上皮性卵巢肿瘤患者血管内皮生长因子水平的检测

目的检测卵巢上皮性肿瘤患者血管内皮生长因子(VEGF)水平的变化,探讨其临床价值。方法采用酶联免疫吸附试验双抗体夹心法(ELISA)检测20例上皮性卵巢肿瘤、12例交界性上皮性卵巢肿瘤和48例上皮性卵巢癌患者血中VEGF水平。结果良性、交界性及恶性卵巢上皮肿瘤患者血VEGF水平依次升高分别为(686±150)、(750±90)和(1183±293)ng/L,P<0.01;血VEGF在浆液性卵巢腺癌、低分化卵巢腺癌中水平明显升高(P=0.046和P=0.029),而与临床分期无关。结论检测血VEGF水平有助于恶性卵巢上皮性肿瘤的术前诊断及估计预后。     

骨骺损伤修复过程中骺板形态及VEGF表达的变化

 目的 探讨大鼠骨骺损伤修复过程中骺板形态结构及骺板内血管内皮细胞生长因子(vascular endothelial growth factor, VEGF)表达的变化。方法 取4~5 周龄SD大鼠30 只, 制作胫骨近端骨骺损伤动物模型。随机分为五组, 每组6 只, 分别于术后2、4、6、10、21 天处死, 取双侧胫骨。测量双侧胫骨长度, 计算实验侧长度和对照侧长度比值。行双侧胫骨X 线摄片、Micro CT扫描, 观察骺板大体形态及骺板内骨桥形成情况。通过组织切片HE 染色及免疫组织化学染色, 观察骺板内软骨细胞变化及VEGF表达。结果 双侧肢体长度于术后第4 天出现差异, 第10 天差异最大, 第21 天肢体长度差异缩小。术后第6 天开始骺板内逐渐出现纤细骨质, 终至骨桥形成。HE 染色提示伤后早期出现静止区软骨细胞聚集成团, 细胞规律性排列丧失、分化加速。免疫组织化学染色显示术后生长板内出现VEGF 高反应区, 阳性表达区逐渐扩大, 跨越骺板全层, 直后血管长入、骨化。结论 骨骺损伤修复过程中, 骺板形态早中期表现为干骺端肥大, 肢体短缩, 骺板内纤细骨质形成;随修复进展, 骺板内VEGF 表达反应性增强, 出现贯通骺板的表达带, 与骨桥形成相关。     

Promotion of acute‐phase skin wound healing by Pseudomonas aeruginosa C4‐HSL

A Pseudomonas aeruginosa quorum‐sensing system, which produces N‐(3‐oxododecanoyl)‐l ‐homoserine lactone (3‐oxo‐C₁₂‐HSL) and N‐butanoyl‐l ‐homoserine lactone (C₄‐HSL), regulates the virulence factors. In our previous study, 3‐oxo‐C₁₂‐HSL, encoded by lasI gene, was shown to promote wound healing. However, the effect of C₄‐HSL, encoded by rhlI gene, remains to be elucidated. We addressed the effect of C₄‐HSL on wounds in P. aeruginosa infection. Wounds were created on the backs of Sprague–Dawley SD rats, and P. aeruginosa PAO1 (PAO1) or its rhlI deletion mutant (ΔrhlI) or lasI deletion mutant (ΔlasI) was inoculated onto the wound. Rats were injected intraperitoneally with anti‐C₄‐HSL antiserum or treated with C₄‐HSL at the wound surface. PAO1 inoculation led to significant acceleration of wound healing, which was associated with neutrophil infiltration and TNF‐α synthesis. These responses were reversed, except for TNF‐α production, when ΔrhlI was inoculated instead of PAO1 or when rats were co‐treated with PAO1 and anti‐C₄‐HSL antiserum . In contrast, the healing process and neutrophil infiltration, but not TNF‐α synthesis, were accelerated when C₄‐HSL was administered in the absence of PAO1. This acceleration was not affected by anti‐TNF‐α antibody. These results suggest that C₄‐HSL may be involved in the acceleration of acute wound healing in P. aeruginosa infection by modifying the neutrophilic inflammation.     

