Macrophage-derived matrix metalloproteinase-1 enhances aortic aneurysm formation in transgenic rabbits

Increased expression of matrix metalloproteinase-1 (MMP-1) has been observed in the lesions of atherosclerosis and aneurysms; however, it is not fully understood whether macrophage-derived MMP-1 affects these diseases. To investigate whether macrophage-derived MMP-1 participates in the development of vascular diseases, we generated transgenic (Tg) rabbits expressing human MMP-1 in the monocyte/macrophage lineage under the control of the human scavenger receptor enhancer/promoter. Tg rabbits exhibited no visible abnormalities throughout their bodies. Western blotting analysis revealed that the amount of MMP-1 proteins in the conditioned media secreted from peritoneal macrophages of Tg rabbits was up to 3-fold higher than that in non-Tg rabbits. For the first experiment, Tg and non-Tg rabbits were fed a cholesterol diet for 16 weeks, and aortic and coronary atherosclerosis were evaluated. The gross lesion area of aortic atherosclerosis in Tg rabbits was not significantly different from that in non-Tg rabbits, but Tg rabbits had marked destruction of the medial elastic lamina of the aortic lesions on microscopic examination. For the second experiment, we generated aortic aneurysms by incubating with elastase. Compared with non-Tg rabbits, Tg rabbits exhibited a significantly greater aortic dilation. Increased macrophage-derived MMP-1 led to increased medial destruction in both aortic atherosclerosis and aneurysms. These results demonstrate that MMP-1 plays a different role in the pathogenesis of atherosclerosis and aneurysms.     

转染VEGF165基因抑制兔动脉损伤后内膜增生

目的：观察局部转染血管内皮生长因子165（VEGF165）基因对损伤动脉内膜增生的影响并探讨可能机制。 方法： 采用显微外科手术方法，建立兔右髂外动脉损伤模型。将105只新西兰大白兔随机分为3组，每组35只。A组为生理盐水对照组，B组为脂质体介导的pBudCE4.1转染组，C组为脂质体介导的pBudCE4.1/VEGF165转染组。用微注射器将各种转染液注入损伤的血管壁，每组按实验终点（术后2、3、7、14和28 d）分为5个亚组，每个亚组7只兔。于术后各实验终点，取损伤段的血管用于病理组织学检查、电镜观察、RT-PCR和免疫组化染色检查。 结果： 术后各时点C组血管内膜厚度和内膜面积均显著小于A组和B组（P<0.01），术后28 d时C组管腔狭窄率平均比A组和B组低51.6%和49.8％。术后各时点C组血管壁的VEGF165基因的mRNA表达量和血管壁VEGF165阳性细胞率明显高于A组和B组（P<0.01）。C组血管壁血管内皮细胞（VEC）修复和生长增殖速度明显快于A组和B组。A组和B组间无明显差异。 结论： 局部转染VEGF165基因可抑制血管新生内膜增生及血管再狭窄，为将来血管内膜增生的基因治疗奠定基础。     

脂质体介导的VEGF_(165)基因转染对内皮细胞生长的作用

构建血管内皮细胞生长因子基因VEGF165真核表达载体pcDNA3 VEGF165,以阳离子脂质体介导的基因转染技术 ,将基因转入原代培养的人脐静脉内皮细胞中。结果表明 ,基因转染 1h后细胞内就有VEGFDNA存在 ,VEGFmRNA水平显著上升 ;基因转染 2d后培养液上清VEGF蛋白表达显著上升 (135 5 12± 6 2 34)pg/ml和 (19 2 7± 2 96 )pg/ml,P <0 0 1。转染VEGF165基因 2d的内皮细胞再经程序降温冷冻保存复苏后 ,其存活率显著高于对照组 (pcDNA3 组 ) (90 13%± 2 84 %和81 5 2 %± 2 15 % ,P <0 0 5 ) ,凋亡率显著低于对照组 (7 15 %± 0 4 2 %和 17 6 1%± 1 5 6 % ,P <0 0 5 )。MTT法显示转染VEGF165基因能促进内皮细胞VEGF蛋白的表达 ,促进细胞增殖 ,抑制细胞凋亡。在治疗心脏及下肢动脉缺血性疾病中 ,VEGF165基因治疗可能具有重要的意义。     

