Ephrin-B2 has been shown to participate in angiogenesis, but the underlying mechanisms involved remain unclear. In this study, a rat model of local cerebral ischemia was prepared by focal middle cerebral artery occlus...Ephrin-B2 has been shown to participate in angiogenesis, but the underlying mechanisms involved remain unclear. In this study, a rat model of local cerebral ischemia was prepared by focal middle cerebral artery occlusion, followed by 24-hour reperfusion. Then, ephrin-B2 protein was administered intracerebroventricularly for 3 consecutive days via a micro-osmotic pump. Western blot assay and quantitative real-time reverse transcription PCR demonstrated the expression levels of angiopoietin-1 （Ang-1） mRNA and protein in the penumbra cortex of the ephrin-B2 treated group were decreased at day 4 after reperfusion, and increased at day 28, while the expression levels of angiopoietin-2 （Ang-2） were highly up-regulated at all time points tested. Double immunofluorescent staining indicated that Ang-1 and Ang-2 were both expressed in vascular endothelial cells positive for CD31. These findings indicate that ephrin-B2 influences the expressions of Ang-1 and Ang-2 during angiogenesis following transient focal cerebral ischemia.展开更多
AIM: To determine the inhibitory effect of the adenovirus- based angiopoietin-1 (Ang-1) targeted small interfering RNA expression system (Ad/Ang-1si) on the expression of the Ang-1 gene, cell growth and apoptosis in h...AIM: To determine the inhibitory effect of the adenovirus- based angiopoietin-1 (Ang-1) targeted small interfering RNA expression system (Ad/Ang-1si) on the expression of the Ang-1 gene, cell growth and apoptosis in human esophageal cancer cell line Eca109. METHODS: siRNA-expressing adenovirus targeting Ang-1 gene was constructed using the Ad Easy System. Cultured Eca109 cells were transfected with Ad/Ang-1si (Eca109/Ang-1si), and Ad/si was used to infect Eca109 cells as control (Eca109/si). Ang-1 gene expression and concentration was determined with RT-PCR and ELISA, respectively. Human umbilical vein endothelial cell (HUVEC) migration and proliferation were analyzed. After s.c. injection into athymic nu/nu mice, the tumor growth, vessel density and apoptosis of each group was also determined. RESULTS: HUVEC migration induced by conditioned medium from Ang-1si-transfected Eca109 cells was significantly less than that induced by conditioned medium from Eca109 cells and control adenovirus- transfected Eca109 cells. Furthermore, after s.c. injection into athymic nu/nu mice, the tumor growth and cell apoptosis of Ad/Ang-1si -expressing Eca109 cells was significantly lower than that of parental or control adenovirus-transfected cells. Vessel density assessed by CD31 immunohistochemical analysis and Ang-1 expression by RT-PCR were also decreased. CONCLUSION: The targeting Ang-1 may provide a therapeutic option for esophageal cancer.展开更多
AIM To evaluate the efficacy of peripheral blood concentrations of angiopoietins(Ang) as cirrhosis biomarkers of chronic hepatitis C(CHC).METHODS Ang1 and Ang2 serum levels were measured by enzyme-linked immunosorbent...AIM To evaluate the efficacy of peripheral blood concentrations of angiopoietins(Ang) as cirrhosis biomarkers of chronic hepatitis C(CHC).METHODS Ang1 and Ang2 serum levels were measured by enzyme-linked immunosorbent assays(ELISA) in samples from 179 cirrhotic and non-cirrhotic CHC patients, classified according to the METAVIR system.Groups were compared by non-parametric MannWhitney U test. Subsequently, the association of peripheral concentrations of angiopoietins with the stage of fibrosis was analyzed using Spearman correlation test. Finally, the accuracy, sensitivity and specificity of circulating angiopoietins for cirrhosis diagnosis were determined by the study of the respective area under the curve of receiver operator characteristics(AUC-ROC).RESULTS Peripheral blood concentrations of Ang1 and Ang2 in CHC patients were significantly related to fibrosis. While Ang1 was decreased in cirrhotic subjects compared to non-cirrhotic(P < 0.0001), Ang2 was significantly increased as CHC progressed to the end stage of liver disease(P < 0.0001). Consequently, Ang2/Ang1 ratio was notably amplified and significantly correlated with fibrosis(P < 0.0001). Interestingly, the individual performance of each angiopoietin for the diagnosis of cirrhosis reached notable AUC-ROC values(above 0.7, both), but the Ang2/Ang1 ratio was much better(AUC-ROC = 0.810) and displayed outstanding values of sensitivity(71%), specificity(84%) and accuracy(82.1%) at the optimal cut-off(10.33). Furthermore, Ang2/Ang1 ratio improved the performance of many other previously described biomarkers or scores of liver cirrhosis in CHC.CONCLUSION Ang2/Ang1 ratio might constitute a useful tool for monitoring the progression of chronic liver disease towards cirrhosis and play an important role as therapeutic target.展开更多
