Differences In Vascular Endothelial Growth Factor Research Articles

High frequency home-based exercise decreases levels of vascular endothelial growth factor in patients with stable angina pectoris

In coronary artery disease (CAD), circulating angiogenic factors have been seen to increase, possibly as a response to ischaemia. Regular physical activity (PA) is recommended for prevention and treatment of CAD, but more research is needed to optimise PA regimes. We investigated the effect of home-based high frequency exercise (HFE) on angiogenic cytokines and cardiac markers in patients with stable CAD. This was a randomised case-control study Sixty-two patients, with stable CAD, were randomised to HFE (n = 33), (aerobic exercise 70% of max, 30 min, five times/week and resistance exercise three times/week), performed at home, or usual lifestyle (control, n = 29). After eight weeks, percutaneous coronary intervention (PCI) was performed in both groups, and the HFE group continued another six months of exercise. Serum vascular endothelial growth factor (VEGF) and stromal derived factor-1 (SDF-1), plasma N-terminal-brain natriuretic peptide (NT-proBNP), high-sensitive troponin T (TnT) and copeptin were analysed. Data are presented as median (25(th), 75(th) percentile) of relative changes (%) from baseline. Values of p are given for the difference between the HFE and controls. HFE decreased circulating VEGF levels, before PCI (-5% (-15%, -2%)), while VEGF levels increased in the control group (5% (-3%, 20%) p = 0.004). A significant difference in VEGF remained at three months post-PCI (HFE (-1%(-12%, 5%), control (7% (0%, 14%), p = 0.04), but not at six months after PCI. SDF-1, NT-proBNP, TnT and copeptin levels did not differ significantly. In addition, VEGF levels were positively correlated to NT pro-BNP. Home-based HFE decreased circulating VEGF in patients with stable CAD, suggesting a reduced ischaemic burden. HFE does not increase markers of cardiac dysfunction, suggesting that it is a safe therapeutic intervention in these patients.

Relationship of serum levels of VEGF and TGF-β1 with radiosensitivity of elderly patients with unresectable non-small cell lung cancer

This study aimed to evaluate the relationship of serum levels of vascular endothelial growth factor (VEGF) and transforming growth factor-β1 (TGF-β1) with radiosensitivity of elderly patients with unresectable non-small cell lung cancer (NSCLC) receiving three-dimensional conformal radiation therapy (3D-CRT). Fifty-eight elderly patients with unresectable NSCLC and 40 healthy controls were enrolled in this study. Serum levels of VEGF and TGF-β1 were detected by the enzyme-linked immunosorbent assay (ELISA) method before and after 3D-CRT. Clinical performances of serum VEGF and TGF-β1 levels in predicting radiosensitivity of NSCLC patients with 3D-CRT were evaluated. Serum VEGF and TGF-β1 levels of NSCLC patients were higher than those of health controls (all p < 0.05). After 3D-CRT treatment, 41 patients achieved effective clinical response (complete response (CR) + partial response (PR)) and 17 patients were ineffective clinical response (stable disease (SD) + progressive disease (PD)). There was no significant difference in the VEGF and TGF-β1 levels between the effective and ineffective groups before 3D-CRT (all p > 0.05). Serum levels of VEGF and TGF-β1 after 3D-CRT in the effective group were lower compared with the levels before 3D-CRT treatment (p < 0.001 and 0.027, respectively). However, no significant differences in serum VEGF and TGF-β1 levels between before and after 3D-CRT in the ineffective group were observed (p = 0.196 and 0.517, respectively). We observed significant differences in serum VEGF and TGF-β1 levels between the effective and ineffective groups after 3D-CRT (p < 0.001 and 0.013, respectively). Sensitivity and specificity of VEGF combined with TGF-β1 in predicting radiosensitivity of NSCLC patients with 3D-CRT were 87.8 and 94.1%, respectively. In conclusion, our results indicate that serum VEGF and TGF-β1 levels may accurately predict radiosensitivity of elderly patients with unresectable NSCLC receiving 3D-CRT.

Angiogenic factors in relation to clinical effect in a phase II trial of weekly paclitaxel

BackgroundSeveral anticancer agents including paclitaxel have an inhibitory effect on angiogenesis. AimsTo compare the overall response rate and time to progression with changes in circulating angiogenic factors during palliative treatment with weekly paclitaxel. Material and methodsPatients with metastatic BC, ECOG 0–2, received weekly paclitaxel, concomitant with trastuzumab if HER2+ BC (n = 7). Circulating vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were determined at base-line and before start of new course. ResultsFifty-five of 63 included patients were evaluable. The overall response rate including stable disease ≥24 weeks (CR + PD + SD) was obtained in 25 of the evaluable patients (45%). The median time to progression (TTP) was 5.3 months and overall survival (OS) 16.7 months. Patients with triple negative breast cancer (TNBC) showed a trend towards higher base-line VEGF compared with hormone receptor positive or HER2+ tumours and had shorter TTP. Significant differences in VEGF and bFGF levels at 12 weeks were found between patients with longer versus shorter TTP (VEGF: p = 0.046, bFGF: p = 0.005) and between patients gaining versus lacking clinical benefit (VEGF: p = 0.05, bFGF: p = 0.02). ConclusionsThe clinical utility of circulating VEGF may be a useful tool for monitoring treatment efficacy.

