Prostaglandin E 2 (PGE 2 ) and prostacyclin are lipid mediators produced by cyclooxygenase and implicated in the regulation of vascular function, wound repair, inflammatory processes, and acute lung injury. Although protective effects of these prostaglandins (PGs) are associated with stimulation of intracellular cAMP production, the crosstalk between cAMP-activated signal pathways in the regulation of endothelial cell (EC) permeability is not well understood. We studied involvement of cAMP-dependent kinase (PKA), cAMP-Epac-Rap1 pathway, and small GTPase Rac in the PGsinduced EC barrier protective effects and cytoskeletal remodeling. PGE 2 and PGI 2 synthetic analog beraprost increased transendothelial electrical resistance and decreased dextran permeability, enhanced peripheral F-actin rim and increased intercellular adherens junction areas reflecting EC barrier-protective response. Furthermore, beraprost dramatically attenuated thrombin-induced Rho activation, MLC phosphorylation and EC barrier dysfunction. In vivo, beraprost attenuated lung barrier dysfunction induced by high tidal volume mechanical ventilation. Both PGs caused cAMPmediated activation of PKA-, Epac/Rap1-and Tiam1/Vav2-dependent pathways of Rac1 activation and EC barrier regulation. Knockdown of Epac, Rap1, Rac-specific exchange factors Tiam1 and Vav2 using siRNA approach, or inhibition of PKA activity decreased Rac1 activation and PGinduced EC barrier enhancement. Thus, our results show that barrier-protective effects of PGE 2 and prostacyclin on pulmonary EC are mediated by PKA and Epac/Rap pathways, which converge on Rac activation and lead to enhancement of peripheral actin cytoskeleton and adherens junctions. These mechanisms may mediate protective effects of PGs against agonist-induced lung vascular barrier dysfunction in vitro and against mechanical stress-induced lung injury in vivo.
Abstract. In this paper, a field test of wake-steering control is presented. The field test is the result of a collaboration between the National Renewable Energy Laboratory (NREL) and Envision Energy, a smart energy management company and turbine manufacturer. In the campaign, an array of turbines within an operating commercial offshore wind farm in China have the normal yaw controller modified to implement wake steering according to a yaw control strategy. The strategy was designed using NREL wind farm models, including a computational fluid dynamics model, Simulator fOr Wind Farm Applications (SOWFA), for understanding wake dynamics and an engineering model, FLOw Redirection and Induction in Steady State (FLORIS), for yaw control optimization. Results indicate that, within the certainty afforded by the data, the wake-steering controller was successful in increasing power capture, by amounts similar to those predicted from the models.
Activation of protein 4.1R exon 16 (E16) inclusion during erythropoiesis represents a physiologically important splicing switch that increases 4.1R affinity for spectrin and actin. Previous studies showed that negative regulation of E16 splicing is mediated by the binding of heterogeneous nuclear ribonucleoprotein (hnRNP) A/B proteins to silencer elements in the exon and that down-regulation of hnRNP A/B proteins in erythroblasts leads to activation of E16 inclusion. This article demonstrates that positive regulation of E16 splicing can be mediated by Fox-2 or Fox-1, two closely related splicing factors that possess identical RNA recognition motifs. SELEX experiments with human Fox-1 revealed highly selective binding to the hexamer UGCAUG. Both Fox-1 and Fox-2 were able to bind the conserved UGCAUG elements in the proximal intron downstream of E16, and both could activate E16 splicing in HeLa cell co-transfection assays in a UGCAUG-dependent manner. Conversely, knockdown of Fox-2 expression, achieved with two different siRNA sequences resulted in decreased E16 splicing. Moreover, immunoblot experiments demonstrate mouse erythroblasts express Fox-2. These findings suggest that Fox-2 is a physiological activator of E16 splicing in differentiating erythroid cells in vivo. Recent experiments show that UGCAUG is present in the proximal intron sequence of many tissue-specific alternative exons, and we propose that the Fox family of splicing enhancers plays an important role in alternative splicing switches during differentiation in metazoan organisms.Alternative splicing of pre-mRNA leads to the synthesis of