AIM： To evaluate the effects of angiopoietin-1 （Ang-1） on adhesion of gastric cancer cell line BGC-823 and expression of integrin β1, CD44V6, urokinase-type plasminogen activator （uPA） and matrix metalloproteina...AIM： To evaluate the effects of angiopoietin-1 （Ang-1） on adhesion of gastric cancer cell line BGC-823 and expression of integrin β1, CD44V6, urokinase-type plasminogen activator （uPA） and matrix metalloproteinase-2 （MMP-2）. METHODS： BGC-823 cells were transfected transiently with adenovirus-Ang-1 （Ad-Ang-1）. Cells transfected transiently with adenovirus-green fluorescent protein （Ad-GFP） and untransfected cells were used as a negative and blank control group, respectively. The cell adhesion rate between cell and extracellular matrix （ECM） was determined by cell adhesion assay. To investigate whether Ang-1 could reinforce gastric carcinoma metastasis, we performed migration and invasion assays in BGC-823 cells. The mRNA and protein expression of integrin β1, CD44V6, uPA and MMP-2 were detected by reverse transcription polymerase chain reaction and Western blotting, respectively. The expression of integrin β1 and CD44V6 was measured by immunohistochemistry. RESULTS： BGC-823 cells were transfected successfully. The adhesion rate increased significantly in the Ad-Ang-1 group （P 〈 0.05）. The Ad-Ang-1-transfected group had a significant increase in migration and invasion compared with that of the mock-transfected and Ad-GFP groups. The mRNA and protein expression of integrin β1, CD44V6, uPA and MMP-2 in the Ad- Ang-1 group was higher than that in the Ad-GFP and blank control groups （P 〈 0.05）. Compared with mocktransfected and Ad-GFP groups, integrin 131 and CD44V6 expression intensity greatly increased （P 〈 0.05）. CONCLUSION： Transfection of Ang-1 into human gastric cancer cell line BGC-823 can significantly increase expression of integrin β1 and CD44V6, by which cell adhesion and metastasis to the ECM are promoted.展开更多
Objectives This study investigated the efficacy of human skeletal myoblasts (SkM) mediated either human vascular endothelial growth factor-165 (hVEGF165) or angiopoietin-1 (Ang-1) on vascular development and myocardia...Objectives This study investigated the efficacy of human skeletal myoblasts (SkM) mediated either human vascular endothelial growth factor-165 (hVEGF165) or angiopoietin-1 (Ang-1) on vascular development and myocardial regional perfusion. Methods A porcine heart model of chronic infarction was created in 28 female swine by coronary artery ligation. The animals were randomized into: (1) group-1, DMEM injected (n=6), (2) group-2, Ad-null transduced SkM transplanted (n=6), (3) group-3, Ad-hVEGF165 transduced SkM transplanted (n=8), and (4) group-4, Ad-Ang-1 transduced SkM (n=8). Three weeks later, 5 ml DMEM containing 3×108 SkM carrying exogenous genes were intramyocardially injected into 20 sites in left ventricle in groups-2, -3 and -4. Animals in group-1 were injected 5 ml DMEM without cells. Animals were kept on 5 mg/kg cyclosporine per day for 6 weeks. Regional blood flow was measured using fluorescent microspheres. The heart was explanted at 2, 6 and 12 weeks after transplantation for histological studies. Results Histological examination showed survival of lac-z expressing myoblasts in host tissue. Capillary density based on Von Willebrand factor-VIII (vWF-VIII) at low power field (×100) was 57.13±11.85 in group-3 at 6 weeks and declined to 32.1±5.21 at 12 weeks, while it was 39.9±10.26 at 6 weeks and increased to 45.14±6.54 at 12 weeks in group-4. The mature blood vessel index was highest in group- 4 at 6 and 12 weeks after transplantation. The regional blood flow in the center and peri-infarct area was significantly increased in animals of groups-3 and -4. Conclusions SkM carrying either hVEGF165 or Ang-1 induced neovascularization with increased blood flow. Ang-1 overexpression resulted in mature and stable blood vessel formation and may be a more potent arteriogenic inducer for neovascularization.展开更多
The expression of angiopoietin- 1 （Ang- 1） and thrombospondin- 1 （TSP- 1） in 5/6 subtotal nephrectomy （STN） rats model, and its correlation to the renal microvasculature injury were investigated. Rat 5/6 STN mod...The expression of angiopoietin- 1 （Ang- 1） and thrombospondin- 1 （TSP- 1） in 5/6 subtotal nephrectomy （STN） rats model, and its correlation to the renal microvasculature injury were investigated. Rat 5/6 STN model was established in adult male SD rats, and the sham-operated group and 5/6 STN group were set up. The renal function and histopathological changes were examined at the 1st, 2nd, 4th, 8th and 12th week after operation. The expression orAng-1, TSP-1 and CD31 in renal tissues was detected by using immunohistochemistry. From 2nd to 8th week after operation, Ang-1 was significantly expressed in glomeruli of rats with STN. Ang-1 staining in glomeruli of STN group was increased significantly as compared with that in sham-operated group at 4th and 8th week after operation, and subsequently decreased after the 12th week. The expression of TSP-1 was increased significantly in STN group. As compared with sham-operated group, the CD31 expression was significantly down-regulated from the 2nd week. The expression of Ang-1 mRNA was detected by using RT-PCR at the same time points. The expression of Ang-1 mRNA in renal tissue of rats with STN was significantly up-regulated at the 2nd, 4th and 8th week after operation as compared with that in STN group at other time points or in sham-operated group at the same time points, while decreased evidently at the 12th week as compared with that in sham-operated group. It is concluded that there are changes in the mRNA expression of Ang-1, and the significant up-regulation of the expression of TSP-1 in renal tissue of rats with STN, which may be involved in the remnant renal microvasculature injury.展开更多
