IMPACTS ON THE EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR IN ISCHEMIC BRAIN TISSUE TREATED WITH CLUSTER PUNC-TURE ON SCALP ACUPOINTS IN RATS

Objective To probe into the impacts on the expression of vascular endothelial growth factor （VEGF） in ischemic brain tissue treated with cluster puncture on scalp acupoints in rats, Methods 128 rats were randomized into pseudo-operation group, model group, scalp-needling group and cluster-needling group, In scalp-needling group, penetration method was used on the focal side from Bǎihuì （百会 GV20） to Qǔbīn （曲鬓 GB7）, and in cluster-needling group, penetration method was used on both sides from GV20 to GB7, and a common needling on GV 20. Needles were punctured 2 mm in depth, constantly rotated for 10min, retained for 2 h. Immunohistrochemical method was applied to determine VEGF expression and microvessel density （MVD）. Results With the intervention of cluster puncture on scalp acupoints, VEGF： expressions on every time-spot after ischemia were enhanced apparently, superior to those in scalp-needling group. On the three time-spots of the 7^th, 14^th and 21^st days, MVD was increased after cluster puncture on scale acupoints, superior to those in scalp-needling group. Conclusion Cluster puncture on scalp acupoints up-regulated VEGF expression and promoted regeneration of microvessel.

Expression of p-STAT3 and vascular endothelial growth factor in MNNG-induced precancerous lesions and gastric tumors in rats

AIM: To investigate the dynamic expression of p-signal transducer and activator of transcription 3 (STAT3) and vascular endothelial growth factor (VEGF) in the formation of gastric tumors induced by drinking water containing N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) in Wistar rats. METHODS: One hundred and twenty Wistar rats were randomly divided into two groups (60 in each group): Control group and Model group. The rats in each group were then randomly divided into three groups (20 in each group): C/M15, C/M25 and C/M40 (15, 25 and 40 represent the number of feeding weeks from termination). Rats in the control group received normal drinking water and rats in the model group received drinking water containing 100 μg/mL MNNG. Stomach tissues were collected at the end of the 15th, 25th and 40th week, respectively, for microscopic measurement using hematoxylin and eosin staining. The expression of p-STAT3 and VEGF in different pathological types of gastric tissue, including normal, inflammation, atrophy, hyperplasia and gastric stromal tumor, was observed by immunohistochemistry and Western blot, and the corelation between p-STAT3 and VEGF was analyzed. RESULTS: (1) The expression of p-STAT3 in tissue with gastritis, atrophy, dysplasia and gastric stromal tumor were significantly increased in the model group compared with the control group (2.5 ± 1.0, 2.75 ± 0.36, 6.2 ± 0.45, 5.67 ± 0.55 vs 0.75 ± 0.36, P = 0.026, 0.035, 0.001, 0.002, respectively); the expression of p-STAT3 in tissue with dysplasia was higher than that in samples with gastritis or atrophy (6.2 ± 0.45 vs 2.5 ± 1.0, P = 0.006; 6.2 ± 0.45 vs 2.75 ± 0.36, P = 0.005, respectively); however, the expression of p-STAT3 in gastritis and atrophy was not significantly different (P > 0.05); (2) the expression of VEGF in tissue with gastritis, atrophy, dysplasia and gastric stromal tumor was significantly increased in the model group compared with normal gastric mucosa; and the expression of VEGF in tissue with dysplasia was higher than that in tissue with inflammation and atrophy (10.8 ± 1.96 vs 7.62 ± 0.25, P = 0.029; 10.8 ± 1.96 vs 6.26 ± 0.76, P = 0.033, respectively); similarly, the expression of VEGF in tissue with gastritis and atrophy was not significantly different (P > 0.05); and (3) the expression of VEGF was positively correlated with p-STAT3. CONCLUSION: p-STAT3 plays an important role in gastric cancer formation by regulating the expression of VEGF to promote the progression of gastric tumor from gastritis.