Antioxidant properties of glutamine and its role in VEGF-Akt pathways in portal hypertension gastropathy.

To investigate the effects of glutamine on oxidative/nitrosative stress and the vascular endothelial growth factor (VEGF)-Akt-endothelial nitric oxide synthase (eNOS) signaling pathway in an experimental model of portal hypertension induced by partial portal vein ligation (PPVL). Portal hypertension was induced by PPVL. The PPVL model consists of a partial obstruction of the portal vein, performed using a 20 G blunt needle as a guide, which is gently removed after the procedure. PPVL model was performed for 14 d beginning treatment with glutamine on the seventh day. On the fifteenth day, the mesenteric vein pressure was checked and the stomach was removed to test immunoreactivity and oxidative stress markers. We evaluated the expression and the immunoreactivity of proteins involved in the VEGF-Akt-eNOS pathway by Western blotting and immunohistochemical analysis. Oxidative stress was measured by quantification of the cytosolic concentration of thiobarbituric acid reactive substances (TBARS) as well as the levels of total glutathione (GSH), superoxide dismutase (SOD) activity, nitric oxide (NO) production and nitrotyrosine immunoreactivity. All data are presented as the mean ± SE. The production of TBARS and NO was significantly increased in PPVL animals. A reduction of SOD activity was detected in PPVL + G group. In the immunohistochemical analyses of nitrotyrosine, Akt and eNOS, the PPVL group exhibited significant increases, whereas decreases were observed in the PPVL + G group, but no difference in VEGF was detected between these groups. Western blotting analysis detected increased expression of phosphatidylinositol-3-kinase (PI3K), P-Akt and eNOS in the PPVL group compared with the PPVL + G group, which was not observed for the expression of VEGF when comparing these groups. Glutamine administration markedly alleviated oxidative/nitrosative stress, normalized SOD activity, increased levels of total GSH and blocked NO overproduction as well as the formation of peroxynitrite. Glutamine treatment demonstrated to reduce oxidative damage but does not reduce angiogenesis induced by PH in gastric tissue, demonstrating a beneficial role for the PI3K-Akt-eNOS pathway.

ASSOCIATION TISSUE KALLIKREIN WITH THE SEVERITY OF CORONARY ARTERY DISEASE

ObjectivesThe objective of this study was to examine the presence of tissue kallikrein (TK) in coronary artery disease (CAD) and its association with inflammation, intraplaque angiogenesis and plaque stability.MethodsPlasma TK...

An Overview of Molecular and Cellular Mechanisms Associated with Porcine Pregnancy Success or Failure

Prenatal mortality remains one of the major constraints for the commercial pig industry in North America. Twenty to thirty per cent of the conceptuses are lost early in gestation and an additional 10-15% is lost by mid-to-late gestation. Research over the last two decades has provided critical insights into how uterine capacity, placental efficiency, genetics, environment, nutrition and immune mechanisms impact successful conceptus growth; however, the exact cause and effect relationship in the context of foetal loss has yet to be determined. Similar to other mammalian species such as the human, mouse, rat, and primates, immune cell enrichment occurs at the porcine maternal-foetal interface during the window of conceptus attachment. However, unlike other species, immune cells are solely recruited by conceptus-derived signals. As pigs have epitheliochorial placentae where maternal and foetal tissue layers are separate, it provides an ideal model to study immune cell interactions with foetal trophoblasts. Our research is focused on the immune-angiogenesis axis during porcine pregnancy. It is well established that immune cells are recruited to the maternal-foetal interface, but their pregnancy specific functions and how the local milieu affects angiogenesis and inflammation at the site of foetal arrest remain unknown. Through a better understanding of how immune cells modulate crosstalk between the conceptus and the mother, it might be possible to therapeutically target immune cells and/or their products to reduce foetal loss. In this review, we provide evidence from the literature and from our own work into the immunological factors associated with porcine foetal loss.