Doxorubicin （Dox） is a major anticancer chemotherapeutic agent. However, it causes cardiomyopathy due to the side effect of cardiomyocyte apoptosis. We have previously reported that angiopoietin-1 significantly redu...Doxorubicin （Dox） is a major anticancer chemotherapeutic agent. However, it causes cardiomyopathy due to the side effect of cardiomyocyte apoptosis. We have previously reported that angiopoietin-1 significantly reduced myocardial infarction after ischemic injury and protected cardiomyocytes from oxidative stress-induced apoptosis. It is hypothesized that angiopoietin-1 may protect cardiomyocytes from Dox-induced apoptosis. Cardiomyocytes H9C2 were transfected with adenovirus expressing angiopoietin-1 （Ad5-Ang-1） 24 h before the cells were chal- lenged with Dox at a concentration of 2 ~tmol/L. Ad5-GFP served as the vector control. Cardiomyocyte apoptosis was evaluated using Annexin V-FITC staining and caspase-3 and caspase-8 activity was determined by Western blotting. The results showed that Dox treatment significantly induced cardiomyocyte apoptosis as evidenced by the greater number of Annexin V-FITC stained cells and increases in caspase-3 and caspase-8 activity. In contrast, overexpression of angiopoietin-1 significantly prevented Dox-induced cardiomyocyte apoptosis. To elucidate the mechanisms by which angiopoietin-1 protected cells from Dox-induced apoptosis, we analyzed both extrinsic and intrinsic apoptotic signaling pathways. We observed that angiopoietin-1 prevented Dox-induced activation of both extrinsic and intrinsic apoptotic signaling pathways. Specifically, angiopoietin-1 prevented DOX-induced in- creases in FasL and Bax levels and cleaved caspase-3 and caspase-8 levels in H9C2 cells. In addition, overexpres- sion of angiopoietin-1 also activated the pro-survival phosphoinositide-3 kinase （PI3K）/Akt signaling pathway and decreased Dox-induced nuclear factor-kappaB （NF-~：B） activation. Our data suggest that promoting the expression of angiopoietin-1 could be a potential approach for reducing Dox-induced cardiomyocyte cytoxicity.展开更多
Introduction: Endometriosis affects up to 1 every 5 women at their reproductive age, with variable and complex symptomatology. Patients may be asymptomatic but may have pain episodes or subfertility. Its negative impa...Introduction: Endometriosis affects up to 1 every 5 women at their reproductive age, with variable and complex symptomatology. Patients may be asymptomatic but may have pain episodes or subfertility. Its negative impact is on patients’ health and quality of life. Objective: it was to investigate the serum and peritoneal fluid (PF) concentrations of Angiopoietin- 2, Interleukin-1β, and Vascular Endothelial Growth Factor, aiming to evaluate their diagnostic performance in endometriosis. Methods: Serum and peritoneal fluid samples were taken from 112 women undergoing laparoscopy for infertility, pelvic pain or adnexal masses. 61 diagnosed with endometriosis and 51 controlled. Primary outcome was to estimate serum and PF concentrations of Angio-2, IL-1β and VEGF and secondarily correlate these concentrations to disease stages thus assuming their diagnostic potential. Results: Significant differences were found between patients and control as regards serum and PF concentration of all studied markers except serum IL-1β. Serum Angio-2 and PF VEGF showed a significantly higher level in more advanced stages of endometriosis. PF VEGF showed a positively significant correlation with the stage of the disease, spearman coefficient t = 0.442 p = 0.014. PF concentrations of Angio-2 and Serum VEGF did not show significant pattern changes with stage-related levels. Diagnostic potential of serum and PF concentrations of the 3 markers were assessed by the ROC curve. Angio-2 proved an excellent diagnostic ability for endometriosis. PF and serum VEGF proved an equal diagnostic performance, whereas, PF IL-1β was the least efficient. Based on the results, we suggested preliminary serum threshold values for these markers to be used as diagnostic or follow-up landmarks with relatively acceptable sensitivity, specificity, positive and negative predictive values. Conclusion: Non-invasive predictive biomarkers for endometriosis were Serum Angio-2, IL-1β, and VEGF independently or in combination with the estimated threshold values. Serum Angio-2 merit is considered as a novel marker for endometriosis due to its diagnostic power.展开更多
