Endostatin Levels Research Articles

Effects of lentivirus-mediated endostatin on endothelial progenitor cells

Endothelial progenitor cells (EPCs) are candidates for gene therapies against retinal neovascularization (NV). The aim of present study was to investigate the effects of endostatin transfection on EPC function. In the present study, the EPCs were infected with lentivirus overexpressing endostatin. The transfection effects of endostatin overexpression on the proliferation, migratory, differentiation, apoptosis and the cell cycle of this cell line were determined. The real-time quantitative polymerase chain reaction (RT-qPCR) and western blot assays showed high expression levels of endostatin. A cell counting kit-8 assay showed that endostatin overexpression inhibited EPC proliferation. The transwell assay indicated that endostatin overexpression could suppress EPC migration. Furthermore, endostatin overexpression enhanced apoptosis (as showed by AnnexinV-FITC/propidiumiodide staining analysis), induced differentiation and blocked the cell cycle. As compared with negative control group, EPC viability significantly decreased in gene transfection group. In conclusion, present study determined the feasibility of lentivirus-mediated endostatin gene transfer, and indirectly proved the effect of endostatin secretion on EPCs. Also our study provided a new opportunity for the potential application of gene therapy in retinal NV.

Endostatin: a promising biomarker in the cardiovascular continuum?

The current review article aims to provide an up-to-date summary of previous studies in humans that have reported the association between circulating endostatin levels and different cardiovascular phenotypes. We also aim to provide suggestions for future directions of future research evaluating endostatin as a clinically relevant cardiovascular biomarker. With a few exceptions, higher circulating levels of endostatin seem to reflect vascular and myocardial damage, and a worsened prognosis for cardiovascular events or mortality in individuals with hypertension, diabetes, kidney disease, cardiovascular disease, as well as in the general population. Circulating endostatin seems to be a promising biomarker for cardiovascular pathology, but there is not enough evidence to date to support the use of endostatin measurements in clinical practice.

Assessment of plasma endostatin to predict acute kidney injury in critically ill patients.

We evaluated whether plasma endostatin predicts acute kidney injury (AKI), need for renal replacement therapy (RRT), or death. Prospective, observational, multicenter study from 1 September 2011 to 1 February 2012 with data from 17 intensive care units (ICUs) in Finland. A total of 1112 patients were analyzed. We measured plasma endostatin within 2 h of ICU admission. Early AKI (KDIGO stage within 12 h of ICU admission) was found in 20% of the cohort, and 18% developed late AKI (KDIGO criteria > 12 h from ICU admission). Median (IQR) admission endostatin was higher in the early AKI group, 29 (19.1, 41.9) ng/ml as compared to 22.4 (16.1, 30.1) ng/ml for the late AKI group, and 18 (14.0, 23.6) ng/ml for non-AKI patients (P < 0.001). Endostatin level increased with increasing KDIGO stage. Significantly higher endostatin levels were found in patients with sepsis as compared to those without. Predictive properties for AKI, RRT, and mortality were low with corresponding areas under the receiver operating characteristic curve (AUC) of 0.62, 0.67, and 0.59. Sensitivity analyses among patients with chronic kidney disease or sepsis did not improve the predictive ability of endostatin. Adding endostatin to a clinical AKI prediction model (illness severity score, urine output, and age) insignificantly changed the AUC from 0.67 to 0.70 (P = 0.14). Endostatin increases with AKI severity but has limited value as a predictor of AKI, RRT and 90-day mortality in patients admitted to ICU. Moreover, endostatin does not improve AKI risk prediction when added to a clinical risk model.

Effect of Niacin on Inflammation and Angiogenesis in a Murine Model of Ulcerative Colitis

Butyrate and niacin are produced by gut microbiota, however butyrate has received most attention for its effects on colonic health. The present study aimed at exploring the effect of niacin on experimental colitis as well as throwing some light on the ability of niacin to modulate angiogenesis which plays a crucial role of in the pathogenesis of inflammatory bowel disease. Rats were given niacin for 2 weeks. On day 8, colitis was induced by intrarectal administration of iodoacetamide. Rats were sacrificed on day 15 and colonic damage was assessed macroscopically and histologically. Colonic myeloperoxidase (MPO), tumour necrosis factor (TNF)-α, interleukin (IL)-10, vascular endothelial growth factor (VEGF), angiostatin and endostatin levels were determined. Niacin attenuated the severity of colitis as demonstrated by a decrease in weight loss, colonic wet weight and MPO activity. Iodoacetamide-induced rise in the colonic levels of TNF-α, VEGF, angiostatin and endostatin was reversed by niacin. Moreover, niacin normalized IL-10 level in colon. Mepenzolate bromide, a GPR109A receptor blocker, abolished the beneficial effects of niacin on body weight, colon wet weight as well as colonic levels of MPO and VEGF. Therefore, niacin was effective against iodoacetamide-induced colitis through ameliorating pathologic angiogenesis and inflammatory changes in a GPR109A-dependent manner.

