Beta-nerve growth factor promotes neurogenesis and angiogenesis during the repair of bone defects

We previously showed that the repair of bone defects is regulated by neural and vascular signals. In the present study, we examined the effect of topically applied β-nerve growth factor（β-NGF） on neurogenesis and angiogenesis in critical-sized bone defects filled with collagen bone substitute. We created two symmetrical defects, 2.5 mm in diameter, on either side of the parietal bone of the skull, and filled them with bone substitute. Subcutaneously implanted osmotic pumps were used to infuse 10 μgβ-NGF in PBS（β-NGF ＋ PBS） into the right-hand side defect, and PBS into the left（control） defect, over the 7 days following surgery. Immunohistochemical staining and hematoxylin-eosin staining were carried out at 3, 7, 14, 21 and 28 days postoperatively. On day 7, expression of β III-tubulin was lower on the β-NGF ＋ PBS side than on the control side, and that of neurofilament 160 was greater. On day 14, β III-tubulin and protein gene product 9.5 were greater on the β-NGF ＋ PBS side than on the control side. Vascular endothelial growth factor expression was greater on the experimental side than the control side at 7 days, and vascular endothelial growth factor receptor 2 expression was elevated on days 14 and 21, but lower than control levels on day 28. However, no difference in the number of blood vessels was observed between sides. Our results indicate that topical application of β-NGF promoted neurogenesis, and may modulate angiogenesis by promoting nerve regeneration in collagen bone substitute-filled defects.

Vascular endothelial growth factor induced angiogenesis following focal cerebral ischemia/reperfusion injury in rabbits

BACKGROUND： Therapeutic angiogenesis has opened up new pathway for the treatment of ischemic cerebrovascular disease in recent years. The exploration of the effect of vascular endothelial growth factor （VEGF） on inducing angiogenesis following ischemia/reperfusion injury can provide better help for the long-term treatment of cerebrovascular disease in clinic. OBJECTIVE： To observe the effect of VEGF on inducing angiogenesis following focal cerebral ischemia /reperfusion injury in rabbits through the angiogenesis of microvessels reflected by the expression of the factors of vascular pseudohemophilia. DESIGN： A randomized controlled animal tria SETTNG： Department of Medical Imaging, Second Hospital of Hebei Medical University MATERIALS： Sixty-five healthy male New Zealand rabbits of clean degree, weighing （2.6±0.2） kg, aged 4.5-5 months, were used. The polyclonal antibody against vascular pseudohemophilia （Beijing Zhongshan Company）, recombinant VEGF165 （Peprotech Company, USA）, biotinylated second antibody and ABC compound （Wuhan Boster Company） were applied. METHODS： The experiments were carried out in the Laboratory of Neuromolecular Imaging and Neuropathy, Second Hospital of Hebei Medical University from May to August in 2005. （1） The rabbits were randomly divided into three groups： sham-operated group （n=15）, control group （n=25） and VEGF-treated group （n=-25）. In the control group and VEGF-treated group, models were established by middle cerebral artery occlusion （MCAO） induced focal cerebral ischemia/reperfusion. In the VEGF-treated group, VEGF165 （2.5 mg/L） was stereotactically injected into the surrounding regions of the infarcted sites immediately after the 2-hour ischemia/reperfusion; Saline of the same dosage was injected in the control group. But the rabbits in the sham-operated group were only drilled but not administrated. （2） The experimental indexes were observed on the 3^rd 7^th, 14^th, 28^th and 70^th days of the experiment respectively, 3 rabbits in the sham-operated group and 5 in the control group and VEGF-treated group were observed at each time point. The brain tissues in the surrounding regions of the infarcted sites were collected. The positive expressions of the factors of vascular pseudohemophilia in vascular endothelial cells were analyzed with immunohistochemical method. The microvessels in unit statistical field were counted with the imaging analytical software. MAIN OUTCOME MEASURES： The changes of microvascular density in the brain tissue and the positive expressions of the factors of vascular pseudohemophilia in the surrounding regions of the infarcted sites were observed on the 3^rd 7^th, 14^th, 28^th and 70^th days of the experiment. RESULTS： All the 65 New Zealand rabbits were involved in the analysis of results without deletion. Changes of the number of microvessels at different time points in each group： There were no obvious changes at different time points in the sham-operated group. The numbers of microvessels at 7 and 14 days were obviously more in the control group than in the sham-operated group [（6.0±1.1）, （9.0±0.9） microvessels; （3.0±1.1）, （3.0±1.1） microvessels; P〈 0.05-0.01], and those at 3, 7, 14 and 28 days were obviously more in the VEGF-treated group than in the control group [（8.3±2.0）, （13.4±1.4）, （15.5±2.3）, （6.8± 1.0） microvessels; （3.4±0.6）, （6.0±1.1）, （9.0±0.9）, （3.2±0.8） microvessels; P 〈 0.01]. （2） Positive expressions of the factors of vascular pseudohemophilia in the surrounding regions of infarcted sites： There were no obvious changes at different time points in the sham-operated group. In the control group, the changing law of the expressions was the same as that for the number of microvessels that the expression began to mildly increase at 7 days, reached the peak value at 14 days, and began to reduce at 28 days. In the VEGF-treated group, the expression was obviously increased at 3 days, also reached the peak value at 14 days, and reduced to the normal level at 70 days, but the expressions were obviously stronger than those in the control group at the same time points. CONCLUSION： Angiogenesis can be obviously induced in rabbits after the focal cerebral ischemia/reperfusion injury is treated with VEGF for 18 days.