Angiogenesis in ischemic tissue is a complex and multi-gene event. In the study, we constructed hypoxic re-sponse elements （HRE） and the Tet-On advanced double-controlled systems and investigated their effects on th...Angiogenesis in ischemic tissue is a complex and multi-gene event. In the study, we constructed hypoxic re-sponse elements （HRE） and the Tet-On advanced double-controlled systems and investigated their effects on the expression of hVEGF165 and angiopoietin-1 （Ang-1） genes in rat cardiomyocytes exposed to hypoxia and pharma-cologic induction. We infected neonatal rat cardiomyocytes with recombinant rAAV-rtTA-Rs-M2/rAAV-TRE-Tight-Ang-1 and rAAV-9HRE- hVEGF165. Our results indicated that the viral titer was 1×1012 vg /mL and the viral purity exceeded 98%. hVEGF165 expression was induced by hypoxia, but not by normoxia （P 0.001）. Ang-1 expression was evident under doxycycline induction, but undetectable without doxycycline induction （P 0.001）. Immunofluorescence staining showed that positively stained hVEGF165 and Ang-1 protein appeared only under both hypoxia and doxycycline induction. We demonstrate here that HRE and the recombinant Tet-On advanced double gene-controlled systems sensitively regulate the expression of hVEGF165 and Ang-1 genes in an altered oxygen environment and under pharmacological induction in vitro.展开更多
Objective： To study the expression and the significance of Angiopoietin-1 （Ang-1） through observing the correlations among the expression of Ang-1 to clinicopathologic characteristics and microvessel density （MVD...Objective： To study the expression and the significance of Angiopoietin-1 （Ang-1） through observing the correlations among the expression of Ang-1 to clinicopathologic characteristics and microvessel density （MVD） in oral squamous cell carcinoma （OSCC）. Methods： Expressions of Angiopoietin-1 and CD34 in 41 human OSCC tissues, 30 adjacent noncancerous oral tissues and 10 normal oral mucosas were detected by immunohistochemical SABC method. MCD was also assessed. Results： Of the 41 OSCC tissues, 41.46% （17/41） was Ang-1 positive. The expression of Ang-1 was significantly lower in OSCC than that in adjacent noncancerous oral tissues （P〈0.05） and normal oral mucosa （P〈0.05）. The Ang-1 expression was significantly higher in high differentiated tumor than that in moderately differentiated tumor （P〈0.05）. The MVD was significantly higher in Ang-1-negative OSCC than in Ang-1-positive OSCC （P〈0.01）, and negatively correlated with the expression of Ang-1 （r=-0.32, P〈0.05）. Conclusion： Down-regulated expression of Ang-1 may play a crucial role in the development of OSCC. It negatively regulated the angiogenesis of tumor.展开更多
Angiopoietin-1/tyrosine kinase with immunoglobulin and epidermal growth factor homology domains 2 （Tie-2） is a newly discovered signaling pathway of angiogenesis. Angiogenesis benefits recovery of neurological funct...Angiopoietin-1/tyrosine kinase with immunoglobulin and epidermal growth factor homology domains 2 （Tie-2） is a newly discovered signaling pathway of angiogenesis. Angiogenesis benefits recovery of neurological functions such as swallowing. In the present study, a rat model of dysphagia following stroke was induced by middle cerebral artery occlusion to investigate the influence of low frequency electrical stimulus with bidirectional square waves and triangular waves on angiopoietin-1/-13e-2 mRNA expression. Reverse transcription-polymerase chain reaction results showed that low frequency electrical stimulus significantly improved the neurological scores of the model rats, and increased angiopoietin-1/＇13e-2 mRNA expression. This demonstrates that low frequency electrical stimulation can ameliorate neurological function in rats with focal brain ischemia, potentially through regulation of angiopoietin-1/-13e-2 expression in the angiogenesis pathway.展开更多
Angiopoietin (Ang),as a cytokine found in recent years that can promote angiogenesis,belongs to a growth factor family including Ang 1,Ang 2,Ang 3 and Ang 4.Most research focus on Ang 1 and its receptor Tie 2.Differen...Angiopoietin (Ang),as a cytokine found in recent years that can promote angiogenesis,belongs to a growth factor family including Ang 1,Ang 2,Ang 3 and Ang 4.Most research focus on Ang 1 and its receptor Tie 2.Different from vascular endothelial growth factor,Ang 1/Tie 2 affect the end of angiogenesis,induce cell migration and maintain survive of endothelial cell and play an important role in angiogenesis and maintenance of the stability of the neoformative capillary network.Ang 1 is composed of 498 amino acids.The homology between human and mouse is 96.7%.Ang 1 is extensively distributed in tissues containing rich blood vessels such as muscle,prostate,ovary,uterus,but little is in tissues which have no or a few blood vessel.Adenovirus vector is able to transfect effectively Ang 1 gene to MSC.Previous studies have demonstrated that MSCAng 1 cell can highly express and secrete Ang 1 protein.MSCAng 1 could survive in the cardiac muscle tissue of acute myocardial infarction rat,and express exogenous Ang 1 protein for a period time.In acute myocardial infarction,MSCAng 1 cell have more potent effect on prompting angiogenesis than MSC cell.Moreover,MSCAng 1 cell could further reduce infarct size,increase thickness of cardiac ventricle wall,and improve cardiac muscle reconstitution.This study aim to find differential expressed protein related with Ang 1 using proteomic technologies.Protein spots showing significant difference by gel image analysis and statistics analysis were selected for identification using ESI MS/MS.We hope this work can provide new clues for the study on mechanism of action of Ang 1.展开更多