Angiopoietin-1 and vascular endothelial growth factor expression in human esophageal cancer

Angiopoietin-1 (Ang-1) has been shown to act as an angiogenic promoter in embryonic angiogenesis by promoting vascular branching, pericyte recruitment and endothelial survival. Ang-1 expression has not been examined in human esophageal cancer. We examined Ang-1 and vascular endothelial growth factor (VEGF) gene expression in tumors from 45 esophageal cancer patients who underwent surgical resection. Forty (88.9%) of the 45 esophageal cancers revealed Ang-1 gene expression. VEGF121, VEGF165 and VEGF189 isoforms were detected in 93.3 (42/45), 55.6 (25/45) and 26.7% (12/45) of the cases, respectively. Ang-1 gene expression was significantly correlated with VEGF121 and VEGF165 gene expression (P=0.0289 and P=0.0127, respectively, Fisher's test). The results suggest that Ang-1 is associated with neovascularization in the cancer stroma through VEGF net-works in esophageal cancer.     

Assessment of the biological activity of an improved naked-DNA vector for angiogenesis gene therapy on a novel non-mammalian model

From the basic expression vector p/hVEGF165, containing a cDNA sequence encoding the 165-amino-acid isoform of human vascular endothelial growth factor (VEGF165), we generated an improved construct (p163/hVEGF165) by subcloning at the 5' exact end of the VEGF165 cDNA a 163-bp IRES element belonging to the 1,014-bp, 5'-untranslated region of the murine VEGF gene. This IRES structure caused enhanced synthesis and increased secretion of the mature protein both in HEK-293 and in COS-7 cells, when compared to the basic construct. Both p/hVEGF165 and p163/hVEGF165 vectors were tested for in vivo angiogenic activity on a novel hirudinean model. As expected, the p/hVEGF165 vector injected as naked DNA was able to induce angiogenesis in the non-vascularized muscular tissue of Hirudo medicinalis. This model also allowed us to monitor intracellular synthesis of VEGF165 and subsequent interstitial secretion from muscle cells. Interestingly, significantly larger muscle tissue areas underwent marked angiogenesis when the improved vector p163/hVEGF165 was injected in H. medicinalis. It thus appears that the p163/hVEGF165 construct allows enhanced expression of the human VEGF165 gene, which in turn is responsible for increased secretion of biologically active growth factor by transduced cells. Since a naked-DNA vector very similar to p/hVEGF165 was recently found to be very active in humans for treatment of heart and limb ischemia, we suggest that our improved construct might be further tested in view of potential therapeutic applications.     

人VEGF165基因真核表达质粒的构建及其对血管内皮细胞增殖的影响

目的 :构建人VEGF16 5基因的真核表达质粒pBudCE4 .1/VEGF16 5 ,观察其在血管内皮细胞 (VEC)中的表达和表达产物对VEC增殖的影响。方法 :用RT PCR法从引产胎儿心脏组织中克隆VEGF16 5基因 ,并将其克隆至真核表达质粒pBud CE4 .1中 ,对重组真核表达质粒pBudCE4 .1/VEGF16 5进行酶切鉴定和测序。以脂质体转染法将pBudCE4 .1/VEGF16 5导入VEC中 ,用Northernblot和免疫细胞化学染色法 ,分别从mR NA水平和蛋白质水平检测它们在转染的VEC中的表达 ,并检测表达产物对VEC增殖的影响。结果 :人VEGF16 5基因的RT PCR产物为 5 76bp。测序结果显示 ,扩增的VEGF16 5基因的序列与基因文库中登录的序列完全一致。经HindⅢ和BamHI酶切鉴定证实 ,VEGF16 5基因已成功地克隆至真核表达质粒pBudCE4 .1中。以其转染血管内皮细胞 (VEC)后 ,经Northernblot杂交和免疫细胞化学染色法检测均证实VEGF16 5基因表达。表达产物对VEC增殖有明显的促进作用。结论 :所构建的pBudCE4 .1/VEGF16 5真核表达质粒可在VEC中表达 ,表达产物可明显促进VEC增殖 ,为通过VEGF16 5基因转染防治移植器官内血管的狭窄奠定了基础     