High levels of FLT3-ligand in bone marrow and peripheral blood of patients with advanced multiple myeloma.

IntroductionMultiple myeloma (MM) is still incurable due to resistance against various therapies. Thus, the identification of biomarkers predicting progression is urgently needed. Here, we evaluated four biomarkers in bone marrow and peripheral blood of MM patients for their prognostic significance.Materials & methodsBone marrow- and peripheral blood plasma levels of FLT3-L, soluble TIE2, endostatin, and osteoactivin were determined in patients with monoclonal gammopathy of undetermined significance (MGUS, n = 14/n = 4), patients with newly diagnosed MM (NDMM, n = 42/n = 31) and patients with relapsed/refractory MM (RRMM, n = 27/n = 16) by sandwich ELISA.ResultsMedian FLT3-L expression increased from MGUS (58.77 pg/ml in bone marrow; 80.40 pg/ml in peripheral blood) to NDMM (63.15 pg/ml in bone marrow; 85.05 pg/ml in peripheral blood) and was maximal in RRMM (122 pg/ml in bone marrow; 160.47 pg/ml in peripheral blood; NDMM vs. RRMM p<0.001). A cut-off value of FLT3-L >92 pg/ml in bone marrow and >121 pg/ml in peripheral blood was associated with relapse or refractoriness in MM patients. FLT3-L was found to be a high predictive marker for discrimination between NDMM and RRMM as well in bone marrow as in peripheral blood (AUC 0.75 in bone marrow; vs 0.84 in peripheral blood).ConclusionHigh levels of FLT3-L in bone marrow and peripheral blood of MM patients identify patients with progressive disease and are associated with relapse or refractoriness in MM patients. FLT3-L could be useful as a marker to identify RRMM patients and should be evaluated as target for future therapies.

Angiogenic and angiostatic factors in renal scleroderma-associated vasculopathy

BackgroundThe angiogenesis in systemic sclerosis (SSc) is impaired. An imbalance of pro-angiogenic factors and angiogenesis inhibitors has been implicated in the progression of peripheral microvascular damage, defective vascular repair and fibrosis. Intrarenal resistance index are considered markers of renal vasculopathy. The aim of the study is to evaluate angiogenic and angiostatic factors (VEGF and endostatin) in SSc patients and to correlate with intrarenal hemodynamic parameters. Methods91 SSc patients were enrolled in this study. Serum VEGF and endostatin levels were determined. All patients underwent a renal Doppler ultrasound ResultsA significant positive correlation was observed between endostatin and renal Doppler parameters (p<0.0001). A negative correlation was observed between serum levels of endostatin and eGFR (p<0.01). In SSc patients with high resistive index, serum levels of endostatin were significantly (p<0.01) higher than in SSc patients with normal resistive index. The serum levels of endostatin significantly increased with progression of nailfold videocapillaroscopy damage (p<0.01) and were significantly (p<0.05) higher in SSc patients with digital ulcers than in SSc patients without digital ulcers. ConclusionThis is the first study that assess in SSc patients intrarenal hemodynamic parameters and endostatin. In SSc patients, endostatin represents a marker of renal scleroderma-associated vasculopathy.

Unexpected and striking effect of heparin-free dialysis on cytokine release

Heparin (both unfractionated and low molecular weight) is not only a potent anticoagulant but also has many pleiotropic effects, some of which are mediated by cytokine release. We compared the effect of hemodialysis (HD) with enoxaparin as an anticoagulant and without systemic anticoagulation (heparin-grafted membrane—Evodial) on the release of monocyte chemoattractant protein 1 (MCP-1), endostatin (ES) and activin A (Act-A). Nineteen stable HD patients were dialyzed with or without heparin, and plasma levels of MCP-1, ES and Act-A were measured after such a dialysis. During HD with Evodial, the intradialytic levels of all three cytokines were 2–3 folds lower. The between-anticoagulant differences were significant over time for all three cytokines: MCP-1 (P < 0.001), ES (P < 0.001) and Act-A (P < 0.001). This striking effect of heparin-free dialysis with Evodial membrane may be beneficial not only because it reduces the possibility of bleeding complications but also because it might reduce proinflammatory cytokine concentration and therefore contribute to the improvement in endothelial function. Further studies are needed to determine whether it has a positive effect on morbidity and mortality of maintenance HD patients.

