血管内皮生长因子基因C(—634)G多态性及血浆血管内皮生长因子水平与糖尿病视网膜病变的相关性研究

目的探讨血管内皮生长因子(VEGF)基因C(-634)G多态性及血浆中VEGF水平与糖尿病视网膜病变(DR)的关系。方法在昆明地区汉族人中检测96名正常对照者(NC组)和285例T2DM患者的VEGF基因C(-634)G多态性及血浆中VEGF水平。结果 DR组CC基因型和C等位基因频率显著高于非DR(NDR)组(P均0.05),DR组中CC基因型者血浆VEGF水平高于CG及GG基因型者和NC组(P0.05或P0.01)。结论 VEGF基因C(-634)G多态性是昆明地区汉族T2DM患者发生DR的危险因素。     

血管内皮生长因子基因多态性与糖尿病视网膜病变相关性

目的 探讨血管内皮生长因子(VEGF)+450基因的多态性与糖尿病视网膜病变(DR)的关系.方法 运用PCR-RFLP技术检查2型糖尿病(T2DM)患者249例,单纯T2DM患者120例,DR患者129例,及体检的98例对照者的基因型,比较各组的基因型和等位基因的频率.结果 DR组CC基因型及C等位基因频率显著高于T2DM组和健康对照组(P＜0.01,P＜0.05);CC基因糖尿病(DM)患者VEGF产量高.结论 VEGF+450C/G基因多态性可能与DR的发生发展有关,C等位基因可能是DR的易感基因.     

血管内皮生长因子基因多态性与糖尿病肾病相关研究

运用聚合酶链反应-限制性片段长度多态性(PCR-RFLP)技术检查T2DM患者249例(其中DN患者129例)及体检的98例健康对照者的基因型.结果(1)DN组CC基因型及C等位基因频率显著高于T2DM组和正常对照组;结论VEGF 936C/T基因多态性可能与DN的发生发展有关,C等位基因可能是DN的易感基因.     

血管内皮生长因子基因多态性与糖尿病视网膜病变的相关性

目的探讨血管内皮生长因子（VEGF）-460C/T基因多态性与糖尿病视网膜病变（DR）的相关性。方法病史超过10年的2型糖尿病（T2DM）患者204例,分为非增殖型视网膜病变组（NP-DR,65例）、增殖型视网膜病变组（PDR,64例）及单纯2型糖尿病组（DM,75例）。用PCR-RFLP方法检测各组基因型,比较各组基因型和等位基因的频率。结果DR组VEGF-460位点TT基因型频率显著低于DM组（P〈0.01）,C等位基因频率显著高于DM组（P〈0.01）;NPDR组与PDR组基因型和等位基因频率差异无统计学意义（P〉0.05）。DM中CC、CT、TT基因型的DR发生率分别为69.8%、68.9%和42.2%,CC和CT基因型的DR发生率显著高于TT基因型（P〈0.01）。结论VEGF-460C/T多态性与DR的发生发展有关,C等位基因可能是DR的易感基因。     

血管内皮生长因子基因SNPrs3025039多态性与2型糖尿病相关性的研究

目的探讨T2DM患者血管内皮生长因子(VEGF)基因SNPrs3025039多态性。方法运用PCR-RFLP检测170例T2DM患者(T2DM组)和100名健康体检者(NC组)的VEGF基因SNPrs3025039多态性。结果 (1)两组的基因型和等位基因频率差异无统计学意义(P>0.05),但糖尿病慢性肾脏疾病(CKD)组CT、TT基因型频率和T等位基因频率高于糖尿病非CKD(NCKD)组(P<0.05);CC基因型和C等位基因频率低于NCKD组(P<0.05);(2)T2DM组BMI、SBP、DBP、HbA1c、FPG、2hPG、TG水平高于NC组(P<0.01),HDL-C水平较NC组降低(P<0.01)。结合FPG、TG、TC、LDL-C及HDL-C进行多重Logistic回归分析显示,CT和TT基因型与CKD的发生呈正相关(OR=2.75,P<0.05)。结论 T2DM患者VEGF基因SNPrs3025039多态性与正常人群比较差异无统计学意义,但CT、TT基因型与CKD的发病风险有关。     