S100A4和血管内皮生长因子C、D在胃癌组织中的表达及其与淋巴结转移和预后的关系

目的 观察人胃癌S100A4和血管内皮生长因子(VEGF)-C、VEGF-D的表达,探讨其与临床病理特征及预后的关系.方法 应用免疫组织化学方法检测108例人胃癌及20例癌旁组织中S100A4和VEGF-C、VEGF-D的表达.分析S100A4和VEGF-C、VEGF-D表达与患者年龄、性别、肿瘤大小、病理类型、浸润深度、淋巴结转移和肿瘤TNM分期的关系,并作预后分析.结果 S100A4和VEGF-C、VEGF-D在胃癌组织中的阳性率明显高于癌旁组织(P＜0.05).S100A4的表达与肿瘤大小和淋巴结转移有关(P＜0.05),VEGF-C的表达与淋巴结转移和TNM分期有关(P＜0.05),VEGF-D的表达与淋巴结转移和TNM分期无明显相关(P＞0.05),肿瘤组织内S100A4和VEGF-C两者表达呈正相关(P＜0.05).S100A4和VEGF-C、VEGF-D阳性胃癌患者5年生存率分别低于阴性患者,差异无统计学意义(P＞0.05).结论 S100A4和VEGF-C的表达与胃癌的淋巴结转移有关,S100A4可能参与VEGF-C淋巴管生成通路,在胃癌的淋巴结转移中发挥重要作用.     

Effects of transcutaneous topical injection of oxygen on vascular endothelial growth factor gene into the healing ligament in rats.

The effects of intermittent exposure to oxygen injection on an experimentally induced ligament tear were studied in the right hind limb of 17 male Sprague-Dawley rats. Two rats were used for monitoring the partial oxygen pressure (pO(2)) of subcutaneous tissue and 15 rats were divided into the following three groups of 5 after an experimentally induced ligament tear: Group A, control group; Group B, injection of 0.5 ml hyaluronan to the wound transcutaneously; Group C, injection of 0.5 ml hyaluronan mixed with haemoglobin and oxygen (n=5). At 7 days post-ligament injury, we compared the ligaments of the three treatment groups for gross appearance, histology and expression of vascular endothelial growth factor (VEGF) mRNA by RT-PCR. Our results indicate that the pO(2) was immediately elevated to 334.6 mmHg by topical oxygen injection and this method was effective in promoting vessel formation in comparison to the control group (p<0.01). However, the expression of VEGF mRNA in the topical oxygen injection group (Group C) was lower than that in control group (p<0.05). Our results suggest that oxygen is able to accelerate vessel formation in spite of its effect of decreasing VEGF mRNA. Our method of using topical injection proved to be useful in healing the ligament and the wound.     

人血管内皮生长因子基因工程生物膜愈合大鼠皮肤创伤的初步研究

目的：观察含有人血管内皮生长因子（hVEGF）重组质粒的成纤维细胞种植的基因工程生物膜对于大鼠创面新生血管的生成以及创伤愈合的影响,为基因工程生物膜用于临床提供进一步的资料.方法：实验大鼠分为3组3个时相,分别为实验组Ⅴ：创面覆盖含有hVEGF重组质粒的成纤维细胞种植的基因工程生物膜后常规包扎;对照组B：创面覆盖生物膜后常规包扎;对照组O：创面进行常规包扎.术后7 d、14 d、29 d分别取创伤皮肤组织进行创面微血管密度（microvessel density,MVD）计数大鼠皮肤组织创面的新生血管量,同时观察创伤愈合的过程、通过Western印迹实验检测实验大鼠组织的VEGF蛋白表达.结果：Western印迹证实应用hVEGF制备的基因工程生物膜能使大鼠皮肤创面持续表达VEGF,表达量高于对照组;MVD计数表明创面覆盖含有hVEGF重组质粒的成纤维细胞种植的基因工程生物膜后新生血管明显多于对照组（P＜0.01）.结论：提示我室研制的基因工程生物膜能促进创面新生血管生成,以达到减少瘢痕形成、促进伤口愈合的目的.     

