Platelet-activating factor mediates acid-induced lung injury in genetically engineered mice

Adult respiratory distress syndrome (ARDS) is an acute lung injury of high mortality rate, and the molecular mechanisms underlying it are poorly understood. Acid aspiration-induced lung injury is one of the most common causes of ARDS, characterized by an increase in lung permeability, enhanced polymorphonuclear neutrophil (PMN) sequestration, and respiratory failure. Here, we investigated the role of platelet-activating factor (PAF) and the PAF receptor (PAFR) gene in a murine model of acid aspiration-induced lung injury. Overexpression of the PAFR gene in transgenic mice enhanced lung injury, pulmonary edema, and deterioration of gas exchange caused by HCl aspiration. Conversely, mice carrying a targeted disruption of the PAFR gene experienced significantly less acid-induced injury, edema, and respiratory failure. Nevertheless, the efficiency of PMN sequestration in response to acid aspiration was unaffected by differences in PAFR expression level. The current observations suggest that PAF is involved in the pathogenesis of acute lung injury caused by acid aspiration. Thus, inhibition of this pathway might provide a novel therapeutic approach to acute lung injury, for which no specific pharmaceutical agents are currently available.     

Soluble vascular endothelial growth factor receptor-1 protects mice in sepsis

OBJECTIVE: To determine the putative role in the modulation of inflammation of a soluble form of Flt-1 (sFlt), a potent vascular endothelial growth factor antagonist, in experimental endotoxemia and sepsis. DESIGN: Randomized prospective experimental study. SETTING: University medical laboratory. SUBJECTS: Male C56BL/6 strain mice. INTERVENTIONS: We investigated the expression patterns and the effects of vascular endothelial growth factor and soluble Flt-1 in experimental endotoxic shock and sepsis. The possible anti-inflammatory mechanism of soluble Flt-1 was also evaluated. MEASUREMENTS AND MAIN RESULTS: Both vascular endothelial growth factor and sFlt-1 were rapidly released from macrophages activated in vitro by lipopolysaccharide and in the plasma of endotoxemic mice. Administration of vascular endothelial growth factor enhanced proinflammatory cytokine production and mediated a dramatic increase in mortality in endotoxemic mice. Treatment with sFlt-1 attenuated inflammatory responses, inhibited recruitment of inflammatory cells into the peritoneal cavity, and improved survival in a lethal endotoxemia and cecal ligation and puncture-induced sepsis model, even when administered as late as 24 hrs after the onset of sepsis. CONCLUSIONS: These findings support a critical protective role of sFlt-1 in endotoxic shock and sepsis. sFlt-1 may therefore have utility as an adjunctive agent for the treatment of sepsis syndrome.     

Placental growth factor mediates mesenchymal cell development, cartilage turnover, and bone remodeling during fracture repair

Current therapies for delayed- or nonunion bone fractures are still largely ineffective. Previous studies indicated that the VEGF homolog placental growth factor (PlGF) has a more significant role in disease than in health. Therefore we investigated the role of PlGF in a model of semi-stabilized bone fracture healing. Fracture repair in mice lacking PlGF was impaired and characterized by a massive accumulation of cartilage in the callus, reminiscent of delayed- or nonunion fractures. PlGF was required for the early recruitment of inflammatory cells and the vascularization of the fracture wound. Interestingly, however, PlGF also played a role in the subsequent stages of the repair process. Indeed in vivo and in vitro findings indicated that PlGF induced the proliferation and osteogenic differentiation of mesenchymal progenitors and stimulated cartilage turnover by particular MMPs. Later in the process, PlGF was required for the remodeling of the newly formed bone by stimulating osteoclast differentiation. As PlGF expression was increased throughout the process of bone repair and all the important cell types involved expressed its receptor VEGFR-1, the present data suggest that PlGF is required for mediating and coordinating the key aspects of fracture repair. Therefore PlGF may potentially offer therapeutic advantages for fracture repair.     

Comparison of vascular endothelial growth factor and basic fibroblast growth factor on angiogenesis in SCID mice.