Correlation of Plasma Vascular Endothelial Growth Factor and Endostatin Levels with Symptomatic Intra- and Extracranial Atherosclerotic Stenosis in a Chinese Han Population

Symptomatic intracranial atherosclerotic stenosis (ICAS) and extracranial atherosclerotic stenosis (ECAS) are different in many aspects. Here, we explored the association between the location or severity of atherosclerotic stenosis and pro- or antiangiogenic factors, specifically vascular endothelial growth factor (VEGF) and endostatin (ES). We evaluated 198 consecutive patients with acute ischemia stroke: 132 with large-artery atherosclerosis (LAA) and 66 with small-artery occlusion (small-vessel occlusion). The LAA group was subclassified into 102 patients with ICAS and 30 with ECAS. Independent associations of VEGF, ES levels, and VEGF/ES ratio with the location of cerebral stenosis and the severity or short-term prognosis (14th day modified Rankin Scale) of ICAS were evaluated. Plasma concentrations of VEGF and ES were lower (P < .05) in ICAS (38.07, 32.76-46.28 pg/mL and 58.95, 55.04-59.77 ng/mL) than those in ECAS (45.00, 34.30-83.34 pg/mL and 140.74, 85.63-231.21 ng/mL). Logistic regression analysis showed that VEGF concentrations and dyslipidemia were independently associated with ICAS, with odds ratios of .987 [95% CI = (.976, .998)] and .265 [95% CI = (.103, .792)], respectively. Moreover, plasmatic VEGF levels increased gradually along with the severity of ICAS (P = .003), and lower levels of ES (P = .040) or a higher VEGF/ES ratio (P = .048) were related to unfavorable short-term prognosis of ICAS. Lower VEGF levels are associated with the presence of symptomatic ICAS, but not with ECAS. Furthermore, the severity of ICAS is positively correlated with the levels of VEGF, and lower ES levels or a predominance of VEGF over ES are predictors of poor short-term prognosis of ICAS.

Abstract TMP28: Aging Predicts Levels of Antiangiogenic Factors and Stroke in Intracranial Atherosclerosis

Introduction: Stroke risk more than doubles each decade after the age of 55. In patients with intracranial atherosclerosis (ICAS), the SAMMPRIS study showed that individuals 60 years or older had a higher risk of recurrent stroke or death (17%) than younger patients (12%) despite intensive medical management (IMM). In prior studies, an antiangiogenic profile has been associated with higher rates of failure to IMM in stroke. The purpose of this study is to evaluate the hypothesis that aging is an independent predictor of higher circulating levels of antiangiogenic factors endostatin and angiostatin in ICAS. Methods: Prospective observational longitudinal study of circulating angiogenic factors in 74 patients with ICAS treated medically. Patients with ICAS greater than 70% had angiostatin and endostatin measured in duplicate via ELISA at baseline and were followed for 6 months. Mixed model analysis was performed for the outcome variables endostatin and angiostatin. Predictor variables included age, gender, smoking history, hypertension (HTN), hyperlipidemia (HLD), Diabetes 2, obesity, coronary artery disease (CAD) and peripheral artery disease (PAD). Classification and regression tree (CART) analysis were used to select the threshold age for the longitudinal model. Results: Age was a significant independent predictor of higher levels of endostatin (p=0.002) and angiostatin (p&lt;0.001). Age older than 52 was significantly associated with higher levels of endostatin (p=0.03), and age older than 70 with higher levels of angiostatin (p=0.005). Additional significant factors associated with higher levels of antiangiogenic circulating factors included PAD, HTN, HLD, and obesity. Conclusion: Aging is a significant independent predictor of elevated antiangiogenic circulating factors, which are associated to failure to IMM in ICAS. Antiangiogenesis can potentially represent a common path for multimorbidity of advanced age.