血管内皮生长因子和碱性成纤维细胞生长因子对大鼠外周血内皮祖细胞培养的影响

目的 探讨不同培养条件对大鼠外周血来源内皮祖细胞（EPC）生长情况的影响。 方法 密度梯度离心法获得大鼠外周血单个核细胞,根据培养基中是否添加血管内皮生长因子（VEGF）、碱性成纤维细胞生长因子（bFGF）及培养板是否预铺纤连蛋白（FN）分组培养。观察记录细胞生长情况并进行统计分析,以免疫组化和免疫荧光法进行鉴定。 结果 大鼠外周血来源的单个核细胞在体外呈现贴壁生长,培养第7天各组细胞数及细胞集落数提示,在相同培养条件下,预铺FN可以促进EPC的贴壁增殖（t = 4.43,P < 0.05;t = 3.70,P < 0.05）。在同样预铺或未铺FN的情况下,在培养液中加入生长因子可促使单个核细胞更好地向EPC分化（t = －10.96,P < 0.01;t = －13.22,P < 0.01）。免疫组化及免疫荧光结果显示,细胞培养第4、7、10天,细胞表面CD34、CD133表达不断增强[（35.7±4.2）%、（60.1±3.8）%、（81.8±6.4）%;（3.2±0.9）%、（18.4±7.3）%、（32±3.8）%],第14天下降[（32.1±5.4）%,（1.9±2.7）%];而Flk-1表达在第4、7、10、14天均不断增强[（31.2±3.5）%、（40.6±5.3）%、（71.2±8.4）%、（81.5±4.1）%]。 结论 FN有利于内皮祖细胞的贴壁生长和增殖。VEGF及bFGF促进其增殖分化。内皮祖细胞的体外成功培养将为其应用于血管组织工程提供足够数量的种子细胞来源,并为外周血干细胞移植治疗多种疾病提供新的思路。     

血管内皮细胞生长因子诱导肝癌转移的实验研究

目的　研究血管内皮细胞生长因子(VEGF)与肝癌细胞转移的相互关系。方法　运用BodyenChamber膜侵袭培养系统培养VEGF肝癌细胞株HepG     

粒细胞-单核巨噬细胞集落刺激因子对人皮肤成纤维细胞血管内皮细胞生长因子表达的影响

目的探讨粒细胞-单核巨噬细胞集落刺激因子（granulocyte-macrophage colony stimulating factor,GM—CSF）对人皮肤成纤维细胞血管内皮细胞生长因子（vascular endothelial cell growth factor,VEGF）表达的影响。方法分别用含GM—CSF（GM—CSF组）和不含GM—CSF（对照组）培养液,孵育离体培养的人皮肤成纤维细胞,作用不同时间后,采用逆转录-聚合酶链反应（RTPCR）、蛋白质印迹法（Western印迹法）、酶联免疫吸附试验（EI,1SA）分别检测人皮肤成纤维细胞VEGF mRNA表达和蛋白表达。结果GM—CSF作用1、3、6、12h后,人皮肤成纤维细胞VEGF mRNA表达均较对照组显著增强（P〈0．05）,其中GM—CSF作用6h后VEGFmRNA表达最强;与对照组比较,GM—CSF作用12、24、48h后。成纤维细胞内VEGF蛋白表达旺著增强,其中24h表达最强;细胞培养上清液中VEGF蛋白分泌堪随时间延长逐渐增加,各时相点GM-CSF组VEGF蛋白分泌量高于对照组（P〈0．01）。结论GM—CSF可促进人皮肤成纤维细胞VEGF的表达,这可能是GM-CSF加速创面血管新生化的机制之一。     

Extensive in vivo angiogenesis following controlled release of human vascular endothelial cell growth factor: implications for tissue engineering and wound healing