Therapeutic angiogenesis is a promising approach to treat patients with cardiovascular disease, and will likely be critical to engineering large tissues. Many growth factors have been found to play significant roles in angiogenesis, and vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are the most extensively investigated angiogenic factors to date. However, the appropriate dose to obtain a desired response and the effectiveness of each factor, relative to the other, in promoting angiogenesis at a specific site in the body remains unclear. We have used alginate hydrogels as localized delivery vehicles for VEGF and bFGF, and compared the ability of these factors to promote new blood vessel formation in the subcutaneous tissue of severe combined immunodeficient (SCID) mice. We have found that the thickness of a granulation tissue layer formed around the gel and the number of blood vessels in the layer increased with the dose of VEGF in the gel, but the density of new blood vessels remained relatively constant. Sustained and localized delivery of bFGF from the gels, while similarly leading to an increase in the density of blood vessels in the granulation tissue, did not lead to as high of a blood vessel density as VEGF. The results of this study support previous studies demonstrating the utility of both VEGF and bFGF in promoting angiogenesis, and suggest VEGF is more appropriate for creating a dense bed of new blood vessels in this model.     

The critical role of vascular endothelial growth factor in pulmonary vascular remodeling after lung injury

The pulmonary vascular endothelial cell plays a crucial role in the regulation of the pulmonary vascular tone and in the maintenance of the barrier function and integrity of the alveolar-capillary membrane. It also plays a major role in coagulation, fibrinolysis, and angiogenesis and participates in inflammatory reactions. Vascular endothelial growth factor (VEGF) is a central growth and survival factor for the endothelial cell. Particularly high levels of VEGF are expressed in the lungs, reflecting the critical role of VEGF for lung development and structural integrity of the adult lung. Vascular endothelial growth factor exerts a variety of physiological and pathophysiological actions in the lung. Recent evidence suggests its involvement in the pathogenesis of lung diseases such as bronchopulmonary dysplasia, acute lung injury, emphysema, and pulmonary hypertension. To summarize the critical effects of VEGF on the pulmonary endothelial cell in the pathogenesis of these diseases, the purposes of this review are to (1) discuss the biological activities and intracellular signaling pathways of VEGF in the lung; (2) summarize the regulatory mechanisms involved in VEGF expression; (3)address the effects of VEGF on endothelial cells in hyperoxia-induced and other forms of lung injury; (4) highlight the endothelial effects of VEGF in the pathogenesis of emphysema; and (5) explore the role of VEGF in the pathogenesis of pulmonary arterial hypertension.     

Increased oxidant activity mediates vascular dysfunction in vibration injury

Occupational exposure to hand-operated vibrating tools causes a spectrum of pathological changes in the vascular, neurological, and musculoskeletal systems described as the hand-arm vibration syndrome (HAVS). Experiments were performed to determine the effects of acute vibration on the function of digital arteries. Rats paws were exposed to a vibrating platform (4 h, 125 Hz, constant acceleration of 49 m/s(2) root mean squared), and digital artery function was assessed subsequently in vitro using a pressure myograph system. Constriction to phenylephrine or 5-hydroxytryptamine was reduced in digital arteries from vibrated paws. However, after endothelium denudation, constriction to the agonists was no longer impaired in vibrated arteries. Inhibition of nitric-oxide synthase (NOS) with N(omega)-nitro-l-arginine methyl ester (l-NAME) increased constriction to phenylephrine or 5-hydroxytryptamine in vibrated but not control arteries and abolished the vibration-induced depression in constrictor responses. However, nitric oxide (NO) activity, determined using the NO-sensitive probe 4-amino-5-methylamino-2', 7'-difluorofluorescein, was reduced in vibrated compared with control arteries. Endogenous levels of reactive oxygen species (ROS), determined using the ROS-sensitive probe 5-(and 6)-chloromethyl-2',7'-dichlorodihydro-fluorescein, were increased in vibrated compared with control arteries. The increased ROS levels were abolished by L-NAME or by catalase, which degrades extracellular hydrogen peroxide. Catalase also increased constriction to phenylephrine or 5-hydroxytryptamine in vibrated but not control arteries and abolished the vibration-induced depression in constrictor responses. The results suggest that acute vibration causes vascular dysfunction in digital arteries by increasing ROS levels, which is probably mediated by uncoupling of endothelial NOS. Therefore, therapeutic strategies to inhibit ROS or augment NO activity may be beneficial in HAVS.     