血管内皮细胞生长因子基因多态性与糖尿病肾病的相关性

目的研究血管内皮细胞生长因子（VEGF）基因多态性与糖尿病肾病（DN）的关系。方法单纯2型糖尿病（DM）组76例,DN组81例,健康对照（NC）组60例。UNIQ-10柱提取全血基因组DNA。标本基因型的判断用聚合酶链反应-限制性酶切片断长度多态性技术。Hardy-Weinberg平衡法检验各组基因频率的群体代表性。结果（1）DN组VEGF-460和＋405CC基因型频率和C等位基因频率明显高于DM组和NC组。（2）-460位点CC基因型DN患病率明显高于CT和TT基因型。＋405位点CC基因型DN患病率明显高于CG和GG基因型。（3）显示VEGF-460和＋405基因多态性均为DN发生的独立危险因素。结论（1）VEGF-460C/T基因多态性与DN发生有关。C等位基因可能是DN易感基因。（2）VEGF＋405G/C基因多态性与DN发生有关。C等位基因可能是DN的易感基因。     

IL-6基因启动子区-634C/G多态性与糖尿病肾病的相关性

目的 探讨白细胞介素6（IL-6）基因启动子区-634C／G多态性与糖尿病肾病（DN）的关系。方法 在昆明地区汉族人中，应用PCR—RFLP方法和等位特异PCR（ASPCR）方法，对246例2型糖尿病（T2DM）患者（正常白蛋白尿组88例，微量白蛋白尿组93例，大量白蛋白尿组65例）和101例健康对照者（NC组）的IL-6基因启动子区-634C／G多态性进行检测。结果 （1）微量白蛋白尿组、大量白蛋白尿组、微量、大量白蛋白尿组的G／G基因型频率均高于正常白蛋白尿组（P=0．001），大量白蛋白尿组亦高于微量白蛋白尿组（P=0．045）。（2）大量白蛋白尿组和大量、微量白蛋白尿组的G等位基因频率均高于正常白蛋白尿组（P=0．031），大量白蛋白尿组亦高于微量白蛋白尿组（P=0．005）。（3）T2DM组中，GG基因型组UAER明显高于CG、CC基因型组（P〈0．01）。（4）Logistic回归分析表明：IL-6基因启动子区-634G／G基因型、病程是DN发生的危险因素。结论在昆明地区汉族人中，IL-6基因启动子区-634G／G基因型可能是DN发生的危险因素，此基因型携带者UAER明显增高；G等位基因可能是DN发展的危险因素之-。     

DR患者血浆血管内皮生长因子的变化

92例T2DM患者：无NDR31例、背景型BDR32例、增殖型PDR29例,NC40例。分别检测各组的血浆VEGF等指标。结果与对照组（NC）相比,DM组血浆VEGF水平升高（P〈0．01）。由NDR到PDR呈逐渐增高的趋势（P〈0．01）。结论DR患者的血浆VEGF水平明显增高,且和HbA1C呈正相关。     

血管内皮生长因子基因3'-非翻译区936C/T多态性与2型糖尿病外周神经病变相关

目的 探讨血管内皮生长因子(VEGF)基因3'-非翻译区936C/T多态性与山东地区汉族人2型糖尿病合并周围神经病变(DPN)之间的关系.方法 194例糖尿病患者分为单纯糖尿病组(n=92)和糖尿病神经病变组(n=102),另120名健康个体设为健康对照组.采用PCR-限制性片段长度多态性(RFLP)方法确定全部个体的基因型;对不同基因型间及病例组间的临床与生化参数、血清VEGF浓度以及VEGF基因936C/T多态性进行了统计分析.结果 糖尿病神经病变组C等位基因及CC基因型频率显著高于对照组(x2为9.406和9.677,P＜0.05)和糖尿病组(x2为5.578和5.614,P＜0.05),而携带T等位基因的基因型(CT+TT)频率及T等位基因频率显著低于对照组(x2为9.406和9.677,P＜0.05)和糖尿病组(x2为5.578和5.614,P＜0.05).Logistic多元回归分析显示血清低密度脂蛋白胆固醇(LDL-C)、总胆固醇、HbA1c水平以及VEGF浓度与DPN发生呈正相关,而VEGF基因936C/T多态性与糖尿病周围神经病变发病危险呈负相关(β=-1.046,OR=0.457,P=0.006,95%CI:0.166～0.741).结论 中国山东地区汉族人群中存在VEGF基因936C/T多态性,C等位基因及CC基因型患者可能是糖尿病易于发生神经病变危险性的遗传标志,而T等位基因和携带T等位基因的基因型(936TF基因型和936CT基因型)可能是降低糖尿病发生神经病变风险的遗传标志.