文摘Ephrin-B2 has been shown to participate in angiogenesis, but the underlying mechanisms involved remain unclear. In this study, a rat model of local cerebral ischemia was prepared by focal middle cerebral artery occlusion, followed by 24-hour reperfusion. Then, ephrin-B2 protein was administered intracerebroventricularly for 3 consecutive days via a micro-osmotic pump. Western blot assay and quantitative real-time reverse transcription PCR demonstrated the expression levels of angiopoietin-1 （Ang-1） mRNA and protein in the penumbra cortex of the ephrin-B2 treated group were decreased at day 4 after reperfusion, and increased at day 28, while the expression levels of angiopoietin-2 （Ang-2） were highly up-regulated at all time points tested. Double immunofluorescent staining indicated that Ang-1 and Ang-2 were both expressed in vascular endothelial cells positive for CD31. These findings indicate that ephrin-B2 influences the expressions of Ang-1 and Ang-2 during angiogenesis following transient focal cerebral ischemia.
基金The Natural Science Foundation of China, No.30471718
文摘AIM: To determine the inhibitory effect of the adenovirus- based angiopoietin-1 (Ang-1) targeted small interfering RNA expression system (Ad/Ang-1si) on the expression of the Ang-1 gene, cell growth and apoptosis in human esophageal cancer cell line Eca109. METHODS: siRNA-expressing adenovirus targeting Ang-1 gene was constructed using the Ad Easy System. Cultured Eca109 cells were transfected with Ad/Ang-1si (Eca109/Ang-1si), and Ad/si was used to infect Eca109 cells as control (Eca109/si). Ang-1 gene expression and concentration was determined with RT-PCR and ELISA, respectively. Human umbilical vein endothelial cell (HUVEC) migration and proliferation were analyzed. After s.c. injection into athymic nu/nu mice, the tumor growth, vessel density and apoptosis of each group was also determined. RESULTS: HUVEC migration induced by conditioned medium from Ang-1si-transfected Eca109 cells was significantly less than that induced by conditioned medium from Eca109 cells and control adenovirus- transfected Eca109 cells. Furthermore, after s.c. injection into athymic nu/nu mice, the tumor growth and cell apoptosis of Ad/Ang-1si -expressing Eca109 cells was significantly lower than that of parental or control adenovirus-transfected cells. Vessel density assessed by CD31 immunohistochemical analysis and Ang-1 expression by RT-PCR were also decreased. CONCLUSION: The targeting Ang-1 may provide a therapeutic option for esophageal cancer.
基金Supported by the Ministerio de Ciencia e Innovación(SAF:2010/21805,partially)CIBERehd(Instituto de Salud CarlosⅢ,Madrid)+1 种基金Fundación Mutua Madrile■a(to Moreno-Otero R)a grant from Asociación Espa■ola Contra el Cáncer(AIO 2010,AECC,to Sanz-Cameno P)
文摘AIM To evaluate the efficacy of peripheral blood concentrations of angiopoietins(Ang) as cirrhosis biomarkers of chronic hepatitis C(CHC).METHODS Ang1 and Ang2 serum levels were measured by enzyme-linked immunosorbent assays(ELISA) in samples from 179 cirrhotic and non-cirrhotic CHC patients, classified according to the METAVIR system.Groups were compared by non-parametric MannWhitney U test. Subsequently, the association of peripheral concentrations of angiopoietins with the stage of fibrosis was analyzed using Spearman correlation test. Finally, the accuracy, sensitivity and specificity of circulating angiopoietins for cirrhosis diagnosis were determined by the study of the respective area under the curve of receiver operator characteristics(AUC-ROC).RESULTS Peripheral blood concentrations of Ang1 and Ang2 in CHC patients were significantly related to fibrosis. While Ang1 was decreased in cirrhotic subjects compared to non-cirrhotic(P < 0.0001), Ang2 was significantly increased as CHC progressed to the end stage of liver disease(P < 0.0001). Consequently, Ang2/Ang1 ratio was notably amplified and significantly correlated with fibrosis(P < 0.0001). Interestingly, the individual performance of each angiopoietin for the diagnosis of cirrhosis reached notable AUC-ROC values(above 0.7, both), but the Ang2/Ang1 ratio was much better(AUC-ROC = 0.810) and displayed outstanding values of sensitivity(71%), specificity(84%) and accuracy(82.1%) at the optimal cut-off(10.33). Furthermore, Ang2/Ang1 ratio improved the performance of many other previously described biomarkers or scores of liver cirrhosis in CHC.CONCLUSION Ang2/Ang1 ratio might constitute a useful tool for monitoring the progression of chronic liver disease towards cirrhosis and play an important role as therapeutic target.