Levels and values of circulating endothelial progenitor cells, soluble angiogenic factors, and mononuclear cell apoptosis in liver cirrhosis patients

BackgroundThe roles of circulating endothelial progenitor cell (EPC) and mononuclear cell apoptosis (MCA) in liver cirrhosis (LC) patients are unknown. Moreover, vascular endothelial growth factor (VEGF) and stromal cell-derived factor (SDF)-1α are powerful endogenous substances enhancing EPC migration into circulation. We assessed the level and function of EPCs [CD31/CD34 (E1), KDR/CD34 (E2), CXCR4/CD34 (E3)], levels of MCA, VEGF and SDF-1α in circulation of LC patients.MethodsBlood sample was prospectively collected once for assessing EPC level and function, MCA, and plasma levels of VEGF and SDF-1α using flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively, in 78 LC patients and 25 age- and gender-matched healthy controls.ResultsNumber of EPCs (E1, E2, E3) was lower (all p < 0.0001), whereas SDF-1α level and MCA were higher (p < 0.001) in study patients compared with healthy controls. Number of EPCs (E2, E3) was higher but MCA was lower (all p < 0.05) in Child's class A compared with Child's class B and C patients, although no difference in VEGF and SDF-1α levels were noted among these patients. Chronic hepatitis B and esophageal varices bleeding were independently, whereas chronic hepatitis C, elevated aspartate aminotransferase (AST), and decompensated LC were inversely and independently correlated with circulating EPC level (all p < 0.03). Additionally, angiogenesis and transwell migratory ability of EPCs were reduced in LC patients than in controls (all p < 0.001).ConclusionThe results of this study demonstrated that level, angiogenic capacity, and function of circulating EPCs were significantly reduced, whereas plasma levels of SDF-1α and circulating MCA were substantially enhanced in cirrhotic patients.

Oral supplementation with sulodexide inhibits neo-angiogenesis in a rat model of peritoneal perfusion

Peritoneal dialysis (PD) is associated with functional and morphological alterations of the peritoneal membrane (PM). It is hypothesized that vascular endothelial growth factor (VEGF) plays a role in this process. Sulodexide is a glycosaminoglycan with effects on vascular biology. Therefore, the impact of oral sulodexide on PM function and morphology in a rat model of peritoneal perfusion was evaluated. Rats received 10 mL peritoneal dialysate fluid (PDF) twice daily via a tunnelled PD catheter. The test-PD group (Sul) received 15 mg/kg/day oral sulodexide versus none in the control-PD group (Con). A third group received no PDF (Sham). After 12 weeks, a peritoneal equilibration test was performed and the PM was sampled. Neo-angiogenesis was evaluated using immunostaining with von Willebrand, and epithelial-to-mesenchymal transition (EMT) using co-localization of cytokeratin and α-smooth muscle actin. VEGF was determined in the dialysate by enzyme-linked immunosorbent assay. PD induced loss of ultrafiltration, also in the sulodexide group. Creatinine and glucose transport were better preserved, and sodium dip was more pronounced in the sulodexide group versus control. Submesothelial thickness, neo-angiogenesis and EMT were more pronounced in the Con versus Sul versus Sham group. VEGF in the dialysate, corrected for diffusion was higher in Con and Sul versus Sham. Oral sulodexide administration diminishes neo-vascularization, submesothelial thickening and EMT induced by exposure to PDF in a rat model. As there was no difference in VEGF at the protein level in the dialysate, we hypothesize that oral sulodexide inhibits VEGF locally by binding.

Aging Disrupts The Balance Between Positive And Negative Angiogenic Factors In Skeletal Muscle

Skeletal muscle microvessel density has been found to decrease with age in both rats and humans, but the mechanisms for this process are still poorly understood. Physiologic regulation of angiogenesis is thought to be controlled by a balance between positive and negative proteins, among which vascular endothelial growth factor (VEGF) and thrombospondin‐1 (TSP‐1) are emerging as two important factors. We hypothesized that age‐related reduction in skeletal muscle capillarity would correlate with lower VEGF and/or higher TSP‐1 levels. We measured protein expression of VEGF and TSP‐1 in the hind limb muscles, plantaris (PLT), soleus (SOL), and gastrocnemius (GA), of 12 week‐ (young, N=5–6) and 20 week‐ old (aged, N=3–6) lean Zucker rats. In the GA, aged animals had a 570% increase in TSP‐1 (p=0.014) and no difference in VEGF. In the PLT, aged animals had a no significant difference in TSP‐1 and a 60% decrease in VEGF (p=0.049). In the SOL, aged animals had a 198% increase in TSP‐1 (p=0.003), and no difference in VEGF. These results suggest that an altered balance between VEGF and TSP‐1 may be responsible for capillary rarefaction in aged muscle. Research supported by NIH 5T32‐HL090610 and AHA 10BGIA3630002.