Abstract：

Objective To elucidate the relationship between a 936C/T mutation at 3'-untranslated region of human vascular endothelial growth factor(VEGF) gene and diabetic peripheral neuropathy ( DPN ). Methods All subjects recruited in this study were assigned into DM (n = 92, diabetes without neuropathy, retinopathy or nephropathy), DPN (n = 102, diabetes with peripheral neuropathy only ), and healthy control (n = 120 ) groups,respectively. The gene polymorphism was determined by PCR-RFLP, as well as the other clinical parameters including serum VEGF by ELISA. Results The frequencies of both genotype CC and allele C were significantly higher in DPN group than those in either DM group(x2 = 5.578 and 5.614, P＜0. 05 ) or control group (x2 = 9. 406 and 9. 677, P＜0. 05 ). However, the frequencies of genotype(CT+TT) and allele T were significantly lower in DPN group than that in either DM group(x2 =5.578 and 5.614, P＜0. 05) and control group (x2=9.406 and 9.677, P＜0.05). The multivariate logistic regression analysis showed that the levels of HbA1c, total cholesterol, low-density lipoproteincholesterol( LDL-C ), and serum VEGF positively correlated with DPN, while the 936C/T polymorphism of VEGF gene negatively correlated with DPN(β= -1. 046, OR=0. 457, P=0. 006, 95% CI: 0. 166-0. 741 ). Conclusions Allele 936C of VEGF gene may serve as a genetic marker susceptible to DPN, while allele 936T may be a protective genetic marker of DPN.     

蒙族高血压患者内皮型一氧化氮合酶(eNOS)基因多态性研究

目的旨在探讨一氧化氮合酶(eNOS)基因(G894T、T786C)多态性与中国蒙古族高血压患者的相关性。方法采用聚合酶链反应和限制性酶切的片段长度多态分析方法检测蒙族高血压患者100例和健康人50例的一氧化氮合酶(eNOS)基因(G894T、T786C)多态性。结果一氧化氮合酶基因T894G(GT、TT)、T786C(CT、CC)基因型及T、C等位基因频率在高血压组显著高于对照组(P <0．05)。同时具有eNOS894TT、786CC和894TG、786TC基因型者高血压组比对照组多,差异有显著性(P<0．05)。结论一氧化氮合酶基因T894G、T786C基因多态性与蒙族高血压相关,但尚需在更大的人群中进一步验证。     

Molecular biology of the VEGF and the VEGF receptor family

Vascular endothelial growth factor (VEGF) is the founding member of a still growing family of endothelial cell growth factors. The diverse functions of VEGF and its homologues (PIGF, VEGF-B, VEGF-C, VEGF-D, and VEGF-E) can be explained by their differential binding to the three signaling VEGF receptors. The VEGF family members PIGF and VEGF-B with exclusive binding capacities to the VEGFR-1 can influence monocyte activation and differentiation. The VEGFR-2 and VEGFR-3 binding VEGF homologues, VEGF-C and VEGF-D, are mitogens for both vascular and lymphatic endothelial cells. The orf virus encoded VEGF-E homologue binds and activates only the VEGFR-2 and thus may be the prototype of a vascular endothelial cell-specific growth factor. Further specific activities of VEGF and its homologues result from receptor-specific signaling and differential expression of ligands or receptors. A naturally occurring soluble form of the VEGFR-1 suggests a regulatory role for this receptor. Finally, the production and activation of factors involved in the coagulation/fibrinolytic system provide further evidence for the hypothesis that processes of hemostasis are involved in angiogenesis.     

