Endothelial Growth Factor Gene Research Articles

Mesh-like electrospun membrane loaded with atorvastatin facilitates cutaneous wound healing by promoting the paracrine function of mesenchymal stem cells

BackgroundFunctional electrospun membranes are promising dressings for promoting wound healing. However, their microstructure and drug loading capacity need further improvements. It is the first time to design a novel mesh-like electrospun fiber loaded with atorvastatin (ATV) and investigated its effects on paracrine secretion by bone marrow-derived mesenchymal stem cells (BMSCs) and wound healing in vivo.MethodsWe fabricated a mesh-like electrospun membrane using a copper mesh receiver. The physical properties of the membranes were evaluated by SEM, FTIR spectroscopy, tensile strength analysis, and contrast angle test. Drug release was measured by plotting concentration as a function of time. We tested the effects of conditioned media (CM) derived from BMSCs on endothelial cell migration and angiogenesis. We used these BMSCs and performed RT-PCR and ELISA to evaluate the expressions of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (b-FGF) genes and proteins, respectively. The involvement of FAK and AKT mechanotransduction pathways in the regulation of BMSC secretion by material surface topography was also investigated. Furthermore, we established a rat model of wound healing, applied ATV-loaded mesh-like membranes (PCL/MAT) seeded with BMSCs on wounds, and assessed their efficacy for promoting wound healing.ResultsFTIR spectroscopy revealed successful ATV loading in PCL/MAT. Compared with random electrospun fibers (PCL/R) and mesh-like electrospun fibers without drug load (PCL/M), PCL/MAT induced maximum promotion of human umbilical vein endothelial cell (HUVEC) migration. In the PCL/MAT group, the cell sheet scratches were nearly closed after 24 h. However, the cell sheet scratches remained open in other treatments at the same time point. The PCL/MAT promoted angiogenesis and led to the generation of longer tubes than the other treatments. Finally, the PCL/MAT induced maximum gene expression and protein secretion of VEGF and b-FGF. As for material surface topography effect on BMSCs, FAK and AKT signaling pathways were shown to participate in the modulation of MSC morphology and its paracrine function. In vivo, PCL/MAT seeded with BMSCs significantly accelerated healing and improved neovascularization and collagen reconstruction in the wound area compared to the other treatments.ConclusionsThe mesh-like topography of fibrous scaffolds combined with ATV release creates a unique microenvironment that promotes paracrine secretion of BMSCs, thereby accelerating wound healing. Hence, drug-loaded mesh-like electrospun membranes may be highly efficacious for wound healing and as artificial skin. It is a promising approach to solve the traumatic skin defect and accelerate recovery, which is essential to developing functional materials for future regenerative medicine.

A study of the genotyping and vascular endothelial growth factor polymorphism differences in diabetic and diabetic retinopathy patients

BackgroundRetinopathy is one of the major causes of visual impairment which is the most severe microvascular complication of diabetes mellitus (DM). The aim of this study was to evaluate the association between diabetic retinopathy (DR) and two SNPs (− 152G > A and − 165C > T) located in the promoter region of the vascular endothelial growth factor (VEGF) gene in a small sample from Egyptian population. One hundred diabetic patients without retinopathy (DWR) and two hundred diabetic patients with retinopathy were included in this study. Genotype analysis for the two SNPs (− 152G > A and − 165C > T) was assessed by using the PCR–RFLP technique. In addition, the serum protein level of VEGF was measured by ELISA assay.ResultsThe results showed a significant relationship between − 152G > A (rs13207351) polymorphism and both proliferative and non-proliferative retinopathy in genotypes (GG, GA, AA). The risk factor increment in the mutant heterozygous genotype (GA) was significantly increased in NPDR compared to PDR (OR = 16.3, 95%CI = 0.80–331.7); (OR = 20.4, 95%CI = 1.08–385.3), respectively. There was no significance between VEGF − 165C > T (rs79469752) gene polymorphism and retinopathy. Moreover, the serum protein level of VEGF showed a highly significant increase (P = 0.0001) in PDR (Mean ± SD = 3691 ± 124.9) when compared to both DWR (Mean ± SD = 497.3 ± 18.51) and NPDR (Mean ± SD = 1674.5 ± 771.7). These results were supported by the increased level of VEGF in serum protein which is positively correlated with the severity of retinopathy. Measuring VEGF protein level in DR patients would help as a biomarker in early diagnosis.ConclusionThe increase in the mutant heterogeneous GA genotype in VEGF − 152G > A SNP could be a risk factor for the progression of severe retinopathy in diabetic patients.

