Endothelium : journal of endothelial cell research最新文献

Although the etiology underlying scar formation is not well understood, previous studies revealed that endothelial cells play a role in the pathogenesis of scar development. Recently, the authors developed a reliable technique to obtain endothelial cells from hypertrophic scars that involved separation of cells from the scar tissue matrix and isolation from other cell types. Using phase-contract and electron microscopy, the cells were observed to have a characteristic morphology consistent with cells of endothelial origin. The cells were further characterized as endothelial cells by assessment of endothelin (ET)-1 and intercellular adhesion molecule (ICAM) mRNA expression, and the presence of factor VIII antigen, CD34, CD31, and VE-cadherin. This isolation method provides a simple method for culturing endothelial cells from hypertrophic scar tissue and should prove useful for studying the role of endothelial cell involvement in scar formation.

Sinusoidal endothelial liver cells (SECs) have a key role in the pathophysiology of chronic liver disease. Leptin is an important profibrogenic and proinflammatory cytokine whose expression in sinusoidal endothelial liver has not been documented. The authors studied the potential of rat SECs to express the leptin and leptin receptor genes. Two cell lines of rat SECs were generated from a male rat liver by pronase-collagenase perfusion and dilution cloning. They were characterized according to morphology, ploidy, von Willebrand antigen immunoreactivity, CD31 transcription, matrix metalloproteinase secretion, and pseudocapillary formation. Expression of the leptin and leptin receptor genes was studied using qualitative reverse transcriptase-polymerase chain reaction. Both cell lines fulfilled the accepted criteria for consideration as being derived from the liver sinusoidal endothelium. Confluent monolayers of both cell lines transcribed leptin and leptin receptor genes. This work demonstrated that SECs can transcribe the leptin gene in vitro, cotranscribing with the leptin receptor gene. Leptin production and signaling at this level could be of paramount importance in liver physiopathology; further studies of this issue are warranted because it represents a potential intervention point during chronic liver diseases.

Oxidative stress induced by superoxide plays an important role in pathogenesis of cardiovascular diseases. NAD(P)H oxidase is a principal enzymatic origin for superoxide in vasculature. Recently, novel homologues of cytosolic components of NAD(P)H oxidase, Nox organizer 1 (NOXO1) and Nox activator 1 (NOXA1), are identified. On the other hand, oxidized low-density lipoprotein (ox-LDL) generates reactive oxygen species (ROS) in endothelial cells via lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). In the present investigation, the authors examined the expression, the regulation, and the role of NOXA1 in the generation of ROS in endothelial cells. The expression of NOXA1 was confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR). Dihydroethidium method showed that ox-LDL and angiotensin II increased the generation of intracellular ROS. Once the expression of p22(phox) or NOXA1 was suppressed by siRNA, the generation of ROS induced by ox-LDL and angiotensin II were potently decreased. Moreover, the expression of NOXA1 was increased by ox-LDL in a time-and dose-dependent manner. In conclusion, endothelial NOXA1 plays an essential role in generation of ROS. Ox-LDL not only increased the generation of ROS via LOX-1, but also enhanced the expression of NOXA1 in endothelial cells. NOXA1 is likely a key player that links ox-LDL with the activation of endothelial NAD(P)H oxidase.

Napoleone Ferrara and his colleagues at Genentech were the first to isolate and clone vascular endothelial growth factor (VEGF) in 1989. His laboratory has investigated many aspects of VEGF biochemistry and molecular biology. In 1993, Ferrara reported that inhibition of VEGF-induced angiogenesis by specific monoclonal antibodies resulted in dramatic suppression of the growth of a variety of tumors in vivo. These findings provided an important evidence that inhibition of angiogenesis may suppress tumor growth and blocking VEGF action could have therapeutic value for a variety of malignancies and validate the notion introduced in 1971 by Judah Folkman that inhibition of tumor angiogenesis might be a valid approach to control tumor growth. A further development was the design in a rational fashion in 1997 of a humanized anti-VEGF monoclonal antibody (Avastin), now in clinical trials as a treatment for several solid tumors and also outside of cancer, for example, in the treatment of age-related macular degeneration.