Functions of the VEGF/VEGF receptor system in the vascular system

The vascular endothelial growth factor (VEGF)/VEGF receptor system plays a central regulatory role in physiological and pathological angiogenesis. During embryogenesis, the VEGF/VEGF receptor system is critically involved in the formation of the vascular system by regulating both the growth and the survival of blood vessels. In the vasculature of the adult organism, the high-affinity signaling VEGF receptor-2 (VEGFR-2) is downregulated but is reinduced during transient phases of physiological angiogenesis. Moreover, a variety of pathological conditions are associated with the upregulation of VEGF and the VEGF receptors. VEGF stimulates angiogenesis and the survival of endothelial cells in tumors, thereby enabling tumor expansion and metastasis. VEGF is also upregulated in ischemic diseases, such as coronary heart disease or stroke, and is thought to stimulate the--often insufficient--compensatory formation of blood vessels. The implication of VEGF in these pathological processes has opened up promising new therapeutic strategies. In malignancies, attempts are made to inhibit VEGF-mediated signaling and angiogenesis. In ischemic disease, the exogenous application of VEGF may enhance the formation of collaterals. However, considering the complexity of the regulatory pathways involved in the formation of new blood vessels under physiological conditions, a treatment relying on VEGF as the sole angiogenic factor may be insufficient, and the combination with other factors may improve the functionality of newly formed blood vessels and the efficacy of therapeutic angiogenesis.     

Soluble VEGF receptors

The three human VEGF receptors 1–3 mediate biological signals important for new blood vessel formation and lymphangiogenesis. Soluble VEGF receptors contain all the information necessary for high affinity ligand binding and have been used as experimental tools and regulators in several angiogenic in vitro and in vivo models. Recombinant receptor molecules can be used for specific inhibition of VEGF mediated signal transduction and for blocking tumor angiogenesis by limiting the amount of VEGF secreted from tumor cells or stroma cells. A naturally occurring soluble VEGFR-1 has been discovered in the supernatant from endothelial cells and at present appears to be the key regulator for the availability of VEGF secreted from different cells and tissues. The exact physiological role has not yet been demonstrated. This revised version was published online in June 2006 with corrections to the Cover Date.     

Association of the placental VEGF promoter polymorphisms and VEGF mRNA expression with preeclampsia

Background: Preeclampsia (PE), is a pregnancy-specific complication with the placental origin which associated with altered expression of angiogenic factors. Vascular Endothelial Growth Factor (VEGF), is a growth and hypoxia-induced factor which contributes to the regulation of various processes. The present study has investigated the association of the placental VEGF ?634G/C (rs2010963), ?1154G/A (rs1570360), and ?2549 I/D (18bpindel) polymorphisms in the promoter region with VEGF mRNA expression in PE women and control group.

Methods: This case-control study was performed on the placenta of 84 PE women and 103 controls after delivery. Genotyping of the VEGF polymorphisms was done by PCR or PCR, PCR-RFLP or sequencing methods. The mRNA expression levels were measured by Quantitative Real-Time PCR.

Results: The relative mRNA expression of VEGF gene was significantly higher in PE women compared to controls. The relative mRNA expression of VEGF gene was significantly higher in women with ?634CC genotype compared to CG + GG genotypes in PE women and total studied women but not in control women. However, there was no association between the placental VEGF ?1154G/A and ?2549 I/D polymorphisms and VEGF mRNA expression neither in PE nor in control groups.

Conclusion: The current study found higher mRNA expression of placental VEGF gene in PE women. The mRNA expression of the placental VEGF gene has been up-regulated in the placenta of women with ?634CC genotype. No association was found between the placental VEGF ?1154G/A and ?2549 I/D polymorphisms and VEGF mRNA expression.     

喉癌血清VEGF水平与癌组织中VEGF表达及MVD的关系

目的 探讨喉癌患者血清血管内皮生长因子(sVEGF)水平与喉癌血管生成的关系.方法 采用酶联免疫吸附试验(ELISA)检测50例喉癌、20例喉良性病变组织的sVEGF水平,免疫组化SABC法检测相应喉癌和喉良性病变组织中VEGF及微血管密度(MVD)表达水平,并分析sVEGF水平与VEGF及MVD表达的关系.结果 喉癌组sVEGF、VEGF及MVD的表达均明显高于喉良性病变组(P<0.01或P<0.05).喉癌患者sVEGF水平与癌组织中VEGF及MVD表达均呈正相关(P均<0.01).结论 喉癌患者sVEGF水平与癌组织中VEGF及MVD表达呈正相关,sVEGF水平或许可作为判断喉部疾病良恶性及预后的一个辅助参考指标.     