Effects of Sirt1 on proliferation,migration,and apoptosis of endothelial progenitor cells in peripheral blood of SD rats with chronic obstructive pulmonary disease

Objective:To explore the effect of Sirt1 on the function of endothelial progenitor cells(EPCs)in rats with chronic obstructive pulmonary disease(COPD).Methods:A rat COPD model was established via smoking and endotoxin administration for three months.The peripheral circulating EPCs were isolated by gradient centrifugation,and their functions,cell cycle distribution,apoptosis,and Sirt1 expression were examined.The function changes of EPCs in the presence or absence of Sirt1 agonist and inhibitor were estimated;meanwhile,the expressions of Sirt1,FOXO3a,NF-κB,and p53 were also evaluated.Results:The proliferation,adhesion,and migration of EPCs decreased while the apoptosis rate was increased in the COPD rats.The expression of Sirt1 protein in EPCs of the COPD group was significantly lower than that in the control group(P<0.01).The overexpression of the Sirt1 gene using a gene transfection technique or Sirt1 agonists(SRT1720)improved the proliferation,migration,and adhesion,and decreased the apoptosis of EPC.However,Sirt1 inhibitor(EX527)decreased EPC functions in the COPD group.The effect of Sirt1 expression on EPC function may be related to reduction of FOXO3a and increase of NF-κB and p53 activity.Conclusions:Increased expression of Sirt1 can improve the proliferation and migration of EPCs and reduce their apoptosis in COPD rats.This change may be related to FOXO3a,NF-κB,and p53 signaling pathways.

The expression of vascular endothelial growth factor and Factor VIII related anti-gen in the border zone after intracer ebral hemorrhage in rats

AIM:To observe the expression of vascula r endothelial growth factor(VEGF)and Factor VIII related antigen(FVIII-R Ag )in rats brain tissue in the bor-der zone of the hematoma,and explore the relationship between the expres-sion of VEGF and the brain angionesis after intracerebral hmorrhage(ICH).METHODS:Rat of model of ICH was induced by inje ction of collagenase physiological saline into the right caudate nucleus.The immunohistoch emical methods were performed by using diff erent antisera of VEGF and FVIII-R Ag in serial sections of rats brain at 12h,1d,3d and 7d after ICH respec-tively.RESULTS:T he amount of VEGF positive cells was markedly in-creased,and reached the highest at 7th day in the border zone after ICG(P<0.01),but there were no significant diffe rence among the mean value of A of cells at different time points(P>0.05).Compared with the controls,both the FVIII-R Ag positive endothelial cells and the A of cells were signific antly different in experimental ICH goups after 1day.CONCLUSION:The up-regulated expression of VEGF might i nduce proliferation of endothelial cell and angiogenesis in the border zone a fter ICH in rats.

Dan Huang Ming Mu Recipe Suppresses the Progression of Streptozotocin-induced Diabetic Retinopathy After Retinal Laser Photocoagulation in Brown Norway Rats via Down-regulating Vascular Endothelial Growth Factor and Up-regulating Pigment Epithelium-derived

Objective To analyze the effects of Dan Huang Ming Mu Recipe(DHMMR)(a pharmaceutical preparation from herbs and having the function of replenishing vital essence,removing heat,promoting blood circulation and excreting pathogenic water) on diabetic retinopathy(DR) after retinal laser photocoagulation through regulating the expression of vascular endothelial growth factor(VEGF) and pigment epithelium-derived factor(PEDF).Methods Forty male Brown Norway(BN) rats were randomly divided into blank group(group A,10 BN rats) and model group(30 BN rats).DR models were induced by 40 mg/kg streptozotocin(STZ) and the body weight and blood glucose of rats were monitored.After 12-week injection,fluorescein fundus angiography(FFA) and histopathological examination were detected to confirm the successful establishment of DR models.Subsequently,the right eyes of model group rats were conducted retinal laser photocoagulation with the left eyes having no retinal laser photocoagulation and rats of the model group were randomly divided into group B(the model control group),group C(the positive control group),and group D(the DHMMR group),with 10 rats in each group.Rats of the group A and the group B were given vehicle,and the group C were given calcium dobesilate suspension by gavage,and the group D were given DHMMR by gavage.After 4-week gavage,FFA was carried out to observe the fundus microvascular change.Then,the rats were sacrificed to do histopathological examinations and the serum levels of P-selectin,cAMP,cGMP,and the relative expression of the protein of VEGF and PEDF in retina were detected.Results After 12-week STZ injection,the blood glucose of the model group were pronouncedly higher than the blank group(P < 0.01).Fundus micro-hemangioma changes were observed in the rats of the model group through FFA,and the rats of the blank group did not see any changes.The pictures of HE staining showed that the retinal structure of the model group was more disordered than the blank group.After 4-week gavage,treatments with DHMMR showed dramatic reduction of serum levels of Pselectin,cAMP and cGMP compared with the group B(P < 0.01).FFA and histopathological examinations of DHMMR-treated rats revealed significantly suppression of retinal edema,fundus microvascular destruction and retinal destruction distinguishing from the group B.And the relative expression of VEGF protein of the group D and the group A was markedly lower than that of the group B(P < 0.01).The relative expression of PEDF protein of the group D and the group A was dramatically higher than that of the group B to the contrary(P < 0.01).Conclusions DHMMR demonstrates pronounced suppressive effects on the progression of DR after retinal laser photocoagulation,through down-regulating VEGF and upregulating PEDF.