Coronary stent thrombosis remains an important problem after the implantation of different stent types. This study investigates the risk of stent thrombosis associated with the use of drug-eluting stents (DESs), bare-metal stents (BMSs) compared to balloon angioplasty. A meta-analysis of 28 randomized trials involving 5612 versus 7639 versus 2994 patients with coronary heart disease treated with DES, BMS, or balloon angioplasty was therefore performed. Comparing the implantation of DES versus BMS, DES was not found to increase the hazard for thrombosis up to 15 months (odds ratio [OR] = 0.86, 95% confidence interval [CI] 0.58 to 1.3, p < .48). There was also no significant difference in the hazard for subacute thrombosis (SAT) or late stent thrombosis (LST) in the DES versus BMSs group (OR = 0.86, 95% CI 0.50 to 1.5, p < .6 and OR = 0.92, 95% CI 0.50 to 1.68, p < .78, respectively). Comparing incidences of stent thromboses in patients receiving balloon angioplasty or implantation of BMS, the rate of SAT in the balloon angioplasty group (1.7% SAT) versus BMS group (1.8% SAT) was also similar (OR = 0.93, 95% CI 0.61 to 1.4, p < .71). Finally, there was no significant difference in the occurrence of stent thrombosis for the different coatings of DESs. In conclusion, the use of DES was not observed to have a significant effect on stent thrombosis events, compared with the implantation of BMS or balloon angioplasty.

The development of drug-eluting stents to combat the problem of in-stent restenosis has revolutionized interventional cardiology. However, concerns have emerged about the risk of late angiographic stent thromboses associated with drug-eluting stents. It has been shown that noncytotoxic concentrations of paclitaxel exert an antiangiogenic effect, suggesting that paclitaxel and similar agents may inhibit key cellular functions in a threshold-independent manner. In this study, the effect of vinblastine, an antimitotic drug, on endothelial cells is analyzed. It is investigated whether noncytotoxic concentrations of the drug could exert an antirestenotic effect. The change in levels of cell proliferation, activity, and viability in human umbilical vein endothelial cells was measured at a range of concentrations and over a number of time points. Also, the level of apoptotic activity in response to vinblastine was analyzed. This study shows that the concentration of vinblastine most appropriate in restenosis treatment would be between 0.1 and 1 nM. At this concentration, vinblastine exerts a distinct biological effect without causing an increase in apoptotic activity. These results emphasize the importance of finding an appropriate concentration window in order to minimize the risk of delayed endothelialization and thrombosis.

Membrane type 1-matrix metalloproteinase (MT1-MMP) plays a key role in extracellular matrix remodeling, endothelial cell (EC) migration, and angiogenesis. Whereas cyclic strain (CS) increases MT1-MMP expression, shear stress (SS) decreases MT1-MMP expression. The aim of this study was to determine if changes in levels of Sp1 phosphorylation induced by protein kinase Czeta (PKCzeta) in ECs exposed to SS but not CS are important for MT1-MMP expression. The results showed that SS increased Sp1 phosphorylation, which could be inhibited by pretreatment with PKCzeta inhibitors. In the presence of PKCzeta inhibitors, the SS-mediated decrease in MT1-MMP protein expression was also abolished. These data demonstrate that increased affinity of Sp1 for MT1-MMP's promoter site occurs as a consequence of PKCzeta-induced phosphorylation of Sp1 in response to SS, increasing Sp1 binding affinity for the promoter site, preventing Egr-1 binding, and consequently decreasing MT1-MMP expression.

