3,147 research outputs found
Angiotensin II induces soluble fms-Like tyrosine kinase-1 release via calcineurin signaling pathway in pregnancy
No full textMaternal endothelial dysfunction in preeclampsia is associated with increased soluble fms-like tyrosine kinase-1 (sFlt-1), a circulating antagonist of vascular endothelial growth factor and placental growth factor. Angiotensin II (Ang II) is a potent vasoconstrictor that increases concomitant with sFlt-1 during pregnancy. Therefore, we speculated that Ang II may promote the expression of sFlt-1 in pregnancy. Here we report that infusion of Ang II significantly increases circulating levels of sFlt-1 in pregnant mice, thereby demonstrating that Ang II is a regulator of sFlt-1 secretion in vivo. Furthermore, Ang II stimulated sFlt-1 production in a dose- and time-dependent manner from human villous explants and cultured trophoblasts but not from endothelial cells, suggesting that trophoblasts are the primary source of sFlt-1 during pregnancy. As expected, Ang II-induced sFlt-1 secretion resulted in the inhibition of endothelial cell migration and in vitro tube formation. In vitro and in vivo studies with losartan, small interfering RNA specific for calcineurin and FK506 demonstrated that Ang II-mediated sFlt-1 release was via Ang II type 1 receptor activation and calcineurin signaling, respectively. These findings reveal a previously unrecognized regulatory role for Ang II on sFlt-1 expression in murine and human pregnancy and suggest that elevated sFlt-1 levels in preeclampsia may be caused by a dysregulation of the local renin/angiotensin system
THE ROLE OF ANG/TIE SIGNALING IN LYMPHANGIOGENESIS
No full textThe angiopoietin/Tie system plays a keyrole in remodeling and maturation of bloodvessels as well as lymphatic vessels. The angiopoietinfamily includes four ligands (Ang-1,Ang-2 and Ang-3/4) and two correspondingtyrosine kinase receptors (Tie1 and Tie2). Thebest characterized angiopoietins are Ang-1and Ang-2. Ang-1 acts as an obligate agonistof the Tie2 receptor. Binding of Ang-1 to Tie2induces its autophosphorylation and promotesvascular stability and integrity. Ang-1 induceslymphatic vessel enlargement, sprouting andproliferation in a VEGFR-3-dependentmanner. In contrast, whether Ang-2 isagonistic or antagonistic is dependent on doseand context. Ang-2 modulates angiogenesis ina cooperative manner with another importantangiogenic factor, vascular endothelial growthfactor A. In the presence of VEGF-A, Ang-2promotes vascular sprouting. When in theabsence of VEGF-A, Ang-2 induces vascularregression. However, genetic studies haverevealed that Ang-2-deficient mice exhibitmore severe defects in the lymphatic vasculaturethan in blood vessels. Ang-2 seems to beinvolved in the remodeling and stabilizationof lymphatic vessels. Although the Ang/Tiesystem is essential for blood and lymphaticvessel remodeling and maturation, defining itsprecise role in blood and lymphatic developmenthas been a major challenge. This reviewprovides an update on our current understandingof the angiopoietin/Tie system inlymphangiogenesis
In-situ monitoring of the thermal desorption of alkanethiols with Surface Plasmon Resonance Spectroscopy (SPRS)
No full textThermal desorption investigations on self-assembled monolayers (SAMs) had previously been carried out using techniques such as thermal desorption spectroscopy (TDS), scanning tunneling microscopy (STM) and X-ray photo-electron spectroscopy (XPS). In this paper, the thermal dissociation of alkanethiols (C(n)H(2n+1)SH) at various chain lengths (n = 6, 12, 18) on sputtered gold layers was monitored in-situ using the Kretschmann surface plasmon resonance configuration on a spectroscopic ellipsometer. We found that the longest alkanethiol (C(18)) exhibits the greatest thermal stability, manifested by the least amount of angular shift, during heating, in the resonant spectral features. Predictions of desorption temperatures from SPAS for the longer chain thiols are in good agreement with XPS measurements
Paris / Par Ad. Joanne ; dessiné par Ang.; gravé par F. Lefèvre
No full text1 mapa. Datat entre el 1859 i 1870 aproximadament.27 x 40 c
Paris / Par Ad. Joanne ; dessiné par Ang.; gravé par F. Lefèvre
No full text1 mapa. Datat entre el 1859 i 1870 aproximadament.27 x 40 c
Elucidation of the activity, pharmacokinetics, and toxicity profiles of fasciocidal drug candidates 1,2,4-trioxolane OZ78 and 1,2,4,5-tetraoxane MT04
No full textAbstract: Fascioliasis is a disease belonging to the most neglected tropical diseases. Causative agents are the food borne trematodes Fasciola hepatica and Fasciola gigantica, whereas F. hepatica is especially of human concern. F. hepatica and F. gigantica are setteled in the liver, bile ducts, and gallbladder and are therefore termed as liver flukes.
About 2.4 to 17 million people are infected worldwide with F. hepatica leading to a global burden of about 35’206 disability-adjusted life years (DALYs). Fascioliasis is a disease not only of human concern, but affects also farm animals, especially sheep and cattle.