Phosphorothioate oligonucleotide inhibits tissue factor expression in endothelial cells induced by blood flow shear stress in rats

Objective： To determine the effect of antiparallel phosphorothioate triplex-forming oligonucleotide （apsTFO）, which was designed according to shear stress response element （SSRE） in tissue factor （TF） gene promoter region, on the expression of endothelial TF in carotid artery stenosis rats. Methods： Rat model of severe carotid artery stenosis were inflicted by silica gel tube ligation. Half an hour before the model infliction, GT20-apsTFO, GT20-psTFO and GT21-apsTFO labeled with green fluorescence （FITC） were injected into the vena caudalis of rat at a dose of 0.5 mg/kg. Half an hour, 4 or 9 h after the ligation, the distribution of TFO in the common carotid artery, the liver and the kidney was detected with aid of fluorescence microscopy. And the mRNA and protein expressions of TF, Egr-1 and Spl in the above-mentioned organs were determined with in situ hybridization and immunohistochemical assay respectively in 6 h after the model establishment, and the results were analyzed with an image analysis system. Results： Only in 1 h after TFO injection, fluorescent granules appeared in the liver, the kidney and the vascular wall and lumen of carotid artery, and then in 4.5 h, they still deposited in above sites except the vascular lumen. GT20-apsTFO and GT21-apsTFO significant down-regulated the mRNA and protein expressions of TF compared to the rats without treatment （P〈0.05）, and the former apsTFO had a more stronger effect than the later （P〈0.05）. GT20-psTFO had no such effect （P〉0.05）. The 3 TFOs had no inhibition on the mRNA and protein expressions of Egr-1 and Spl. Conclusion： Pretreated apsTFO can partly come into the vascular endothelial cells, and inhibit TF expression induced by shear stress, but had no effect on Egr-1 and Spl gene expressions.

The activities of endothelial elastase and cathepsin G in rats at oxidative stress caused by heavy metals salts

CoCl2 introduction increased cathepsin G activity in the heart and liver as well as endothelial elastase (EEl) in kidney that indicated the development of destructive processes. CoCl2 introduction decreased EEl and cathepsin G activities in blood serum and cathepsin G in lungs. HgCl2 injection decreased EEl in blood serum, heart, liver, kidney and cathepsin G in blood serum. These decreasing of proteinases activities may be caused by cytotoxic effects of heavy metals and/or the inclusion of these proteases in the destructive processes and absence of their synthesis and/or release.

Effect of Buyang Huanwu Decoction on the proliferation,migration and adhesion of endothelial progenitor cells in rats with acute myocardial infarction

Objective:To investigate the intervention effect of Buyang Huanwu Decoction on endothelial progenitor cell function(EPCs),and to explore the therapeutic mechanism of Buyang Huanwu Decoction.Methods:This research take 54 rats were divided into blank group,the control group,model group,Chinese medicine,western medicine group and combine traditional Chinese and western medicine group,tonifying Yang also five decoction and atorvastatin calcium for acute myocardial infarction(AMI)rats group intervention,by using the method of density gradient centrifugation separation training each rat endothelial progenitor cells,using tetramethyl azo salt trace enzyme reaction colorimetry,used in the determination of adhesion ability and improved Boyden chamber,the method of analysis and comparison between groups of endothelial progenitor cells proliferation,adhesion and migration.Results:In 7 days,14 days and 4 weeks,acute myocardial infarction model of rat peripheral blood EPCs count,proliferation,migration and adhesion ability were compared with control group were significantly decline,tonifying Yang also five decoction group,western medicine control group and combine traditional Chinese and western medicine group of peripheral blood EPCs count,proliferation,migration and adhesion ability compared with model group were significantly increased(P<0.05),and combine traditional Chinese and western medicine group is the traditional Chinese medicine and western medicine group on the improvement of the function of EPCs effect more significantly(P<0.05).Conclusion:Statins combined with Buyang Huanwu Decoction can better improve the endothelial function of AMI rats than traditional Chinese medicine or western medicine.Further clinical studies on statins may better improve the endothelial function of AMI patients,and provide a new research entry point for improving the long-term prognosis of AMI patients.