文摘AIM： To evaluate the effects of angiopoietin-1 （Ang-1） on adhesion of gastric cancer cell line BGC-823 and expression of integrin β1, CD44V6, urokinase-type plasminogen activator （uPA） and matrix metalloproteinase-2 （MMP-2）. METHODS： BGC-823 cells were transfected transiently with adenovirus-Ang-1 （Ad-Ang-1）. Cells transfected transiently with adenovirus-green fluorescent protein （Ad-GFP） and untransfected cells were used as a negative and blank control group, respectively. The cell adhesion rate between cell and extracellular matrix （ECM） was determined by cell adhesion assay. To investigate whether Ang-1 could reinforce gastric carcinoma metastasis, we performed migration and invasion assays in BGC-823 cells. The mRNA and protein expression of integrin β1, CD44V6, uPA and MMP-2 were detected by reverse transcription polymerase chain reaction and Western blotting, respectively. The expression of integrin β1 and CD44V6 was measured by immunohistochemistry. RESULTS： BGC-823 cells were transfected successfully. The adhesion rate increased significantly in the Ad-Ang-1 group （P 〈 0.05）. The Ad-Ang-1-transfected group had a significant increase in migration and invasion compared with that of the mock-transfected and Ad-GFP groups. The mRNA and protein expression of integrin β1, CD44V6, uPA and MMP-2 in the Ad- Ang-1 group was higher than that in the Ad-GFP and blank control groups （P 〈 0.05）. Compared with mocktransfected and Ad-GFP groups, integrin 131 and CD44V6 expression intensity greatly increased （P 〈 0.05）. CONCLUSION： Transfection of Ang-1 into human gastric cancer cell line BGC-823 can significantly increase expression of integrin β1 and CD44V6, by which cell adhesion and metastasis to the ECM are promoted.
文摘Objectives This study investigated the efficacy of human skeletal myoblasts (SkM) mediated either human vascular endothelial growth factor-165 (hVEGF165) or angiopoietin-1 (Ang-1) on vascular development and myocardial regional perfusion. Methods A porcine heart model of chronic infarction was created in 28 female swine by coronary artery ligation. The animals were randomized into: (1) group-1, DMEM injected (n=6), (2) group-2, Ad-null transduced SkM transplanted (n=6), (3) group-3, Ad-hVEGF165 transduced SkM transplanted (n=8), and (4) group-4, Ad-Ang-1 transduced SkM (n=8). Three weeks later, 5 ml DMEM containing 3×108 SkM carrying exogenous genes were intramyocardially injected into 20 sites in left ventricle in groups-2, -3 and -4. Animals in group-1 were injected 5 ml DMEM without cells. Animals were kept on 5 mg/kg cyclosporine per day for 6 weeks. Regional blood flow was measured using fluorescent microspheres. The heart was explanted at 2, 6 and 12 weeks after transplantation for histological studies. Results Histological examination showed survival of lac-z expressing myoblasts in host tissue. Capillary density based on Von Willebrand factor-VIII (vWF-VIII) at low power field (×100) was 57.13±11.85 in group-3 at 6 weeks and declined to 32.1±5.21 at 12 weeks, while it was 39.9±10.26 at 6 weeks and increased to 45.14±6.54 at 12 weeks in group-4. The mature blood vessel index was highest in group- 4 at 6 and 12 weeks after transplantation. The regional blood flow in the center and peri-infarct area was significantly increased in animals of groups-3 and -4. Conclusions SkM carrying either hVEGF165 or Ang-1 induced neovascularization with increased blood flow. Ang-1 overexpression resulted in mature and stable blood vessel formation and may be a more potent arteriogenic inducer for neovascularization.
文摘The expression of angiopoietin- 1 （Ang- 1） and thrombospondin- 1 （TSP- 1） in 5/6 subtotal nephrectomy （STN） rats model, and its correlation to the renal microvasculature injury were investigated. Rat 5/6 STN model was established in adult male SD rats, and the sham-operated group and 5/6 STN group were set up. The renal function and histopathological changes were examined at the 1st, 2nd, 4th, 8th and 12th week after operation. The expression orAng-1, TSP-1 and CD31 in renal tissues was detected by using immunohistochemistry. From 2nd to 8th week after operation, Ang-1 was significantly expressed in glomeruli of rats with STN. Ang-1 staining in glomeruli of STN group was increased significantly as compared with that in sham-operated group at 4th and 8th week after operation, and subsequently decreased after the 12th week. The expression of TSP-1 was increased significantly in STN group. As compared with sham-operated group, the CD31 expression was significantly down-regulated from the 2nd week. The expression of Ang-1 mRNA was detected by using RT-PCR at the same time points. The expression of Ang-1 mRNA in renal tissue of rats with STN was significantly up-regulated at the 2nd, 4th and 8th week after operation as compared with that in STN group at other time points or in sham-operated group at the same time points, while decreased evidently at the 12th week as compared with that in sham-operated group. It is concluded that there are changes in the mRNA expression of Ang-1, and the significant up-regulation of the expression of TSP-1 in renal tissue of rats with STN, which may be involved in the remnant renal microvasculature injury.
