• Title, Summary, Keyword: endostatin

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Protein Kinase B Inhibits Endostatin-induced Apoptosis in HUVECs

  • Kang, Hee-Young;Shim, Dong-Hwan;Kang, Sang-Sun;Chang, Soo-Ik;Kim, Hak-Yong
    • BMB Reports
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    • v.39 no.1
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    • pp.97-104
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    • 2006
  • Endostatin is a tumor-derived angiogenesis inhibitor, and the endogenous 20 kDa carboxyl-terminal fragment of collagen XVIII. In addition to inhibiting angiogenesis, endostatin inhibits tumor growth and the induction of apoptosis in several endothelial cell types. However, the mechanisms that regulate endostatin-induced apoptotic cell death are unclear. Here, we investigated apoptotic cell death and the underlying regulatory mechanisms elicited of endostatin in human umbilical vein endothelial cells (HUVECs). Endostatin was found to induce typical apoptotic features, such as, chromatin condensation and DNA fragmentation in these cells. Thus, as the phosphoinositide 3-OH kinase (PI3K)/protein kinase B (PKB) signaling pathway has been shown to prevent apoptosis in various cell types, we investigated whether this pathway could protect cells against endostatin induced apoptosis. It was found that the inhibition of PI3K/PKB significantly increased endostatin-induced apoptosis, and that endostatin-induced cell death is physiologically linked to PKB-mediated cell survival through caspase-8.

Anti-Angiogenic Activity of Mouse N-/C-terminal deleted Endostatin

  • Cho, Hee-Yeong;Kim, Woo-Jean;Lee, Sae-Won;Kim, Young-Mi;Choi, Eu-Yul;Park, Yong-Suk;Kwon, Young-Guen;Kim, Kyu-Won
    • BMB Reports
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    • v.34 no.3
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    • pp.206-211
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    • 2001
  • Endostatin, a proteolytic fragment of collagen XVIII, is a potent inhibitor of angiogenesis and the growth of several primary tumors. However, the opinions on the activity of endostatin derivatives deleted N- or C- terminal are still controversial. In this regard, we produced mouse endostatin and its derivatives in the prokaryotic system, and studied their anti-tumor activity. The [$^3H$]-thymidine incorporation assay demonstrated that N-terminal deleted mouse endostatin, and a C- and N-terminal deleted mutant, effectively inhibited the proliferation of human umbilical vein endothelial cells (HUVECs). The biological activity of endostatin was also shown by its in vivo anti-angiogenic ability on the chorioallantoic membrane (CAM) of a chick embryo. Treatment of $200\;{\mu}g$ of mouse endostatin, or N-terminal deleted mouse endostatin, inhibited capillary formation of CAM 45 to 71%, which is comparative to a 80% effect of positive control, $1\;{\mu}g$ of retinoic acid. An in vivo mouse tumor growth assay showed that N-terminal deleted mouse endostatin, and the N-/C-terminal deleted mutant, significantly repressed the growth of B16F10 melanoma cells in mice as did the full-length mouse endostatin. According to these results, N-and N-/C-terminal deleted mouse endostatins are the potent inhibitors of tumor growth and angiogenesis.

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Prognostic Values of VEGF and Endostatin with Malignant Pleural Effusions in Patients with Lung Cancer

  • Zhang, Yu;Yu, Li-Ke;Lu, Guo-Jun;Xia, Ning;Xie, Hai-Yan;Hu, Wei;Hao, Ke-Ke;Xu, Chun-Hua;Qian, Qian
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.19
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    • pp.8435-8440
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    • 2014
  • Aims: Angiogenesis is important in malignant pleural effusion (MPE) formation and it is regulated by a number of pro- and anti-angiogenic cytokines. The purpose of this study was to evaluate the prognostic value of angiogenic factor vascular endothelial growth factor (VEGF) and angiogenesis inhibitor endostatin in lung cancer patients with MPE, and investigate the relationship between these two kinds of agent. Methods: Using enzyme-linked immunoadsorbent assay, the concentrations of VEGF and endostatin were measured in pleural effusions (PE) and serum from a total of 70 lung cancer patients with MPE and 20 patients with tuberculosis. Results: Compared to patients with tuberculosis, the levels of VEGF and endostatin in both PE and serum were significantly higher in patients with lung cancer. There were statistically significant correlations between VEGF levels in PE and serum (r=0.696, p<0.001), endostatin levels in PE and serum (r=0.310, p=0.022), and VEGF and endostatin levels in PE (r=0.287, p=0.019). Cox multivariate analysis revealed that elevated pleural VEGF and endostatin levels and serum endostatin level were independent predictors of shorter overall survival. Conclusion: Both pro- and anti-angiogenic factors are likely contributors to PE formation. Our results suggest that the levels of VEGF and endostatin in PE, together with endostatin in serum, may be potential prognostic parameters for lung cancer patients with MPE.