Tumor necrosis factor mediates hypertriglyceridemia during thermal injury in mice genetically susceptible to lipopolysaccharides.

A rise in plasma triglycerides has been noted after thermal injury in a number of animal species including humans. In this study we identified a factor, tumor necrosis factor, which was responsible for increased plasma triglycerides during thermal injury that was induced by scalding. Two strains of mice that differed genetically in susceptibility to lipopolysaccharides were used. These were CH3HEB/HeJ (LPS-) and CH3HEB/FeJ (LPS+). A 15% total body surface area was burned; this resulted in an increase of plasma triglycerides of 126% of preburn levels in the LPS+ strain 24 hours after burn injury. No change in triglycerides was noted in the LPS- mice at any time after burn injury. Sera from LPS+ mice at 1 to 2 hours after burn injury was injected into nonburned animals of the same strain; this caused a 62% +/- 5% increase in plasma triglycerides 24 hours after injection. When thermally injured LPS+ mice were injected with anti-tumor necrosis factor-alpha at 1 hour after injury, they did not show a rise in plasma triglycerides at any time between 24 to 72 hours after injury. Hepatic secretion of triglycerides was also measured 1 day after burn; the average secretion of triglycerides was significantly reduced (2.69 +/- 0.36 mg/kg/hr, compared with 3.83 +/- 0.15 mg/kg/hr for the control). We conclude that tumor necrosis factor, a cytokine that inhibits lipoprotein lipase, causes hypertriglyceridemia during thermal injury in spite of a decreased secretion of triglycerides. This is the first report that demonstrates that hypertriglyceridemia that is secondary to thermal injury is induced by tumor necrosis factor.     

高脂血症小鼠动脉粥样硬化形成与血管内皮生长因子的关系

背景高脂血症的长期存在可促进动脉粥样硬化( atherosclerosis,AS)的形成和发展,在此过程中生长因子所起的作用越来越受重视,但对血管内皮生长因子 ( vascular endothelial growth factor, VEGF)的作用还不十分清楚.目的探讨 VEGF在 AS形成过程中的作用. 设计随机对照的实验研究. 地点、材料和干预本实验在沈阳体育学院研究生实验室完成.实验动物为 C57BL/6J(易感 AS型)纯系小黑鼠 20只.实验动物被随机分两组对照组 10只,脂肪乳组 10只.对照组喂以常备标准饲料;脂肪乳组除正常喂养外,每日以脂肪乳灌喂方式按 10 mL/kg进行高脂膳食. 主要观察指标高脂血症与非高脂血症小鼠主动脉组织光镜下形态学改变、主动脉壁的 VEGF蛋白表达、血脂水平比较. 结果对照组主动脉壁 VEGF蛋白表达均为阴性,仅外膜残存脂质 VEGF阳性.脂肪乳组血栓中、局部增生的新内膜均有 VEGF蛋白表达.血脂测定结果脂肪乳组的总胆固醇、三酰甘油、低密度脂蛋白胆固醇( low density lipoprotein cholesterol,HDL-C)分别为( 4.97± 0.10),( 2.24± 0.26),( 2.93± 0.17) mmol/L较对照组 [分别为( 4.17± 0.29),( 1.22± 0.10 ),( 2.44± 0.33) mmol/L]明显升高 (t=2.364, 2.335, 2.939, P< 0.01);脂肪乳组 HDL-C[(1.29± 0.24) mmol/L]较对照组 [(1.50± 0.16) mmol/L]明显降低 (t=2.231,P< 0.05) 结论高脂血症状态下,内源性 VEGF参与血栓形成及内膜增生,促进 AS发生发展.     

