Tehran University Medical Journal

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Tumor angiogenesis shown by Microvessel Count (MVC) or Microvessel Density (MVD), is assessed by several studies as prognostic factor in some types of tumors, and also in colorectal carcinoma. This article is payed to correlation between clincopathologic factors and tumor angiogenesis. In this study, immunohistochemical techniques are used for vascular evaluation in specimens from twenty-nine colorectal carcinoma, and stained for Factor VIII-Related Antigen (F8RA) by using monoclonal antibody. Uni and multivariate analysis disclosed that total MVC was higher in tumor [76.3±33 (×100=2.5 mm²/field) and 29.8±11 (×200=0.785 mm²/field)] than in normal tissue [37.7±15.8 (×100) and 17.6±7.8 (×200)], (P=0.022, P=0.000009). Microvessel quantification was significantly higher in stage D (115±36.6, ×100 and 26.7±6.4, ×200, P=0.002 and P=0.04). In this study MVD has correlation with vascular invasion (P=0.024, ×100 and P=0.007, ×200), the mean tumor vessel count although was increased with clinicophatologic findings such as age<60 years, male, right colon involvement, infiltrating type, mucinous carcinoma, transmural penetration, grade III, lymphatic and perineural invasion, but was not statistically significant. Lymph node and hematogenous metastasis and size of tumor also, was not important. As a conclusion, MVD was increased in tumor and has shown correlation with metastasis, and vascular invasion. Resulting angiogenesis increase risk of metastasis.

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Background: The important of angiogenesis for the progressive growth and viability of solid tumors is well established. Only few data are available for hematologic neoplasms.

Materials and Methods: To investigate the role of angiogenesis in the acute myloid leukemia (AML) bone marrow biopsies from 30 adults with newly diagnosed, untreated AML(day 0) were evaluated. Further studies were done after completion on remission induction of treatment (day 35 of 7×3 regimen n=13, complete remission in AML (m3) treat with arsenic trioxide n=17). Micro-vessels were scored in at least 3 areas of highest micro-vessel density in representative section of each bone marrow specimen using immunohistochemistry for Von Willbrand factor.

Results: Median micro-vascular density (MVD) were in AMLM3 patients before treatment, %6.81±3.58 and after treatetment %3.48±3.06 (p<0.0001). In other AML patients MVD were befor treatment %3.38 and after treatment %3.6.

Conclusion: In conclusion, there is evidence of increased micro-vessel density in the bone marrow of patients with AML, which supports the hypothesis of an important role of angiogenesis in AML. MVD was reduced with chemotherapy and arsenic. Furthermore , these finding suggest that antiangiogenesis therapy might constitute a novel strategy for the treatment of AML.

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Background: Tumor cells need food and oxygen supply for growth and division. Therefore one of the most promising areas of cancer therapy focuses on using agents that inhibit tumor angiogenesis. Inhibition of angiogenesis prevents cell growth, division and metastasis. Previous studies showed that plasminogen related Protein-B has an anti-tumor activity in mice. This protein has a high level of homology with preactivation Peptide (PAP) of human plasminogen. According to this high homology, antiangiogeneic activity of PAP was investigated in an in vitro angiogenesis model.

Methods: PAP encoding region of human plasminogen gene was isolated by Polymerase Chain Reaction and ‎cloned in pGEX-2T vector. This plasmid was expressed in Escherichia coli as a fusion protein (GST-PAP). ‎GST-PAP was expressed as inclusion body and purified by affinity chromatography on GSH-sepharose ‎resin after refolding. antiangiogenic effects of purified protein were surveyed with Matrigel assay‏.‏‎ ‎

Results: The GST-PAP was expressed and purified and its accuracy was confirmed by SDS-PAGE analysis ‎and immunoblotting. Microscopic studies showed that GST-PAP inhibited angiogenesis in Matrigel system ‎which is shown by shrinking the length of capillary like structures and a decrease in the number of tubule. ‎While applying concentarations of 25μg/ml of GST-PAP and concentrations above that, antiangiogenic ‎activity of GST-PAP was significant comparing to the controls. ‎

Conclusion: Finding shows that GST-PAP can inhibit network formation in Matrigel system. This findings ‎support the theory that PAP is a potent angiogenesis inhibitor.‎

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Background: Plasminogen has a central role in fibrinolyrtic system can activate through various activators (PAs) to its active form plasmin and perfoem its vital function that is fibrin clot lysis. Furthermore the fibrinolyrtic system plays a major role in angiogenesis. The fibrinolyrtic system activation control cell migration and invasion. In addition to this, plasmin regulates tumor growth. Monoclonal antibodies, as biological tools, play an important role in basic researches.

Methods: In the first step the effects of antibodies on the activation of fibrinolyrtic system with PAs were evaluated with several methods including macroscopic observation, quantitative measurement of DD/E fragments by D-dimer assay and activation of plasminogen by S-2251 synthetic substrate (ELISA method), subsequently we studied the effect of antibodies on angiogenesis process in an in- vitro model.

Results: Results showed that MC2B8 that is an inhibitor of plasminogen activation in presence of plasminogen activators can inhibit angiogenesis process: A1D12 that is against N-terminal domain of Glu-plasminogen, in addition to activation of fibrinolyrtic system in presence of plasminogen activators, can activate in vitro angiogenesis process.

Conclusion: Plasmin formation is a critical step in invasion and migration of endothelial cells to form new vessels. Plasmin directly participates in angiogenesis by direct fibrin and other matrix components degradation, and indirectly by activating matrix degrading metalloproteinase and angiogenic growth factors. According to the in- vitro results, MC2B8 and A1D12 monoclonal antibodies play roles in this process in a dose dependent manner.