Genetic association of vascular endothelial growth factor (VEGF) gene variants with the risk for diabetic retinopathy: a meta-analysis

The vascular endothelial growth factor (VEGF) significantly contributes to the manifestation of neovascularization in the retina which progressively develops retinopathy in diabetic patients. The aim of this study was to assess the role of VEGF polymorphisms in the pathogenesis of diabetic retinopathy (DR). This meta-analysis comprised a total of six case-control and cohort studies published in the last 6 years. The selection of studies was done by sorting in reliable database searches: NCBI, Ensemble, GenBank, Embase, UCSC Genome Browser. After the extraction of data, the Q test of heterogeneity was performed. The Stata Software version 13.1 was used as the statistical package. Fixed and random effect models were applied for forest plot depiction while funnel plot and Egger’s test were carried out for the evaluation of publication bias. A significant relationship was found between the risk allele of VEGF polymorphism rs3025039 with predisposition of DR (OR = 1.45, 95%CI = 1.04–1.86) in the absence of heterogeneity by three studies. Furthermore, significant association was also observed for four polymorphisms of VEGF (rs833061, s13207351, rs1570360, and rs2010963) with higher susceptibility of DR among the population of Pakistan (OR = 1.46, 95%CI = 1.21–1.71). The meta-analysis suggested the substantial role of VEGF polymorphism rs3025039 as a possible biomarker for the assessment of DR risk. However, genome-wide association study (GWAS) is required in future to elucidate the multi-SNP effect of these polymorphisms in VEGF gene.

Correlation of gene polymorphisms of vascular endothelial growth factor with grade and prognosis of lung cancer

BackgroundVascular endothelial growth factor (VEGF) gene is highly polymorphic, and single nucleotide polymorphisms (SNP) of VEGF gene are associate with cancer prognosis. This study aimed to analyze the correlation of VEGF gene polymorphisms with grade and prognosis of lung cancer.MethodsA total of 458 Chinese patients with primary lung cancer were enrolled from September 2008 to October 2013. The genotypes of −2578C > A, −1154G > A, − 460 T > C, and + 405G > C were analyzed in white blood cells from patients using polymerase chain reaction based restriction fragment length polymorphism.ResultsOur data showed that –1154G > A polymorphism was significantly associated with tumor stages, but all four tested VEGF gene polymorphisms had no significant effect on survival.ConclusionsVEGF polymorphisms may relate to stage of lung cancer in Chinese population.

Dietary Supplemental Glutamine Enhances the Percentage of Circulating Endothelial Progenitor Cells in Mice with High-Fat Diet-Induced Obesity Subjected to Hind Limb Ischemia.

This study investigated whether glutamine (GLN) pretreatment can enhance circulating endothelial progenitor cells (EPCs) and attenuate inflammatory reaction in high-fat diet-induced obese mice with limb ischemia. Mice were assigned to a normal control (NC), high-fat control (HC), limb ischemia (HI), and GLN limb ischemia (HG) groups. The NC group provided chow diet and treated as a negative control. Mice in the HC and HI groups were fed a high-fat diet which 60% energy provided by fat for 8 weeks. Mice in the HG group were fed the same diet for 4 weeks and then transferred to a high-fat diet with 25% of total protein nitrogen provided as GLN to replace part of the casein for the subsequent 4 weeks. After feeding 8 weeks, mice in the HC group were sham-operated, while the HI and HG groups underwent an operation to induce limb ischemia. All mice except the NC group were euthanized on either day 1 or 7 after the operation. The results showed that the 8 weeks' high-fat diet feeding resulted in obesity. The HG group had higher circulating EPCs on day 1 while muscle vascular endothelial growth factor, matrix metalloproteinase-9, and hypoxia-inducible factor-1 gene expressions were higher on day 7 postischemia than those of the HI group. The superoxide dismutase activity and reduced glutathione content in affected muscles were higher, whereas mRNA expressions of interleukin-6 and tumor necrosis factor-α were lower in the HG than those in the HI group. These findings suggest that obese mice pretreated with GLN-supplemented high-fat diet increased circulating EPC percentage, enhanced the antioxidant capacity, and attenuated inflammatory reactions in response to limb ischemia.

