-PR or AML-NR(P<0.05).TGFβ1 expression in the AML group was 0.33±0.05.Patients with AML-CR showed a higher TGFβ1 expression than those with AML-PR or AML-NR(P<0.05).VEGF and TGFβ1 expression in patients with AML was significantly correlated with the counts of leukocytes,platelets,hemoglobin,and peripheral blood immature cells(P<0.05);Kaplan-Meier survival analysis revealed that patients with high TGFβ1 expression had better OS and DFS than those with low TGFβ1 expression(P<0.05),whereas patients with low VEGF levels showed better OS and DFS than those with high VEGF levels(P<0.05).VEGF,TGFβ1,and platelet count were identified by the Cox proportional hazards model as independent risk factors for OS(P<0.05),while VEGF,TGFβ1,and white blood cell count were independent risk factors for DFS(P<0.05).CONCLUSION Decreased VEGF expression and increased TGFβ1 expression in patients with AML provide valuable references for determining and individualizing clinical treatment strategies.

Diagnostic value of serum vascular endothelial growth factor and interleukin-17 in primary hepatocellular carcinoma

BACKGROUND Despite significant advancements in the medical treatment of primary hepato-cellular carcinoma(PHC)in recent years,enhancing therapeutic effects and im-proving prognosis remain substantial challenges worldwide.AIM To investigate the expression levels of serum vascular endothelial growth factor(VEGF)and interleukin(IL)-17 in patients with PHC and evaluate their diagnostic value while exploring their relationship with patients’clinical characteristics.METHODS The study included 50 patients with confirmed PHC who visited Wuhan Han-yang Hospital from January 2021 to January 2022,and 50 healthy individuals from the same period served as the control group.Serum VEGF and IL-17 levels in both groups were measured by Enzyme-Linked Immunosorbent Assay,and their diagnostic value was assessed using receiver operating characteristic(ROC)curves.Pearson correlation analysis was performed to examine the relationship between serum VEGF and IL-17 levels.Pathological data of the PHC patients were analyzed to determine the relationship between serum VEGF and IL-17 levels and pathological characteristics.RESULTS Serum VEGF and IL-17 levels were significantly higher in the study group com-pared to the control group(P<0.05).No significant association was observed between serum VEGF and IL-17 levels and gender,age,combined cirrhosis,tumor diameter,or degree of differentiation(P>0.05).However,there was a significant relationship between clinical TNM stage,tumor metastasis,and serum VEGF and IL-17 levels(P<0.05).Correlation analysis revealed a positive correlation between serum VEGF and IL-17(P<0.05).ROC analysis demonstrated that both serum VEGF and IL-17 had good diagnostic efficacy for PHC.CONCLUSION Serum VEGF and IL-17 levels were significantly higher in PHC patients compared to healthy individuals.Their levels were closely related to pathological features such as tumor metastasis and clinical TNM stage,and there was a significant positive correlation between VEGF and IL-17.These biomarkers may serve as valuable reference in-dicators for the early diagnosis and treatment guidance of PHC.

Nomogram for predicting short-term response to anti-vascular endothelial growth factor treatment in neovascular age-related macular degeneration:An observational study

BACKGROUND Anti-vascular endothelial growth factor(anti-VEGF)therapy is critical for managing neovascular age-related macular degeneration(nAMD),but understanding factors influencing treatment efficacy is essential for optimizing patient outcomes.AIM To identify the risk factors affecting anti-VEGF treatment efficacy in nAMD and develop a predictive model for short-term response.METHODS In this study,65 eyes of exudative AMD patients after anti-VEGF treatment for≥1 mo were observed using optical coherence tomography angiography.Patients were classified into non-responders(n=22)and responders(n=43).Logistic regression was used to determine independent risk factors for treatment response.A predictive model was created using the Akaike Information Criterion,and its performance was assessed with the area under the receiver operating characteristic curve,calibration curves,and decision curve analysis(DCA)with 500 bootstrap re-samples.RESULTS Multivariable logistic regression analysis identified the number of junction voxels[odds ratio=0.997,95%confidence interval(CI):0.993-0.999,P=0.010]as an independent predictor of positive anti-VEGF treatment outcomes.The predictive model incorporating the fractal dimension,number of junction voxels,and longest shortest path,achieved an area under the curve of 0.753(95%CI:0.622-0.873).Calibration curves confirmed a high agreement between predicted and actual outcomes,and DCA validated the model's clinical utility.CONCLUSION The predictive model effectively forecasts 1-mo therapeutic outcomes for nAMD patients undergoing anti-VEGF therapy,enhancing personalized treatment planning.