Vascular endothelial cell growth factor (VEGF) has strong stimulating effects on vascularization. Though very potent, VEGF is rapidly degraded due to its short half-life and when administrated by uncontrolled and nonspecific methods; however, its systemic administration in large doses can cause harmful side effects. Controlled release technology would allow delivering desired levels of bioactive VEGF within extended periods and permit examination of the in vivo effects of the compound in a broader way. The objective of this study was to determine the in vitro release behavior of VEGF from calcium alginate microspheres and the potency of this controlled release system in promoting localized neovascularization at the subcutaneous site of the rat model. In vitro release of human VEGF165 (2 and 4 microg/cm3 microsphere) was studied for 3 weeks under static conditions at 25 degrees C, and daily hormone release was measured using a competitive enzyme immunoassay. Following an uncontrolled release within the first 4 days, a quite constant zero-order VEGF release of 50 to 90 and 70 to 120 ng/day was achieved from 2 and 4 microg/cm3 polymer loaded microspheres respectively. In vivo angiogenesis was studied for a period of 8 weeks and evaluated using immunoperoidase staining and histopathological measurements. In vivo studies with rats (n = 24) showed a considerable level of capillary network formation at the epigastric groin fascia of VEGF microsphere-implanted rats starting from the first week. The most extensive neovascularization was observed in the group with 3 week postimplanted 4 microg VEGF containing microspheres; this level of vascularization was quite similar after 8 weeks. While the control group showed no evidence of angiogenesis, the difference in VEGF-induced neovascularization is statistically significant (p < 0.03). Immunostaining of the specimens showed a strong relationship between the release of human VEGF and neovascularization. The controlled VEGF release system described here promotes vigorous angiogenesis and has applicability for tissue engineering and wound healing studies.     

Alveolar and plasma concentrations of interleukin-8 and vascular endothelial growth factor following oesophagectomy

The acute respiratory distress syndrome occurs in approximately 10% of all patients undergoing elective oesophagectomy. Local increases in lung pro-inflammatory cytokines have been previously detected in high-risk patients before the development of the acute respiratory distress syndrome. We hypothesised that similar changes would occur following oesophagectomy. Two groups of patients were studied. In the collapsed lung group (n = 11), interelukin-8 and vascular endothelial growth factor were measured in bronchoalveolar lavage samples obtained from the intra-operative collapsed lung after operation. In the ventilated lung group (n = 10), bronchoalveolar lavage was performed after operation from the ventilated lung and cytokines measured. Cytokines were also measured in peripheral blood samples before and after operation. Bronchoalveolar lavage cytokine levels in both lungs were of an order of magnitude greater than in peripheral blood. Pulmonary pro-inflammatory cytokine release occurs following oesophageal surgery and may indicate subclinical lung injury.     

成骨细胞移植促进老龄鼠骨质疏松性骨折愈合过程中VEGF的动态变化

目的：动态观察在幼龄成骨细胞移植促进骨质疏松性骨折愈合过程中，VEGF在不同时相的表达及其生物学意义。方法：通过建立老龄骨质疏松SD大鼠骨折的动物模型，并将体外培养的SD雄性乳鼠成骨细胞移动到SD雌性鼠老年骨质疏松性骨缺损部位，利用免疫组化及原位杂交检测骨折愈合过程中不同时间相的移植标本VEGF、VEGFmRNA的表达，并作图像分析、绘出其动态变化图。结果：实验组VEGF、VEGFmRNA均在7d左右可见有阳性表达的细胞，14d有分泌,高峰其中以软骨细胞中阳性最强，21 d分泌量开始下降，56d后基本消失。而对照且未见明显分泌高峰。结论；成骨细胞细胞促进老龄鼠骨质疏松性骨折愈合，其机制可能是通过促进VEGF的转录和表达，从而促进骨折部位建立良好的血液循环，加速骨形成。     

普萘洛尔治疗增生期血管瘤患儿血清中血管内皮生长因子及基质金属蛋白酶-9的变化

目的 研究普萘洛尔口服治疗增生期血管瘤,及对血清中血管内皮生长因子（vascular endothelial growth factor,VEGF）、基质金属蛋白酶-9(matrix metalloproteinase-9,MMP-9)水平的影响。方法 采用ELISA法检测患者服药4、8周后血清VEGF、MMP-9水平,并分析其与预后的关系。结果 治疗前血清VEGF水平最高,而服药后4周(255.7±130.4)和服药后8周(224.2±120.6)VEGF水平呈逐渐下降趋势,治疗后8周VEGF水平显著低于治疗前(P＜0.05)。MMP-9的表达变化与VEGF一致,并与VEGF呈正相关。结论 普萘洛尔可以通过降低患者血清VEGF和MMP-9水平,从而治疗增生期血管瘤。     