Preferential targeting cancer-related i-motif DNAs by the plant flavonol fisetin for theranostics applications

The relationship of i-motif DNAs with cancer has prompted the development of specific ligands to detect and regulate their formation. Some plant flavonols show unique fluorescence and anti-cancer properties, which suggest the utility of the theranostics approach to cancer therapy related to i-motif DNA. We investigated the effect of the plant flavonol, fisetin (Fis), on the physicochemical property of i-motif DNAs. Binding of Fis to the i-motif from the promoter region of the human vascular endothelial growth factor (VEGF) gene dramatically induced the excited state intramolecular proton transfer (ESIPT) reaction that significantly enhanced the intensity of the tautomer emission band of Fis. This unique response was due to the coincidence of the structural change from i-motif to the hairpin-like structure which is stabilized via putative Watson-Crick base pairs between some guanines within the loop region of the i-motif and cytosines in the structure. As a result, the VEGF i-motif did not act as a replication block in the presence of Fis, which indicates the applicability of Fis for the regulation of gene expression of VEGF. The fluorescence and biological properties of Fis may be utilised for theranostics applications for cancers related to a specific cancer-related gene, such as VEGF.

Expression Level of TGFβ1 and VEGF Gene in Acute Myeloid Patients and Its Clinical Prognostic Value

To study the expression level of TGFβ1 and VEGF gene in patients with acute myeloid leukemia (AML) and its clinical prognostic value. Seventy-eight AML patients treated in our hospital from July 2016 to September 2018 were selected. After isolation of bone marrow mononuclear cells from the patients, the levels of TGFβ1 and VEGF genes were detected by RT-PCR, and the correlation of TGFβ1 with VEGF genes and clinical characteristics of AML patients was analyzed. OS and EFS of the patients were evaluated by Kaplan-Meier, and Cox risk ratio model was used to analyze the prognostic risk factors of AML patients. The relative expression level of TGFβ1 gene in AML patients was 0.32±0.04, which was significantly lower than that in control group (P<005). The relative expression level of vascular endothelial growth factor(VEGF) gene in the patients was 2.65±0.15, which was significantly higher than that in the control group (P<0.05). The levels of TGFβ1 and VEGF genes significantly correlated with leukocyte count, hemoglobin, platelet and peripheral blast levels in AML patients (P<0.05). The level of TGFβ1 in AML patients with complete remission was higher than that in patients with partial remission or non-remission (P<0.05). The level of TGFβ1 in AML patients with partial remission was significantly higher than that in patients with non-remission (P<0.05). The level of VEGF in AML patients with complete remission was lower than at in patients with partial remission or non-remission (P<0.05). The level of VEGF in AML patients with partial remission was significantly lower than that in patients with non-remission (P<0.05). Kaplan-Meier survival analysis showed that OS and DFS in AML patients with high expression of TGFβ1 were better than those in patients with low expression of TGFβ1 (P<0.05), OS and DFS in AML patients with low expression of VEGF were better than those in patients with high expression of VEGF (P<0.05). Multivariate Cox regression analysis showed that platelet, TGFβ1 and VEGF gene were independent influencing factors of OS (P<0.05). Leukocyte, TGFβ1 and VEGF gene were independent influencing factors of DFS (P<0.05). Decreased expression of TGFβ1 and increased expression of VEGF gene in AML patients closely relate to the poor prognosis of AML patients, which can provide reference for improving clinical efficacy of AML patients.

Shenqi Fuzheng Injection (SFI) Enhances IFN-α Inhibitory Effect on Hepatocellular Carcinoma Cells by Reducing VEGF Expression: Validation by Gene Silencing Technique.

Shenqi Fuzheng Injection (SFI) is a traditional Chinese medicine injection with anticancer properties and is mainly composed of ginseng and astragalus. Its efficacy has been confirmed in clinical trials, but the mechanism remains unclear. We investigated the effect of SFI on vascular endothelial growth factor (VEGF) gene expression in hepatocellular carcinoma (HCC) cells and identified its possible mechanism of synergistic effects when combined with the chemotherapeutic drug interferon (IFN-) α. An MTT assay was used to measure the inhibition effects of low-dose IFN-α (6000 IU) with or without SFI (0.5 g/L) on the HCC cell line MHCC97. VEGF-silenced MHCC97L-mir200 cell lines were prepared using lentiviral vectors and evaluated by real-time PCR to determine the inhibition effect. We examined MHCC97L-mir200 and MHCC97L cells by MTT assay, using IFN-α alone or in combination with SFI. The inhibition ratio of IFN-α (6000 IU) was -29.5%, while that for IFN-α (6000 IU) + SFI (0.5 g/L) was 17.0%, which was significantly higher than that for the IFN-α group (P < 0.01). The VEGF gene was silenced successfully in MHCC97-L cells. After interference of VEGF, the inhibition by SFI and IFN-α in MHCC97L-mir200 did not differ from that in MHCC97-L cells (P > 0.05). SFI can reduce the expression of VEGF in HCC, which can increase the efficacy of IFN-α, providing a theoretical basis for clinical application.