In vitro and in vivo study of the expression vector encoding vascular endothelial growth factor

Vascular endothelial growth factor (VEGF) is an angiogenic cytokine with potential therapeutic applications in human diseases. It is a mitogen primarily for endothelial cells. The transfer of the cDNA encoding VEGF to ischemic tissues, which cannot be revascularized otherwise, represents a novel and promising approach to the treatment of vascular disorders. In this work the VEGF165 cDNA was cloned into the expression vector pSecTag2B. The activity of the construct was studied in cell culture as well as in vivo. Western blotting study showed that the cells transfected with the vector secreted significantly higher amounts of VEGF to the culture medium than the non-transfected cells. In vivo study revealed an increased number of new vessels in animals injected with vector encoding VEGF as compared with empty plasmid. Also, tumor cells transfected with the VEGF plasmid exhibited extensive vascularization.     

Transfection of VEGF(165) genes into endothelial progenitor cells and in vivo imaging using quantum dots in an ischemia hind limb model

Endothelial progenitor cells (EPCs) were transfected with fluorescently labeled quantum dot nanoparticles (QD NPs) with or without VEGF(165) plasmid DNA (pDNA) to probe the EPCs after in?vivo transplantation and to test whether they presented as differentiated endothelial cells (ECs). Bare QD NPs and QD NPs coated with PEI or PEI?+?VEGF(165) genes were characterized by dynamic light scattering, scanning electron microscopy, and atomic force microscopy. Transfection of EPCs with VEGF(165) led to the expression of specific genes and proteins for mature ECs. A hind limb ischemia model was generated in nude mice, and VEGF(165) gene-transfected EPCs were transplanted intramuscularly into the ischemic limbs. At 28 days after transplantation, the VEGF(165) gene-transfected EPCs significantly increased the number of differentiated ECs compared with the injection of medium or bare EPCs without VEGF(165) genes. Laser Doppler imaging revealed that blood perfusion levels were increased significantly by VEGF(165) gene-transfected EPCs compared to EPCs without VEGF(165). Moreover, the transplantation of VEGF(165) gene-transfected EPCs increased the specific gene and protein expression levels of mature EC markers and angiogenic factors in the animal model.     

激素性股骨头坏死模型兔血管内皮细胞生长因子表达与普伐他汀的干预

背景：有研究表明他汀类药物可使体外培养的人脐静脉内皮细胞的血管内皮细胞生长因子呈高表达,血管内皮细胞生长因子可促进内皮细胞增殖,明显提高成骨细胞活性并加速骨形成。 目的：观察普伐他汀对激素性股骨头坏死兔模型血管内皮生长因子的蛋白表达的影响。 方法：将新西兰白兔随机分为对照组,模型组和普伐他汀组。模型组和普伐他汀组制备兔激素性股骨头缺血坏死模型。普伐他汀组建模成功后以普伐他汀(1.2 mg/kg)灌胃,1次/d,模型组和对照组以等体积的蒸馏水灌胃。于造模后8,12,16周截取各组股骨头行免疫组织化学检测血管内皮生长因子蛋白的表达情况。 结果与结论：对照组不同时间点股骨头血管内皮生长因子蛋白表达均为阳性。造模后8周,模型组血管内皮生长因子蛋白表达呈阴性表达,普伐他汀组呈弱阳性表达。造模后12周,模型组和普伐他汀组血管内皮生长因子蛋白表达呈阳性表达,16周呈弱阳性表达。结果证实,普伐他汀可有效促进早期激素性股骨头坏死兔模型坏死股骨头内源性血管内皮生长因子的蛋白表达。     