Overexpression of a caspase-resistant form of Bcl-2 (D34A) in human umbilical vein endothelial cells (ECs) implanted into immunodeficient mice promotes the maturation of human EC-lined microvessels invested by vascular smooth muscle cells (VSMCs) of mouse origin. In contrast, EC implants not overexpressing Bcl-2 form only simple, uncoated EC tubes. Here the authors compare the phenotypes of vessels formed in vivo and the transcriptomes in vitro of EC expressing different forms of Bcl-2. Wild-type Bcl-2, like the caspase-resistant D34A Bcl-2 mutant, is antiapoptotic in vitro and promotes VSMC recruitment in vivo, whereas a G145E mutant that has diminished antiapoptotic activity in vitro does not promote vessel maturation in vivo. The D34A and wild-type forms of Bcl-2, but not the G145E mutant form of Bcl-2, significantly regulate RNA transcripts previously associated with EC-VSMC interactions and VSMC biology, including matrix Gla protein, insulin-like growth factor-binding protein (IGFBP)-2, matrix metalloproteinase (MMP)-14, ADAM17, stanniocalcin-1, and targets of the nuclear factor (NF)-kappa B, cAMP response element-binding (CREB), and activator protein 1 (AP1) transcription factor families. These effects of Bcl-2 on the transcriptome are detected in ECs cultured as angiogenic three-dimensional (3-D) tubes but are attenuated in ECs cultured as 2-D monolayers. Bcl-2-regulated transcription in ECs may contribute to vascular maturation, and support design of tissue engineering strategies using EC.

Stent thrombosis remains an important problem after the implantation of different stent types. A potential solution to this problem may be vasoactive agents with dual effects on different cell types like C-type natriuretic peptide (CNP). Therefore, in vitro and in vivo effects of CNP were investigated in a porcine restenotic model. Gene transfer of CNP in cultures of porcine vascular cells revealed up to 30% reduction of growth of smooth muscle cells (p<.05), but no suppression of endothelial growth using CNP. Applied in vivo, angiography revealed a trend of reduced restenosis formation in balloon-injured porcine arteries treated with CNP gene or beta-galactosidase (beta-Gal) control gene after three months (2.59 +/- 2.04-fold reduction, p = n.s.). Histologically, morphometry revealed significantly reduced neointima formation after treatment with CNP plasmid (7.26 +/- 1.44-fold reduction, p < .05). Evans blue staining demonstrated complete endothelial repair already 3 weeks after intervention using CNP. Transfer of CNP gene resulted in a significant inhibition of neointima formation without compromising endothelial repair. Therefore, use of the CNP gene may offer a solution to suppress restenosis formation while preventing subacute or late thrombosis.

The purpose of this investigation was to test the hypothesis that chronic N(G)-nitro-l-arginine methyl ester (l-NAME) treatment produces differential effects on conduit artery and resistance arteriole relaxation responses to endothelium-dependent and -independent vasodilators in arteries that perfuse skeletal muscle of swine. To test this hypothesis, conduit skeletal muscle arteries and second-order skeletal muscle (2A) arterioles were harvested from 14 Yucatan swine that were chronically administered l-NAME and from 16 controls. In vitro assessments of vasorelaxation to increasing doses of acetylcholine (ACH), bradykinin (BK), and sodium nitroprusside (SNP) were performed in both conduit and 2A arterioles. l-NAME treatment produced a significant reduction in both BK and ACH relaxation responses in the conduit arteries. In contrast, the relaxation response and/or sensitivity to SNP were significantly greater in the intact, but not denuded, conduit arterial rings from chronically l-NAME-treated swine. There were no significant effects of chronic l-NAME treatment on vasodilation of skeletal muscle arterioles. These findings suggest (1) that unlike arterioles, skeletal muscle conduit arteries do not functionally compensate for a lack of NO through the upregulation of alternative vasodilator pathways; (2) that the greater relaxation response in conduit arteries of chronically l-NAME-treated swine to SNP can be explained by alterations to the endothelium.