The anti-invasive activity of Robinia pseudoacacia L. and Amorpha fruticosa L. on breast cancer MDA-MB-231 cell line

The paper investigates anticancer effects of methanol extracts of invasive plant species Robinia pseudoacacia L. (RpE) and Amorpha fruticosa L. (AfE) on breast cancer MDA-MB-231 and healthy MRC-5 cells. The anticancer activity was evaluated through examination of cytotoxic effects, anti-invasive potential and impact on redox status in comparative analysis using their chemical composition. According to the IC50 values, the investigated plants had no significant cytotoxic effects either on healthy cell line MRC-5 or on MDA-MB-231 cancer cells, but they showed great anti-invasive potential by suppressing all investigated parameters of tumor invasion and metastases (Matrix Metalloproteinases (MMP), protein concentration and MMP-9, C-X-C Motif Chemokine Ligand 12 (CXCL-12), Vascular Endothelial Growth Factor (VEGF-A) and Hypoxia-Inducible Factor (HIF-1α) gene expression) in MDA-MB-231 cells. Based on their remarkable anti-invasive potential, RpE and AfE are suitable for use as potential supplements in anticancer therapy or as nutritional food supplements.

Effects of vascular endothelial growth factor transfected endothelial progenitor cells transplantation on erectile function for mice

Objective To investigate the the effect of vascular endothelial growth factor (VEGF) gene transfected endothelial progenitor cell (EPCs) transplantation on penile for erectile dysfunction (ED) mice. Methods 24 male Wistar mice, 8 weeks, were made from animal models for ED by feeding high fat forage. They were randomly divided into 3 groups: group Ⅰ (control), group Ⅱ (EPCs transplantation) and group Ⅲ (VEGF gene transfection EPCs transplantation). EPCs and VEGF gene transfection EPCs were administered for group Ⅱ and group Ⅲ respectively. Saline was given to group Ⅰ. Drug erectile experiment, newly born capillaries, ICAM-1and VEGF detection was performed in 28 days. Results The expression of ICAM-1 in corpus cavernosum penis in group Ⅲ was (1 090.44±158.38) pg/ml, which was lower than that in group Ⅱ [(1 654.27±232.14) pg/ml, P=0.045]. And which of group Ⅱ was lower than that of group Ⅰ [(3 860.54±456.18) pg/ml, P=0.015]. The expression of VEGF in corpus cavernosum penis in group Ⅲ was (768.13±134.37) pg/ml, it was higher than that in group Ⅱ [(579.79±68.62) pg/ml, P=0.025], and it was higher than that of group Ⅰ [(54.45±19.37) pg/ml , P=0.048]. The erectile times in group Ⅲ was 2.30±0.67 and greater than which of group Ⅱ (1.83±0.87, P=0.033). The number of newly born capillaries in group Ⅲ was 143.32±49.27 and more than which of group Ⅱ (73.69±23.32, P=0.001). The number of newly born capillaries in group Ⅱ was bigger than which of group Ⅰ (46.31±11.17, P=0.024). Conclusion VEGF gene transfection endothelial progenitor cell transplantation can promote the expression of VEGF in orpus cavernosum penis, regeneration of newly born capillaries, vascular endothelial function and erectile function for ED mice. Key words: Erectile dysfunction; Endothelial progenitor cells; Vascular endothelial growth factor

Association between VEGF Gene Polymorphisms and the Susceptibility to Lung Cancer: An Updated Meta-Analysis.