人血管内皮生长因子基因的克隆、表达及生物活性分析

目的 克隆人血管内皮细胞生长因子165(VEGF165)基因,构建真核表达载体,观察其对脐静脉内皮细胞的增殖作用和血管新生的影响。方法 利用RT-PCR方法,从人扁桃体组织中扩增人VEGF165 cDNA完整编码区,并构建成pcDNA3．1( )／VEGF165(简称pcDNA／V)重组体;应用脂质体介导的基因转移技术将构建的真核表达载体pcDNA／V体外转染至人脐静脉内皮细胞(HUVEC),MTT法检测其对内皮细胞增殖的影响。建立家兔下肢缺血模型,注射重组质粒pcDNA／V,pcDNA3．1( )空质粒作对照,选取不同时间点,行血管造影。结果 构建的真核表达载体pcDNA／V的酶切电泳分析和测序表明结果正确。pcDNA／V转染HUVEC能明显促进内皮细胞的分裂增殖。血管造影显示,术后基因治疗组远端动脉充盈早于对照组,新生血管数目也明显多于同时期对照组。结论 成功克隆了人VEGF165基因,构建了其真核表达载体。体内外生物学活性研究证实,重组质粒的表达产物具有刺激HUVEC增殖和促进缺血肢体侧枝循环建立的功能。     

Vascular endothelial growth factor-165 overexpression stimulates angiogenesis and induces cyst formation and macrophage infiltration in human ovarian cancer xenografts

Vascular endothelial growth factor (VEGF) is suggested to be an important regulator of angiogenesis in ovarian cancer. We have evaluated the effects of VEGF overexpression on the histology and growth rate of human ovarian cancer xenografts. OVCAR-3 human ovarian cancer cells were stably transfected with an expression vector encoding the 165-amino acid isoform of VEGF. As subcutaneous xenografts, moderately and highly VEGF(165)-overexpressing OVCAR-3 cells formed tumors with large cysts. Immunohistochemistry demonstrated an increase in the number of CD31-positive microvessels, some of which were larger in diameter than those in the parental tumors, as well as extensive vascular rimming around the cysts. Weakly VEGF(165)-overexpressing tumors also contained an increased number of CD31-positive microvessels and occasional vascular rimming, but cysts were not present. Immunohistochemistry further revealed the presence of monocytes and macrophages in both parental and VEGF(165)-overexpressing xenografts. Interestingly, the number of monocytes/macrophages was greatly increased in moderately and highly VEGF(165)-overexpressing xenografts and large areas populated with monocytes/macrophages were detected within the tumor stroma. Although the higher number of CD31-positive cells would suggest a better vascularization pattern in VEGF(165)-overexpressing xenografts, tumor growth rates were not increased when compared with that of parental xenografts. These data provide functional evidence for a role of VEGF(165) in cyst formation and monocyte/macrophage infiltration.     

Evidence for intramolecular B-cell epitope spreading during experimental immunization with an immunogenic thyroglobulin peptide

Thyroglobulin (Tg) is a target autoantigen in autoimmune thyroid diseases, such as Graves' disease (GD) and Hashimoto's thyroiditis. In a previous study we identified three 20mer Tg peptides bearing epitopes of autoantibodies associated with GD (TgP15, TgP26 and TgP41: sequences 2339-2358, 2471-2490 and 2651-2670 of human Tg, respectively). In the present study, we investigated the antigenicity of the above peptides in experimental immunization with Tg, the immunogenicity of antigenic peptides and the possibility of intramolecular B-cell epitope spreading during peptide immunization. For this purpose, two rabbits were injected with human Tg in CFA six times, every three weeks. Two control animals were injected only with CFA. Testing of antisera and of affinity-purified antibodies, by ELISA against the three peptides, revealed reactivity only to TgP41. This synthetic peptide was subsequently administered to two rabbits, in its free form (100 micro g in CFA six times, every two weeks). A strong serological response was developed not only against TgP41, but also to intact human and rabbit Tg. Immunization with TgP41 induced intramolecular B-cell epitope spreading, i.e. production of antibodies to sites on Tg other than that corresponding to TgP41, as revealed by immunoadsorption and competitive ELISA. Histopathological studies did not reveal any infiltration in thyroid glands. We conclude that peptide TgP41 encompasses not only an epitope of disease-associated autoantibodies, but also a dominant immunogenic epitope of experimentally induced Tg-specific antibodies, able to drive B-cell epitope spreading.     