5-aza-2'-deoxycytidine in the regulation of antioxidant enzymes in retinal endothelial cells and rat diabetic retina

AIM: To investigate the roles of a DNA methyltransferase(DNMT) inhibitor 5-aza-2'-deoxycytidine(5-aza-dC) in the regulation of antioxidant enzymes in diabetic retinopathy(DR) models. METHODS: DNMTs expressions and activity, and changes of two key antioxidant enzymes in DR, MnSOD(encoded by SOD2 gene) and glutathione S-transferase theta 1(GSTT1), were quantified in the isolated human retinal endothelial cells(HRECs) exposed to high glucose(HG) with or without 5-aza-dC treatment. The downstream exacerbating factors including vascular endothelial growth factor(VEGF), intercellular adhesion molecule 1(ICAM-1) and matrix metalloproteinase 2(MMP2), which are implicated in the pathogenesis of DR and closely related to oxidative stress were also analyzed. The key parameters were confirmed in the retina from streptozotocin(STZ) diabetic rats. RESULTS: DNMTs expression and DNMT activity was induced in HRECs exposed to HG. Hyperglycemia decreased MnSOD and GSTT1 expression. 5-aza-dC administration effectively suppressed DNMTs expression and activity and reversed the MnSOD and GSTT1 expression under HG condition. VEGF, ICAM-1 and MMP2 induced by HG were also suppressed by 5-aza-dC treatment. Similar results were observed in the retina from STZ diabetic rats. CONCLUSION: Our findings suggest that DNA methylation may serves as one of the mechanisms of antioxidant defense system disruption in DR progression. Modulation of DNA methylation using pharmaceutic means such as DNMT inhibitors could help maintain redox homeostasis and prevent further progression of DR.

Trimethylamine N-oxide aggravates vascular permeability and endothelial cell dysfunction under diabetic condition:in vitro and in vivo study

AIM:To provide the direct evidence for the crucial role of trimethylamine N-oxide(TMAO)in vascular permeability and endothelial cell dysfunction under diabetic condition.METHODS:The role of TMAO on the in vitro biological effect of human retinal microvascular endothelial cells(HRMEC)under high glucose conditions was tested by a cell counting kit,wound healing,a transwell and a tube formation assay.The inflammation-related gene expression affected by TMAO was tested by real-time polymerase chain reaction(RT-PCR).The expression of the cell junction was measured by Western blotting(WB)and immunofluorescence staining.In addition,two groups of rat models,diabetic and non-diabetic,were fed with normal or 0.1%TMAO for 16wk,and their plasma levels of TMAO,vascular endothelial growth factor(VEGF),interleukin(IL)-6 and tumor necrosis factor(TNF)-αwere tested.The vascular permeability of rat retinas was measured using FITC-Dextran,and the expression of zonula occludens(ZO)-1 and claudin-5 in rat retinas was detected by WB or immunofluorescence staining.RESULTS:TMAO administration significantly increased the cell proliferation,migration,and tube formation of primary HRMEC either in normal or high-glucose conditions.RT-PCR showed elevated inflammation-related gene expression of HRMEC under TMAO stimulation,while WB or immunofluorescence staining indicated decreased cell junction ZO-1 and occludin expression after high-glucose and TMAO treatment.Diabetic rats showed higher plasma levels of TMAO as well as retinal vascular leakage,which were even higher in TMAO-feeding diabetic rats.Furthermore,TMAO administration increased the rat plasma levels of VEGF,IL-6 and TNF-αwhile decreasing the retinal expression levels of ZO-1 and claudin-5.CONCLUSION:TMAO enhances the proliferation,migration,and tube formation of HRMEC,as well as destroys their vascular integrity and tight connection.It also regulates the expression of VEGF,IL-6,and TNF-α.