Overexpression of vascular endothelial growth factor enhances the neuroprotective effects of bone marrow mesenchymal stem cell transplantation in ischemic stroke

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.

Serum vascular endothelial growth factor and cortisol expression to predict prognosis of patients with hypertensive cerebral hemorrhage

BACKGROUND Cerebral hemorrhage is a common and severe complication of hypertension in middle-aged and elderly men.AIM To investigate the correlation between vascular endothelial growth factor(VEGF)and cortisol(Cor)and the prognosis of patients with hypertensive cerebral hemorrhage.METHODS A hundred patients with hypertensive intracerebral hemorrhage were enrolled from January 2020 to December 2022 and assigned to the hypertensive intracerebral hemorrhage group.Another 100 healthy people who were examined at our hospital during the same period were selected and assigned to the healthy group.Peripheral venous blood was collected,and serum Cor and VGEF levels were measured through enzyme linked immunosorbent assay.RESULTS A statistically significant difference in serum Cor and VGEF levels was observed among patients with varying degrees of neurological impairment(P<0.05).Serum Cor and VGEF levels were significantly higher in the severe group than in the mild-to-moderate group.Cor and VEGF levels were significantly higher in patients with poor prognoses than in those with good prognoses.Multiple logistic regression analysis revealed that serum Cor and VGEF levels were independent factors affecting hypertensive intracerebral hemorrhage(P<0.05).CONCLUSION Cor and VGEF are associated with the occurrence and development of hypertensive cerebral hemorrhage and are significantly associated with neurological impairment and prognosis of patients.

Vascular endothelial growth factor protein and gene delivery by novel nanomaterials for promoting liver regeneration after partial hepatectomy

Partial hepatectomy(PH)can lead to severe complications,including liver failure,due to the low regenerative capacity of the remaining liver,especially after extensive hepatectomy.Liver sinusoidal endothelial cells(LSECs),whose proliferation occurs more slowly and later than hepatocytes after PH,compose the lining of the hepatic sinusoids,which are the smallest blood vessels in the liver.Vascular endothelial growth factor(VEGF),secreted by hepatocytes,promotes LSEC proliferation.Supplementation of exogenous VEGF after hepatectomy also increases the number of LSECs in the remaining liver,thus promoting the reestablishment of the hepatic sinusoids and accelerating liver regeneration.At present,some shortcomings exist in the methods of supplementing exogenous VEGF,such as a low drug concentration in the liver and the reaching of other organs.Moreover,VEGF should be administered multiple times and in large doses because of its short half-life.This review summarized the most recent findings on liver regeneration and new strategies for the localized delivery VEGF in the liver.

Vascular endothelial growth factor B improves impaired glucose tolerance through insulin-mediated inhibition of glucagon secretion