Expression of vascular endothelial growth factor by human renal cancer cells enhances angiogenesis of primary tumors and production of ascites but not metastasis to the lungs in nude mice

We determined the role that vascular endothelial growth factor (VEGF), also known as vascular permeability factor (VPF), plays in the progression of human renal cell cancer in nude mice. Low metastatic and low VEGF/VPF-expressing human renal cancer cells SN12C were transfected with the VEGF165 cDNA or plasmid alone as control. VEGF165-transfected SN12C cells produced large amounts of biologically active VEGF in culture that did not affect cell doubling time or confluence. Subsequent to implantation into the renal subcapsule of nude mice, the VEGF165-transfected SN12C cells produced fast-growing (PCNA labeling), large tumors that expressed high levels of VEGF/VPF and were well vascularized (CD3-positive vessels). The tumors produced hyperpermeability of peritoneal blood vessels (Evans blue dye-leak assay), bloody ascites, and short survival time. Parental or control transfected SN12C cells produced less vascularized, slower growing tumors with no ascites. Regardless of in vivo expression level of VEGF, the incidence of spontaneous lung metastasis was low, suggesting that in itself, the expression of VEGF/VPF by renal cancer cells is not sufficient to produce metastasis. This revised version was published online in July 2006 with corrections to the Cover Date.     

人血管内皮生长因子的克隆表达及免疫学检测工作标准品的制备

目的 :克隆表达人血管内皮生长因子 16 5 (humanvascularendothelialgrowthfactor 16 5 ,hVEGF16 5 )基因 ,制备VEGF免疫学检测工作标准品。方法 :用RT -PCR从肝癌组织扩增目的基因VEGF16 5 ,插入到pUCm -T质粒中并测序鉴定。构建原核表达重组质粒 pET32a -hVEGF16 5 ,转化入大肠杆菌 ,经IPTG诱导 ,固化Ni2 吸收色谱纯化得到重组人VEGF16 5 (recombinanthumanVEGF16 5 ,rhVEGF16 5 )。用WsternBlot检测rhVEGF16 5免疫原性 ,并通过HUVEC增殖试验检测rhVEGF16 5生物学活性。按标准品的要求制备VEGF检测工作标准品 ,最后用ELASA试剂盒标定。结果 :PCR产物为 6 0 0bp ,测序结果表明其序列正确 ,人VEGF16 5可在大肠杆菌内可溶性表达 ,rhVEGF16 5蛋白相对分子质量为 38kD ,具有生物学活性。工作标准品浓度标定为 5 0 5pmol/L。结论 :成功地克隆了有生物学活性的VEGF16 5基因 ,并制备出VEGF免疫学检测工作标准品。     

Delivery of high dose VEGF plasmid using fibrin carrier does not influence its angiogenic potency

Delivery of DNA mixed with a degradable matrix carrier was supposed to improve transgene expression. Using a rabbit hind-limb ischemia model, we tested the angiogenic potency of plasmid encoding human vascular endothelial growth factor (pSG5-VEGF165) entrapped in fibrin sealant. Animals were injected intramuscularly with 500 microg of pSG5-VEGF165 or control plasmid, dissolved in saline (PBS) or fibrin glue. After 14 days, presence of delivered constructs and expression of transgene was confirmed in injected muscles of all animals. There were no significant differences in the levels of human VEGF mRNA and protein between VEGF-PBS and VEGF-fibrin groups (Mann-Whitney test). Accordingly, pSG5-VEGF165 regardless of the way of delivery, induced similar increases in capillary density within treated muscles (ANOVA). Control plasmid did not show any effects. In conclusion, injection of pSG5-VEGF165 into ischemic adductor muscle leads to synthesis of human VEGF and increases the number of capillaries. Fibrin carrier does not influence its angiogenic potential.     

