Antiangiogenic effect of soluble vascular endothelial growth factor receptor-1 in placental angiogenesis

Differential splicing of the flt-1 mRNA generates soluble variant of vascular endothelial growth factor (VEGF) receptor-1 (sVEGFR-1, also known as sFlt-1). The action of VEGF is antagonized by sVEGFR-1. Soluble VEGFR-1 binds to VEGF with a high affinity and therefore works to modulate VEGF and VEGF signaling pathway. In this study, the authors tested the hypothesis that VEGF-mediated endothelial cell angiogenesis is tightly modulated by the release of sVEGFR-1 and placental expression of sVEGFR-1 is upregulated by hypoxia. Immunolocalization studies showed progressively intense staining for sVEGFR-1 and VEGF in the trophoblast of placental villous explants throughout gestation. Endothelial cell migration studies using a modified Boyden's chamber showed a significant increase in cell migration in response to VEGF that was significantly attenuated in the presence of exogenous sVEGFR-1. Furthermore, stimulation of endothelial cells with VEGF led to a dose-dependent increase in the release of sVEGFR-1 as determined by enzyme-linked immunosorbent assay (ELISA). Exposure of normal placental villous explants to hypoxia (1% pO2) increased trophoblast expression of sVEGFR-1 when compared with tissue normoxia (5% pO2). In addition, conditioned media from hypoxia treated placental villous explants induced a significant increase in endothelial cell migration that was significantly reduced in presence of sVEGFR-1. Our study demonstrates that hypoxia positively regulates sVEGFR-1 protein expression in ex vivo trophoblasts, which control VEGF-driven angiogenesis.

Resveratrol inhibits the release of soluble fms-like tyrosine kinase (sFlt-1) from human placenta

Objective - Soluble vascular endothelial growth factor receptor–1 (also know as soluble fms-like tyrosine kinase [sFlt]-1) is a key causative factor of preeclampsia. Resveratrol, a plant phytoalexin, has antiinflammatory and cardioprotective properties. We sought to determine the effect of resveratrol on sFlt-1 release. Study Design - Human umbilical vein endothelial cells, transformed human trophoblast-8 (HTR/SVneo)-8/SVneo trophoblast cells, or placental explants were incubated with cytokines and/or resveratrol. Conditioned media were assayed for sFlt-1 by enzyme-linked immunosorbent assay and cell proteins used for Western blotting. Results - Resveratrol inhibited cytokine-induced release of sFlt-1 from normal placental explants and from preeclamptic placental explants. Preincubation of human umbilical vein endothelial cells or HTR-8/SVneo cells with resveratrol abrogated sFlt-1 release. Resveratrol prevented the up-regulation of early growth response protein-1 (Egr-1), a transcription factor necessary for induction of the vascular endothelial growth factor receptor–1 gene and caused up-regulation of heme oxygenase–1, a cytoprotective enzyme found to be dysfunctional in preeclampsia. Conclusion - In summary, resveratrol can inhibit sFlt-1 release and up-regulate heme oxygenase–1; thus, may offer therapeutic potential in preeclampsia.

Heparin elevates circulating soluble fms-like tyrosine kinase-1 immunoreactivity in pregnant women receiving anticoagulation therapy

Background—Alterations in circulating levels of pro- and antiangiogenic factors have been associated with adverse pregnancy outcomes. Heparin is routinely administered to pregnant women, but without clear knowledge of its impact on these factors. Methods and Results—We conducted a longitudinal study of 42 pregnant women. Twenty-one women received prophylactic heparin anticoagulation, and 21 healthy pregnant women served as controls. Compared with gestational age-matched controls, heparin treatment was associated with increased circulating levels of soluble fms-like tyrosine kinase-1 (sFlt-1) in the third trimester (P<0.05), in the absence of preeclampsia, placental abruption, or fetal growth restriction. Heparin had no effect on circulating levels of vascular endothelial growth factor, placenta growth factor, or soluble endoglin as assessed by ELISA. In vitro, low-molecular weight and unfractionated heparins stimulated sFlt-1 release from placental villous explants, in a dose- and time-dependent manner. This effect was not due to placental apoptosis, necrosis, alteration in protein secretion, or increased transcription. Western blot analysis demonstrated that heparin induced shedding of the N-terminus of Flt-1 both in vivo and in vitro as indicated by a predominant band of 100–112 kDa. By using an in vitro angiogenesis assay, we demonstrated that serum of heparin-treated cases inhibited both basal and vascular endothelial growth factor-induced capillary-like tube formation. Conclusions—Heparin likely increases the maternal sFlt-1 through shedding of the extracellular domain of Flt-1 receptor. Our results imply that upregulation of circulating sFlt-1 immunoreactivity in pregnancy is not always associated with adverse outcomes, and that heparin's protective effects, if any, cannot be explained by promotion of angiogenesis.