BACKGROUND Impaired glucose tolerance(IGT)is a homeostatic state between euglycemia and hyperglycemia and is considered an early high-risk state of diabetes.When IGT occurs,insulin sensitivity decreases,causing a reduction in insulin secretion and an increase in glucagon secretion.Recently,vascular endothelial growth factor B(VEGFB)has been demonstrated to play a positive role in improving glucose metabolism and insulin sensitivity.Therefore,we constructed a mouse model of IGT through high-fat diet feeding and speculated that VEGFB can regulate hyperglycemia in IGT by influencing insulin-mediated glucagon secretion,thus contributing to the prevention and cure of prediabetes.AIM To explore the potential molecular mechanism and regulatory effects of VEGFB on insulin-mediated glucagon in mice with IGT.METHODS We conducted in vivo experiments through systematic VEGFB knockout and pancreatic-specific VEGFB overexpression.Insulin and glucagon secretions were detected via enzyme-linked immunosorbent assay,and the protein expression of phosphoinositide 3-kinase(PI3K)/protein kinase B(AKT)was determined using western blot.Further,mRNA expression of forkhead box protein O1,phosphoenolpyruvate carboxykinase,and glucose-6 phosphatase was detected via quantitative polymerase chain reaction,and the correlation between the expression of proteins was analyzed via bioinformatics.RESULTS In mice with IGT and VEGFB knockout,glucagon secretion increased,and the protein expression of PI3K/AKT decreased dramatically.Further,in mice with VEGFB overexpression,glucagon levels declined,with the activation of the PI3K/AKT signaling pathway.CONCLUSION VEGFB/vascular endothelial growth factor receptor 1 can promote insulin-mediated glucagon secretion by activating the PI3K/AKT signaling pathway to regulate glucose metabolism disorders in mice with IGT.

Role of vascular endothelial growth factor B in nonalcoholic fatty liver disease and its potential value

Nonalcoholic fatty liver disease(NAFLD)refers to fatty liver disease caused by liver injury factors other than alcohol.The disease is characterized by diffuse fat infiltration,including simple steatosis(no inflammatory fat deposition),nonalcoholic fatty hepatitis,liver fibrosis,and so on,which may cause liver cirrhosis,liver failure,and even liver cancer in the later stage of disease progression.At present,the pathogenesis of NAFLD is still being studied.The"two-hit"theory,represented by lipid metabolism disorder and inflammatory reactions,is gradually enriched by the"multiple-hit"theory,which includes multiple factors,such as insulin resistance and adipocyte dysfunction.In recent years,vascular endothelial growth factor B(VEGFB)has been reported to have the potential to regulate lipid metabolism and is expected to become a novel target for ameliorating metabolic diseases,such as obesity and type 2 diabetes.This review summarizes the regulatory role of VEGFB in the onset and development of NAFLD and illustrates its underlying molecular mechanism.In conclusion,the signaling pathway mediated by VEGFB in the liver may provide an innovative approach to the diagnosis and treatment of NAFLD.

Expression of vascular endothelial growth factor and its role in oncogenesis of human gastric carcinoma

AIM: To establish the role of vascular endothelial growth factor (VEGF) in the oncogenesis of human gastric carcinoma more directly. METHODS: The expression of VEGF and its receptor kinase-domain insert containing receptor (KDR) in human gastric cancer tissue were observed by immunohistochemical staining. VEGF levels were manipulated in human gastric cancer cell using eukaryotic expression constructs designed to express the complete VEGF(165) complimentary DNA in either the sense or antisense orientation. The biological changes of the cells were observed in which VEGF was up-regulated or down-regulated. RESULTS: VEGF-positive rate was 50%, and VEGF was mainly localized in the cytoplasm and membrane of the tumor cells, while KDR was mainly located in the membrane of vascular endothelial cells in gastric cancer tissues and peri-cancerous tissue. In 2 cases of 50 specimens, the gastric cancer cells expressed KDR, localized in both the cytoplasm and membrane. Introduction of VEGF(165) antisense into human gastric cancer cells (SGC-7901, immunofluorescence intensity, 31.6%)) resulted in a significant reduction in VEGF-specific messenger RNA and total and cell surface VEGF protein (immunofluorescence intensity, 8.9%) (P【0.05). Conversely, stable integration of VEGF(165) in the sense orientation resulted in an increase in cellular and cell surface VEGF (immunofluorescence intensity, 75.4%) (P【0.05). Lowered VEGF levels were associated with a marked decrease in the growth of nude mouse xenografted tumor (at 33 days postimplantation, tumor volume: 345.40 +/- 136.31 mm3)(P【0.05 vs control SGC-7901 group: 1534.40 +/- 362.88 mm3), whereas up-regulation of VEGF resulted in increased xenografted tumor size (at 33 days postimplantation, tumor volume: 2350.50 +/- 637.70 mm3) (P【0.05 vs control SGC-7901 group). CONCLUSION: This study provides direct evidence that VEGF plays an important role in the oncogenesis of human gastric cancer.