Xuebijing improves intestinal microcirculation dysfunction in septic rats by regulating the VEGF-A/PI3K/Akt signaling pathway

BACKGROUND:This study aims to explore whether Xuebijing(XBJ) can improve intestinal microcirculation dysfunction in sepsis and its mechanism.METHODS:A rat model of sepsis was established by cecal ligation and puncture(CLP).A total of 30 male SD rats were divided into four groups:sham group,CLP group,XBJ + axitinib group,and XBJ group.XBJ was intraperitoneally injected 2 h before CLP.Hemodynamic data(blood pressure and heart rate) were recorded.The intestinal microcirculation data of the rats were analyzed via microcirculation imaging.Enzyme-linked immunosorbent assay(ELISA) kits were used to detect the serum levels of interleukin-6(IL-6),C-reactive protein(CRP),and tumor necrosis factor-α(TNF-α) in the rats.Histological analysis and transmission electron microscopy were used to analyze the injury of small intestinal microvascular endothelial cells and small intestinal mucosa in rats.The expression of vascular endothelial growth factor A(VEGF-A),phosphoinositide 3-kinase(PI3K),phosphorylated PI3K(p-PI3K),protein kinase B(Akt),and phosphorylated Akt(p-Akt) in the small intestine was analyzed via Western blotting.RESULTS:XBJ improved intestinal microcirculation dysfunction in septic rats,alleviated the injury of small intestinal microvascular endothelial cells and small intestinal mucosa,and reduced the systemic inflammatory response.Moreover,XBJ upregulated the expression of VEGF-A,p-PI3K/total PI3K,and p-Akt/total Akt in the rat small intestine.CONCLUSION:XBJ may improve intestinal microcirculation dysfunction in septic rats possibly through the VEGF-A/PI3K/Akt signaling pathway.

Protective Effect of Naringenin on Acute Myocardial Ischemia-reperfusion Injury in Rats

[Objectives]To investigate the protective mechanism of naringenin on acute myocardial ischemia-reperfusion injury(AMI-RI)in Sprague-Dawley(SD)rats.[Methods]A total of 32 SD rats with AMI-RI model construction were randomly divided into AMI-RI model control group and citrus pigment A/B/C groups(n=8).The naringenin A,B,and C groups were administrated 20,40 and 80 mg/(kg•d)for 10 d.The AMI group served as the negative control and was not treated.At the conclusion of the treatment regimen,a sample of intraventricular blood was collected for the purpose of measuring lactate dehydrogenase(LDH),glutathione peroxidase(GLH-PX),nitric oxide(NO),and superoxide dismutase(SOD)levels.Additionally,myocardial tissue was identified within the ischemic region.The content of malondialdehyde(MDA)was determined by inducing nitric oxide synthase(iNOS)and endodermal nitric oxide synthase(eNOS)positive cells in the left anterior descending coronary artery.[Results]Following citrus treatment,the contents of GLH-PX and SOD in ventricular blood of the citrus B group were found to be significantly elevated,while the contents of NO and LDH in myocardial MDA and ventricle were observed to be significantly reduced.The number of eNOS-positive cells was significantly increased,while the number of iNOS-positive cells was significantly decreased.The difference was statistically significant when compared with the AMI-RI group(P<0.05).The changes observed in the above indicators in the citrus C group were more pronounced than those observed in the citrus B group.The difference between the citrus C and the B group was statistically significant(P<0.05),indicating that this effect is concentration dependent.[Conclusions]In addition to its ability to inhibit myocardial lipid peroxidation during AMI-RI by increasing SOD activity,naringenin may also affect the synthesis and release of NO by regulating eNOS and iNOS,thereby achieving protection against AMI-RI.One effect is enhanced as the dose of the drug increases.