MMP-9 gene deletion mitigates microvascular loss in a model of ischemic acute kidney injury

Microvascular rarefaction following an episode of acute kidney injury (AKI) is associated with renal hypoxia and progression toward chronic kidney disease. The mechanisms contributing to microvascular rarefaction are not well-understood, although disruption in local angioregulatory substances is thought to contribute. Matrix metalloproteinase (MMP)-9 is an endopeptidase important in modifying the extracellular matrix (ECM) and remodeling the vasculature. We examined the role of MMP-9 gene deletion on microvascular rarefaction in a rodent model of ischemic AKI. MMP-9-null mice and background control (FVB/NJ) mice were subjected to bilateral renal artery clamping for 20 min followed by reperfusion for 14, 28, or 56 days. Serum creatinine level in MMP-9-null mice 24 h after injury [1.4 (SD 0.8) mg/dl] was not significantly different from FVB/NJ mice [1.5 (SD 0.6) mg/dl]. Four weeks after ischemic injury, FVB/NJ mice demonstrated a 30-40% loss of microvascular density compared with sham-operated (SO) mice. In contrast, microvascular density was not significantly different in the MMP-9-null mice at this time following injury compared with SO mice. FVB/NJ mice had a 50% decrease in tissue vascular endothelial growth factor (VEGF) 2 wk after ischemic insult compared with SO mice. A significant difference in VEGF was not observed in MMP-9-null mice compared with SO mice. There was no significant difference in the liberation of angioinhibitory fragments from the ECM between MMP-9-null mice and FVB/NJ mice following ischemic injury. In conclusion, MMP-9 deletion stabilizes microvascular density following ischemic AKI in part by preserving tissue VEGF levels.

Exercise Increases Gene Expression of Vascular Endothelial Growth Factor in Mice on a High-fat Diet

Obesity is associated with elevated white adipose tissue hypoxia. Hypoxia inducible factor-1 a (HIF-1a) can upregulate vascular endothelial growth factor (VEGF) to prevent cell death during hypoxia. However, the effects of exercise on markers of angiogenesis and hypoxia within visceral adipose tissue are unknown. PURPOSE: To determine if high- fat (HF) diet and/or exercise training (EX) will increase gene expression of angiogenic and hypoxia markers in juvenile mice. METHODS: C57BL/6 male mice (5 wks old) were randomized into 4 groups (n=10/group): normal chow (NC)/sedentary (SED), NC/EX, HF/SED, and HF/EX. The HF diet was 45% fat, 35% carbohydrate, and 20% protein. The mice were placed on the 6 wk diet and exercise intervention concomitantly. Treadmill exercise was performed 5d/wk at 12 m/min and12% grade for 40 min/d. HIF-1a and VEGF gene expression were assessed using quantitative real- time PCR. NC/SED group was the referent group. RESULTS: Body weights of the HF groups were greater than the NC groups (p<0.001). The epididymal fat pad weights were greater in the HF groups (p<0.001), with the HF/SED having the greatest fat pad weight (p<0.001); this effect remained after adjusting for body weight. No differences were observed between groups for gene expression of HIF-1a. However, VEGF mRNA was greatest in the HF/EX group vs. the NC groups (p<0.05). No differences in VEGF were observed between the HF groups, nor between HF/SED and the NC groups. CONCLUSION: Mice exposed to a HF diet while exercising exhibited an increase in gene expression of a known marker of angiogenesis, VEGF, without changes in HIF-1a mRNA. This suggests that other mechanisms may be involved in driving exercise-induced adipose tissue adaptations in obesity.Table

Expression of Vascular Endothelial Growth Factor and Hypoxia Inducible Factor‐1α in Cervical Neoplasia

Altered angiogenesis is an important phenotype of high-grade cervical lesions and invasive cervical carcinomas. Many researchers, including us, have shown that oncoproteins of human papillomavirus could enhance the vascular endothelial growth factor (VEGF) expression. We investigated the change in VEGF and hypoxia-inducible factor-1alpha (HIF-1alpha) expression patterns that occur during the carcinogenesis and progression of cervical cancer. Expressions of HIF-1alpha and VEGF were evaluated by immunohistochemistry using paraffin-embedded specimens obtained from 41 patients with a normal cervix, 39 patients with cervical intraepithelial neoplasia (CIN 1, 10; CIN 2/3, 29), and 36 patients with cervical cancer. The VEGF and HIF-1alpha expressions were higher in CIN and invasive cancer than in normal cervix (P= 0.021, P < 0.001, respectively). It is interesting to note that there was no significant difference in VEGF and HIF-1alpha overexpressions between CIN 2/3 and cervical cancer and between nonmetastatic and metastatic cancers. In addition, there was a significant correlation between the immunohistochemical score of HIF-1alpha and that of VEGF expression (Spearman's correlation coefficient 0.275, P= 0.003, n= 116). Taken together, the results of our study suggest that expressions of VEGF and HIF-1alpha could be involved in cervical carcinogenesis. In addition, the weak correlation between VEGF and HIF-1alpha expressions suggests that regulatory mechanisms other than HIF-1alpha may be involved in the expression of VEGF during cervical carcinogenesis.

