Yunpeng Shang

Resveratrol reduces endothelial progenitor cells senescence through augmentation of telomerase activity by Akt-dependent mechanisms

Recent studies have shown that resveratrol increased endothelial progenitor cells (EPCs) numbers and functional activity. However, the mechanisms remain to be determined. Previous studies have demonstrated that increased EPC numbers and activity were associated with the inhibition of EPC senescence, which involves activation of telomerase. Therefore, we investigated whether resveratrol inhibits the onset of EPC senescence through telomerase activation, leading to potentiation of cellular activity.

Effects of nicotine on the number and activity of circulating endothelial progenitor cells.

Recently, some studies have shown that nicotine increased neovascularization, which involves endothelial progenitor cells (EPCs). The effects of nicotine on EPCs are still unclear at present. Therefore, the authors investigated whether nicotine had influences on EPC number and activity. The EPCs were stimulated with nicotine (to make a series of final concentrations: 10−12 mol/L, 10−10 mol/L, 10−8 mol/L, 10−6 mol/L, 10−4 mol/L) or vehicle control for the respective time points(12, 18, 24, 32, and 48 hours). The EPCs were characterized as adherent cells double positive for DiLDL uptake and lectin binding by direct fluorescent staining under a laser‐scanning confocal microscope. They were further documented by demonstrating the expression of KDR, VEGFR‐2, and AC133 with flow cytometry. The EPC proliferation, migration, and in vitro vasculogenesis activity were assayed with the 3‐(4, 5‐dimethylthiazol‐2‐yl)‐2, 5‐diphenyltetrazolium bromide assay; the modified Boyden chamber assay; and the in vitro vasculogenesis kit, respectively. The EPC adhesion assay was performed by replating those on fibronectin‐coated dishes and then counting the adherent cells. As a result, nicotine dose dependently increased the EPC number and the proliferative, migratory, adhesive, and in vitro vasculogenesis capacity at nicotine concentrations of 10−12 to 10−8 mol/L. The peak effects on EPCs were observed at concentrations of nicotine 10−8 mol/L, similar to those in the blood of smokers. In addition, nicotine (10−8 mol/L) time dependently increased the EPC number and activity. However, cytotoxicity was seen at higher nicotine concentrations (> 10−6 mol/L). In conclusion, nicotine had complex effects on EPCs: nicotine might induce the augmentation of EPCs with enhanced functional activity at relatively low concentrations. However, cytotoxicity was seen at higher nicotine concentrations.

Effects of ox-LDL on number and activity of circulating endothelial progenitor cells.

Backgrounds: Endothelial dysfunction is thought to play a crucial role in the pathogenesis of atherosclerosis induced by ox‐LDL. Recently, a variety of evidence suggested that endothelial progenitor cells (EPCs) participated in neovascularization and reendothelialization. However, effects of ox‐LDL on EPCs number and activity are ill understood. Methods: Total mononuclear cells (MNCs) were isolated from peripheral blood by Ficoll density gradient centrifugation, and then the cells were plated on fibronectin‐coated culture dishes. After 7 days culture, attached cells were stimulated with ox‐LDL (to make a series of final concentrations: 25 µg/mL, 50 µg/mL, 100 µg/mL, 200 µg/mL), native LDL (100 µg/mL) or vehicle control for the respective time points (6 h, 12 h, 24 h and 48 h). EPCs were characterized as adherent cells double positive for DiLDL‐uptake and lectin binding by direct fluorescent staining under a laser scanning confocal microscope. EPCs were further documented by demonstrating the expression of KDR, VEGFR‐2 and AC133 with flow cytometry. Proliferation, migration and in vitro vasculogenesis activity of EPCs were assayed by MTT assay, modified Boyden chamber assay and in vitro vasculogenesis kit, respectively. EPCs adhesion assay was performed by replating those on fibronectin‐coated dishes, and then counting adherent cells. Results: Incubation of isolated human EPCs with ox‐LDL decreased the number of EPCs in concentration‐dependent manner, maximum at 200 µg/mL (approximately 70% reduction, P < 0.001). In time‐course experiments performed with an ox‐LDL concentration of 100 µg/mL, decrease of EPCs number became apparent at 12 hours and reached the maximum at 24 hours (approximately 50% reduction, P < 0.01). In addition, ox‐LDL dose and time dependently impaired EPC proliferative, migratory, adhesive and in vitro vasculogenesis capacity. Conclusion: The results of the present study defined a novel mechanism of action of ox‐LDL: the reduction of EPCs with decreased functional activity.

Proteomic analysis of the serum in patients with idiopathic pulmonary arterial hypertension.

Idiopathic pulmonary arterial hypertension (IPAH) is a rare disease of unknown etiology. The exact pathogenesis of pulmonary arterial hypertension is still not well known. In the past decades, many protein molecules have been found to be involved in the development of IPAH. With proteomic techniques, profiling of human plasma proteome becomes more feasible in searching for disease-related markers. In present study, we showed the protein expression profiles of the serum of IPAH and healthy controls after depleting a few high-abundant proteins in serum. Thirteen spots had changed significantly in IPAH compared with healthy controls and were identified by LC-MS/MS. Alpha-1-antitrypsin and vitronectin were down-regulated in IPAH and may be valuable candidates for further explorations of their roles in the development of IPAH.