基金supported by the National Natural Science Foundation of China (No. 30971258)the National 973 project of China(NO.2012CB517503)+1 种基金the Key Project of Natural Science Foundation of Jiangsu Province (No. 11KJA310004)Jiangsu "Six Personnel Peak" Talent-Funded Projects
文摘Doxorubicin （Dox） is a major anticancer chemotherapeutic agent. However, it causes cardiomyopathy due to the side effect of cardiomyocyte apoptosis. We have previously reported that angiopoietin-1 significantly reduced myocardial infarction after ischemic injury and protected cardiomyocytes from oxidative stress-induced apoptosis. It is hypothesized that angiopoietin-1 may protect cardiomyocytes from Dox-induced apoptosis. Cardiomyocytes H9C2 were transfected with adenovirus expressing angiopoietin-1 （Ad5-Ang-1） 24 h before the cells were chal- lenged with Dox at a concentration of 2 ~tmol/L. Ad5-GFP served as the vector control. Cardiomyocyte apoptosis was evaluated using Annexin V-FITC staining and caspase-3 and caspase-8 activity was determined by Western blotting. The results showed that Dox treatment significantly induced cardiomyocyte apoptosis as evidenced by the greater number of Annexin V-FITC stained cells and increases in caspase-3 and caspase-8 activity. In contrast, overexpression of angiopoietin-1 significantly prevented Dox-induced cardiomyocyte apoptosis. To elucidate the mechanisms by which angiopoietin-1 protected cells from Dox-induced apoptosis, we analyzed both extrinsic and intrinsic apoptotic signaling pathways. We observed that angiopoietin-1 prevented Dox-induced activation of both extrinsic and intrinsic apoptotic signaling pathways. Specifically, angiopoietin-1 prevented DOX-induced in- creases in FasL and Bax levels and cleaved caspase-3 and caspase-8 levels in H9C2 cells. In addition, overexpres- sion of angiopoietin-1 also activated the pro-survival phosphoinositide-3 kinase （PI3K）/Akt signaling pathway and decreased Dox-induced nuclear factor-kappaB （NF-~：B） activation. Our data suggest that promoting the expression of angiopoietin-1 could be a potential approach for reducing Dox-induced cardiomyocyte cytoxicity.
文摘Introduction: Endometriosis affects up to 1 every 5 women at their reproductive age, with variable and complex symptomatology. Patients may be asymptomatic but may have pain episodes or subfertility. Its negative impact is on patients’ health and quality of life. Objective: it was to investigate the serum and peritoneal fluid (PF) concentrations of Angiopoietin- 2, Interleukin-1β, and Vascular Endothelial Growth Factor, aiming to evaluate their diagnostic performance in endometriosis. Methods: Serum and peritoneal fluid samples were taken from 112 women undergoing laparoscopy for infertility, pelvic pain or adnexal masses. 61 diagnosed with endometriosis and 51 controlled. Primary outcome was to estimate serum and PF concentrations of Angio-2, IL-1β and VEGF and secondarily correlate these concentrations to disease stages thus assuming their diagnostic potential. Results: Significant differences were found between patients and control as regards serum and PF concentration of all studied markers except serum IL-1β. Serum Angio-2 and PF VEGF showed a significantly higher level in more advanced stages of endometriosis. PF VEGF showed a positively significant correlation with the stage of the disease, spearman coefficient t = 0.442 p = 0.014. PF concentrations of Angio-2 and Serum VEGF did not show significant pattern changes with stage-related levels. Diagnostic potential of serum and PF concentrations of the 3 markers were assessed by the ROC curve. Angio-2 proved an excellent diagnostic ability for endometriosis. PF and serum VEGF proved an equal diagnostic performance, whereas, PF IL-1β was the least efficient. Based on the results, we suggested preliminary serum threshold values for these markers to be used as diagnostic or follow-up landmarks with relatively acceptable sensitivity, specificity, positive and negative predictive values. Conclusion: Non-invasive predictive biomarkers for endometriosis were Serum Angio-2, IL-1β, and VEGF independently or in combination with the estimated threshold values. Serum Angio-2 merit is considered as a novel marker for endometriosis due to its diagnostic power.