Upregulation of urotensin II receptor in preeclampsia causes in vitro placental release of soluble vascular endothelial growth factor receptor 1 in hypoxia

Preeclampsia is a hypertensive disorder of pregnancy caused by abnormal placental function, partly because of chronic hypoxia at the utero-placental junction. The increase in levels of soluble vascular endothelial growth factor receptor 1, an antiangiogenic agent known to inhibit placental vascularization, is an important cellular factor implicated in the onset of preeclampsia. We investigated the ligand urotensin II (U-II), a potent endogenous vasoconstrictor and proangiogenic agent, for which levels have been reported to increase in patients with preeclampsia. We hypothesized that an increased sensitivity to U-II in preeclampsia might be achieved by upregulation of placental U-II receptors. We further investigated the role of U-II receptor stimulation on soluble vascular endothelial growth factor receptor 1 release in placental explants from diseased and normal patients. Immunohistochemistry, real-time PCR, and Western blotting analysis revealed that U-II receptor expression was significantly upregulated in preeclampsia placentas compared with controls (P<0.01). Cellular models of syncytiotrophoblast and vascular endothelial cells subjected to hypoxic conditions revealed an increase in U-II receptor levels in the syncytiotrophoblast model. This induction is regulated by the transcriptional activator hypoxia-inducible factor 1a. U-II treatment is associated with increased secretion of soluble vascular endothelial growth factor receptor 1 only in preeclamptic placental explants under hypoxia but not in control conditions. Interestingly, normal placental explants did not respond to U-II stimulation.

Activation of proteinase-activated receptor 2 stimulates soluble vascular endothelial growth factor receptor 1 release via epidermal growth factor receptor transactivation in endothelial cells

The proteinase-activated receptor 2 (PAR-2) expression is increased in endothelial cells derived from women with preeclampsia, characterized by widespread maternal endothelial damage, which occurs as a consequence of elevated soluble vascular endothelial growth factor receptor-1 (sVEGFR-1; commonly known as sFlt-1) in the maternal circulation. Because PAR-2 is upregulated by proinflammatory cytokines and activated by blood coagulation serine proteinases, we investigated whether activation of PAR-2 contributed to sVEGFR-1 release. PAR-2–activating peptides (SLIGRL-NH2 and 2-furoyl-LIGRLO-NH2) and factor Xa increased the expression and release of sVEGFR-1 from human umbilical vein endothelial cells. Enzyme-specific, dominant-negative mutants and small interfering RNA were used to demonstrate that PAR-2–mediated sVEGFR-1 release depended on protein kinase C-ß1 and protein kinase C-e, which required intracellular transactivation of epidermal growth factor receptor 1, leading to mitogen-activated protein kinase activation. Overexpression of heme oxygenase 1 and its gaseous product, carbon monoxide, decreased PAR-2–stimulated sVEGFR-1 release from human umbilical vein endothelial cells. Simvastatin, which upregulates heme oxygenase 1, also suppressed PAR-2–mediated sVEGFR-1 release. These results show that endothelial PAR-2 activation leading to increased sVEGFR-1 release may contribute to the maternal vascular dysfunction observed in preeclampsia and highlights the PAR-2 pathway as a potential therapeutic target for the treatment of preeclampsia.