Expression of VEGF and Cadherin in Gastric Cancer

Background/Aims: Cadherin preserves the cellular network which stabilizes cell-to-cell adherens junction. Vascular endothelial growth factor (VEGF) loosens the tightness of adherens junctions and induces proliferation of endothelium, contributes to tubulogenesis and increases vascular permeability. This study examined the expression of VEGF and endothelial specific VE (vascular endothelial)-cadherin and intestine specific LI (liver-intestine)-cadherin, to determine the relationship between their expressions and the clinicopathologic parameters of gastric cancer. Methods: Sixty-six gastric cancer patients at Ewha Womans University Mokdong Hospital, Seoul, Korea were enrolled. Four endoscopic biopsies were taken from the cancer and non-cancerous (normal) mucosa, respectively. The mRNA concentration of VEGF, VE-cadherin and LI-cadherin were analyzed by a semiquantitative polymerase chain reaction with GAPDH as the internal standard. Results: Among the 66 patients (M: F=41: 25, mean age, 66 ± 12 years), 47 patients had gastrectomy. The VEGF mRNA level in the cancers with well/moderate differentiation was significantly higher than in the poorly differentiated/signet ring cell carcinomas (p<0.01). No significant differences in VEGF, VE-cadherin and LI-cadherin were noted based on the TNM stage. Eleven patients (16.7%) had liver metastasis and six patients (9.1%) had peritoneal metastasis. There were no significant differences in the expression of VEGF, VE-cadherin and LI-cadherin by occurrence of distant metastases. Vascular invasion was noted in 26 patients (55.3%) and lymphatic invasion was noted in 31 patients (66.0%) among 47 patients who had gastrectomy; there was a significantly lower expression of VE-cadherin in the cases with lymphatic invasion (p=0.039). There was a negative correlation between VEGF and VE-cadherin (r=-0.291, p=0.018) and a positive correlation between VE-cadherin and LI-cadherin expression was observed (r=0.249, p=0.04). However, there was no significant relationship between VEGF and LI-cadherin. There was no significant relationship between LI-cadherin and the clinicopathologic parameters. Conclusions: VE-cadherin was decreased in cancers with lymphatic invasion and there was a positive correlation between VEGF and VE-cadherin. These findings may suggest that VEGF induces loss of VE-cadherin that lossens cell-to-cell adherens and makes endothelial cell to migrate. There was no significant relationship between LI-cadherin and the clinicopathologic parameters.

Expression of Vascular Endothelial Growth Factor and its Receptors in Canine Lymphoma

Vascular endothelial growth factor (VEGF) stimulates endothelial cell proliferation and has a pivotal role in tumour angiogenesis. The expression of VEGF and its receptors VEGFR-1 and VEGFR-2 was examined immunohistochemically in 43 specimens of canine lymphoma and in six normal lymph nodes. Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR) were performed to detect VEGF protein and mRNA, respectively. VEGF protein was expressed by 60% of the tumours with diffuse cytoplasmic labelling of the neoplastic cells. Endothelial cells, macrophages and plasma cells were also immunolabelled. VEGFR-1 was expressed by variable numbers of neoplastic cells in 54% of lymphoma specimens. VEGFR-1 was also expressed by macrophages, plasma cells, reticulum cells, and vascular endothelial cells. Macrophages and lymphocytes in germinal centres of normal lymph nodes were also immunoreactive with anti-VEGF and VEGFR-1. Most tumours did not express VEGFR-2 but in 7% of sections there was focal labelling of neoplastic and endothelial cells, with a cytoplasmic and perinuclear pattern. The observed variability in expression of VEGF and its receptors probably relates to the fact that lymphoma is a heterogeneous lymphoproliferative tumour. Individual differences in VEGF and VEGFR expression must be taken into account when VEGF and VEGFR-targeted approaches for anti-angiogenic therapy are considered in dogs.