Angiogenesis in hereditary hemorrhagic telangiectasia: VEGF165 plasma concentration in correlation to the VEGF expression and microvessel density

Angiogenesis defines the physiologic process of capillary blood vessel formation. It is a multistep process, which is controlled by a large number of pro- and anti-angiogenic factors. Vascular endothelial growth factors (VEGF) are the best characterized pro-angiogenic factors, which play a key role in angiogenesis. Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant, multi-systemic disorder of angiogenesis, clinically characterized by severe and recurrent hemorrhages. We assume in HHT patients with increased VEGF plasma levels a high VEGF expression also correlates with the degree of microvesssel density (MVD). In 41 HHT patients and 47 healthy patients, the VEGF165 plasma concentration was determined by standard ELISA technique. Cryostat sections of 13 HHT patients were immunostained for VEGF and endothelial cells by an anti-vWF monoclonal antibody using a standard streptavidinbiotin complex procedure. The degree of vessel density was quantified by light microscopy (x200) within 'hot spot' areas of the HHT-tissue. All HHT cryostat sections showed a medium to strong staining for VEFG compared to healthy control tissue. The VEGF expression correlated with the VEGF165 plasma concentration and the mean MVD in HHT patients. HHT patients with medium VEGF staining revealed significantly lower VEGF165 plasma concentrations and a lower mean MVD (204+/-12 vessels/per microscopic field) than HHT patients with strong VEGF staining (327+/-76 vessels/per microscopic field). High VEGF expression in patients with HHT in correlation to their VEGF165 plasma levels and the microvessel density may support the theory that VEGF functions as an important regulator and key protein of angiogenesis, even in HHT.     

携带人血管内皮生长因子165慢病毒表达载体的构建与表达*

背景：已有研究证实血管内皮生长因子在正常肝脏肝部分切除后余肝的再生过程中发挥着重要的作用,但关于其对肝硬化肝脏是否也有相同作用国内外鲜有报道。 目的：构建携带人血管内皮生长因子165基因的慢病毒载体,体外转染BLR 3A大鼠肝细胞并观察该细胞中人血管内皮生长因子165基因的表达。 方法：采用DNA重组技术将人血管内皮生长因子165基因克隆入pLenti6/V5-D-TOPO慢病毒表达载体中,筛选出阳性克隆与慢病毒包装系统ViraPowerTM Packaging Mix共转染293T细胞产生病毒颗粒,通过实时定量-聚合酶链反应法测定病毒滴度;携带人血管内皮生长因子165基因的慢病毒载体体外转染BLR 3A大鼠肝细胞72 h后,利用反转录-聚合酶链反应及Western blot法检测细胞中人VEGF165 mRNA及蛋白的表达。 结果与结论：成功构建了表达人VEGF165基因的慢病毒载体pLenti6/V5-D-TOPO-VEGF165,测得病毒滴度为1.18×     107 VP/mL。重组慢病毒载体转染BLR 3A大鼠肝细胞72 h后,荧光蛋白表达率超过80%,反转录-聚合酶链反应及Western blot法测得转染组人血管内皮生长因子165 mRNA及血管内皮生长因子165蛋白表达阳性。提示构建的携带人血管内皮生长因子165的慢病毒载体可有效转染BLR 3A大鼠肝细胞,并促使该细胞表达人血管内皮生长因子165 mRNA及蛋白。 关键词：人血管内皮生长因子165;慢病毒载体;BLR 3A大鼠肝细胞;转染;基因治疗 doi:10.3969/j.issn.1673-8225.2012.11.007     

Expression of vascular endothelial growth factor and vascular endothelial growth factor receptor-2 (KDR/Flk-1) in ischemic skeletal muscle and its regeneration