Increased expressions of vascular endothelial growth factor(VEGF),VEGF-C and VEGF receptor-3 in prostate cancer tissue are associated with tumor progression

Aim： To investigate the differences in microvessel densities （MVD） and the expressions of vascular endothelial growth factor （VEGF）, VEGF-C and VEGF receptor-3 （VEGFR-3） between prostate cancer （PCa） tissues and adjacent benign tissues, and to explore the correlations among MVD, Jewett-Whitmore staging, Gleason scores and expressions of VEGF, VEGF-C and VEGFR-3 in the progression of PCa. Methods： An immunohistochemical approach was adopted to detect the expressions of CD34, VEGF, VEGF-C and VEGFR-3 in both cancer areas and peripheral benign areas of 71 primary prostatic adenocarcinoma specimens. A statistic analysis was then performed according to the experimental and clinic data. Results： Significantly upregulated expressions of VEGF, VEGF-C and VEGFR-3 were all found in malignant epithelium/cancer cells compared with adjacent benign epithelium （P 〈 0.01）. Patients in stage D had a significantly higher score than patients in stage A, B or C when comparing the expression of VEGF-C or VEGFR-3 in the tumor area （P 〈 0.01）. In addition, significant correlations were observed between Jewett-Whitmore staging and VEGF-C （rs = 0.738, P 〈 0.01）, clinical staging and VEGFR-3 （rs = 0.410, P 〈 0.01）, VEGF-C and Gleason scores （rs = 0.401, P 〈 0.01）, VEGFR-3 and Gleason scores （rs = 0.581, P 〈 0.001） and MVD and VEGF （rs = 0.492, P 〈 0.001）. Conclusion： Increased expressions of VEGF and VEGF-C were closely associ- ated with progression of PCa. The main contribution of increased VEGF expression for PCa progression was to upregulate MVD, which maintained the growth advantage of tumor tissue. However, the chief role of increased expressions of VEGF-C and VEGFR-3 was to enhance lymphangiogenesis and provide a main pathway for cancer cells to disseminate. （Asian J Androl 2006 Mar; 8： 169-175）

IMPACTS ON THE EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR IN ISCHEMIC BRAIN TISSUE TREATED WITH CLUSTER PUNC-TURE ON SCALP ACUPOINTS IN RATS

Objective To probe into the impacts on the expression of vascular endothelial growth factor （VEGF） in ischemic brain tissue treated with cluster puncture on scalp acupoints in rats, Methods 128 rats were randomized into pseudo-operation group, model group, scalp-needling group and cluster-needling group, In scalp-needling group, penetration method was used on the focal side from Bǎihuì （百会 GV20） to Qǔbīn （曲鬓 GB7）, and in cluster-needling group, penetration method was used on both sides from GV20 to GB7, and a common needling on GV 20. Needles were punctured 2 mm in depth, constantly rotated for 10min, retained for 2 h. Immunohistrochemical method was applied to determine VEGF expression and microvessel density （MVD）. Results With the intervention of cluster puncture on scalp acupoints, VEGF： expressions on every time-spot after ischemia were enhanced apparently, superior to those in scalp-needling group. On the three time-spots of the 7^th, 14^th and 21^st days, MVD was increased after cluster puncture on scale acupoints, superior to those in scalp-needling group. Conclusion Cluster puncture on scalp acupoints up-regulated VEGF expression and promoted regeneration of microvessel.