multiple protein isoforms from a single gene. It is an important mechanism for regulating gene expression and may be utilized by 40 -60% of human genes (1-4). Thus, the estimated 25,000 to 30,000 genes of the human genome can generate a much larger number of proteins. Regulation of alternative splicing occurs in both a tissue-and development-specific manner, resulting in alterations in the structure and function of critical proteins. Altered splicing regulation can also be of widespread importance in the etiology of human disease (5-7).The protein 4.1 gene family serves as an excellent model for investigating the regulation of alternative splicing. The four genes that comprise the family (4 .1R, 4.1G, 4.1B, and 4.1N) display a remarkable array of highly regulated, tissue-specific splicing events. These alternative splicing events facilitate expression of distinct isoforms of 4.1 protein in cells of erythroid, epithelial, neural, and muscle origin (8 -14); thus, they provide opportunities for understanding the mechanisms that regulate alternative splicing in several different cell types. To date, mechanistic studies have focused predominantly on erythroid cells, in which 4.1R protein is a structural component of the erythrocyte plasma membrane and is important for structural integrity and stability of the membrane skeleton. In differentiating erythroid progenitor cells, a dramatic switch in pre-mRNA splicing result...
The statins, hydroxy-3-methylglutaryl-CoA reductase inhibitors that lower serum cholesterol, exhibit myriad clinical benefits, including enhanced vascular integrity. One potential mechanism underlying increased endothelial cell (EC) barrier function is inhibition of geranylgeranylation, a covalent modification enabling translocation of the small GTPases Rho and Rac to the cell membrane. While RhoA inhibition attenuates actin stress fiber formation and promotes EC barrier function, Rac1 inhibition at the cell membrane potentially prevents activation of NADPH oxidase and subsequent generation of superoxides known to induce barrier disruption. We examined the relative regulatory effects of simvastatin on RhoA, Rac1, and NADPH oxidase activities in the context of human pulmonary artery EC barrier protection. Confluent EC treated with simvastatin demonstrated significantly decreased thrombin-induced FITC-dextran permeability, a reflection of vascular integrity, which was linked temporally to simvastatin-mediated actin cytoskeletal rearrangement. Compared with Rho inhibition alone (Y-27632), simvastatin afforded additional protection against thrombin-mediated barrier dysfunction and attenuated LPS-induced EC permeability and superoxide generation. Statin-mediated inhibition of both Rac translocation to the cell membrane and superoxide production were attenuated by geranylgeranyl pyrophosphate (GGPP), indicating that these effects are due to geranylgeranylation inhibition. Finally, thrombininduced EC permeability was modestly attenuated by reduced Rac1 expression (small interfering RNA), whereas these effects were made more pronounced by simvastatin pretreatment. Together, these data suggest EC barrier protection by simvastatin is due to dual inhibitory effects on RhoA and Rac1 as well as the attenuation of superoxide generation by EC NADPH oxidase and contribute to the molecular mechanistic understanding of the modulation of EC barrier properties by simvastatin.
Highly luminescent inks are desirable for various applications such as decorative coating, art painting, and anticounterfeiting, to name a few. However, present inks display low photoluminescent efficiency requiring a strong excitation light to make them glow. Here, we report a highly luminescent ink based on the copper-iodide/1-Propyl-1,4-diazabicyclo[2.2.2]octan-1ium (Cu 4 I 6 (pr-ted) 2 ) hybrid cluster with a quantum efficiency exceeding 98%. Under the interaction between the Cu 4 I 6 (pr-ted) 2 hybrid cluster and polyvinylpyrrolidone (PVP), the highly luminescent Cu 4 I 6 (pr-ted) 2 /PVP ink can be facilely prepared via the onepot solution synthesis. The obtained ink exhibits strong green light emission that originates from the efficient phosphorescence of Cu 4 I 6 (pr-ted) 2 nanocrystals. Attractively, the ink displays high conversion efficiency for the ultraviolet light to bright green light emission due to its wide Stokes shift, implying great potential for anticounterfeiting and luminescent solar concentrator coating.