Abstract 67: Blood Biomarkers for Prediction of Cardiac Complications in the Stroke Unit

Introduction: Cardiac complications such as acute coronary syndrome (ACS) or acute congestive heart failure (ACHF) account for 2-6%of mortality after stroke. Early detection of the patients at the highest risk to suffer from cardiac complications could be of interest to indicate a closer monitoring or therapeutic measures. We aimed to test whether a panel of blood biomarkers could predict the development of cardiac complications during stroke unit admission. Methods: The StrokeChip was a prospective, observational study, conducted at six Hospitals in Catalonia. Patients with suspected stroke were enrolled at Emergency Departments. Blood samples were obtained within the first six hours after symptom onset to measure a 21-biomarker panel. In-hospital development of cardiac complications (ACS and ACHF) was recorded. The association between the measured biomarkers and the development of complications was assessed by logistic regression analysis. Results: From August-2012 to December-2013, 941 out of 1308 patients enrolled were ischemic strokes. Cardiac complications were developed by 25 patients, including 19 ACHF and six cases of ACS. Due to the small number of ACS, just ACHF were further analyzed. Patients with ACHF had higher baseline levels of D-dimer, endostatin, VAP-1, NT-proBNP, FasL and vWF, and lower levels of IGFBP-3. After logistic regression analysis, NT-proBNP [OR=3.56(1.17-10.84)], FasL [OR=4.94(1.60-15.26)], VAP-1 [OR=6.56(2.21-19.46)] and IGFBP-3 [OR=0.21(0.07-0.67)] were independently associated with ACHF after adjustment by clinical predictors such as stroke severity, previous disability or history of coronary disease. Addition of the biomarker panel over clinical predictors resulted in improved accuracy (AUC from 0.808 to 0.925, p=0.05). Conclusions: Baseline measurement of blood biomarkers in acute stroke could be useful to predict the development of cardiac complications during stroke unit admission.

Changes in serum vascular endothelial growth factor and endostatin concentrations associated with circulating endothelial progenitor cells after acute ischemic stroke.

Angiogenesis is an important pathophysiological response to cerebral ischemia, and can be modulated by vascular endothelial growth factor (VEGF) and endostatin. Circulating endothelial progenitor cells (EPCs) also play an important role as an endogenous repair mechanism for ischemic injury. We sought to investigate early changes in the expression of VEGF and endostatin in serum and the circulating EPCs in patients with acute ischemic stroke (AIS) and analyzed the relations between them. The peripheral blood and serum samples were obtained from 30 patients at 1, 3, 5 and 7 d after AIS. Flow cytometry was used to quantify EPCs, and VEGF and endostatin were measured by enzyme linked immunosorbent assay. Correlation analysis was performed to assess the relations between them. Receiver operating characteristic (ROC) curve was used to appraise the value of EPCs levels in predicting the 90-day prognosis after AIS. Compared with control subjects, circulating EPCs numbers increased from a very lower initial level (P < 0.001) until 7 d after AIS. Serum VEGF and endostatin levels increased and peaked at 3 d and 5 d post-stroke (both P < 0.001), respectively. A significant correlation (P = 0.001) was found between peak serum VEGF concentration and peak endostatin concentration. VEGF/endostatin ratio at day 1 and day 3 after AIS significantly correlated with circulating EPCs numbers at day 5 (P < 0.001) and day 7 post-stroke (P < 0.001). ROC curve analysis suggested that circulating EPCs number at day 7 had a significantly predictive power for good prognosis. VEGF and endostatin may mediate EPCs proliferation in the early phase of ischemic stroke, and the circulating EPCs levels can be a predictor of clinical outcome in AIS.

Diagnostic Values of sVEGFR-1 and Endostatin in Malignant Pleural Effusions in Patients with Lung Cancer.