Reduction of tumorigenicity of SMMC-7721 hepatoma cells by vascular endothelial growth factor antisense gene therapy

AIM: To test the hypothesis to block VEGF expression of SMMC-7721 hepatoma cells may inhibit tumor growth using the rat hepatoma model. METHODS: Amplify the 200 VEGF cDNA fragment and insert it into human U6 gene cassette in the reverse orientation transcribing small antisense RNA which could specifically interact with VEGF165, and VEGF121 mRNA. Construct the retroviral vector containing this antisense VEGF U6 cassette and package the replication-deficient recombinant retrovirus. SMMC-7721 cells were transduced with these virus and positive clones were selected with G418. PCR and Southern blot analysis were performed to determine if U6 cassette integrated into the genomic DNA of positive clone. Transfected tumor cells were evaluated for RNA expression by ribonuclease protection assays. The VEGF protein in the supernatant of parental tumor cells and genetically modified tumor cells was determined with ELISA. In vitro and in vivo growth properties of antisense VEGF cell clone in nude mice were analyzed. RESULTS: Restriction enzyme digestion and PCR sequencing verified that the antisense VEGF RNA retroviral vector was successfully constructed.After G418 selection, resistant SMMC-7721 cell clone was picked up. PCR and Southern blot analysis suggested that U6 cassette was integrated into the cell genomic DNA. Stable SMMC-7721 cell clone transduced with U6 antisense RNA cassette could express 200 bp small antisense VEGF RNA and secrete reduced levels of VEGF in culture condition. Production of VEGF by antisense transgene-expressing cells was 65+/-10 ng/L per 10(6) cells, 42045 ng/L per 10(6) cells in sense group and 485+/-30 ng/L per 10(6) 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 VEGF RNA in SMMC-7721 cells could decrease the tumorigenicity, and antisense-VEGF gene therapy may be an adjuvant treatment for hepatoma.

Effects of endostatin on expression of vascular endothelial growth factor and its receptors and neovascularization in colonic carcinoma implanted in nude mice

AIM:To investigate the antiangiogenic effects of endostatin on colonic carcinoma cell line implanted in nude mice and its mechanism. METHODS:Nude mice underwent subcutaneous injection with LS-174t colonic carcinoma cell line to generate carcinoma and were randomly separated into two groups.Mice received injection of vehicle or endostatin every day for two weeks. After the tumor was harvested,the tumor volumes were determined,and the expressions of CD34,VEGF and FIk-1 were examined by immunohistochemical method. RESULTS:Tumor volume was significantly inhibited in the endostatin group(84.17%)and tumor weight was significantly inhibited in the endostatin group(0.197±0.049) compared to the control group(1.198±0.105)(F=22.56, P=0.001),microvessel density(MVD)was significantly decreased in the treated group(31.857±3.515)compared to the control group(100.143±4.290)(F=151.62,P<0.001). Furthermore,the expression of FIk-1 was significantly inhibited in the treated group(34.29%) ompared to the control group(8.57%)(X^2=13.745,P=0.001).However no significant decrease was observed in the expression of vascular endothelial growth factor(VEGF)between these two groups(X^2=0.119,P=0.730). CONCLUSION:Endostatin can inhibit tumor growth and angiogenesis by blocking Vegf/FIk-1 pathway.This experiment provides the theory basis for developing a new anti-carcinoma drug through studying the properties of anti-angiogenesis inhibitors.

Serum Vascular Endothelial Growth Factor-C and Vascular Endothelial Growth Factor Level in Patients with Colorectal Carcinoma and Clinical Significance