Flavin-Containing Monooxygenase (FMO) Protein Expression and Its Activity in Rat Brain Microvascular Endothelial Cells

The aim of this study was to examine whether flavin-containing monooxygenase (FMO) protein was expressed in cultured rat brain microvascular endothelial cells (BMECs), which constitute the blood-brain barrier (BBB), and whether N-oxide from the tertiary amine, d-chlorpheniramine, was formed by FMO in rat BMECs. BMECs were isolated and cultured from the brains of three-week-old male Wistar rats. The expression of FMO1, FMO2 and FMO5 proteins was confirmed in rat BMECs by western blotting analysis using polyclonal anti-FMO antibodies, but FMO3 and FMO4 proteins were not found in the rat BBB. Moreover, N-oxide of d-chlorpheniramine was formed in rat BMECs. The intrinsic clearance value for N-oxidation at pH 8.4 was higher than that at pH 7.4. Inhibition of N-oxide formation by methimazole was found to be the best model of competitive inhibition yielding an apparent Ki value of 0.53 μmol/L, suggesting that N-oxidation was catalyzed by FMOs in rat BMECs. Although FMO activity in rat BMECs was lower than that in SD rat normal hepatocytes (rtNHeps), we suggest that rat BMECs enzymes can convert substrates of exogenous origin for detoxification, indicating that BMECs are an important barrier for metabolic products besides hepatic cells.

Rosiglitazone uppresses lipopolysaccharide-induced matrix metalloproteinase-2 activity in rat aortic endothelial cells via rasMEK1/2 signaling

ix metalloproteinase(MMPs) plays a key role in the pathogenesis of chronic inflammatory disease,such as atherosclerosis.Among MMPs,MMP-2 is regarded as a major proteinase in atherosclerotic plaque lesions.Peroxisome proliferator activated receptor-gamma(PPARg) ameliorates oxidative stress and the inflammatory response.The aim of the present study was to evaluate the effect of Rosiglitazone on Lipopolysaccharide(LPS)-induced MMP-2 activation as well as its possible mechanism.LPS-induced MMP-2 activity was inhibited by Rosiglitazone(PPARg agonist) in the rat aortic endothelial cells(RAEC).LPS-induced MMP-2 activation was diminished no matter exposure to NF-kB Activation Inhibitor II(JSH-23)or Ras inhibitor,farnesylthiosalicylic acid(FTS). Further study shows that LPS-induced activation of Phospho-Rho A and Phospho-MEKl/2 were significantly inhibited by Rosiglitazone.The activation of NF-kB p65 in the nuclear extract of cells was also significantly suppressed by Rosiglitazone, moreover,the expression of NF-κB p65 was partly activated by GW9662(PPARg antagonist).NF-kB DNA binding activity was also demolished by Rosiglitazone.In summary,our data showed that PPARg agonist,Rosiglitazone suppresses LPS-activated MMP-2 secretion via Ras-MEK1/2 signaling pathways and NF-kB activation.PPARg agonist and Ras-MEK1/2 pathway may be another potential therapeutic target for the disease induced by chronic inflammation.

Influence of hepatic arterial blockage on blood perfusion and VEGF,MMP-1 expression of implanted lver cancer in rats

AIM: To investigate the influence of hepatic arterial blockage on blood perfusion of transplanted cancer in rat liver and the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase-1 (MMP-1), and to explore the mechanisms involved in transarterial embolization (TAE)-induced metastasis of liver cancer preliminarily. METHODS: Walker 256 carcinosarcoma was transplanted into rat liver to establish the liver cancer model. Hepatic arterial ligation (HAL) was used to block the hepatic arterial blood supply and simulate TAE. Blood perfusion of tumor in control, laparotomy control, and HAL group was analyzed by Hoechst 33342 labeling assay, the serum VEGF level was assayed by ELISA, the expression of VEGF and MMP-1 mRNA was detected by in situ hybridization. RESULTS: Two days after HAL, the number of Hoechst 33342 labeled cells which represent the blood perfusion of tumor directly and hypoxia of tumor indirectly in HAL group decreased significantly compared with that in control group (329+/-29 vs 384+/-19, P【0.01). The serum VEGF level in the HAL group increased significantly as against that of the control group (93 ng.L(-1)+/-44 ng.L(-1) vs 55 ng.L(-1)+/-19 ng.L(-1), P【0.05). The expression of VEGF and MMP-1 mRNA in the tumor tissue of the HAL group increased significantly compared with that of the control and the laparotomy control groups (P【0.05). The blood perfusion data of the tumor, represented by the number of Hoechst 33342 labeled cells, showed a good linear inverse correlation with the serum VEGF level (r=-0.606, P【0.05) and the expression of VEGF mRNA in the tumor tissue ( r =-0.338, P【0.01). CONCLUSION: Blockage of hepatic arterial blood supply results in decreased blood perfusion and increased expression of metastasis-associated genes VEGF and MMP-1 of transplanted liver cancer in rats. Decreased blood perfusion and hypoxia may be the major cause of up-regulated expression of VEGF.