The diagnosis of malignant pleural effusion (MPE) remains a common clinical challenge because of the sensitivity of conventional cytology for the detection is insufficient. Thus, a sensitive clinical marker for diagnosis is required. The aim of this study was to assess the role of two anti-angiogenic cytokines, soluble vascular endothelial growth factor receptor-1 (sVEGFR-1) and endostatin, in diagnosing MPE. Effusion samples from 44 patients with MPE caused by lung cancer and from 36 patients with benign pleural effusion (BPE) were collected. The concentrations of sVEGFR-1 and endostatin in pleural fluid were determined by an enzyme-linked immunosorbent assay (ELISA). The diagnostic performance was measured by receiver operating characteristic curves (ROCs). The levels of sVEGFR-1 and endostatin in MPE due to lung cancer were significantly higher than those in BPE (p < 0.05). The sensitivity and specificity of endostatin were 52.27% and 86.11%, respectively, while for sVEGFR-1, the sensitivity was 88.64% and the specificity was 58.33%. Interestingly, the combination of sVEGFR1 and endostatin produced better sensitivity and specificity of 72.73% and 83.33%, respectively. In addition, the levels of sVEGFR-1 and endostatin were significantly related to each other (p < 0.05), and the levels of endostatin in bloody effusions were significantly higher than those in non-bloody effusions (p < 0.05). Our study indicated that the levels of sVEGFR-1 and endostatin were significantly elevated in MPE. The combined detection of sVEGFR-1 and endostatin may be useful in the diagnosis of MPE.

The Relation between Serum Endostatin Level and Carotid Atherosclerosis in Healthy Residents of Japan: Results from the Kyushu and Okinawa Population Study (KOPS).

Aim: To examine the association between the serum endostatin levels and subclinical atherosclerosis independent of traditional risk factors in a healthy Japanese population.Methods: Among 1,057 residents who attended free public physical examinations between 2010 and 2011, we evaluated the data of 648 healthy residents for whom the serum endostatin level and common carotid intima-media thickness (IMT) were successfully measured.Results: The median endostatin level was 63.7 ng/mL (interquartile ranges: 49.7–93.2 ng/mL), and the mean carotid IMT was 0.68 ± 0.12 mm. Residents with above median endostatin had significantly higher carotid IMT than did those with below median endostatin (0.71 ± 0.14 vs. 0.65 ± 0.09 mm, P < 0.001). Multiple linear regression analysis demonstrated that increased serum endostatin is significantly associated with carotid IMT (above vs. below median endostatin level; beta = 0.11, P = 0.03), independent of the known covariates of age, sex, body mass index, drinking and smoking status, systolic blood pressure, diastolic blood pressure, hemoglobin A1c, low density lipoprotein cholesterol, estimated glomerular filtration rate, and log-transformed high sensitive C-reactive protein.Conclusions: A higher serum endostatin level reflected subclinical atherosclerosis in this Japanese population.

Endostatin a Potential Biomarker for Heart Failure with Preserved Ejection Fraction.

BackgroundEndostatin is a circulating endogenous angiogenesis inhibitor preventing neovascularization. Previous studies demonstrated the prognostic value of Endostatin among patients with heart failure with reduced ejection fraction (HFrEF). However, the role of Endostatin among patients with heart failure with preserved ejection fraction (HFpEF) remains unclear.ObjectiveThis study aimed to investigate the association between serum Endostatin levels, natriuretic peptide levels and the severity of left ventricular diastolic dysfunction and the diagnosis of HFpEF.MethodsEndostatin serum concentrations were measured in 301 patients comprising 77 HFpEF patients, 169 patients with asymptomatic left ventricular diastolic dysfunction (ALVDD), and 55 controls with normal cardiac function.ResultsEndostatin serum levels were significantly elevated in patients with HFpEF (median/interquartile range 179.0 [159-220]) and ALVDD (163.8 [145.4-191.3]) compared to controls (149.1 [130.6-176.9]), p < 0.001 and p = 0.004, respectively) and significant correlated with N-terminal pro B-type natriuretic peptide (NT-proBNP).ConclusionsThis hypothesis-generating pilot study gives first evidence that Endostatin correlates with the severity of diastolic dysfunction and may become a novel biomarker for HFpEF. We hypothesize a rise in Endostatin levels may reflect inhibition of adaptive angiogenesis and adverse cardiac remodeling.