Vascular endothelial growth factor (VEGF) is a hypoxia-inducible endothelial cell mitogen and survival factor. Its receptor VEGFR-2 (KDR/Flk-1) mediates these effects. We studied the expression of VEGF and VEGFR-2 in ischemic human and rabbit skeletal muscle by immunohistochemistry and in situ hybridization. Human samples were obtained from eight lower limb amputations because of acute or chronic critical ischemia. In chronically ischemic human skeletal muscle VEGF and VEGFR-2 expression was restricted to atrophic and regenerating skeletal myocytes, whereas in acutely ischemic limbs VEGF and VEGFR-2 were expressed diffusely in the affected muscle. Hypoxia-inducible factor-1alpha was associated with VEGF and VEGFR-2 expression both in acute and chronic ischemia but not in regeneration. Hindlimb ischemia was induced in 20 New Zealand White rabbits by excising the femoral artery. Magnetic resonance imaging and histological sections revealed extensive ischemic damage in the thigh and leg muscles of ischemic rabbit hindlimbs with VEGF expression similar to acute human lower limb ischemia. After 1 and 3 weeks of ischemia VEGF expression was restricted to regenerating myotubes and by 6 weeks regeneration and expression of VEGF was diminished. VEGFR-2 expression was co-localized with VEGF expression in regenerating myotubes. Macrophages and an increased number of capillaries were associated with areas of ischemic muscle expressing VEGF and VEGFR-2. In conclusion, two patterns of VEGF and VEGFR-2 expression in human and rabbit ischemic skeletal muscle are demonstrated. In acute skeletal muscle ischemia VEGF and VEGFR-2 are expressed diffusely in the affected muscle. In chronic skeletal muscle ischemia and in skeletal muscle recovering from ischemia VEGF and VEGFR-2 expression are restricted to atrophic and regenerating muscle cells suggesting the operation of an autocrine pathway that may promote survival and regeneration of myocytes.     

Vascular endothelial growth factor enhances glomerular capillary repair and accelerates resolution of experimentally induced glomerulonephritis

Vascular endothelial growth factor (VEGF) regulates angiogenesis through endothelial cell proliferation and plays an important role in capillary repair in damaged glomeruli. We tested the hypothesis that VEGF might be beneficial in rats with severe glomerular injury in glomerulonephritis (GN) based on its angiogenic and vascular remodeling properties. Acute GN with severe glomerular destruction was induced in rats by injection of anti-Thy-1.1 antibody (day 0) and Habu-snake venom (day 1). Rats were intraperitoneally injected with recombinant human VEGF(165) (10 microg/100 g body wt/day) or vehicle from day 2 to day 9, and monitored changes in glomerular capillaries, development of glomerular inflammation, and progression to glomerular sclerosis after acute glomerular destruction in both groups. Rats that received anti-Thy-1.1 antibody and Habu-snake venom showed severe mesangiolysis and marked destruction of capillary network on day 2. VEGF was expressed on glomerular epithelial cells, proliferating mesangial cells, and some infiltrating leukocytes, and VEGF(165) protein levels increased in damaged glomeruli during day 5 to day 7. Normal, damaged, and regenerating glomerular endothelial cells expressed VEGF receptor flk-1. However, endothelial cell proliferation and capillary repair was rare in vehicle-treated rats with severe glomerular damage, which progressed to global sclerosis and chronic renal failure by week 8. In contrast, in the VEGF-treated group, VEGF(165) significantly enhanced endothelial cell proliferation and capillary repair in glomeruli by day 9 (proliferating endothelial cells: VEGF(165), 4.3 +/- 1.1; control, 2.2 +/- 0.9 cells on day 7, P < 0.001; and glomerular capillaries: VEGF(165), 24.6 +/- 4.8; control, 16.9 +/- 3.4 capillaries on day 7, P < 0.01). Thereafter, damaged glomeruli gradually recovered after development of capillary network by week 8, and significant improvement of renal function was evident in the VEGF-treated group during week 8 (creatinine: VEGF(165), 0.3 +/- 0.1; control, 2.6 +/- 0.9 mg/dl, P < 0.001; proteinuria: VEGF(165), 54 +/- 15; control, 318 +/- 60 mg/day, P < 0.001). We conclude that the beneficial effect of VEGF(165) in severe glomerular injury in GN emphasizes the importance of capillary repair in the resolution of GN, and may allow the design of new therapeutic strategies against severe GN.