Soluble Expression of Human Angiostatin and Endostatin by Maltose Binding Protein (MBP) Fusion in E. coli (Maltose Binding Protein 융합단백질에 의한 인간유래의 앤지오스타틴과 앤도스타틴의 대장균에서 수용성 단백질발현)

  • Paek, Seon-Yeol;Choi, Shin-Geon
    • Journal of Industrial Technology
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    • v.28 no.B
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    • pp.59-63
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    • 2008
  • Rapid production of therapeutic proteins such as angiostatin and endostatin angiogenic inhibititors has been highly demanded for cancer treatment. In this regard, recombinant human angiostatin and endostatin were successfully expressed as soluble forms by maltose binding protein (MBP)-mediated fusion expression in Escherichia coli. PCR amplified, angiostatin and endostatin genes from human placenta cDNA library were inserted into an expression vector pMAL-c2e to construct prokaryotic expression vectors, pMAL-c2e/AS and pMAL-c2e/ES, respectively. Recombinant angiostatin and endostatin were efficiently expressed in E. coli origami (DE3) after IPTG induction and protein expression were confirmed by SDS-PAGE analyses. The expressed recombinant proteins were purified near homogenity using an amylose affinty column chromatography. In contrast that previous E. coli expressions were all insoluble, our results first time demonstrated that MBP fused human angiostatin and endostatin were soluble in E. coli.

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Comparative Study on Trichoplusia ni Tn 5B1-4 Cells and Bombyx mori BmN Cells for Recombinant Endostatin Production

  • Sohn, Bong-Hee;Lee, Jong-Min;Kang, Pil-Don;Lee, Sang-Uk;Kim, Yong-Soon;Chung, In-Sik
    • International Journal of Industrial Entomology
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    • v.7 no.2
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    • pp.197-201
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    • 2003
  • The recombinant plasmids harboring a heterologous gene coding mouse endostatin were transfected and expressed stably in Trichoplusia ni Tn 5B1-4 cells and Bombyx mori BmN cells, respectively. Recombinant endostatin expressed in the stably transformed Tn 5B1-4 and BmN cells was secreted into the medium. BmN cells are relatively lower in maximum cell growth and recombinant endostatin production than Tn 5B 1-4 cells. Recombinant endostatin was also purified to homogeneity using a simple one-step ${Ni^2+}$ affinity fractionation method. Purified recombinant endostatin inhibited endothelial cell proliferation in a dose-dependent manner. The concentration at half-maximum inhibition $({ED_50})$ for recombinant endostatin was approximately 0.35 ${\mu}g$/ml.

Influence of vascular endothelial growth factor (VEGF) and endostatin on coronary artery lesions in Kawasaki disease (가와사끼병에서 혈청 VEGF 및 endostatin의 변화가 관상동맥병변의 발생에 미치는 영향)

  • Kim, Seon A;Jung, Bum Suk;Yoon, Jong Seo;Han, Ji Whan;Lee, Joon Sung
    • Korean Journal of Pediatrics
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    • v.50 no.12
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    • pp.1241-1246
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    • 2007
  • Purpose : Recently, there has been several studies to clarify the pathogenesis of Kawasaki disease (KD) and the relations of VEGF and endostatin that act on vascular endothelial cells to the coronary artery complications. In this report, we measured serum levels of VEGF and endostatin in acute and subacute phases of KD to assess the change of these levels and the relations to the development of coronary artery lesions (CAL). Methods : Twenty six patients were diagnosed and treated for KD between January, 2001 and July, 2005 at Kangnam St. Mary's Hospital, the Catholic University of Korea. They were divided into those with and without CAL. Serum levels of VEGF and endostatin were measured during acute and subacute phases and compared to those measured in healthy and disease control groups. Results : Serum levels of VEGF were increased in KD but no differences were noted in KD with and without CAL. Serum levels of endostatin were decreased in the acute phase of KD, however they were recovered in the subacute phase of KD, regardless of CAL. The VEGF/endostatin ratio was increased in KD. KD without CAL showed a relative decrease in this ratio during the subacute phase. Significant positive correlations were found between serum VEGF and WBC count, VEGF and ESR, VEGF/endostatin ratio and ESR in the acute phase of KD. Conclusion : Analysis of factors influencing the vascular endothelium such as VEGF and endostatin will help to clarify the etiology of KD and the pathogenesis of CAL.