Platelets from glioblastoma patients promote angiogenesis of tumor endothelial cells and exhibit increased VEGF content and release

Glioblastoma multiforme (GBM) is the most common and fatal intracranial cancer in humans and exhibits intense and aberrant angiogenesis that sustains its malignancy and involves several angiogenic signals. Among them, vascular endothelial growth factor (VEGF) plays a key role and is overexpressed in GBM. Different cells appear to act as triggers of the aberrant angiogenesis, and, among them, platelets act as key participants. In order to provide further insights into the platelet features and angiogenic role in GBM, this study investigated the effects of platelet releasate on GBM-derived endothelial cells (GECs) and the levels of VEGF and endostatin, as pro- and anti-angiogenic components of platelet releasate from GBM patients. We demonstrate for the first time that: 1) platelet releasate exerts powerful pro-angiogenic effect on GECs, suggesting it might exert a role in the aberrant angiogenesis of GBM; 2) ADP and thrombin stimulation leads to significantly higher level of VEGF, but not of endostatin, in the releasate of platelets from GBM patients than those from healthy subjects; and 3) the intraplatelet concentrations of VEGF were significantly elevated in GBM patients as compared to controls. Moreover, we found a direct correlation between platelet-released VEGF and overall survival in our patient cohort. Although preliminary, these findings prompt further investigations to clarify the biologic relevance of platelet VEGF in GBM and prospective studies for screening GBM patients for anti-VEGF therapy and/or to optimize this treatment.

Filamin A interacting protein 1-like expression inhibits progression in colorectal cancer.

Filamin A interacting protein 1-like (FILIP1L) expression, which is decreased in various cancers, may inhibit carcinogenesis. In this study, we evaluated the effects of FILIP1L on oncogenic behavior and prognosis in colorectal cancer. siRNA-mediated FILIP1L knockdown enhanced tumor cell migration and invasion and inhibited apoptosis and cell cycle arrest in COLO205 cells. pcDNA-myc vector-mediated FILIP1L overexpression suppressed tumor cell migration and invasion and induced apoptosis and cell cycle arrest in HCT116 cells. FILIP1L knockdown enhanced angiogenesis by increasing VEGF-A and HIF-1α levels and decreasing angiostatin level. FILIP1L overexpression suppressed angiogenesis by decreasing VEGF-A and -D l level and increasing angiostatin and endostatin levels. Phosphorylated β-catenin levels decreased and phosphorylated Akt and GSK-3β levels increased following FILIP1L knockdown. FILIP1L overexpression had the opposite effects. FILIP1L expression was associated with reductions in tumor size, cell differentiation, lymphovascular invasion, stage, invasion depth and lymph node metastasis, and with longer overall survival. Mean Ki-67 labeling indexes and microvessel density values were lower in FILIP1L-positive tumors than in FILIP1L-negative tumors. These results indicate that FILIP1L suppresses tumor progression by inhibiting cell proliferation and angiogenesis in colorectal cancer.

The association of an elevated serum endostatin level and carotid atherosclerosis: Results from the Kyushu and Okinawa Population Study (KOPS)

The association of an elevated serum endostatin level and carotid atherosclerosis: Results from the Kyushu and Okinawa Population Study (KOPS)

Relationship between thrombospondin-1, endostatin, angiopoietin-2, and coronary collateral development in patients with chronic total occlusion.

This study is aimed to investigate whether serum angiostatic factors (thrombospondin-1 [TSP-1] and endostatin) or angiogenic factors (angiopoietin-2 [Ang-2]) are related to coronary collateral vessel development in patients with chronic total occlusion (CTO).A total of 149 patients were enrolled in the study, and 39 patients with coronary artery disease but without significant stenosis were included in control group. In 110 patients with CTO lesion, 79 with Rentrop grades 2 to 3 collaterals were grouped as good collateral, while 31 with Rentrop grades 0 to 1 collaterals were grouped as poor collateral. Serum TSP-1, endostatin, and Ang-2 levels were studied.Serum endostatin level was significantly higher in poor collateral group compared with control group and good collateral group, respectively (96.2 ± 30.4 vs 77.8 ± 16.5 ng/mL, P = 0.007; 96.2 ± 30.4 vs 81.2 ± 30.4 ng/mL, P = 0.018). In multivariate analysis, decreased serum endostatin level was independently related to good coronary collateral development. Serum TSP-1 level was lower in patients with CTO compared with control group. However, no difference in TSP-1 level was detected between poor and good collateral group. The serum Ang-2 level did not show a significant difference among 3 groups.Circulatory endostatin may be a useful biomarker for coronary collateral development and potential target for therapeutic angiogenesis in patients with CTO.