闭合性颅脑损伤血浆血管内皮细胞生长因子的变化及意义

目的 为探讨闭合性颅脑损伤后血浆血管内皮细胞生长因子(VEGF)的变化规律和脑水肿、继发性脑损伤酌关系。方法 31例闭合性颅脑损伤患者伤后当天、2、3、7d分别取静脉血2ml，离心取上清液，ELISA法测定VEGF，同时记录血小板(PLT)、GCS评分，多次头颅CT检查。对照组30例健康成人用同样方法测定血浆VEGF，并记录PLT。结果 颅脑损伤组与正常对照组年龄、性别、各时点PLT比较差异无显著性意义。颅脑损伤后血浆VEGF逐步升高，3d时最高，至7d有所回降，但与对照组比较差异无显著性意义。VEGF与继发性脑损伤显著相关(r=0．198，P=0．031)，3d时VEGF与脑水肿发生与否显著相关(r=-0．566，P=0．016)。结论 颅脑损伤后，血浆VEGF的表达水平与脑水肿、继发性脑水肿的发生显著相关，利用抗VEGF抗体来调节VEGF在脑损伤后的作用可能是治疗脑水肿，减少继发性脑损伤的有效途径。     

血管内皮生长因子在大鼠急性肺损伤中表达的变化

目的:探讨油酸诱导大鼠急性肺损伤(ALI)中血管内皮生长因子(VEGF)表达的变化及黄芪对VEGF和ALI的影响。方法:将18只Wistar大鼠随机分成3组:正常组、油酸组、黄芪组。ELISA方法检测肺组织及血浆VEGF的表达,观察肺组织病理形态学变化。结果:VEGF在ALI大鼠肺组织的表达较正常组和黄芪组显著降低(P<0.05)、而血浆中VEGF的表达显著增加(P<0.05),肺组织与血浆VEGF的表达有显著负相关(r=-0.49,P<0.05)。结论:VEGF在油酸诱导大鼠ALI形成过程中起重要作用,黄芪对ALI具有一定的治疗作用。     

合并脑损伤时骨折愈合过程中血管内皮细胞生长因子的表达

背景:大量临床病例显示,骨折合并脑损伤患者骨痂量明显增多,骨折愈合速度明显加快.但对合并脑损伤时骨折愈合过程中血管内皮细胞生长因子的表达及作用机制缺乏前瞻性对照研究,对其潜在的机制尚未阐明.目的:对比观察SD大鼠在骨折合并脑损伤以及单纯骨折时骨折愈合过程中骨痂内血管内皮细胞生长因子的表达变化.方法:将SD大鼠以数字表法随机分为3组:正常对照组、单纯骨折组、骨折合并脑损伤组.脑损伤模型采用改进的Marmarou自由落体装置撞击大鼠颅骨制作弥漫性中度脑损伤模型;于左侧胫骨髁间处钻孔,插入无菌克氏钢针,在胫骨中上1/3处横向折断胫骨,制作骨折模型.术后X射线摄片观察骨折修复过程和效果;RT-PCR检及免疫组化染色观察骨痂标本血管内皮细胞生长因子表达.结果与结论:骨折后3,7,14d,单纯骨折组、骨折合并脑损伤组X射线表现无明显差别.与单纯骨折组相比,骨折后24d脑损伤合并骨折组骨折线变得模糊,形成较多骨痂量;42d后骨折线消失,伤肢基本愈合.单纯骨折组骨痂中血管内皮细胞生长因子骨折后7d开始出现,表达逐渐增强,约3周时达高峰,42d已经不见血管内皮细胞生长因子表达;脑损伤合并骨折后3d即可见血管内皮细胞生长因子表达,两三周达高峰期,其表达高峰提前且持续时间较长.说明脑损伤后大鼠骨折愈合加速,骨痂量增多,表明脑损伤对骨折愈合有促进作用,这可能与脑损伤后大鼠体内生长因子血管内皮细胞生长因子表达高峰提前且持续时间较长有关.     