Systemic and intracardiac vascular endothelial growth factor and angiopoietin‐1 and ‐2 levels in coronary artery disease: Effects of angioplasty

Background. Vascular growth factors are involved in the pathophysiology of human atherosclerotic vascular disease and plaque destabilization. We hypothesized that in stable patients with coronary artery disease (CAD), plasma levels of vascular endothelial growth factor (VEGF) and angiopoietins 1 and 2 (as indices of angiogenesis) would be no higher in coronary sinus blood when compared to the aortic root, coronary ostium, and peripheral femoral vein. Secondly, we hypothesized that percutaneous coronary intervention (PCI; angioplasty±stenting) would increase intracardiac levels of these indices, perhaps by destabilizing coronary plaques.Methods. Patients undergoing elective diagnostic coronary angiography (n = 70; mean age 58.8±11.2 years) of which 37 proceeded to PCI were recruited. Blood samples were obtained from the aortic root, coronary ostium, coronary sinus, and femoral vein. Plasma VEGF, angiopoietin‐1 and angiopoietin‐2 levels were measured by immunoassays.Results. There were no significant differences in VEGF, angiopoietin‐1 and angiopoietin‐2 levels when aortic root, coronary ostium, coronary sinus, and femoral vein samples were compared (P = not significant (NS)). In patients undergoing PCI, peripheral angiopoietin‐2 levels were increased significantly post PCI (P = 0.01). There was also a difference in intracardiac gradient (that is, aortic root‐coronary sinus difference) in angiopoietin‐1 (P = 0.02) following PCI. No significant changes in VEGF with PCI were noted.Conclusion. There were no differences in indices of angiogenesis when aortic root, coronary ostium, coronary sinus, and femoral vein levels of VEGF and angiopoietins are compared, suggesting that peripheral blood measurements of these indices are comparable to intracardiac levels. Although no immediate effects were observed in soluble VEGF levels, PCI affected intracardiac angiopoietin‐1 with a systemic release of angiopoietin‐2. Further investigations are necessary to determine the relative systemic and intracardiac effects of the angiopoietins in vascular remodelling post PCI.

Immunohistochemical Expression of Vascular Endothelial Growth Factor Correlates with Microvessel Density in Ph− Chronic Myeloproliferative Disorders.

There is increasing evidence that neovascularization may play an important role in hematological malignancies (particularly lymphomas, acute leukemia and myelodysplastic syndromes), but only a few studies have been performed in Ph− chronic myeloproliferative disorders (CMPDs). Increased angiogenesis has been reported in chronic idiopathic myelofibrosis (CIMF), whereas high serum levels of vascular endothelial growth factor (VEGF) have been described in Ph− CMPDs. These findings have led to the use of new therapeutic approaches aimed at directly targeting endothelial cells or VEGF. The aim of this study was to evaluate microvessel density (MVD) and immunohistochemical VEGF expression in the different categories of Ph− CMPD. We examined the bone marrow biopsies (BMBs) of 98 Ph− CMPD patients, classified as follows on the basis of the WHO criteria: 29 cases of essential thrombocythaemia (ET), 39 cases of CIMF (11 CIMF-0, 11 CIMF-1, 7 CIMF-2 and 10 CIMF-3) and 30 cases of polycythemia vera (PV) (20 polycythemic phase [pp-PV] and 10 post-polycythemic myelofibrosis [post-PVMF]. We also analysed 20 normal BMBs of subjects not suffering from hematological disease. MVD was analysed using the “hot-spot” method and an anti-CD34 antibody. Immunohistochemical VEGF expression was expressed as a VEGF index based on the formula [VEGF(i) = VEGF(+) cells x BMB cellularity/100]. Data were statistically tested at the 5% significance level (p<0.05; one-way ANOVA; Tukey's HSD test; Pearson test). MVD was significantly greater in the CIMF and PV patients (respectively 25.6 ± 6.3 and 20.7 ± 10.2) than in the ET patients (10.1 ± 4.5) or controls (7.5 ± 3.6) (p<0,01). No differences were found in relation to the CMF grading score (CIMF-0: 24.7 ± 6.5; CIMF-1: 26.1 ± 6.4; CIMF-2: 25.3 ± 4.9; CIMF-3: 26.2 ± 7.5), but post- PVMF patients had higher microvessel counts than the pp-PV patients (31.9 ± 7.3 vs 15.1 ± 5.9; p<0.01). There were also significant differences in VEGF(i) between the CIMF and PV patients (respectively 0.29 ± 0.15 and 0.28 ± 0.20) and the ET patients (0.12 ± 0.05) or controls (0.08 ± 0.04). There were no significant differences among the different grades of fibrosis (CIMF-0: 0.25 ± 0.15; CIMF-1: 0.32 ± 0.17; CIMF-2: 0.36 ± 0.16: CIMF-3: 0.27 ± 0.14), but a remarkable difference was found between the pp-PV and post-PVMF patients (0.21 ± 0.15 vs 0.49 ± 0.19; p<0.01). Finally, there was a strong correlation between MVD and VEGF(i) in the patients with CIMF (0.4006; p=0.013) and those with PV (0.7923; p<0.01). In conclusion, our findings show that angiogenesis was not different in ET patients and normal controls, whereas both MVD and VEGF levels were significantly higher in the patients with CIMF or PV, and showed a correlation between them. If confirmed, these results could serve as a rational guide for the anti-angiogenetic therapy of Ph− CMPD.