Background and Objective The association between vascular endothelial growth factor (VEGF) gene polymorphisms (-2578C/A, +936C/T, and -460C/T) and lung cancer risk has been extensively studied in the last decades, but currently available results remain controversial or ambiguous. Therefore, we conducted a meta-analysis to assess whether the relationship between the VEGF gene and lung cancer susceptibility exists. Methods The meta-analysis was conducted by searching the databases PubMed, Embase, and Web of Science covering all eligible studies published up to October 1, 2017. The pooled odds ratios (ORs) as well as their 95% confidence intervals (CIs) were utilized to evaluate the possible associations. Publication bias of relevant studies was examined via Begg's funnel plots and Egger's regression tests. Results This meta-analysis included 13 published case–control studies covering 4477 patients with lung cancer and 4346 healthy controls, who had been accrued from December 1992 to July 2012. For the overall eligible data collected in our meta-analysis, it indicated that VEGF +936C/T, -460C/T, and -2578C/A polymorphisms did not correlate with the elevated lung cancer risk in all genetic comparison models. Moreover, VEGF +460T/C polymorphism was found to be significantly associated with susceptibility to lung cancer in these models (allele model: pooled OR = 1.12, 95% CI: 1.00–1.26, P = 0.184; homozygote model: pooled OR = 1.51, 95% CI: 1.12–2.03, P = 0.821), but no significant results were detected in Caucasian populations. Conclusions VEGF +936C/T, -460C/T, and -2578C/A polymorphisms were not associated with the risk of lung cancer. The VEGF +460T/C polymorphism might be a risk factor for lung cancer only in Asian populations.

An analysis of association between growth factor gene polymorphisms and a risk of bronchial asthma in children

The aim of this study was to assess growth factor genes involvement in the pathogenesis of asthma. Methods. Associations between Arg25Pro polymorphic loci of transforming growth factor β growth (TGF β1) gene, A2073T epidermal growth factor receptor (EGFR) gene, C634G vascular endothelial growth factor (VEGFA) gene and a risk of bronchial asthma development were analyzed. DNA samples were isolated from peripheral blood leukocytes of 30 children with asthma and 27 healthy subjects. Gene polymorphisms were determined by the allele-specific polymerase chain reaction. Results. The study revealed an association between Arg25Pro polymorphism of TGF β1 gene, C634G polymorphism of VEGFA gene and increased risk of asthma in children. The majority of patients were homozygous for 2073T allele of EGFR gene or carriers of ArgArg genotype of TGFβ1 gene. Frequencies of C634G polymorphism of VEGFA gene did not differed significantly between healthy children and asthma patients. Conclusion. The results of this study could help to select patients predisposed to asthma and to develop preventive measures taking into account individual characteristics of each patient.

Abstract A127: The identification and development of selective natural product inhibitors of hypoxia inducible factor-2α for the treatment of renal cell carcinoma

Abstract Clear cell renal cell carcinoma (ccRCC) is the most prevalent form of kidney cancer and is frequently associated with loss of von Hippel Lindau (VHL) gene function, resulting in the aberrant accumulation of the hypoxia inducible factor-alpha subunit (HIF-α), which contributes to tumor growth, angiogenesis, and metastasis. Recent studies suggest that this normoxic stabilization of HIF-1α is not sufficient to reproduce tumorigenesis, and HIF-1α is thought to act as a tumor suppressor in ccRCC. Conversely, HIF-2α acts as an oncogene, turning on transcription of a large number of HIF-regulated genes, and upregulation of HIF-2α is often associated with poor prognosis. To this end, two HIF-2α selective inhibitors are currently in clinical trials for RCC. To isolate additional HIF-2α selective compounds for use as research tools and drug development leads, a cell-based high-throughput screen (HTS) of the NCI Natural Products Repository was performed. Stably transformed clones of the human ccRCC-derived cell line 786-0 expressing a luciferase reporter construct downstream of the human vascular endothelial growth factor (VEGF) gene promoter were used to identify natural product samples with inhibitory activity and minimal cellular toxicity. HTS leads were chromatographically separated into component structures yielding ~40 pure compounds with micromolar or submicromolar IC50 values, &amp;gt;80% inhibition, and &amp;lt;10% cell toxicity. These hits were then screened for their ability to inhibit VEGF-A protein secretion by VHL- and HIF1A-negative 786-0 cells, and several compounds showed &amp;gt;90% inhibition of VEGF secretion at micromolar doses under normoxic conditions. Patterns of mRNA expression of HIF-1α, HIF-2α, and several HIF target genes in cell lines engineered to express either HIF alone or both were studied to help identify HIF-2α-selective inhibitors. Several lead compounds reduced VEGF transcription and secretion, and downregulated HIF-2α target genes. To further characterize the HTS leads, ICM-Pro protein/small-molecule docking software was used to dock the ligands of interest to high-resolution HIF-2α and HIF-1α 3D structures. The HIF-2α selective inhibitor currently undergoing clinical investigation, PT2385, has been found to bind directly to a hydrophobic pocket of the HIF-2α:ARNT dimerization. Using the PT2385 binding mode as a benchmark, docking studies of the HTS leads were performed and the results have been linked to empirical data in an effort to explain selectivity for HIF-2α over HIF-1α, reveal key favorable interactions between the protein and ligand, define mode of action, and aid in future drug development studies. Citation Format: Molly M. Lee, Daniel C. Rabe, Tawnya C. McKee, Girma M. Woldemichael, James R. Vasselli, James B. McMahon, W. Marston Linehan, Donald P. Bottaro. The identification and development of selective natural product inhibitors of hypoxia inducible factor-2α for the treatment of renal cell carcinoma [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A127.