Keratinocyte growth factor gene transduction ameliorates acute lung injury and mortality in mice

At present there is no known effective pharmacological therapy for acute lung injury (ALI). Because keratinocyte growth factor (KGF) promotes epithelial cell growth, intratracheal administration of KGF has the possibility of restoring lung tissue integrity in injured lungs and improving patient outcomes. However, treatment using recombinant KGF protein is limited by its short effective duration. Thus, we investigated the effectiveness of intratracheal KGF gene transduction using adenoviral vector in ALI. We constructed an adenoviral vector expressing mouse KGF (mKGF), and 1.0 x 10(9 ) plaque-forming units of mKGF cDNA-expressing (Ad-KGF) and control (Ad-1w1) adenoviral vector was intratracheally instilled, using a MicroSprayer, into anesthetized BALB/c mice. Three days later, the mice were exposed to >90% oxygen for 72 hr, and the effect of KGF on hyperoxia-induced lung injury was examined. In the Ad-KGF group, KGF was strongly expressed in the airway epithelial cells, while peribronchiolar and alveolar inflammation caused by adenoviral vector instillation was minimal. The KGF overexpression not only induced proliferation of surfactant protein C-positive cuboidal cells, especially in the terminal bronchiolar and alveolar walls, but also prevented lung injury including intraalveolar exudation/hemorrhage, albumin permeability increase, and pulmonary edema. The arterial oxygen tension and the survival rate were significantly higher in the KGF-transfected group. These findings suggest that KGF gene transduction into the airway epithelium is a promising potential treatment for ALI.     

转化生长因子β1在血管损伤后再狭窄形成中的作用

转化生长因子β1是一种高度多效、多功能性的生长与分化因子,它广泛地调节机体的生长、发育、炎症、修复和免疫等许多生理和病理过程。当血管损伤后,局部产生的细胞因子引起炎症反应是血管再狭窄的主要原因。其中,转化生长因子β1在局部水平上调刺激平滑肌细胞增殖和迁移、促进动脉外膜细胞的表型改变和迁移以及局部细胞外基质蛋白的产生和沉积,从而促进血管再狭窄。本文从转化生长因子β1的生物学特性、血管损伤后再狭窄的病理学、转化生长因子β1与血管损伤后再狭窄相关性及转化生长因子β1对血管损伤后再狭窄的作用机制等方面进行综述,说明转化生长因子β1在血管损伤后再狭窄病理过程中所起的作用。     

血管内皮生长因子及血管内皮生长因子受体-2在肾脏疾病中的研究新进展

血管内皮生长因子(VEGF)是一种内皮细胞的特殊生长因子,可以促进内皮细胞的增生、分化和生存,还可以提高血管的通透性,促进血管的形成。血管内皮生长因子受体-2(VEGFR-2),是VEGF的特异性受体,VEGF/VEGFR-2信号系统的激活是刺激血管内皮细胞的主要机制。本文首先对VEGF及VEGFR进行概述,其次介绍VEGF/VEGFR-2肾脏的旁分泌和自分泌机制。最后,对VEGF/VEGFR-2在各种肾脏疾病中的表达及其作用以及靶向VEGF/VEGFR-2信号通路对新药的研发和临床应用进行综述。     

Activation of vascular endothelial growth factor through reactive oxygen species mediates 20-hydroxyeicosatetraenoic acid-induced endothelial cell proliferation

20-Hydroxyeicosatetraenoic acid (20-HETE) is formed by the omega-hydroxylation of arachidonic acid by cytochrome P450 4A and 4F enzymes, and it induces angiogenic responses in vivo. To test the hypothesis that 20-HETE increases endothelial cell (EC) proliferation via vascular endothelial growth factor (VEGF), we studied the effects of WIT003 [20-hydroxyeicosa-5(Z),14(Z)-dienoic acid], a 20-HETE analog on human macrovascular or microvascular EC. WIT003, as well as pure 20-HETE, stimulated EC proliferation by approximately 40%. These proliferative effects were accompanied by increased VEGF expression and release that were observed as early as 4 h after 20-HETE agonist addition. This was accompanied by increased phosphorylation of the VEGF receptor 2. The proliferative effects of 20-HETE were markedly inhibited by a VEGF-neutralizing antibody. Polyethylene glycol-superoxide dismutase (PEG-SOD) markedly inhibited both the increases in VEGF expression and the proliferative effects of 20-HETE. In contrast, administration of the NAD(P)H oxidase inhibitor apocynin had no effect to the proliferative response to 20-HETE. The 20-HETE agonist markedly increased superoxide formation as reflected by an increase in dihydroethidium staining of EC, and this increase was inhibited by PEG-SOD but not by apocynin. 20-HETE also increased the phosphorylation of p42/p44 mitogen-activated protein kinase (MAPK) in EC, whereas an inhibitor of MAPK [U0126, 1,4-diamino-2,3-dicyano-1,4-bis(2-aminophenylthio)butadiene] suppressed the proliferative and the VEGF changes but not the pro-oxidant effects of 20-HETE. These data suggest that 20-HETE stimulates superoxide formation by pathways other than apocynin-sensitive NAD(P)H oxidase, thereby activating MAPK and then enhancing VEGF synthesis that drives EC proliferation. Thus, 20-HETE may be involved in the regulation of EC functions, such as angiogenesis.     