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Background: Despite advances in cancer diagnosis and treatment, survival rate of patients suffering from glioblastoma multiform (GBM) has not been significantly improved. Therefore, novel therapeutic adjuncts to routine therapies have been suggested over time. Inhibition of angiogenesis by antiangiogenic drugs is one of the new approaches to inhibit the growth of malignant cells. Microvessel density (MVD) assay is a technique performed by counting immunohistochemically-stained blood vessels. Nowadays, athymic nude mice are widely used for the establishment of xenograft tumor models in cancer research. The aim of this study was to evaluate the MVD of autochthonous xenograft models of GBM isolated from Iranian patients for use in pharmaceutical research on antiangiogenic drugs.Methods: Fresh tumor samples of GBM were obtained from three patients in Cancer Institute of Tehran University of Medical Sciences in Fall of 2010 and Winter of 2011. After preliminary processing, minced tumor samples were implanted heterotopically on flanks of athymic nude mice. Two months later, the animals were sacrificed and the xenograft tumor samples were sent to the pathology laboratory. After establishing the proof of the xenograft tumor type, MVD-CD34, an endothelial cell marker, was assessed by counting hot spot areas in 22 samples.Results: The mean number of microvessels in these xenograft tumor models was 30±2.1. Conclusion: These autochthonous xenograft models of GBM can be used in preclinical settings for research on antiangiogenic drugs regarding a pharmacogenomics-based treatment regimen for the Iranian population. Moreover, such models can be used in future studies for determining the sensitivity or resistance to antiangiogenic drugs in individualized cancer therapy.

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Background: Colorectal cancer is a major cause of morbidity and mortality throughout the world, and its treatments include surgery, chemo-radiotherapy. Despite improvements in clinical outcomes of patients with this tumor over the past decades, prognosis remains poor with a 5-year survival rate of <10%. Angiogenesis inhibitor agents have been recently added to the treatment regimen of this disease. In the past two decades, it has been recognized that selective inhibitors of the cyclooxygenase -2 (Cox-2) enzyme result in the regression in the size of colorectal tumor, and one of its reasons is attributed to angiogenesis inhibition. The present study aimed at identifying the molecular pathways of angiogenesis inhibition by celecoxib. Methods: HCT-116, which is one of the cell lines of Colorectal cancer (separated from human colorectal adenocarcinoma) was provided by the National Cell bank of Iran (NCBI) affiliated to Pasteur Institute. It was then cultured in DMEM (high glucose) culture medium containing 10% FBS, and then treated in the active substance of celecoxib at pharmacological concentrations of 50 mM (C50) and 100 mM (C100). Afterwards, RNA was extracted and cDNA was prepared. The oligonucleotide of HIF-1 Alpha gene (angiogenesis initiator) was prepared and the level of HIF-1 alpha gene expression was assessed with a real-time PCR device in three control, C50 and C100 groups. Results: HIF-1 alpha gene expression significantly decreased in the celecoxib treatment group (compared with control group) with the concentration of C100 (P< 0.001), but no change was observed in the concentration of C50. Conclusion: Angiogenesis is a key factor in the carcinogenesis process and FDA today approved bevacizumab as a first-line treatment for patients with metastatic colorectal cancer. The results of this study showed one of the causes of angiogenesis reduction in celecoxib-treated colorectal cancer. According to clinical findings and basic studies, celecoxib will be hopefully used as a first-line therapy along with chemotherapy in the near future in colorectal cancer. The advantages of this treatment method include its low cost and low side effects.

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Angiogenesis, as the process of new vessel formation from pre-existing vessels is dependent on a delicate equilibrium between endogenous angiogenic and antiangiogenic factors. However, under pathological conditions, this tight regulation becomes lost which can result in the formation of the different diseases such as cancer. Angiogenesis is a complex process that includes many gene products that are produced by different cells. Each of the processes influenced by specific genes that their expression can be regulated by hypoxi inducible factor-1 (HIF-1). Hypoxia, the imbalance between the oxygen in need and the oxygen available, usually occurs in tumors and ischemic cardiovascular diseases. In order to overcome this challenge, tumors regulate and control the expression of genes related to angiogenesis, cell cycle and metabolism using hypoxia-inducible factor 1 (HIF-1). HIF-1 was first recognized as a transcription factor involved in hypoxia-induced erythropoietin expression. As angiogenesis pathway molecules are being described, this factor has been characterized as a key transcription regulator for these molecules. In this review article, after discussing HIF-1 structure and characterization, the role of this important factor in angiogenesis and cancer as a pathological case and finally, the clinical applications has been evaluated. Articles related to the key words of hypoxia, HIF-1 and angiogenesis were searched from valid databases such as Springer Link, google scholar, Pubmed and Sciencedirect. Then, the articles related to the role of hypoxia and HIF-1 in activation of genes that are involved in angiogenesis and cancer were searched and selected for this study. Studies show that, HIF-1 activation of genes including vascular endothelial growth factor (VEGF), angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2), etc., induced angiogenesis in the tumor cells. Furthermore, the activation of genes such as insulin-like growth factor 2 (IGF2), transforming growth factor &alpha; (TGF-&alpha;) and MAPK and PI3K signaling pathway will also enable the survival and proliferation of tumor cells. HIF-1 by activating genes involved in angiogenesis and also activates signaling pathways associated with cell survival and proliferation plays an important role in the stability and growth of tumors. Therefore, better understanding of molecular mechanisms associated with this factor can be effective in the treatment of cancer.

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