Triclabendazole is the only known drug active against juvenile and adult F. hepatica infection. There are no alternative drugs available characterized by a similar activity and safety profile as triclabendazole. Resistance against triclabendazole has already been described in sheep and is spreading over the world. However, to date no resistance in humans has been detected. Nonetheless, screening for new drugs is urgent.
OZ78 was originally synthesized for malaria drug discovery and is a peroxidic compound. This synthetic peroxide has shown promising in vitro and in vivo activity against juvenile and adult F. hepatica. Moreover, OZ78 has shown to be active against a triclabendazole resistant F. hepatica strain (Oberon isolate).
In the framework of this PhD thesis preclinical investigations were carried out to further strengthen our knowledge on the potential of the synthetic compounds OZ78 and the related derivative MT04 for the treatment of F. hepatica infection. The aim was to find a lead candidate for further studies and to get a better understanding of the drug properties (pharmacokinetic / pharmacodynamic) of these substances.
Different promising trioxanes and tetraoxanes were tested in vitro and in vivo in more detail with regard to F. hepatica activity and results compared to OZ78 (1,2,4-trioxolane). MT04 (1,2,4,5-tetraoxane) was found to be even more active in vitro and in vivo (in rats) against adult and juvenile F. hepatica infection than OZ78. In contrast to OZ78, MT04 has two peroxide bridges. A dose of 50 mg/kg was enough to reach a complete worm burden reduction of 100% against adult F. hepatica in rats, and for juvenile treatment, a dose of 100 mg/kg was required.
In parallel, OZ78 has been tested in sheep according to activity, tolerability, and pharmacokinetic profiles. OZ78 failed to cure fascioliasis in sheep after an application of 50 mg/kg orally or subcutaneously - no reduction in faecal egg count or worm burden could be measured.
To elucidate pharmacokinetic parameters of OZ78 in sheep plasma, a liquid chromatography mass spectrometry (LC/MS) method was developed and validated in terms of accuracy, precision, stability, and selectivity. OZ78 and OZ352 (Internal standard, I.S.) had to be detected in negative mode, according to the mass of deprotonated parent compounds. Protein precipitation was used for sample clean up, recoveries of three concentrations of OZ78 (5, 1.25, and 0.3125 µg/ml) and I.S. (2 µg/ml) were 84.4%, 69.6%, 78.6% and 85.8%, respectively. Calibration line ranged from 5 µg/ml to 0.15625 µg/ml. The developed method demonstrated to be accurate, precise, and selective.
The pharmacokinetic profile of OZ78 following oral application was characterised by a maximal plasma concentration (Cmax) of 45.8±13 µg/ml after 1 h (Tmax). An estimated elimination half-life (t1/2) of 1.0±0.1 h and a mean area under the plasma time curve (AUC) of 116.2±47 µg h/ml were calculated for the oral administration. On the other hand, following subcutaneous treatment with OZ78, Cmax and Tmax were 13.7±6.1 µg/ml and 0.9±0.4 h, respectively. The α and β half-lives were 4.5±4.3 h and 56.5±36 h, respectively and the mean AUC was 219.1±74 µg h/ml.
Further studies were needed to determine the difference in therapeutic outcomes between rats and sheep. The LC/MS method for OZ78 in sheep plasma was adapted and validated for rat plasma measurements. In addition the OZ78 LC/MS method could be approximated, validated, and used for MT04 measurement in sheep and in rat plasma.
OZ78 and MT04 were applied at a dose of 50 mg/kg to rats, and after different time-points, samples were collected. Studies with MT04 according to efficacy and pharmacokinetics in sheep are outstanding, but are planned.
Following oral administration of 50 mg/kg in rats, the Cmax of MT04 and OZ78 were 49.8±9.1 and 70.1±19.1 µg/ml after 2.7±1.2 h and 1.6±1.6 h (Tmax), respectively. The AUCs were estimated at 31’258.8±6’232.7 and 29’794.1±3’990.6 µg min/ml for MT04 and OZ78, respectively. Mean elimination half-lives (t1/2) of MT04 and OZ78 were 6.4±5.7 h and 2.5±1.5 h, respectively.
Toxicity of OZ78 and MT04 was determined on a liver cell line (HepG2). Different viability and toxicity assays were done with OZ78 or MT04 combined with different iron containing solutions (haemin, Fe(II), and Fe(III)) to get a better understanding of the toxicity profiles of these substances. Reactive oxygen species (ROS) production of MT04 and OZ78 was measured in combination with these iron-solutions. OZ78 was only moderately toxic combined with iron, whereas the combination of MT04 with iron resulted in a significant toxicity. Additionally, a high ROS signal could be measured especially in combination with haemin for both substances.