Autoantibody from women with preeclampsia induces soluble Fms-like tyrosine kinase-1 production via angiotensin type 1 receptor and calcineurin/nuclear factor of activated T-cells signaling

Preeclampsia is a pregnancy-specific hypertensive syndrome that causes substantial maternal and fetal morbidity and mortality. Recent evidence indicates that maternal endothelial dysfunction in preeclampsia results from increased soluble Fms-like tyrosine kinase-1 (sFlt-1), a circulating antiangiogenic protein. Factors responsible for excessive production of sFlt-1 in preeclampsia have not been identified. We tested the hypothesis that angiotensin II type 1 (AT1) receptor activating autoantibodies, which occur in women with preeclampsia, contribute to increased production of sFlt-1. IgG from women with preeclampsia stimulates the synthesis and secretion of sFlt-1 via AT1 receptor activation in pregnant mice, human placental villous explants, and human trophoblast cells. Using FK506 or short-interfering RNA targeted to the calcineurin catalytic subunit mRNA, we determined that calcineurin/nuclear factor of activated T-cells signaling functions downstream of the AT1 receptor to induce sFlt-1 synthesis and secretion by AT1-receptor activating autoantibodies. AT1-receptor activating autoantibody–induced sFlt-1 secretion resulted in inhibition of endothelial cell migration and capillary tube formation in vitro. Overall, our studies demonstrate that an autoantibody from women with preeclampsia induces sFlt-1 production via angiotensin receptor activation and downstream calcineurin/nuclear factor of activated T-cells signaling. These autoantibodies represent potentially important targets for diagnosis and therapeutic intervention.

Glutaredoxin-1 up-regulation induces soluble vascular endothelial growth factor receptor 1, attenuating post-ischemia limb revascularization

Glutaredoxin-1 (Glrx) is a cytosolic enzyme that regulates diverse cellular function by removal of GSH adducts from S-glutathionylated proteins including signaling molecules and transcription factors. Glrx is up-regulated during inflammation and diabetes. Glrx overexpression inhibits VEGF-induced endothelial cell (EC) migration. The aim was to investigate the role of up-regulated Glrx in EC angiogenic capacities and in vivo revascularization in the setting of hind limb ischemia. Glrx overexpressing EC from Glrx transgenic mice (TG) showed impaired migration and network formation and secreted higher level of soluble VEGF receptor 1 (sFlt), an antagonizing factor to VEGF. After hind limb ischemia surgery Glrx TG mice demonstrated impaired blood flow recovery, associated with lower capillary density and poorer limb motor function compared to wild type littermates. There were also higher levels of anti-angiogenic sFlt expression in the muscle and plasma of Glrx TG mice after surgery. Non-canonical Wnt5a is known to induce sFlt. Wnt5a was highly expressed in ischemic muscles and EC from Glrx TG mice, and exogenous Wnt5a induced sFlt expression and inhibited network formation in human microvascular EC. Adenoviral Glrx-induced sFlt in EC was inhibited by a competitive Wnt5a inhibitor. Furthermore, Glrx overexpression removed GSH adducts on p65 in ischemic muscle and EC, and enhanced nuclear factor kappa B (NF-kB) activity which was responsible for Wnt5a-sFlt induction. Taken together, up-regulated Glrx induces sFlt in EC via NF-kB -dependent Wnt5a, resulting in attenuated revascularization in hind limb ischemia. The Glrx-induced sFlt may be a part of mechanism of redox regulated VEGF signaling.

Microfluidic-controlled manufacture of liposomes for the solubilisation of a poorly water soluble drug

Besides their well-described use as delivery systems for water-soluble drugs, liposomes have the ability to act as a solubilizing agent for drugs with low aqueous solubility. However, a key limitation in exploiting liposome technology is the availability of scalable, low-cost production methods for the preparation of liposomes. Here we describe a new method, using microfluidics, to prepare liposomal solubilising systems which can incorporate low solubility drugs (in this case propofol). The setup, based on a chaotic advection micromixer, showed high drug loading (41 mol%) of propofol as well as the ability to manufacture vesicles with at prescribed sizes (between 50 and 450 nm) in a high-throughput setting. Our results demonstrate the ability of merging liposome manufacturing and drug encapsulation in a single process step, leading to an overall reduced process time. These studies emphasise the flexibility and ease of applying lab-on-a-chip microfluidics for the solubilisation of poorly water-soluble drugs.

The blurring boundaries between synchronicity and asynchronicity:new communicative situations in work-related Instant Messaging

Instant messaging is one of the most popular communication technologies in virtual teams, enabling interactions to intertwine whole working days, thus creating the sense of copresence for team members who are geographically dispersed. Through close linguistic analyses of naturally occurring data from a virtual team, this article discusses the implications of two novel communicative situations enabled by instant messaging: presence information and the persistence of transcript. The preliminary findings of this study indicate that these new communicative situations require the flouting or rethinking of previously existing interactional norms and that communicative practices employed by the team members are not yet conventionalized/normalized, the expectations and interpretations of interactional rituals and timing vary highly, even within the same virtual team.