Inhibition of β-elemene on the expressions of HIF-lα, VEGF and i NOS in diabetic rats model

AIM: To evaluate the effect of β-elemene on the expressions of hypoxia-inducible factor(HIF)-lα, vascular endothelial growth factor(VEGF) and inducible nitric oxide synthase(i NOS) in a streptozotocin(STZ) induced diabetic SpragueDawley(SD) rat model.METHODS: SD rats were administered an abdominal injection of STZ and induced to a diabetic model. After 6 wk course of diabetes, the treatment groups were given β-elemene through periocular and intravitreous injection separately and the control groups were given blank emulsion injection. HE staining was used to observe the morphology of retina. The m RNA expressions of HIF-1α, VEGF and i NOS was assayed by real-time polymerase chain reaction(PCR) and the protein expression was measured by Western blot and immunocytochemistry methods.RESULTS: The results indicated that the protein and m RNA expressions of HIF-1α, VEGF and i NOS after treated by β-elemene periocularly and intravitreally injections were all found to be reduced compared with the levels in the diabetic rats group(P<0.05). The inhibitory effect of intravitreal injection was more remarkable.CONCLUSION: The results show β-elemene protect the retina of diabetic rats from high glucose damage by downregulating the expression of HIF-1α, VEGF and iNOS.

Time-dependent Expression of PEDF and VEGF in Blood Serum and Retina of Rats with Oxygen-induced Retinopathy

The effects of the balance changes of pigment epithelium growth factor（PEDF） and vascular endothelial growth factor（VEGF） in whole-body and retinal tissue on rats with oxygen-induced retinopathy were investigated. Forty-eight neonatal SD rats at the age of 7 days were randomly divided into 4 groups. The neonatal rats in experimental groups were exposed to 75% to 80% oxygen for 5 days and then to normal air, and those in control groups were kept feeding in normal air. At the age of 17 and 22 days, all the neonatal rats received retina angiography with FITC-dextran and the pathological changes of retinal vessels and perfusion were observed. HE staining of the tissue section and the number counting of endothelial cells extending beyond the inner limiting membrane were performed to evaluate the endothelial proliferation. Immunohistochemistry was applied to detect the expression of PEDF and VEGF in retinal tissue, and ELISA to detect their expression in serum. A hypoxic-ischemic proliferation of retina and more endothelial cells extending beyond the inner limiting membrane were found in the neonatal rats in both experimental groups of 17-day old and 22-day old as compared with those in control group with the difference being statistically significant（P〈0.01）. VEGF staining of the rats in the 17-day old experimental group was significantly stronger, with an increasing positive rate, than that of the rats in the 17-day old control group（P〈0.01）. PEDF staining of the rats of 22 days old was weaker than that of the rats of 17 days old in the experimental groups（P〈0.01）. There was no significant difference in serum VEGF concentration among all groups（P〈0.05）. The serum PEDF concentration in the rats of 17 days old in experimental group was decreased significantly as compared with that in the rats of 17 days old in control group（P〈0.01）, and in experimental groups, the serum PEDF concentration of the rats of 22 days old was increased as compared with that of the rats of 17 days old（P〈0.01）. In conclusion, the obviously decreased serum PEDF concentration and the abnormal enhanced expression of VEGF density in local retinal tissue broke down the balance of PEDF/VEGF in whole-body or local tissues, which might play an important role in retinal vascular proliferation.

Sesamin enhances nitric oxide bioactivity in aortas of spontaneously hypertensive rats