VEGF, VEGFR‐1, and CTGF cell densities in tendon are increased with cyclical loading: An in vivo tendinopathy model

Tendon injuries can occur in athletes and workers whose tasks involve repetitive, high-force hand activities, but the early pathophysiologic processes of tendinopathy are not well known. The purpose of this animal study was to evaluate the effects of cyclical tendon loading on the densities of cells producing growth factors such as vascular endothelial growth factor (VEGF), its receptor, vascular endothelial growth factor receptor 1 (VEGFR-1), and connective tissue growth factor (CTGF) in the Flexor Digitorum Profundus (FDP) tendon at the epicondyle. The FDP muscles of nine New Zealand rabbits were electrically stimulated to contract repetitively for 80 h of cumulative loading over 14 weeks. The contralateral limbs served as controls. The tendons at the medial epicondyle insertion sites were harvested, and sections were immunostained with antibodies directed against VEGF, VEGFR-1, or CTGF. Positive-staining cells were counted in six regions of interest: three along the enthesis, and three corresponding regions 1500 microns distal to the enthesis. VEGF (p = 0.0001), VEGFR-1 (p = 0.046), and CTGF (p = 0.0001) cell densities were increased in the tendon of the loaded limb compared to the nonloaded limb. In addition, regional differences in VEGF, VEGFR-1, and CTGF cell densities were found. VEGF, VEGFR-1, and CTGF are increased in tendon experiencing cyclical loading and may play a role in the early vascular changes in the progression to tendinosis.

Plasma and platelet-derived vascular endothelial growth factor and angiopoietin-1 in hypertension: effects of antihypertensive therapy.

Platelets carry angiogenic growth factors vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang-1). Although platelet-derived growth factors are important in the pathogenesis and metastasis of malignancy, their role in the pathogenesis of complications and the response to treatment in hypertension is less known. To test the hypotheses that there are differences in VEGF and Ang-1 in the plasma and within platelets from patients with hypertension, and that levels change with successful treatment. We recruited 42 previously untreated patients with hypertension (25 male; mean age 53 years) and 30 age- and sex-matched controls. Plasma VEGF, Ang-1 and soluble P-selectin (sPsel, an index of platelet activation), and total platelet [platelet VEGF (pVEGF) and platelet Ang-1 (pAng-1)] were measured by ELISA. The patients were then treated for 6 months with amlodipine-based antihypertensive therapy, achieving a mean blood pressure below 140/80 mmHg. Patients with hypertension had significantly higher levels of plasma sPsel (P =0.01), VEGF (P < 0.001) and Ang-1 (P = 0.01), as well as pVEGF (P < 0.001) and pAng-1 (P =0.02). The levels of plasma and platelet angiogenic growth factors were significantly reduced after antihypertensive treatment (VEGF, P = 0.01; pVEGF, P < 0.001; Ang-1, P < 0.001; pAng-1, P = 0.04). There were no correlations with blood pressure or the levels of sPsel. Levels of plasma and intra-platelet VEGF and Ang-1 are increased in hypertension and are decreased with treatment. Platelet levels of VEGF and Ang-1 may be related to platelet activation but may also involve other mechanisms (for example, the general vascular and haemodynamic changes) that are seen in hypertension.

Vascular endothelial growth factor (VEGF) and other common tissue prognostic indicators in breast cancer: A case-control study

VEGF is a specific mitogen and survival factor for endothelial cells and a key promoter of angiogenesis in physiological and pathological conditions. Nevertheless, VEGF tissue evaluation in cancer patients as a prognostic factor compared to the conventional histological and biological parameters is still controversial. In this case-control study, tissue VEGF was retrospectively determined by immunohistochemistry and related to T, N, ER, PgR, c-erbB-2, p53, MIB-1 and cyclin D1 in 129 breast cancer patients. Seventy-four of these patients had developed distant metastases postoperatively. The remaining 55 patients had remained disease-free >10 years after surgery. In 17 (13%) of the 129 patients (six with distant metastases and eleven disease-free) tissue and plasma VEGF were concomitantly evaluated. In univariate analysis no significant differences in VEGF and tumor size were found between metastatic and disease-free patients, whereas there were significant differences in N, ER, PgR, c-erbB-2, p53, MIB-1 and cyclin D1 (p ranging from 0.001 to 0.0001). In multivariate analysis VEGF showed less significance than N, ER, c-erbB-2, MIB-1 and cyclin D1 (p = 0.012, p = 0.007, p = 0.005, p = 0.005, p = 0.002 and p = 0.001, respectively). VEGF was a significant unfavorable prognostic indicator only in the N+ subset (p = 0.015), while ER (p = 0.05 and p = 0.021) and MIB-1 (p = 0.031 and p = 0.022) were significant in both the N+ and N- subgroups. In multivariate analysis in the 74 metastatic cases VEGF did not show any significance in relation to disease-free interval and overall survival from the time of mastectomy and from the time of relapse, whereas N and PgR did (p ranging from 0.018 to 0.001). In conclusion, tissue VEGF does not seem a suitable candidate to replace conventional histological and other common biological prognostic factors in breast cancer.