Tumoral Necrosis Factor (TNF) mRNA gene expression in relation to Vascular Endothelial Growth Factor (VEGF) and chemokines mRNA gene expression in tumoral microenvironment of endometrial cancer versus normal endometrium: Preliminary results

Tumoral Necrosis Factor (TNF) mRNA gene expression in relation to Vascular Endothelial Growth Factor (VEGF) and chemokines mRNA gene expression in tumoral microenvironment of endometrial cancer versus normal endometrium: Preliminary results

Evaluation of Expression Pattern of Vascular Endothelial Growth Factor (VEGF) and Interleukin-23 (IL-23) Genes in Human Colorectal Tumors

Background: There is a very long interval between the time of tumor initiation and diagnosis of CRCs. So, by early screening by molecular markers, individuals can be diagnosed in a much more convenient time to start the therapies. Among key factors deregulated in advanced CRC, are VEGF (vascular endothelial growth factor) and interleukin-23, which have critical roles in tumor angiogenesis and growth respectively.Objectives: The main purpose of the study was to elucidate the patterns of VEGF-A and IL-23 (IL-23p19) molecular expression in CRC.Material and methods: 47 human colorectal cancer tissues and 47 matched tumor-free margin tissues were gathered during surgery. The mRNA expression levels of VEGF-A and IL-23p19 were examined by real-time quantitative polymers chain reaction (qPCR).Results: The mRNA expression levels of VEGF-A and IL-23p19 were higher in tumor tissues than in the tumor-free margin (control) (p=0.008 and p=0.002, respectively). Though, there was no important association between the mRNA expression levels of VEGF-A and IL-23p19 with clinicopathological features. There was also a positive correlation between these 2 genes expression levels.Conclusion: The analysis of IL23p19 and VEGF-A genes expression could be considered as a biomarker for screening CRC patients and are suggested to be used for diagnosis. Thus, collectively, the high expression level of VEGF-A and IL-23p19 can be considered as a risk factor for CRC carcinogenesis, contributing to high proliferation and invasive properties of the tumor.

Involvement of vascular endothelial growth factor (VEGF) gene polymorphism in hepatocellular carcinoma of HCV patients from local population

Development of Hepatocellular carcinoma (HCC) after Hepatitis C virus (HCV) infection is common cause of poor long-term survival. This study aimed to evaluate the effect of Vascular Endothelial growth factor (VEGF) genes polymorphism as a risk factor of Hepatocellular carcinoma (HCC) in chronic hepatitis C virus in Pakistani population. VEGF regulates angiogenesis so it is mainly involved in progression and development of solid tumor. The results showed that HCC patients had a higher frequency of the VEGF-634 GG genotype. In this study results revealed that VEGF Genotype GG is not associated with the risk of HCC in HCV infected patients in Punjab region of Pakistan. Keywords: HCV; HCC; VEGF Polymorphism http://dx.doi.org/10.19045/bspab.2017.60077

Effect of calcitonin gene-related peptide on osteoporotic fracture healing in vitro