The Effects Of Protease-Activated Receptors 1 And 4 In Human Platelet Activation And Inflammation

Category: Basic Sciences/Biologics Introduction/Purpose: Tendon injuries occur frequently and cost billions of health care dollars annually. Recently, there has been an increase in the use of platelet-rich plasma (PRP) to treat tendon injuries. However, the efficacy of PRP treatment is controversial due to inconsistent results from human clinical trials. It is thought that variations in PRP preparation contribute to these inconsistencies. Specifically, platelets in PRP contain pro-angiogenic (e.g. VEGF) or anti-angiogenic (e.g. endostatin) factors, which may differentially affect the healing of tendon injuries. It is known that these factors are selectively released after platelet activation by specific receptors. Therefore, in this study we investigated the effect of protease-activated receptors 1 and 4 (PAR1 and PAR4) in platelet activation and inflammation. Methods: Platelet preparation – Human blood was obtained from 12 healthy donors and 9 ml of blood was mixed with 1 ml of 3.8% sodium citrate and centrifuged at 500g for 10 min. Then, the supernatant (PRP) without the buffy coat was centrifuged at 1000g for 10 min and the resulting pellet was washed in Tyrodes-HEPES buffer and centrifuged for 10 min at 1000g. Finally, platelets in the pellet was suspended in Tyrodes-HEPES buffer and used in experiments. Platelet activation – About 100 μl of platelet from above was activated with 5 μl 1 mM PAR1-activating peptide (PAR1-AP) or PAR4-activating peptide (PAR4-AP) at 25°C for 10 min. Then, the mixture was centrifuged at 1000g for 10 min, and the levels of VEGF, endostatin, IL-1RA and HMGB-1in the supernatant was determined by ELISA. Platelets without activators were used as controls. Results: PAR1 induced angiogenic effects in human platelets. PAR1 activated platelets released 3 times more VEGF than when activated with PAR4 (Fig. 1A). However, PAR4 activated platelets released 7 times more endostatin than the PAR1 activated platelets (Fig. 1B). Further, PAR1 induced anti-inflammatory effects in human platelets; it did not change IL-1R-A (Fig. 2A) but decreased HMGB-1 levels (Fig. 2B). In contrast, PAR4 stimulated inflammatory effects in human platelets by lowering IL-1-RA and increasing HMGB-1 levels. Conclusion: Our findings indicate that PAR1 induces angiogenetic and anti-inflammatory effects in human platelets, while PAR4 has anti-angiogenetic and inflammatory effects. Of significance is HMGB-1, which is constitutively expressed in the nuclei of most mammalian cells. Under cellular stress, HMGB1 is released into the extracellular matrix and activates the immune response thus acting as a danger-signal. Both PAR1 and PAR4 selectively regulated the release of VEGF and endostatin, and IL-1RA and HMGB-1 from human platelets. Therefore, the role of PAR1 and PAR4 on human platelet activation and inflammation should be considered prior to the use of PRP to treat tendon injuries.

Cnidium officinale extract and butylidenephthalide inhibits retinal neovascularization in vitro and in vivo.

BackgroundRetinal neovascularization, which is the pathological growth of new blood vessels, is associated with retinopathy of prematurity, neovascular age-related macular degeneration, diabetic retinopathy and retinal vein occlusion. In this study, we evaluated the effect of an extract of Cnidium officinale Makino (COE) and its bioactive compound, butylidenephthalide (BP), on the migration and tube formation of human umbilical vein endothelial cells (HUVECs), and on retinal pathogenic neovascularization in the oxygen-induced retinopathy (OIR) mouse model.MethodThe HUVECs were incubated with COE and BP (0.1–10 μg/ml). The mice were exposed to 75 % oxygen for 5 days starting on the 7th postnatal day (P7–P12). Then, the mice were returned to room air and intraperitoneally injected with COE (100 mg/kg) and BP (5 mg/kg) once per day for 5 days (P12–P16). On P17, we measured retinal neovascularization and analyzed the angiogenesis-related proteins expression using protein arrays.ResultsCOE and BP inhibit the HUVECs migration and the tube formation in a dose-dependent manner. In addition, COE significantly decreased retinal neovascularization in the OIR mice. COE reduced the expression levels of AREG, ANG, DLL4, Endostatin, IGFBP-2 and VEGF. Additionally, BP also inhibited the retinal neovascularization and down-regulated the expression of AREG, ANG, DLL4 and VEGF.ConclusionThese results suggest that COE and BP exerts antiangiogenic effects on retinal neovascularization by inhibiting the expression of AREG, ANG, DLL4 and VEGF, indicating that antiangiogenic activities of COE may be in part due to its bioactive compound, BP.