Serum procalcitonin, interleukin-6, soluble intercellular adhesin molecule-1 and IgG to short-chain exocellular lipoteichoic acid as predictors of infection in total joint prosthesis revision

The diagnosis of prosthetic joint infection and its differentiation from aseptic loosening remains problematic. The definitive laboratory diagnostic test is the recovery of identical infectious agents from multiple intraoperative tissue samples; however, interpretation of positive cultures is often complex as infection is frequently associated with low numbers of commensal microorganisms, in particular the coagulase-negative staphylococci (CNS). In this investigation, the value of serum procalcitonin (PCT), interleukin-6 (IL-6) and soluble intercellular adhesion molecule-1 (sICAM-1) as predictors of infection in revision hip replacement surgery is assessed. Furthermore, the diagnostic value of serum IgG to short-chain exocellular lipoteichoic acid (sce-LTA) is assessed in patients with infection due to CNS. Presurgical levels of conventional serum markers of infection including C-reactive protein (CRP), erythrocyte sedimentation rate (ESR) and white blood cell count (WBC) is also established. Forty-six patients undergoing revision hip surgery were recruited with a presumptive clinical diagnosis of either septic (16 patients) or aseptic loosening (30 patients). The diagnosis was confirmed microbiologically and levels of serum markers were determined. Serum levels of IL-6 and sICAM-1 were significantly raised in patients with septic loosening (P=0.001 and P=0.0002, respectively). Serum IgG to sce-LTA was elevated in three out of four patients with infection due to CNS. In contrast, PCT was not found to be of value in differentiating septic and aseptic loosening. Furthermore, CRP, ESR and WBC were significantly higher (P=0.0001, P=0.0001 and P=0.003, respectively) in patients with septic loosening. Serum levels of IL-6, sICAM-1 and IgG to sce-LTA may provide additional information to facilitate the diagnosis of prosthetic joint infection.

World coffee markets and national coffee parastatals since 1989: A comparative study of Colombia and Cote d'Ivoire

The International Coffee Agreements (ICA) involved the majority of nations producing and consuming coffee and provided relative economic stability to the coffee sectors of the exporting Third World countries. This study focuses on the serious impact of the 1989 collapse of the ICA on the domestic coffee sectors of Colombia and Côte d'Ivoire. In particular, the dissertation examines the role of the Colombian and Ivoirian coffee parastatals, the Federacion Nacional de Cafeteros de Colombia and the Caisse de Stabilisation et Soutien des Prix des Produits Agricoles, during the crisis and their transformation by it. ^ The theoretical framework employed in this study is borrowed from the literature on state-society relations. The methodology includes: in-depth analysis of the historical roles of the parastatal agencies in coffee production, state-society relations and economic development in Colombia and Côte d'Ivoire; interviews with parastatal administrators, producers and other knowledgeable informants in both countries; and a comprehensive review of newspaper articles and official statements of coffee policy published in Colombia and Côte d'Ivoire. prior to, during, and after the crisis. ^ The Colombian and Ivoirian coffee sectors and their producers faced serious economic and social problems following the drop in coffee prices. The coffee parastatals in Colombia and Côte d'lvoire first lost some of their responsibilities following the world coffee crisis. The Caisse was in the end eliminated while FEDECAFE struggled to remain in existence. Along the way, both entities faced protests from disgruntled coffee producers, who organized politically for the first time in their nations' histories. I argue that the outcome for the parastatals depended in part on the conditions of their formation, particularly the level of societal involvement in their creation. I also posit that the country's dependence on foreign aid played a key role in the fate of the parastatals. ^ This dissertation concludes that developments in the Colombian and Ivoirian coffee sectors have significantly contributed to the creation of the difficult political and economic conditions of both countries today. ^