RELATIONSHIP BETWEEN VASCULAR ENDOTHELIAL GROWTHFACTOR EXPRESSION AND ANGIOGENESIS AND CELLPROLIFERATION OF MALIGNANTGLIOMAS IN CHILDREN

Objective: To investigate the relation of vascular endothelial growth factor (VEGF) expression with angiogenesis and cell proliferation in malignant glioma in children. Methods: Immunohistochemical technique was used to detect the expression of VEGF, microvessel quantity (MVQ) and PCNA Labeling Index (PCNA LI) in 33 malignant gliomas in children Results: Positive staining for VEGF was obtained in 23 out of the 33 cases (69.7). The MVQ and PCNA LI in VEFG-positive tumors were significantly higher than those in VEGF-negative tumors (P<0.005). The expression of VEGF in tumor tissues was significantly correlative with MVQ and PCNA LI (r=0.52 and 0.37, respectively, P<0.001). Conclusion: VEGF can be synthesized in tumor cells of malignant glioma in children which might play a significant role in angiogenesis and cell proliferation in the tumor.

EXPRESSION DIFFERENCES OF VASCULAR GROWTH FACTORS IN HUMAN LUNG ADENOCARCINOMA CELL LINE A549 AND CISPLATIN-RESISTANT HUMAN LUNG ADENOCARCINOMA CELL LINE A_(549)^(DDP)

Objective:To elucidate the expression differences of vascular growth factors in human lung adenocarcinoma cell line A549 and cisplatin-resistant human lung adenocarcinoma cell line A549^DDP.Methods:RT-PCR and immunohistochemistry was used to detect the mRNA and protein expressions of vascular endothelial growth factor(VEGF)and basic fibroblast growth factor (bFGF) in A549 and A549^DDP.Results:VEGF and bFGF mRNA were expressed in A549 and in A549^DDP.VEGF and bFGF mRNA expression levels in A549^DDP were signifi-cant higher than those in A549(P<0.025).VEGF and bFGF protein expressions were all strong positive in A549 and A549^DDP.Conclusion:There are certain differences between VEGF and bFGF expressions in A549 and A549^DDP.Drug-resistance of lung cancer is associated with those above genes over-expressions.Over-expression of vascular growth factors are related to drug resistance of lung cancer.

Buyanghuanwu decoction promotes angiogenesis after cerebral ischemia/reperfusion injury：mechanisms of brain tissue repair

Buyanghuanwu decoction has been shown to protect against cerebral ischemia/reperfusion injury,but the underlying mechanisms remain unclear.In this study,rats were intragastrically given Buyanghuanwu decoction,15 m L/kg,for 3 days.A rat model of cerebral ischemia/reperfusion injury was established by middle cerebral artery occlusion.In rats administered Buyanghuanwu decoction,infarct volume was reduced,serum vascular endothelial growth factor and integrin αvβ3 levels were increased,and brain tissue vascular endothelial growth factor and CD34 expression levels were increased compared with untreated animals.These effects of Buyanghuanwu decoction were partially suppressed by an angiogenesis inhibitor（administered through the lateral ventricle for 7 consecutive days）.These data suggest that Buyanghuanwu decoction promotes angiogenesis,improves cerebral circulation,and enhances brain tissue repair after cerebral ischemia/reperfusion injury.

Huoxue Anxin Recipe(活血安心方)Promotes Myocardium Angiogenesis of Acute Myocardial Infarction Rats by Up-Regulating miR-210 and Vascular Endothelial Growth Factor