Role of vascular endothelial growth factor receptor 1 and vascular endothelial growth factor receptor 2 in the vasodilator response to vascular endothelial growth factor in the neonatal piglet lung

OBJECTIVE: Vascular endothelial growth factor (VEGF) regulates vascular proliferation and causes vasodilation. In the pulmonary circulation, the vasorelaxing effect of VEGF has been attributed to nitric oxide, whereas in other vascular beds, prostacyclin and other mechanisms are also involved. This vascular effect follows binding to two receptors, VEGF receptor 1 (VEGFR1) and VEGF receptor 2 (VEGFR2), the latter of which is thought to be the main receptor responsible for the vasorelaxing effect of VEGF. The role of VEGFR1 in the neonatal pulmonary vasculature remains to be determined. DESIGN: Prospective randomized laboratory investigation. SETTING: Animal laboratory. SUBJECTS: Newborn Yorkshire-Landrace piglets. INTERVENTIONS: To determine the mechanisms of action of VEGF in the neonatal pulmonary vasculature, the effect of VEGF (10-10 M) was tested in isolated perfused piglet lungs, alone and in the presence of a VEGFR2 kinase inhibitor, N-nitro-l-arginine (L-NNA), indomethacin (Indo), L-NNA + Indo, and GF109203X, a protein kinase C inhibitor. The effect of a VEGFR1 agonist, placenta growth factor (PlGF), was also studied with or without L-NNA. Perfusate was collected, and cyclic guanosine monophosphate (cGMP), as well as 6-keto prostaglandin F1alpha and thromboxane B2, the stable metabolites of prostacyclin and thromboxane, respectively, was measured. MEASUREMENTS AND MAIN RESULTS: VEGF caused vasorelaxation with a concomitant increase in cGMP. PlGF also decreased vascular tone and increased cGMP. VEGFR2 kinase inhibitor did not prevent the reduction in perfusion pressure seen with VEGF but blocked the increase in cGMP. Pretreatment with L-NNA completely inhibited VEGF and PlGF vasodilation and prevented the increase in cGMP seen with both agonists. Pretreatment with Indo or GF109203X did not reduce the dilator response to VEGF. CONCLUSIONS: VEGF vasodilation may follow nitric oxide release in the piglet pulmonary circulation. VEGF vasorelaxation may not only occur through binding to VEGFR2, since PlGF, the specific VEGFR1 agonist, also causes vasodilation. Therefore, vasodilator response to VEGF may involve both types of receptor in the neonatal piglet pulmonary vasculature.     

Nerve growth factor mediates hyperalgesia and cachexia in auto-immune arthritis

Pain and cachexia are two of the most debilitating aspects of rheumatoid arthritis. Despite that, the mechanisms by which they are mediated are not well understood. We provide evidence that nerve growth factor (NGF), a secreted regulatory protein that controls neuronal survival during development, is a key mediator of pain and weight loss in auto-immune arthritis. Function blocking antibodies to NGF completely reverse established pain in rats with fully developed arthritis despite continuing joint destruction and inflammation. Likewise, these antibodies reverse weight loss while not having any effect on levels of the pro-cachectic agent tumor necrosis factor (TNF). Taken together, these findings argue that pathological joint pain and joint destruction are mechanistically independent processes and that NGF regulates an alternative cachexia pathway that is independent or downstream of TNF.     