Objective To explore the effects of calcitonin gene-related peptide (CGRP) on proliferation of vascular endothelial cells (VECs) and osteogenic differentiation of bone mesenchymal stem cells (BMSCs) of osteoporotic rats in vitro, and cell/molecular mechanism of promoting osteoporotic fracture healing. Methods 10 3-month-old SD rats were employed to make osteoporosis model by taking bilateral ovariectomy. Micro-CT analysis was used to determine the success of modeling at 6 months of postoperation. BMSCs and VECs were isolated from osteoporotic rats by blood adherence method. 10-7-10-10 mol/L CGRP were added to the culture medium. CCK-8 method was used to examine the cell proliferation. Effect of CGRP on osteogenic differentiation of BMSCs: Alkaline phosphatase (ALP) activity, and reverse transcription polymerase chain reaction (RT-PCR) were used to detect the expression of ALP, CollagenⅠ, BMP-2, Osteonectin and RunX2 gene. Effect of CGRP on vascularization of VECs: Vascular cell endothelial growth factor (VEGF) protein secretion and gene expression level were tested by ELISA and RT-PCR. Results BMSCs and VECs of osteoporotic rats were isolated successfully.The optical density (OD) values of CCK-8 method increased in a dose-dependent manner, for BMSCs at 7 d, the control group of OD value was 0.58±0.02, the CGRP group 10-7-10-10 mol/L was to 0.80±0.03, 0.79±0.03, 0.74±0.03, 0.69±0.03, difference was significant; for VECs, the control group of OD value was 2.20±0.01, the CGRP group 10-7-10-10 mol/L was to 4.12±0.04, 3.90±0.02, 3.56±0.04, 3.12±0.04, difference was significant. Effect of CGRP on osteogenic differentiation of BMSCs: Alkaline phosphatase (ALP) activity, and reverse transcription polymerase chain reaction (RT-PCR) were used to detect the expression of ALP, CollagenⅠ, BMP-2, Osteonectin and RunX2 gene. Effect of CGRP on vascularization of VECs: Vascular cell endothelial growth factor (VEGF) protein secretion and gene expression level were tested by ELISA and RT-PCR, difference between the control group and CGRP group were all significant. Conclusion 10-7 mol/L, 10-10 mol/L of CGRP can directly promote the proliferation of the BMSCs and VECs of osteoporotic rats, promote the osteogenic differentiation of BMSCs, improve the expression of VEGF gene and protein secretion, promote angiogenesis. Key words: Calcitonin gene-related peptide; Osteoporosis; Fractures, bone

The effects of inner vertical outer spiral complex scaffold in repairing long segment of urethral defect

Objective To explore the possibility of the inner vertical outer spiral complex tubular urethra scaffold vascularization in repairing long segment of urethral defect. Methods From August 2014 to October 2015, 27 clean male New Zealand white rabbits were divided into 3 groups, S1 group was transfected recombinant vascular endothelial growth factor(VEGF) gene lentiviral vector group. S2 group was vascular pedicle transfer tube group. C group was simple stent group. A 3.0 cm inner vertical outer spiral complex scaffold was constructed by using the small intestine acellular matrix (SIS) and polylactic acid copolymer (PLGA) modified by type Ⅰ collagen surface, and adipose-derived mesenchymal stem cells (ADSC) and smooth muscle cells after transformation from New Zealand white rabbits. In S1 group, the seed cells were transfected by recombinant vascular endothelial growth factor (VEGF) gene lentivirus, which express VEGF protein.The complex scaffold was used to repair 3.0 cm rabbit urethral defect In S2 group, the un-transfected cells were seeded into the scaffold and embedded in the skin near the groin artery 3 weeks for repairing urethral defect with vascular pedicle transfer tube. In group C, the unseeded scaffold was used to repair the urethral defect alone.Postoperative observation and urethrography were followed 4, 8 and 24 weeks after implantation. The HE staining, fluorescence tracing, immunohistochemical and scanning electron microscopy were evaluated at the same phase. Results In S1 group, there were one urinary fistula and one urethral stricture-related death, respectively. The urethra was smooth and patent, histological examination showed active hyperplasia of urethral capillary. In S2 group, there were one urinary fistula and two urethral stricture-related deaths, respectively. The urethral was rough, local thinning or dilated. Fat accumulation and mucosal contraction were found in the urethral submucosal, respctively. In C group, there were one urinary fistula, three hypospadias, and three urethral stricture-related deaths. The thickness of the urethra was uneven and stricture bending. The urethral mucosa was poorly repaired and the scar was narrow. HE and CD31 staining showed that S1 and S2 groups were active in the proliferation of urethral capillaries, and the angiogenesis was abundant. VEGF staining showed that the cytoplasm of endothelial cell layer, smooth muscle layer of vascular wall and the urothelial epithelial cell layer were fully expressed at 24 weeks, especially in epithelial cell layer. CKpan staining showed that the epithelium of S1 and S2 group developed to stratified epithelium, and the morphology of urethra was similar to normal urethra at 24 weeks. The urethral epithelial in C group of grew poor as single-level, irregular arrangement, 24 weeks is still a lack of effective stratified epithelium. HE and α-SMA staining showed that the smooth muscle and actin gradually increased in group S1 and S2, α-SMA staining in group C was scarce and increased at 24 weeks. PLGA was encapsulated by the surrounding tissue and the structure of electrospinning was clear after 4 weeks, absorbed and degraded after 8 weeks and absorbed after 24 weeks. Conclusions The inner vertical outer spiral complex tubular urethra scaffold maybe a reasonable method in repairing long segment urethral defects, and the methods of tubular urethra scaffold vascularization by transfected VEGF gene recombinant lentiviral vector and vascular flap deserve more research. Key words: Tissue engineering; Scaffold; Urethra defect; Vascularization