Objective： To investigate the microR NAs（miR NAs） expression profile of acute myocardial infarction（AMI） rats and the regulating effects of Huoxue Anxin Recipe（活血安心方, HAR） on angiogenesis-related miR NAs and genes. Methods： Forty-five Wistar rats were randomly assigned to 3 groups according to a random number table： sham, AMI, and AMI＋HAR groups（15 in each group）. AMI rats were established by ligation of the left descending coronary artery. HAR was intragastrically administered to rats of the AMI＋HAR group for successive 21 days since modeling, meanwhile the same volume of 0.9% normal saline was administered to rats of the sham and AMI groups. Doppler echocardiography was used for noninvasive cardiac function test. Hematoxylin and eosin staining was used to observe the histopathological change. miR NAs expression profile was detected by quantitative realtime polymerase chain reaction（qR T-PCR）. The mR NA and protein expressions of vascular endothelial growth factor（VEGF）, and a target gene of miR-210 was further detected by qR T-PCR and Western blot, respectively. The microvessels density of myocardium was evaluated by CD31 immunostaining. Results： Compared with the sham group, ejection fraction（EF） and fractional shortening（FS） values were decreased significantly in the AMI group（P〈0.01）, while the infarction area and the interstitial collagen deposition were increased obviously. As for the AMI＋HAR group, EF and FS values were increased significantly（P〈0.05 vs. AMI group）, and the infarction area was reduced and the interstitial collagen deposition were alleviated significantly. Total of 23 miR NAs in the AMI group expressed differently by at least 1.5 folds compared with those in the sham group; 5 miR NAs in the AMI＋HAR group expressed differently by at least 1.5 folds compared with those in the AMI group. Among them, miR-210 was low in the AMI group and high in the AMI＋HAR group. The relative mR NA and protein expressions of VEGF were decreased significantly in the AMI group（P〈0.05 vs. sham group）, and increased significantly in the AMI＋HAR group（P〈0.01 vs. AMI group）. CD31 expression area and optical intensity were decreased significantly in the AMI group（P〈0.05 vs. sham group）, and increased significantly in the AMI＋HAR group（P〈0.01 vs. AMI group）. Conclusions： HAR could reduce the infarction area, alleviate the interstitial fibrosis and improve the cardiac function of AMI rats. Those effects could be related to promoting myocardium angiogenesis of HAR by up-regulating miR-210 and VEGF.

Autophagy: a potential target for the treatment of intraocular neovascularization

The formation of neovascularization is a common pathological feature of many ocular vascular diseases, and is an important cause of vision loss in patients. Neovascularization can cause retinal hemorrhage, vitreous hemorrhage, and other serious complications, leading to loss of vision. The treatment of intraocular neovascularization is the focus of ophthalmology research. In recent years, some studies have found that autophagy is closely related to vascular endothelial growth factor and the formation of neovascularization. Autophagy is expected to become a new target for the treatment of intraocular neovascularization. Therefore, this article reviews the research on autophagy and the formation of intraocular neovascularization.

Early Exercise Promotes Angiogenic Response in Mice Model of Myocardial Infarction

Objectives Little is known about the mechanism of exercise-induced angiogenic response in ischemic myocardium. This study was designed to investigate the effects of exercise training on expression of vascular endothelial growth factor and angiogenesis in infarcted heart. Methods Fifty male FVB mice were divided into three subgroups to test various responses to exercise, including time- dependent response of angiogenic factors to exercise training in intact heart (n=10) and infarcted heart (n= 10), as well as exercise-induced angiogenic response in heart with myocardial infarction (MI) (n=30). The mice in the exercise-training groups were allowed to exercise daily at 1 hour per day for 7 days. Results VEGF protein expression was up-regulated by exercise training in time dependent fashion in mice with MI. Angiogenesis was evident by increased myocardial mi- crovessels observed by PECAM-1 immunohistoc-hemi- cal staining in post-MI exercise group (16.5±3.4)/0.4 mm2 versus post-MI sedentary mice (10±2.1)/0.4 mm2 (P < 0.05). Cell proliferation assessment showed significantly higher (P < 0.05 ) number of BrdU positive cells in post MI mice in exercise group as opposed to sedentary post MI mice. 2%TTC staining disclosed a profound difference in the size of MI (18.25±2.93)% in exercise group vs sedentary group (29.26±7.64)% (P< 0.05). Conclusions Activation and up-regulation of VEGF in infarcted mice heart may contributes the angiogenic response to exercise training at the early stage of myocardial infarction. This underscores the impact of exercise on angiogenesis in post myocardial infarction setting.

Effect of calycosin on left ventricular ejection fraction and angiogenesis in rat models with myocardial infarction

OBJECTIVE:To evaluated the effect of calycosin on left ventricular ejection fraction and angiogenesis.METHODS:Adult male Sprague-Dawley rats were randomly assigned into calycosin-treated groups(0.5,1,2,and 4 mg/kg qd),a dimethyl sulfoxide(DMSO),or a sham-operated control group.The myocardial ischaemia(Ml) model was intraperitoneally administered calycosin for 28 days.The survival rates and left ventricular ejection fractions(LVEF)were compared between groups.The expression levels of vascular endothelial growth factor(VEGF)and cluster of differentiation 31(CD31) in ischaemic myocardium were also measured and compared.RESULTS:The construction of MI model resulted in a LVEF reduction of 50% compared with the sham-control.After 28 days,the LVEF value was 10% higher when calycosin(4 mg/kg) was administered compared with the DMSO group.The expression of VEGF and CD31 showed a dose-dependent manner when calycosin was administrated.The calycosin-treated(4 mg/kg) group displayed a twofold increase in VEGF expression at both the mRNA and protein levels compared with the DMSO group.In addition,CD31 expression in the microvascular increased 1.5-fold in the 4 mg/kg calycosin-treated group.CONCLUSION:Calycosin improved left ventricular ejection fraction in the MI rat models,induced VEGF expression in the ischaemic myocardium,increased CD31 expression and promoted angiogenesis.