Sugar and coffee: A history of settler agriculture in nineteenth-century Liberia

This dissertation is about commercial agriculture in nineteenth-century Liberia. Based primarily on the archives of the American Colonization Society (founder of Liberia), it examines the impact of environmental and demographic constraints on an agrarian settler society from 1822 to the 1890s. Contrary to the standard interpretation, which linked the poor state of commercial agriculture to the settlers' disdain for cultivation, this dissertation argues that the scarcity of labor and capital impeded the growth of commercial agriculture. The causes of the scarcity were high mortality, low immigration and the poverty of the American “Negroes” who began to settle Liberia in 1822. ^ Emigration to Liberia meant almost certain death and affliction for many immigrants because they encountered a new set of diseases. Mortality was particularly high during the early decades of colonization. From 1822 to 1843, about 48 percent of all immigrants died of various causes, usually within their first year. The bulk of the deaths is attributed to malaria. There was no natural increase in the population for this early period and because American “Negroes” were unenthusiastic about relocation to Liberia, immigration remained sparse throughout the century. Low immigration, combined with the high death rate, deprived the fledgling colony of its potential human resource, especially for the cultivation of labor-intensive crops, like sugar cane and coffee. Moreover, even though females constituted approximately half of the settlers, they seldom performed agricultural labor. ^ The problem of labor was compounded by the scarcity of draft animals. Liberia is in the region where trypanosomiasis occurs. The disease is fatal to large livestock. Therefore, animal-drawn plows, common in the United States, were never successfully transplanted in Liberia. Besides, the dearth of livestock obstructed the development of the sugar industry since many planters depended on oxen-powered mills because they could not afford to buy the more expensive steam engine mills. ^ Finally, nearly half of the immigrants were newly emancipated slaves. Usually these former bondsmen arrived in Liberia penniless. Consequently, they lacked the capital to invest in large-scale plantations. The other categories of immigrants (e.g., those who purchased their freedom), were hardly better off than the emancipated slaves. ^

Small farmer market knowledge and specialty coffee commodity chains in Western Highlands Guatemala

For producers motivated by their new status as self-employed, landowning, capitalist coffee growers, specialty coffee presents an opportunity to proactively change the way they participate in the international market. Now responsible for determining their own path, many producers have jumped at the chance to enhance the value of their product and participate in the new "fair trade" market. But recent trends in the international coffee price have led many producers to wonder why their efforts to produce a certified Fair Trade and organic product are not generating the price advantage they had anticipated. My study incorporates data collected in eighteen months of fieldwork, including more than 45 interviews with coffee producers and fair trade roasters, 90 surveys of coffee growers, and ongoing participant observation to understand how fair trade certification, as both a market system and development program, meets the expectations of the coffee growers. By comparing three coffee cooperatives that have engaged the Fair Trade system to disparate ends, the results of this investigation are three case studies that demonstrate how global processes of certification, commodity trade, market interaction, and development aid effect social and cultural change within communities. This study frames several lessons learned in terms of (1) socioeconomic impacts of fair trade, (2) characteristics associated with positive development encounters, and (3) potential for commodity producers to capture value further along their global value chain. Commodity chain comparisons indicate the Fair Trade certified cooperative receives the highest per-pound price, though these findings are complicated by costs associate with certification and producers' perceptions of an "unjust" system. Fair trade-supported projects are demonstrated as more "successful" in the eyes of recipients, though their attention to detail can just as easily result in "failure". Finally, survey results reveal just how limited is the market knowledge of producers in each cooperative, though fair trade does, in fact, provide a rare opportunity for producers to learn about consumer demand for coffee quality. Though bittersweet, the fair trade experiences described here present a learning opportunity for a wide range of audiences, from the certified to the certifiers to the concerned public and conscientious consumer.

Beginning to Explore Dashboard Dining

It can be nutritious and healthy if done right. Fruits and vegetables, a granola bar, smoothie, or some fresh squeezed Florida orange juice would be good choices. On the other hand, it can poison you. Perishable protein and dairy products must be packed in a well- insulated cooler with plenty of ice and a refrigerator thermometer kept inside to en-sure the food stays below 40 degrees Fahrenheit. If you are not completely safe, it can kill you. According to Hagerty Insurance of Traverse City, Michigan, the top ten worst foods to consume are coffee, hot soups, tacos, chili, juicy hamburgers, fried chicken, any barbecued food, filled doughnuts, soft drinks, and chocolate. (see Lisa Chin, 2003) It simply takes a sudden scalding spill, an unexpected splash, or dripping condiments, any of which demand your immediate attention, to become an instant fatality.