The molecular mechanism that down-regulation of VEGF gene transcription inhibits nasopharyngeal carcinoma cell proliferation

Objective To investigate down-regulation of vascular endothelial growth factor (VEGF) gene at mRNA level on effects of nasopharyngeal carcinoma (NPC) cell proliferation and analyze the key molecule expressions of extracellular-signal regulated protein kinase/mitogen-activated protein kinase (Erk/MAPK) pathway to explore its molecular mechanism. Methods Highest-efficiency VEGF-siRNA plasmid was successfully constructed and screened, confirmed by sequencing, and then transfected to CNE2 cell line. Interference efficiency was quantitatively evaluated by a fluorescence quantitation polymerase chain reaction assay. Expressions of VEGF, cell cycle regulator cyclin D1, and key molecules in Erk/MAPK pathway were analyzed with Western blot. Cell proliferation was tested by cell counting kit-8 (CCK-8) kit. Clone formation rate was checked with plate clone formation assay. Alteration of cell cycle was measured with flow cytometry. Stable cell clones were screened by G418. Results VEGF expression in NPC cells transfected with specific siRNA were significantly down-regulated at the levels of mRNA or protein, and cell proliferations were inhibited in a time-dependent manner, with significant decreases of colony-forming, cyclinD1, and p-Erk, and with cell cycle arrested at G1 phase. Conclusions Intervencing VEGF gene transcription inhibited NPC cell proliferation through Erk/MAPK signaling pathway, and suggesting that VEGF should be a novel therapeutic target for NPC. Key words: Vascular endothelial growth factors/PD; RNA, small interfering; Nasopharyngeal neoplasms/DT; Cell proliferation; Neoplasm metastasis

There is no correlation between the functional polymorphism −460C&gt;T of vascular endothelial growth factor (VEGF) gene promoter and uncomplicated recurrent urinary tract infection among young women

Abstract Introduction The −460C>T polymorphism in vascular endothelial growth factor (VEGF) gene promoter (rs833061) has robust association with various diseases including renal parenchymal scarring following urinary tract infection (UTI). However, the association of this polymorphism with uncomplicated recurrent UTI (RUTI) is still unclear. Aim The objective of this study was to assess the correlation between VEGF −460C>T polymorphism and uncomplicated RUTI among young women. Material and methods Polymerase chain reaction – restriction fragment length polymorphism (PCR-RFLP) with BstU1 restriction enzyme – and DNA sequencing were used to determine the VEGF −460C>T polymorphism among 34 young adult women with uncomplicated RUTI and 34 healthy controls. Differences in genotype and allele frequencies between case and control groups were analyzed with χ2 test or Fisher's exact test. Results and discussion This study found that there was no significant difference in genotype frequencies between cases (CC 23.5%; CT 26.5%; TT 0%) and controls (CC 85.3%; CT 14.7%; AA 0%). Dominant model analysis found that there was no significant difference between uncomplicated RUTI and normal groups (P = 0.368). Similarly, allele analysis also found that there was no association between VEGF −460C>T polymorphism and uncomplicated RUTI (P = 0.398). Conclusions This study found that there was no correlation between VEGF −460C>T polymorphism and uncomplicated RUTI among young women.