Degradation of soluble VEGF receptor-1 by MMP-7 allows VEGF access to endothelial cells

Vascular endothelial growth factor (VEGF) signaling in endothelial cells serves a critical role in physiologic and pathologic angiogenesis. Endothelial cells secrete soluble VEGF receptor-1 (sVEGFR-1/sFlt-1), an endogenous VEGF inhibitor that sequesters VEGF and blocks its access to VEGF receptors. This raises the question of how VEGF passes through this endogenous VEGF trap to reach its membrane receptors on endothelial cells, a step required for VEGF-driven angiogenesis. Here, we show that matrix metalloproteinase-7 (MMP-7) degrades human sVEGFR-1, which increases VEGF bioavailability around the endothelial cells. Using a tube formation assay, migration assay, and coimmunoprecipitation assay with human umbilical vein endothelial cells (HUVECs), we show that the degradation of sVEGFR-1 by MMP-7 liberates the VEGF(165) isoform from sVEGFR-1. The presence of MMP-7 abrogates the inhibitory effect of sVEGFR-1 on VEGF-induced phosphorylation of VEGF receptor-2 on HUVECs. These data suggest that VEGF escapes the sequestration by endothelial sVEGFR-1 and promotes angiogenesis in the presence of MMP-7.     

靶向肝癌HepG2细胞VEGF基因的siRNA表达载体构建及体外抑制作用

目的: 构建靶向肝癌HepG2细胞VEGF基因的siRNA表达载体并在体外检测其对VEGF基因表达的抑制作用.方法: 设计合成靶向HepG2细胞VEGF基因的siRNA cDNA序列并与pSUPER载体连接, 构建VEGF siRNA表达载体, 经酶切鉴定和测序确认后, 脂质体2000介导VEGF siRNA转染HepG2细胞. RT-PCR及Western blot检测VEGFsiRNA表达载体转染的HepG2细胞中VEGF基因表达.结果: 通过双酶切鉴定和测序分析, 成功构建了靶向HepG2细胞VEGF基因的siRNA表达载体. RT-PCR和Western blot法检测转染VEGFsiRNA表达载体的HepG2细胞VEGF mRNA及VEGF165表达下调, 其抑制率分别为65%和74%. 实验组中的HepG2中VEGF mRNA表达较阴性对照组、空载体组表达量显著下降(0.304±0.062 vs 0.896±0.061, 0.884±0.074,P<0.05).结论: 成功构建了靶向肝癌HepG2细胞VEGF基因的siRNA表达载体, 且该载体在体外能有效抑制HepG2细胞VEGF基因表达.     

Dual effects of heparin on VEGF binding to VEGF receptor-1 and transduction of biological responses

Vascular endothelial growth factor (VEGF) regulates blood vessel formation by binding to the receptor tyrosine kinases VEGF receptor-1 (Flt-1) or VEGF receptor-2 (KDR) and to the structurally unrelated neuropilins. As exon 7-containing isoforms of VEGF bind to heparin, angiogenesis may be modulated by heparin/heparan sulfate. We analyzed the effect of heparin on VEGF₁₆₅-binding and activation of VEGF receptor-1 in porcine aortic endothelial cells, which lack expression of VEGF receptor-2 and neuropilins. Heparin decreased binding of ¹²⁵I-VEGF to 50% at 5 μg/ml and cross-linking of ¹²⁵I-VEGF to VEGF receptor-1 on intact cells was similarly decreased. Schatchard analyses showed that the affinity for binding of ¹²⁵I-VEGF to VEGF receptor-1 was decreased in the presence of heparin. In contrast, VEGF receptor-1 kinase activity was elevated when cells were treated simultaneously with VEGF and heparin. In accordance, VEGF-induced tyrosine phosphorylation of phospholipase Cγ (PLCγ) and DNA synthesis were augmented by heparin. However, basal PLCγ tyrosine phosphorylation and DNA synthesis levels were to some extent increased by incubation of cells with heparin alone. We conclude that although heparin decreases binding of VEGF to VEGF receptor-1, the remaining binding results in more efficient kinase activation. Taken together, there is no loss of VEGF/VEGF receptor-1 function in the presence of heparin. This revised version was published online in June 2006 with corrections to the Cover Date.