神经生长因子和血管内皮生长因子联合应用对兔局灶性脑缺血再灌注损伤神经元的保护时间窗

目的：观察神经生长因子和血管内皮生长因子对兔局灶性脑缺血再灌注损伤的神经元的保护作用，以及发挥作用的有效时间窗。 方法：实验于2005-05／08在河北医科大学第二医院神经分子影像医学和神经病学实验室进行。34只雄性4．5～5月龄新西兰白兔随机数字表法分为假手术组（n=6）、生理盐水组（n=8），再灌注3h因子治疗组（n=10）和再灌注6h因子治疗组（n=10）。采用兔大脑中动脉阻断局灶性脑缺血再灌注模型，假手术组线栓插入的深度不同。因子治疗组在缺血2h再灌注损伤后3h和6h应用微量进样器将2．5μg／L血管内皮生长因子16550μL和神经生长因子400AU（相当于16μg／L）立体定向导入梗死灶周，生理盐水组则注入同等剂量的生理盐水，于再灌注72h，应用MR影像学、红四氮唑染色和流式细胞术评价各组动物脑梗死体积、灶周缺血半暗带细胞凋亡率、表观弥散系数值比率及半胱氨酸蛋白酶3活性表达。在大脑中动脉阻塞2h再灌注后24，72h，采用Purdy评分标准行神经功能缺损评分。总得分最低为2分，表示无神经功能缺陷；总得分最高为11分，表示动物意识丧失或死亡。 结果：在实验过程中，无动物死亡，均进入结果分析。①缺血2h再灌注72h，MR影像测得梗死灶主要限于左侧大脑中动脉供血区的皮质和皮质下白质和部分尾壳核。再灌注3h因子治疗组、再灌注6h因子治疗组脑梗死百分率分别较生理盐水组下降44．0％和33．3％。②缺血2h再灌注72h的再灌注3h因子治疗组、再灌注6h因子治疗组神经功能缺损评分分别较生理盐水组明显减少，差异有显著性意义[（6．4&;#177;0．5），（4．8&;#177;0．8），（5.4&;#177;0.5），P〈0.01）；[（2．8&;#177;0．4），（3．2&;#177;0．8），（4．6&;#177;0．5），P〈0．01]。③再灌注3h因子治疗组、再灌注6h因子治疗组脑组织含水量与生理盐水组相比明显减少，差异均有显著性意义[（79．2&;#177;0．5）％，（79．9&;#177;0．6）％，（81．8&;#177;0．3）％，0．01]。④缺血2h再灌注72h的再灌注3h因子治疗组、再灌注6h因子治疗组灶周皮质区梗死灶周区表观弥散系数值与生理盐水组相比明显升高，差异有显著性意义[（89&;#177;3）％，（83&;#177;3）％，（74&;#177;4）％，P〈0．01]。这两组灶周皮质凋亡率及半胱氨酸蛋白酶3活性表达与生理盐水组相比则明显降低，差异均有显著性[（10．4&;#177;0．7）％（15．5&;#177;1．2）％（20．2&;#177;1．3）％，P〈0．01]；[（17．4&;#177;1．3），（26．1&;#177;1．0），（54．1&;#177;6．9），P〈0．01]。结论：联合神经生长因子和血管内皮生长因子治疗脑缺血再灌注损伤具有明显的神经元护作用，有效时间窗最少是再灌注后6h。     

ROCK1 mediates leukocyte recruitment and neointima formation following vascular injury