Increased but ineffectual angiogenic drive in nonhealing venous leg ulcers.

Our previous work demonstrated that angiogenesis is inhibited in nonhealing venous ulcers. The object of this study was to determine whether local expression of vascular endothelial growth factor (VEGF) and other major regulators of vessel growth are related to healing of venous ulcers. The study included 35 patients with venous ulcers (CEAP 6) and 9 patients whose ulcers had healed (CEAP 5). Control subjects were 18 patients undergoing routine operations (8 with closed suction drains, 10 standard skin biopsies). Healing ulcers were defined as having healed in less than a year from entry to the study; nonhealing ulcers failed to heal in this period. A 1-cm square biopsy specimen was taken from the edge of the ulcer or from a site of lipodermatosclerosis around a healed ulcer. Wound fluids were aspirated from beneath transparent occlusive dressings. Concentrations of VEGF(165) and VEGF-R1 were measured in tissue homogenates with enzyme-linked immunosorbent assay, and results are expressed as mean +/- SEM per milligram of soluble protein (SP). Expression of mRNA transcripts for the VEGF splice variants VEGF(121), VEGF(189), and VEGF(165); the receptors VEGF-R1 and VEGF-R2; the angiopoietins Ang-1 and Ang-2; and their receptor, Tie-2, were measured in biopsy samples with multiplex polymerase chain reaction. Expression of each transcript was normalized to that of the housekeeping gene, GAPDH. Results were analyzed with analysis of variance, t test, and chi(2) test. There was no difference in VEGF(165) protein concentration between biopsy specimens from healing ulcers (2.12 +/- 0.34 ng/mg SP; n = 18) and nonhealing ulcers (2.36 +/- 0.39 ng/mg SP; n = 12), but concentration was higher in all ulcer samples compared with healthy skin (0.57 +/- 0.20 ng/mg SP; n = 10; P <.01)) and healed ulcers (0.33 +/- 0.06 ng/mg SP; n = 9; P <.01). Concentration of VEGF(165) protein in wound fluid was significantly higher in nonhealing venous ulcers (67.17 +/- 13.87 ng/mg SP; n = 13) compared with healing venous ulcers (32.19 +/- 7.90 ng/mg SP; n = 19; P <.05) or acute wounds (12.26 +/- 4.50; n = 8; P <.01). Concentration of VEGF-R1 was similar in wound fluid obtained from healing ulcers (7.18 +/- 1.34 ng/mg SP; n = 13) and nonhealing ulcers (7.02 +/- 1.21 ng/mg SP; n = 19), and acute wounds (7.12 +/- 2.35 ng/mg SP; n = 8). There was a weak but significant correlation between VEGF(165) protein concentration in the ulcer biopsy specimen and wound fluid from the same ulcer (R(2) = 0.2; P =.019; n = 27). Expression of mRNA for VEGF receptors and Tie-2 was poor. VEGF(121) was expressed in all samples, and VEGF(165) in 43 of 48 samples. mRNA expression of VEGF(189) (P =.001), Ang-1 (P =.002), and Ang-2 (P =.026) was found in more samples from unhealed ulcers than from other sites. Healed ulcers had reduced mRNA expression of VEGF(165) (0.181 +/- 0.003) than did healing ulcers (0.307 +/- 0.016; P =.007) or nonhealing ulcers (0.375 +/- 0.033; P =.001). Relative expression of VEGF(165) to Ang-2 was much lower in healed ulcers (0.4236 +/- 0.060) than in healing ulcers (1.382 +/- 0.235; P =.010) and nonhealing ulcers (1.887 +/- 0.280; P =.003). In nonhealing venous ulcers there is a consistently high level of expression of VEGF, at both the gene transcript and protein level. As our previous data demonstrated that angiogenesis is depressed in these poorly healing ulcers, an increase in VEGF production may indicate an increased but ineffectual angiogenic drive. It is also possible that undiscovered inhibitors are released in the ulcer environment.