基金supported by a grant from the National Natural Science Foundation of China (No.30672081)
文摘Angiogenesis in ischemic tissue is a complex and multi-gene event. In the study, we constructed hypoxic re-sponse elements （HRE） and the Tet-On advanced double-controlled systems and investigated their effects on the expression of hVEGF165 and angiopoietin-1 （Ang-1） genes in rat cardiomyocytes exposed to hypoxia and pharma-cologic induction. We infected neonatal rat cardiomyocytes with recombinant rAAV-rtTA-Rs-M2/rAAV-TRE-Tight-Ang-1 and rAAV-9HRE- hVEGF165. Our results indicated that the viral titer was 1×1012 vg /mL and the viral purity exceeded 98%. hVEGF165 expression was induced by hypoxia, but not by normoxia （P 0.001）. Ang-1 expression was evident under doxycycline induction, but undetectable without doxycycline induction （P 0.001）. Immunofluorescence staining showed that positively stained hVEGF165 and Ang-1 protein appeared only under both hypoxia and doxycycline induction. We demonstrate here that HRE and the recombinant Tet-On advanced double gene-controlled systems sensitively regulate the expression of hVEGF165 and Ang-1 genes in an altered oxygen environment and under pharmacological induction in vitro.
基金This work was supported by the Medical Science Grant of Sichuan Province Key Sci & Tech Subjects (No. 20020807) and Guizhou Province Government Century Grant (No.9813).
文摘Objective： To study the expression and the significance of Angiopoietin-1 （Ang-1） through observing the correlations among the expression of Ang-1 to clinicopathologic characteristics and microvessel density （MVD） in oral squamous cell carcinoma （OSCC）. Methods： Expressions of Angiopoietin-1 and CD34 in 41 human OSCC tissues, 30 adjacent noncancerous oral tissues and 10 normal oral mucosas were detected by immunohistochemical SABC method. MCD was also assessed. Results： Of the 41 OSCC tissues, 41.46% （17/41） was Ang-1 positive. The expression of Ang-1 was significantly lower in OSCC than that in adjacent noncancerous oral tissues （P〈0.05） and normal oral mucosa （P〈0.05）. The Ang-1 expression was significantly higher in high differentiated tumor than that in moderately differentiated tumor （P〈0.05）. The MVD was significantly higher in Ang-1-negative OSCC than in Ang-1-positive OSCC （P〈0.01）, and negatively correlated with the expression of Ang-1 （r=-0.32, P〈0.05）. Conclusion： Down-regulated expression of Ang-1 may play a crucial role in the development of OSCC. It negatively regulated the angiogenesis of tumor.
基金the National High-Tech R&D Program of China (863 Program), No.2007AA022Z482
文摘Angiopoietin-1/tyrosine kinase with immunoglobulin and epidermal growth factor homology domains 2 （Tie-2） is a newly discovered signaling pathway of angiogenesis. Angiogenesis benefits recovery of neurological functions such as swallowing. In the present study, a rat model of dysphagia following stroke was induced by middle cerebral artery occlusion to investigate the influence of low frequency electrical stimulus with bidirectional square waves and triangular waves on angiopoietin-1/-13e-2 mRNA expression. Reverse transcription-polymerase chain reaction results showed that low frequency electrical stimulus significantly improved the neurological scores of the model rats, and increased angiopoietin-1/＇13e-2 mRNA expression. This demonstrates that low frequency electrical stimulation can ameliorate neurological function in rats with focal brain ischemia, potentially through regulation of angiopoietin-1/-13e-2 expression in the angiogenesis pathway.
文摘Angiopoietin (Ang),as a cytokine found in recent years that can promote angiogenesis,belongs to a growth factor family including Ang 1,Ang 2,Ang 3 and Ang 4.Most research focus on Ang 1 and its receptor Tie 2.Different from vascular endothelial growth factor,Ang 1/Tie 2 affect the end of angiogenesis,induce cell migration and maintain survive of endothelial cell and play an important role in angiogenesis and maintenance of the stability of the neoformative capillary network.Ang 1 is composed of 498 amino acids.The homology between human and mouse is 96.7%.Ang 1 is extensively distributed in tissues containing rich blood vessels such as muscle,prostate,ovary,uterus,but little is in tissues which have no or a few blood vessel.Adenovirus vector is able to transfect effectively Ang 1 gene to MSC.Previous studies have demonstrated that MSCAng 1 cell can highly express and secrete Ang 1 protein.MSCAng 1 could survive in the cardiac muscle tissue of acute myocardial infarction rat,and express exogenous Ang 1 protein for a period time.In acute myocardial infarction,MSCAng 1 cell have more potent effect on prompting angiogenesis than MSC cell.Moreover,MSCAng 1 cell could further reduce infarct size,increase thickness of cardiac ventricle wall,and improve cardiac muscle reconstitution.This study aim to find differential expressed protein related with Ang 1 using proteomic technologies.Protein spots showing significant difference by gel image analysis and statistics analysis were selected for identification using ESI MS/MS.We hope this work can provide new clues for the study on mechanism of action of Ang 1.