胰腺癌中血管形成因子和细胞粘附分子的表达研究

目的　探讨CD3 4 和血管内皮生长因子 (VEGF)、细胞间粘附分子 (ICAM 1)在胰腺癌(pancreaticadenocarcinoma ,PAC)组织、慢性胰腺炎组织 (chronicpancreatitis ,CP)中的表达及病理意义 ,明确血管形成在胰腺癌演化过程中所起的重要作用。方法　应用免疫组化Envission方法对 2 4例胰腺癌组织、2 4例慢性胰腺炎组织、7例正常胰腺组织 ,进行了CD3 4 、VEGF、ICAM 1的表达情况的检测。对CD3 4 阳性血管进行MVD计数 ,并结合胰腺癌的病理特征进行分析。结果　癌组织 (4 7 2±9 4 )和慢性胰腺炎组织 (4 0 8± 7 93)中微血管密度明显高于对照组 (9 85± 2 86 ) (P <0 0 1)。VEGF在正常胰腺组织中不表达 ,在胰腺癌组织中VEGF主要表达于肿瘤细胞和导管细胞的胞质中 ,在慢性胰腺炎组织中VEGF在叶间及叶内导管细胞均高表达。细胞间粘附分子ICAM 1在正常对照胰腺组织中未见表达 ,仅见于某些内皮细胞着色 ,在胰腺癌组织及慢性胰腺炎组织中 ,除内皮细胞高表达外 ,肿瘤细胞阳性表达亦较高 ,慢性胰腺炎中ICAM 1亦在叶间及小叶内导管细胞中高表达。结论　我们发现微血管形成及其调控因子、细胞粘附分子与慢性胰腺炎、胰腺癌的生物学行为密切相关 ,在胰腺癌的演变中亦起了不可低估的作用 ,抗血管形成治疗无论是对胰腺癌抑     

股骨头缺血性坏死骨质含量与VEGF、bFGF、BMP-2 mRNA表达的相关性研究

目的坏死骨的血管再生及骨修复能力与VEGF、bFGF与BMP-2多种生长因子关系密切,通过观察股骨颈骨折、创伤性及非创伤性股骨头缺血性坏死(avascular necrosis of femoral head,ANFH)的股骨头局部上述因子的表达变化,并与其组织病理学的骨质含量指标进行相关性分析,为进一步探讨ANFH的发病机制及临床针对不同病因进行个性化治疗提供实验依据。方法取59例人工全髋关节置换术患者自愿捐赠的股骨头标本进行观察。创伤性ANFH 22例(A组),Ficat分期:Ⅲ期13例,Ⅳ期9例。非创伤性ANFH 19例(B组),Ficat分期:Ⅲ期11例,Ⅳ期8例;其中激素性10例,酒精性7例,原因不明2例。新鲜股骨颈骨折18例(C组)。3组患者性别、年龄等一般资料比较,差异均无统计学意义(P>0.05)。采用双能X线骨密度仪测量股骨头负重区骨密度;大体观察组织病理学改变,HE染色光镜及扫描电镜下观察其病理变化,计算空骨陷窝百分比及骨小梁面积百分比,并采用原位杂交技术分别对其VEGF、bFGF、BMP-2 mRNA表达进行检测。结果 A、B组骨密度均低于C组,B组低于A组,组间差异均有统计学意义(P<0.05)。A、B组股骨头形态不规则,光镜下见坏死区骨小梁稀疏、不完整,有大量空骨陷窝;健存区A组有较多纤维组织增生,B组髓腔内脂肪细胞增生、肥大。扫描电镜示A、B组大多骨细胞脂肪变性、坏死,骨基质内见脂肪细胞增生。C组均呈正常股骨头结构。A、B组空骨陷窝百分比均高于C组,骨小梁面积百分比均低于C组,差异均有统计学意义(P<0.05);A、B组间仅空骨陷窝百分比差异有统计学意义(P<0.05)。A组与B组VEGF、BMP-2、bFGF mRNA阳性染色面积百分比及吸光度(A)值均明显低于C组(P<0.05);A组BMP-2、bFGF mRNA两指标均较B组高(P<0.05),但VEGF mRNA A、B组间差异无统计学意义(P>0.05)。上述各因子表达强度与骨密度、骨小梁面积百分比成正相关,而与空骨陷窝百分比成负相关。结论创伤性ANFH的股骨头修复能力强于非创伤性ANFH,创伤性与非创伤性ANFH股骨头局部VEGF、bFGF、BMP-2 mRNA表达均降低。     