Although Rho-associated kinase (ROCK) activity has been implicated in cardiovascular diseases, the tissue- and isoform-specific roles of ROCKs in the vascular response to injury are not known. To address the role of ROCKs in this process, we generated haploinsufficient Rock1 (Rock1(+/-)) and Rock2 (Rock2(+/-)) mice and performed carotid artery ligations. Following this intervention, we found reduced neointima formation in Rock1(+/-) mice compared with that of WT or Rock2(+/-) mice. This correlated with decreased vascular smooth muscle cell proliferation and survival, decreased levels proinflammatory adhesion molecule expression, and reduced leukocyte infiltration. In addition, thioglycollate-induced peritoneal leukocyte recruitment and accumulation were substantially reduced in Rock1(+/-) mice compared with those of WT and Rock2(+/-) mice. To determine the role of leukocyte-derived ROCK1 in neointima formation, we performed reciprocal bone marrow transplantation (BMT) in WT and Rock1(+/-) mice. Rock1(+/-) to WT BMT led to reduced neointima formation and leukocyte infiltration following carotid ligation compared with those of WT to WT BMT. In contrast, WT to Rock1(+/-) BMT resulted in increased neointima formation. These findings indicate that ROCK1 in BM-derived cells mediates neointima formation following vascular injury and suggest that ROCK1 may represent a promising therapeutic target in vascular inflammatory diseases.     

神经生长因子和血管内皮生长因子联合应用对兔局灶性脑缺血再灌注损伤神经元的保护时间窗

目的:观察神经生长因子和血管内皮生长因子对兔局灶性脑缺血再灌注损伤的神经元的保护作用,以及发挥作用的有效时间窗。方法:实验于2005-05/08在河北医科大学第二医院神经分子影像医学和神经病学实验室进行。34只雄性4.5～5月龄新西兰白兔随机数字表法分为假手术组(n=6)、生理盐水组(n=8),再灌注3h因子治疗组(n=10)和再灌注6h因子治疗组(n=10)。采用兔大脑中动脉阻断局灶性脑缺血再灌注模型,假手术组线栓插入的深度不同。因子治疗组在缺血2h再灌注损伤后3h和6h应用微量进样器将2.5μg/L血管内皮生长因子16550μL和神经生长因子400AU(相当于16μg/L)立体定向导入梗死灶周,生理盐水组则注入同等剂量的生理盐水,于再灌注72h,应用MR影像学、红四氮唑染色和流式细胞术评价各组动物脑梗死体积、灶周缺血半暗带细胞凋亡率、表观弥散系数值比率及半胱氨酸蛋白酶3活性表达。在大脑中动脉阻塞2h再灌注后24,72h,采用Purdy评分标准行神经功能缺损评分。总得分最低为2分,表示无神经功能缺陷;总得分最高为11分,表示动物意识丧失或死亡。结果:在实验过程中,无动物死亡,均进入结果分析。①缺血2h再灌注72h,MR影像测得梗死灶主要限于左侧大脑中动脉供血区的皮质和皮质下白质和部分尾壳核。再灌注3h因子治疗组、再灌注6h因子治疗组脑梗死百分率分别较生理盐水组下降44.0%和33.3%。②缺血2h再灌注72h的再灌注3h因子治疗组、再灌注6h因子治疗组神经功能缺损评分分别较生理盐水组明显减少,差异有显著性意义犤(6.4±0.5),(4.8±0.8),(5.4±0.5),P<0.01);犤(2.8±0.4),(3.2±0.8),(4.6±0.5),P<0.01犦。③再灌注3h因子治疗组、再灌注6h因子治疗组脑组织含水量与生理盐水组相比明显减少,差异均有显著性意义犤(79.2±0.5)%,(79.9±0.6)%,(81.8±0.3)%,0.01犦。④缺血2h再灌注72h的再灌注3h因子治疗组、再灌注6h因子治疗组灶周皮质区梗死灶周区表观弥散系数值与生理盐水组相比明显升高,差异有显著性意义犤(89±3)%,(83±3)%,(74±4)%,P<0.01犦。这两组灶周皮质凋亡率及半胱氨酸蛋白酶3活性表达与生理盐水组相比则明显降低,差异均有显著性犤(10.4±0.7)%(15.5±1.2)%(20.2±1.3)%,P<0.01犦;犤(17.4±1.3),(26.1±1.0),(54.1±6.9),P<0.01犦。结论:联合神经生长因子和血管内皮生长因子治疗脑缺血再灌注损伤具有明显的神经元护作用,有效时间窗最少是再灌注后6h。