BackgroundEstrogen receptor (ER), progesterone receptor (PgR), HER2, and Ki67 have been increasingly evaluated by core needle biopsy (CNB) and are recommended for classifying breast cancer into molecular subtypes. However, the concordance rate between CNB and open excision biopsy (OEB) has not been well documented.MethodsPatients with paired CNB and OEB samples from Oct. 2009 to Feb. 2012 in Ruijin Hospital were included. ER, PgR, HER2, and Ki67 were determined by immunohistochemistry (IHC). Patients with HER2 IHC 2+ were further examined by FISH. Cutoff value for Ki67 high expression was 14%. Molecular subtypes were constructed as follows: Luminal A, Luminal B, Triple Negative, and HER2 positive.ResultsThere were 298 invasive breast cancer patients analyzed. Concordance rates for ER, PgR, and HER2 were 93.6%, 85.9%, and 96.3%, respectively. Ki67 expression was slightly higher in OEB than in CNB samples (29.3% vs. 26.8%, P = 0.046). Good agreement (κ = 0.658) was demonstrated in evaluating molecular subtypes between CNB and OEB, with a concordance rate of 77.2%. We also used a different Ki67 cutoff value (20%) for determining Luminal A and B subtypes in HR (hormone receptor) +/HER2- diseases and the overall concordance rate was 79.2%. However, using a cut-point of Ki67 either 14% or 20% for both specimens, there will be about 14% of HR+/HER2- specimens that are called Luminal A on CNB and Luminal B on OEB.ConclusionCNB was accurate in determining ER, PgR, and HER2 status as well as non-Luminal molecular subtypes in invasive breast cancer. Ki67 should be retested on OEB samples in HR+/HER2- patients to accurately distinguish Luminal A from B tumors.
Acute respiratory distress syndrome (ARDS) is a severe clinical condition marked by acute respiratory failure and dysregulated inflammation. Pulmonary vascular endothelial cells (PVEC) function as an important pro-inflammatory source in ARDS, suggesting that modulation of inflammatory events at the endothelial level may have therapeutic benefit. Dipeptidyl peptidase-4 (DPP4) inhibitors, widely used for the treatment of diabetes mellitus, have been reported to have possible anti-inflammatory effects. However, the potential anti-inflammatory effects of DPP4 inhibition on PVEC function and ARDS pathophysiology are unknown. Therefore, we evaluated the effects of sitagliptin, a DPP4 inhibitor in wide clinical use, on LPS-induced lung injury in mice and in human lung endothelial cells (EC) in vitro. In vivo, sitagliptin reduced serum DPP4 activity, BAL protein concentration, cell number and pro-inflammatory cytokine levels, after LPS, and alleviated histological findings of lung injury. LPS decreased the expression levels of CD26/DPP4 on pulmonary epithelial cells and PVEC isolated from mouse lungs, and the effect was partially reversed by sitagliptin. In vitro, human lung microvascular EC (HLMVEC) expressed higher levels of CD26/DPP4 than human pulmonary arterial EC. LPS induced release of TNFα, IL-6, and IL-8 by HLMVEC that was inhibited by sitagliptin. LPS promoted proliferation of HLMVEC, and sitagliptin suppressed this response. However, sitagliptin failed to reverse LPS-induced permeability in cultured EC or lung epithelial cells in vitro. In summary, sitagliptin attenuates LPS-induced lung injury in mice and exerts anti-inflammatory effects on HLMVEC. These novel observations indicate DPP4 inhibitors may have potential as therapeutic drugs for ARDS.
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