Clinical Observation and Therapeutic Evaluation of Rh-endostatin Combined with DP Regimen in Treating Patients with Advanced Esophageal Cancer

  • Deng, Wen-Ying;Song, Tao;Li, Ning;Luo, Su-Xia;Li, Xiang
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.16
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    • pp.6565-6570
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    • 2014
  • Objective: To observe the curative effects of rh-endostatin combined with DP regimen in treating patients with advanced esophageal cancer and analyze the correlation of CT perfusion (CTP) parameters and the expression of vascular endothelial growth factor (VEGF). Methods: Twenty patients with esophageal cancer confirmed pathologically were randomly divided into combined treatment (rh-endostatin+DP regimen) group and single chemotherapy group, 10 patients in each group, respectively. All patients were given conventional CT examination and CTP imaging for primary tumor. The level of VEGF, the size of tumor and CTP parameters (BF, BV, PS and MTT) before treatment and after 2 cycles of treatment were determined for the comparison and the correlation between CTP parameters and VEGF expression was analyzed. Results: the therapeutic effect of rh-endostatin+DP regimen group was superior to single chemotherapy group. VEGF level after treatment in rh-endostatin+DP regimen group was obviously lower than single chemotherapy group (P<0.01). The expression of VEGF had positive correlation with BF and BV but negative correlation with MTT. Compared with treatment before for rh-endostatin+DP regimen group, BF, BV and PS decreased while MTT increased after treatment (P<0.05). However, there were no significant differences between treatment before and after treatment in single chemotherapy (P>0.05). Conclusions: Rh-endostatin can down-regulate the expression of VEGF in esophageal cancer, change the state of hypertransfusion and high permeability of tumor vessels and had the better curative effect and slighter adverse reactions when combined with chemotherapy.

The Orientation-Dependent Expression of Angiostatin-Endostatin Hybrid Proteins and Their Characterization for the Synergistic Effects of Antiangiogenesis

  • Paek, Sun-Yeol;Kim, Yong-Seok;Choi, Shin-Geon
    • Journal of Microbiology and Biotechnology
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    • v.20 no.10
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    • pp.1430-1435
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    • 2010
  • Two angiostatic fusion proteins (hAE and hEA), differing in tandem connection manners, were constructed from human angiostatin (hAS) and endostatin (hES) proteins. These fusion proteins were then evaluated for synergistic antiangiogenic properties. The 65 kDa secreted fusion proteins, expressed in Pichia pastoris, were verified by both mass analysis and Western blotting assay. Luciferase reorter gene assay, using a VEGF promoter, revealed that the angiostatin-endostatin fusion protein (hAE), and its corresponding fusion gene delivery on human microvascular endothelial cells (HMEC-1), resulted in a more potent synergistic antiangiogenic effect than the endostatin-angiostatin fusion protein (hEA). These results suggest that the orientation of the fusion genes in hAS and hES might be an important factor in the development of therapeutic proteins.

Efficacy Evaluation of Tissue Inhibitor of Metalloproteinases-2 and Endostatin on Angiogenesis (Tissue Inhibitor of Metalloproteinases-2와 Endostatin의 혈관신생 제어 효능 평가)

  • Kim, Soo-Hyeon;Cho, Young-Rak;Yoon, Hyun-Jae;Ko, Hee-Young;Kim, Pyeung-Hyeun;Seo, Dong-Wan
    • YAKHAK HOEJI
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    • v.54 no.6
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    • pp.488-493
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    • 2010
  • Therapeutic manipulation of angiogenesis, the formation of new vascular sprouts from existing capillaries, is one of the promising strategies for treatment of human diseases such as cancer, arthritis, and cardiovascular disorder. In the present study, we examined the effects and molecular mechanism of tissue inhibitor of metalloproteinases-2 (TIMP-2) and endostatin on fibroblast growth factor-2 (FGF-2)-stimulated endothelial cell proliferation, migration and adhesion in vitro, and angiogenesis in vivo. TIMP-2 and endostatin showed potent anti-angiogenic activity in vitro and in vivo. These effects appear to be mediated through different angiogenic signaling pathways. Collectively, our findings demonstrate that TIMP-2 and endostatin strongly inhibit FGF-2-induced angiogenic responses, and the establishment of fast and reproducible evaluation system in vitro and in vivo for the development of anti-angiogenic biomaterials and therapeutics.

Effect of Transcriptional Terminator Sequences on Recombinant Protein Expression from Drosophila melanogaster S2 Cell (전사 종결 염기 서열이 Drosophila melanogaster Schneider line 2 세포에서 외래 단백질의 발현에 미치는 영향)

  • Hwang, In-Sook;Park, Jong-Hwa;Lee, Youn-Hyung;Yoon, Jae-Seung;Baek, Kwang-Hee;Chung, In-Sik
    • Applied Biological Chemistry
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    • v.44 no.4
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    • pp.211-214
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    • 2001
  • Utilizing the foreign gene expression system of Drosophila melanogaster Schneider line 2(S2) cell, the degree of transient protein and mRNA expression was examined with different terminators. In the case of transient expression, the expression level of green fluorescent protein(GFP) was the highest when the transfection agent was eliminated and then cultivated for 36 to 48 hr. The terminators(SV40 p(A), SV40 small T-antigen and human gastrin 3'UTR) of the expression vector system were each cloned with endostatin; thereafter, the expression levels of the endostatin and its mRNA were compared. When the expression levels of endostatin were compared 36 hr after transfection, the SV40 p(A) terminator showed the highest expression level of endostatin and its mRNA.

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