不同剂量重组牛碱性成纤维细胞生长因子治疗创面的比较

目的　探索外用重组牛碱性成纤维细胞生长因子（bFGF）治疗外伤创面的最佳用药浓度、给药方法与剂量,以指导临床应用。方法　设等渗盐水加灭滴灵治疗为对照组,选择bFGF的不同用药浓度（75Au/cm     

联合检测大肠癌组织中血管内皮生长因子与门静脉血细胞角蛋白20 mRNA的临床意义

目的探讨大肠癌病人肿瘤组织血管内皮生长因子（VEGF）蛋白和门静脉血细胞角蛋白20（cytokeratin,CK20）mRNA作为血行播散标志物的临床意义。方法RT-PCR技术检测63例大肠癌病人门静脉血CK20mRNA表达阳性率,免疫组织化学方法检测肿瘤标本VEGF蛋白的表达。结果随着病情由Dukes A、B期向Dukes C、D期进展,门静脉血CK20 mRNA表达的阳性率由64．9％升高到96．2％（P〈0．01）。高、中分化肿瘤组织VEGF蛋白阳性率为50．0％,低、未分化时升高到92．3％（P〈0．01）。CK20 mRNA与VEGF作为单项指标测定时,阳性率分别为77．8％和58．7％,联合检测时阳性率显著升高（92．1％）。结论大肠癌病人门静脉血中CK20 mRNA和肿瘤组织VEGF作为衡量大肠癌浸润能力的指标,二者表达分别与Dukes分期和肿瘤恶性程度的关系密切。联合检测二者时,敏感性显著提高,可弥补单项指标检测的不足,值得在临床上推广。     

血管内皮细胞生长因子(VEGF)基因转染骨髓间充质干细胞的表达

目的检测兔骨髓间充质干细胞(BMSc)经血管内皮细胞生长因子(VEGF165)基因转染后外源基因的表达，为进一步利用经基因转染的BMSc构建血管化组织工程骨组织打下基础。方法构建VEGF真核细胞表达载体，利用脂质体介导转染兔BMSc，使用原位杂交、免疫组织化学的方法检测VEGF165在BMSc中的表达。结果成功构建VEGF真核细胞表达载体，原位杂交、免疫组化方法显示经基因转染的BMSc中有阳性棕黄色颗粒出现，而未转染组呈现阴性结果。结论采用基因转移技术可以将VEGF转染至．BMSc中，并有外源性基因和蛋白的表达。     

转基因的骨髓间充质干细胞治疗大鼠缺血皮瓣

目的探讨转染血管内皮细胞生长因子(VEGF)基因的大鼠骨髓间充质干细胞(MSCs)同种异体移植促进缺血皮瓣的血管新生,从而提高皮瓣存活率的可能性。方法体外分离、培养、鉴定SD大鼠MSCs,PcDNA3.1(-)/VEGF165质粒转染MSCs,免疫荧光方法检测MSCs体外表达VEGF的情况,CM-DiI标记MSCs。SD大鼠随机分3组:A组[PcDNA3.1(-)/VEGF165质粒转染的MSCs移植]、B组(单纯MSCs移植)、C组(DMEM-F12培养基)。每只大鼠背侧皮下按组分别注射细胞悬液和培养基,注射后ELISA法连续检测大鼠血浆VEGF浓度,注射后第4天掀起1个蒂在尾侧的9 cm×2 cm的随意皮瓣。在术后第14天分别观察皮瓣的存活率、激光多普勒血液监测仪监测血流灌注、CD34免疫组织化学检测皮瓣毛细血管密度、荧光显微镜检测MSCs在皮瓣内的分布和存活状况。结果转染VEGF165基因的MSCs体外和体内检测均高表达VEGF165蛋白。A、B、C三组的皮瓣存活率分别为(83.1±2.6)%、(66.4±6.1)%、(51.5±7.5)%(P< 0.05);A、B、C三组的毛细血管密度(条/mm2)分别为:89.2±6.1、57.1±4.7、28.7±2.8(P< 0.05);血流灌注比值A组高于B、C两组,B组高于C组(P<0.05);转染VEGF165基因的MSCs移植SD大鼠皮瓣后,MSCs存活并参与血管新生。结论转染VEGF基因的大鼠MSCs体外培养后异体移植可促进缺血皮瓣的血管新生,提高存活率。