Puerarin reduces endothelial progenitor cells senescence through augmentation of telomerase activity

Endothelial progenitor cells (EPCs) play an important role in both reendothelialization and neovascularization. Ex vivo expansion of EPCs might be useful for potential clinical cell therapy of ischemic diseases. However, ex vivo cultivation of EPCs leads to rapid onset of EPCs senescence, thereby severely limiting the proliferative capacity and clonal expansion potential. Therefore, we investigated whether puerarin might be able to prevent senescence of EPCs. EPCs were isolated from peripheral blood and characterized. After ex vivo cultivation, EPCs became senescent as determined by acidic beta-galactosidase staining. Puerarin dose dependently prevented the onset of EPCs senescence in culture. Moreover, puerarin increased proliferation of EPCs as assessed by BrdU incorporation assay and colony-forming capacity. To get further insights into the underlying mechanisms of these effects induced by puerarin, we measured telomerase activity and determined the phosphorylation of serine/threonine protein kinase Akt by using western blot. Puerarin significantly increased telomerase activity and phosphorylation of Akt, a downstream effector of phosphoinositide 3-kinase (PI-3K). Moreover, pretreatment with PI-3K blockers, either wortmannin or LY294002, significantly attenuated the puerarin puerarin-induced telomerase activity. Taken together, the results of the present study indicated that puerarin delayed the onset of EPCs senescence, which may be related to the activation of telomerase through the PI-3K/Akt pathway. The inhibition of EPCs senescence by puerarin in vitro may improve the functional activity of EPCs in a way that is important for potential cell therapy.

Level of Pregnancy-associated Plasma Protein-A Correlates With Coronary Thin-cap Fibroatheroma Burden in Patients With Coronary Artery Disease: Novel Findings From 3-Vessel Virtual Histology Intravascular Ultrasound Assessment.

Abstract Pregnancy-associated plasma protein-A (PAPP-A) level is an independent predictor of acute cardiovascular event occurrence. To test the hypothesis that increased PAPP-A levels would be associated with a higher burden of coronary thin-cap fibroatheroma (TCFA) thereby underlying the heightened risk for cardiovascular events in patients with coronary artery disease; 154 patients (462 vessels and 975 plaques) with stable angina or non-ST-segment elevation acute coronary syndrome (NSTE-ACS) referred for percutaneous coronary intervention were assessed using 3-vessel virtual histology (VH)-intravascular ultrasound (IVUS). Thin-cap fibroatheroma virtual histology was defined as focal, necrotic core (NC)-rich (≥10% of cross-sectional area) plaques in contact with the lumen, and plaque burden ≥40%. Pregnancy-associated plasma protein-A levels were determined by sandwich enzyme-linked immunosorbent assay, and patients were divided into 3 groups based on PAPP-A level tertiles. Although the highest PAPP-A level tertile was not associated with 3-vessel plaque number, it was associated with 3-vessel VH-TCFA number and necrotic core volume. Patients with ≥3 VH-TCFAs had a higher PAPP-A level than patients with 1 to 3 VH-TCFAs or without any VH-TCFA (13.3 ± 11.8 versus 7.8 ± 4.7 versus 7.4 ± 4.7 mIU/L, P < 0.001, respectively). Moreover, PAPP-A level was an independent predictor of higher total number of VH-TCFAs (OR 1.18; 95% CI 1.07–1.29, P = 0.001). This VH-IVUS study demonstrated, for the first time to our knowledge, that higher PAPP-A levels are associated with higher 3-vessel TCFA burden in patients with coronary artery disease. Pregnancy-associated plasma protein-A, therefore, might be a useful serum biomarker to predict increased coronary TCFA burden and plaque instability.

Comprehensive assessment of the association of WNK4 polymorphisms with hypertension: evidence from a meta-analysis.

The relationship between with-no-lysine [K] kinase 4 (WNK4) gene polymorphisms and hypertension has been widely investigated, However, the studies yielded contradictory results. To evaluate these inconclusive findings comprehensively, we therefore performed a meta-analysis. Ten articles encompassing 16 independent case-control studies with 6089 hypertensive cases and 4881 normotensive controls were selected for this meta-analysis. Four WNK4 gene polymorphisms were identified (G1155942T, G1156666A, T1155547C, and C6749T). The results showed statistically significant associations of G1155942T polymorphism (allelic genetic model: odds ration or OR = 1.62, 95% confidence interval or CI: 1.11–2.38, P = 0.01; dominant model: OR = 1.85, 95% CI: 1.07–3.19, P = 0.03) and C6749T polymorphism (allele contrast: OR = 2.04, 95% CI: 1.60–2.59, P<0.01; dominant model: OR = 2.04, 95%CI: 1.59–2.62, P<0.01; and homozygous model: OR = 5.01, 95% CI: 1.29–19.54, P = 0.02) with hypertension risk. However, neither C1155547T nor G1156666A was associated significantly with hypertension susceptibility. In conclusion, this meta-analysis suggested that WNK4 G1155942T and C6749T gene polymorphisms may contribute to the susceptibility and development of hypertension. Further well-designed studies with larger sample size are required to elucidate the association of WNK4 gene multiple polymorphisms with hypertension risk.