Circulating vascular endothelial growth factor-C （VEGF-C） and vascular endothelial growth factor （VEGF） levels in patients with colorectal carcinoma were determined in order to assess their clinical significance as a diagnostic tool for monitoring lymph node metastasis. In 66 patients with colorectal carcinoma and 30 healthy controls, circulating VEGF-C and VEGF levels were assessed by using enzyme-linked immunosorbent assay （ELISA）. Serum VEGF-C and VEGF levels were higher in patients with colorectal carcinoma than in healthy controls. Patients with lymph node metastasis had higher serum VEGF-C and VEGF levels than those without lymph node metastasis. The levels of VEGF-C and VEGF were higher in the invasion group than in the non-invasion group. Serum VEGF-C levels reached a sensitivity of 81% and a specificity of 76 % with a cutoff value of 1438.0 pg/mL, whereas VEGF levels reached 72 % sensitivity and 74 % specificity at 240.2 pg/ mL. If 66 patients were divided into 4 groups according to the combined determination of VEGF-C and VEGF levels, the positive predictive value was 85.3 %, the negative predictive value was 94.6 %, and accuracy was 93.7 %. It was suggested that circulating VEGF-C levels might provide additional information for distinguishing the absence from presence of lymph node metastasis in patients with colorectal carcinoma. The combined determination of VEGF-C and VEGF levels could be used as an important index for preoperatively clinical stage of colorectal carcinoma.

Expression of vascular endothelial growth factor and its receptors VEGFR-1 and 2 in gastrointestinal stromal tumors,leiomyomas and schwannomas

AIM： To investigate the role of vascular endothelial growth factor （VEGF/and its receptors VEGFR-1 and 2 in the growth and differentiation of gastrointestinal strornal tumors （GISTs）. METHODS： Thirty-three GISTs, 15 leiomyomas and 6 schwannomas were examined by immunohistochemistry in this study. RESULTS： VEGF protein was expressed in the cytoplasm of tumor cells, and VEGFRol and 2 were expressed both in the cytoplasm and on the membrane of all tumors. Irnrnunohistochernical staining revealed that 26 GISTs （78.8%）, 9 leiornyornas （60.0%） and 3 schwannornas （50.0%/were positive for VEGF; 24 GISTs （72.7%/, 12 leiornyornas （80.0%） and 4 schwannornas （66.7%） were positive for VEGFR-1; 30 GISTs （90.9%/, 5 leiornyornas （33.3%/and 4 schwannornas （66.7%） were positive for VEGFR-2. VEGFR-2 expression was statistically different between GISTs and leiomyomas （P 〈 0.0001）. However, there was no correlation between the expression of VEGF pathway componenets and the clinical risk categories. CONCLUSION： Our results suggest that the VEGF pathway may play an important role in the differentiation of GISTs, leiomyomas and schwannomas.

Streptozotocin induced diabetic retinopathy in rat and the expression of vascular endothelial growth factor and its receptor

·AIM: To establishtherat model of streptozotocin (STZ)- induced diabetic retinopathy (DR), which is the most common cause of visual loss and blindness in patients with diabetes, and observe the gene expression of vascular endothelial growth factor (VEGF) and its receptors during the development of DR. ·METHODS: A rat model of diabetes was established by intraperitoneal injection of STZ. The diabetic rats were housed for 2, 3 and 4 months after the development of diabetes. Retinal histopathological observation was performed. The retinal vessels were observed by immunofluorescence staining by CD31. The mRNA expression of VEGF, VEGF receptor 1 and 2 (VEGFR1/2) in rat retina was detected by reverse transcription - polymerase chain reaction (RT-PCR) analysis. · RESULTS: Retinal histopathological observation showed the morphological changes of inner nuclear layer (INL) and outer nuclear layer (ONL) at any time -point, and also demonstrated the increased new vessels at both 3, 4 months after the development of diabetes. The CD31 staining results showed that the number of vessels was increased in the retinas of diabetic rats at both 3 and 4 months after the development of diabetes. As compared to the normal rats, the mRNA expression of VEGF was increased in retinas of diabetic rats at 3 months after the development of diabetes, while VEGFR1 and VEGFR2 mRNA expression was increased at 2, 3 and 4 months after the development of diabetes.·CONCLUSION:Takentogether,ourresultsdemonstrated that DR was occurred at 3 months after the development of diabetes, and the mRNA expression of VEGF, VEGFR1 and VEGFR2 were increased in the process of DR. The present study further evidenced the involvement of VEGF and its receptors in the process of DR. ·