OBJECTIVE To explore the underlying mechanisms involved in the effect of sesamin on aortic NO bioactivity in spontaneously hypertensive rat(SHR).METHODS Sesamin was orally administered for consecutive 8 weeks in SHR.Systolic blood pressure(SBP)was measured using the tail-cuff method.The aortas were isolated and in vitro vascular reactivity studies were performed.Superoxide anion production in carotid arteries was assessed by dihydroethidium fluorescence staining.The protein expression of endothelial nitric oxide synthase(eNOS),phosphorylated eNOS(P-eNOS),dihydrofolate reductase(DHFR),nicotinamide adenine dinucleotide phosphate(NADPH)oxidase subunit p47 phox and copper,zinc-superoxide dismutase(Cu/Zn-SOD)in aortas was detected by Western blotting.The dimeric form of eNOS in aortas was determined by low-temperature SDS-PAGE.Aortic level of nitrotyrosine and activities of antioxidant enzymes,namely,total SOD(T-SOD),glutathione peroxidase(GPx)and catalase were also detected.RESULTS In SHR,sesamin treatment reduced SBP,improved vascular relaxation induced by acetylcholine and enhanced aortic NO bioactivity.Sesamin treatment enhanced NO biosynthesis in SHR aortas was due to upregulated P-eNOS and suppressed eNOS uncoupling,and the latter effect might be attributed to decreased nitrotyrosine and upregulated DHFR.Sesamin also reducd the NO oxidative inactivation and decreased the superoxide anion production through downregulation of p47 phox and amelioration of eNOS uncoupling.In addition,sesamin treatment did not alter the levels of GPx and catalase activity but obviously reduced the compensatory elevated T-SOD activity and Cu/Zn-SOD protein expression.CONCLUSION Chronic treatment with sesamin could reduce hypertension and improve endothelial dysfunction through enhancement of NO bioactivity in SHRs aortas.

Zinc-deficient diet aggravates ventilation-induced lung injury in rats

We investigated the effects of zinc deficiency on acute lung injury （ALI） induced by mechanical ventilation. Male Sprague-Dawley rats were fed with a zinc-deficient or zinc-proficient diet for 4 weeks, and then received mechanical ventilation at normal frequency and pressure for 30 min. Total protein, cell count, the number of poly- morphonuclear neutrophil （PMN） in the bronchoalveolar lavage （BAL）, and vascular endothelial growth factor （VEGF） expression in the lung were determined. Activation of nuclear factor-t^B （NF-~cB） was detected by exam- ining the phosphorylation of NF-kB （pNF-kB p65） and the expression of inhibitor of NF-kB （pI-kBa）. Compared to the controls, total cell count and the number of PMNs were significantly increased to 160% and 140%, respec- tively, in zinc-deficient rats treated with ventilation. Activation of NF-kB was significantly increased and VEGF was also increased to three-folds. Zinc deficiency aggravated the inflammatory response in rats and was associated with the overexpression of VEGF in response to mechanical ventilation. Zinc supplementation may be beneficial to zinc-deficient patients during mechanical ventilation.

Attenuated blood-brain barrier dysfunction by XQ-1H following ischemic stroke in hyperlipidemic rats

Following ischemic stroke, blood-brain barrier （BBB） is disrupted and is further aggravated with the corresponding incidence of hyperlipidemia. BBB breakdown promotes inflammation infiltration into the brain, which exacerbates cerebral ischemic injury as a result. Here, we report that 10-O-（N,N-dimethylaminoethyl）-ginkgolide B methanesulfonate （XQ-1H） , a novel analog of ginkgolide B, alleviates BBB breakdown in hyperlipidemic rats and protects endothelial cells against inflammatory response. Middle cerebral artery occlusion （MCAO） modeled is- chemic stroke in rats. Before surgery, these rats were fed a cholesterol-rich diet to induce an experimental hyperlip- idemic condition. Additionally, lipopolysaccharide （LPS） incubation with rat brain microvessel endothelial cells （rBMECs） was applied to mimic hyperlipidemia-induced inflammatory injury of BBB. The results indicated more severe infarct size, increased BBB permeability, excessive secretion of pro-inflammatory cytokines, and exaggerated inflammation infiltration of the brain in hyperlipidemic rats following MCAO when compared to rats fed with normal diet. XQ-1H protected BBB integrity, lessoned brain edema and inflammation penetration, down- regulated MMP- 9 and VCMA-1 expressions, and extenuated ischemic infarction. XQ-1H alleviated LPS-induced inflammatory re- sponse in rBMECs, characterized by promoting cell viability, inhibiting TNF-α, IL-1β, and IL-6 releasing, and downregulating NF-KB inflammatory signal and down- stream proteins, such as VCAM-1 and iNOS. In conclusion, the present study shows that XQ-1H stabilizes BBB function following ischemic stroke in hyperlipidemic rats, and the possible mechanisms may be related to inflammation inhibition.