In summary, OZ78 and MT04 are promising lead candidates for the development of anti-Fasciola drugs. OZ78 and MT04 were active against juvenile and adult infection in rats. The therapy failure in sheep after oral treatment can be explained by a too short exposure of the flukes to the drug. Another application has to be tested in sheep. To date, it is not clear if the applied doses are toxic and, more studies especially in animals are needed. However, ROS formation seems to be responsible for activity of these substances. ---------- Zusammenfassung: Fasziolose ist eine Erkrankung, die zu den vernachlässigten Tropenkrankheiten zählt. Verursacher sind die durch Ernährung übertragbaren Trematoden Fasciola hepatica und Fasciola gigantica, wobei F. hepatica besonders für Menschen Besorgnis erregend ist.
F. hepatica und F. gigantica leben in der Leber, Gallengängen und Gallenblase, und werden deshalb auch als Leberegel bezeichnet.
Etwa 2.4 bis 17 Millionen Menschen sind weltweit mit F. hepatica infiziert, was zu einer weltweiten Bürde von etwa 35'206 behinderungsbedingten Lebensjahren führt. Fasziolose ist nicht nur eine Erkrankung von Menschen, sondern auch landwirtschaftliche Nutztiere wie Schafe und Rinder sind betroffen.
Triclabendazol ist die einzige Substanz, die gegen juvenile und adulte F. hepatica Infektion wirkt. Es gibt keine alternativen Medikamente mit gleicher Aktivität und gleichem Sicherheitsprofil wie Triclabendazol. Resistenzen gegen Triclabendazol sind bereits in Schafen vorhanden und verbreiten sich über die Welt. Bis jetzt sind aber noch keine Resistenzen im Menschen beschriebe worden. Trotzdem ist es wichtig nach neuen Medikamenten zu suchen.
OZ78 wurde ursprünglich für die Malariabehandlung synthetisiert und gehört zu den Peroxiden. Dieses synthetische Peroxid hat versprechende Aktivitäten in vitro und in vivo gegen juvenile und adulte F. hepatica gezeigt. Überdies war OZ78 auch gegen einen Triclabendazol resistenten Stamm (Oberon Isolat) aktiv.
Im Rahmen meiner Dissertation, wurden präklinische Untersuchungen durchgeführt um das Wissen über die versprechenden Substanzen OZ78 und MT04 in der Behandlung von F. hepatica Infektion zu erweitern. Das Ziel war eine neue Leitsubstanz für weitere Studien zu finden und die Wirkweise der Substanzen besser zu verstehen (Pharmakokinetik / Pharmakodynamik).
In einer in vitro und in vivo Studie wurden verschiedene Trioxane und Tetraoxane auf ihre F. hepatica Aktivität im Detail getestet und mit OZ78 (1,2,4-Trioxolan) verglichen. Aus dieser Studie kam hervor, dass MT04 (1,2,4,5-Tetraoxan) noch aktiver in vitro und in vivo (in Ratten) gegen adulte und juvenile F. hepatica Infektionen war als OZ78. Im Unterschied zu OZ78, hat MT04 zwei Peroxid-Brücken. Eine Dosis von 50 mg/kg genügte um eine komplette Reduktion der Wurmbelastung von 100% gegen adulte F. hepatica in Ratten zu erreichen, für eine juvenile Behandlung war eine Dosis von 100 mg/kg nötig.
Parallel wurde OZ78 im Schaf getestet und Aktivität, Toleranz und Pharmakokinetik untersucht. Aber OZ78 schaffte es nicht Fasziolose im Schaf nach einer Anwendung von 50 mg/kg OZ78 oral oder subkutan zu heilen, keine Reduktion von Stuhleiern oder Würmern konnte gemessen werden.
Um die pharmakokinetischen Parameter von OZ78 im Plasma zu bestimmen, musste eine Flüssigchromatographische Massenspektrometrische (LC/MS) Methode entwickelt und auf Genauigkeit, Präzision, Stabilität, und Selektivität validiert werden. OZ78 und OZ352 (interner Standard, I.S.) musste im Negativ-Modus nach der Masse der deprotonierten Muttersubstanz detektiert werden. Eine Proteinfällung wurde zur Probenvorbereitung angewendet. Die Wiederfindungen von OZ78 (5, 1.25, und 0.3125 µg/ml) und von I.S. (2 µg/ml) waren 84.4%, 69.6%, 78.6%, und 85.8%. Kalibrationskurve reichte von 5 µg/ml zu 0.15625 µg/ml. Die entwickelte Methode war genau, präzis, und selektiv.
Das Pharmakokinetik-Profil von OZ78 nach oraler Injektion zeigte eine maximale Plasma Konzentration von (Cmax) 45.8±13 µg/ml nach 1 h (Tmax) auf. Die Halbwertszeit (t1/2) betrug 1.0±0.1 h und eine mittlere Fläche unter der Plasma Zeit Kurve (AUC) von 116.2±47 µg h/ml wurde für die orale Anwendung berechnet. Auf der anderen Seite, nach subkutaner Injektion mit OZ78, wurden ein Cmax und ein Tmax von 13.7±6.1 µg/ml und 0.9±0.4 h berechnet. Die α und β Halbwertszeit waren 4.5±4.3 h und 56.5±36 h, und die mittlere AUC war 219.1±74 µg h/ml.
Weitere Studien waren nötig um den Unterschied im Behandlungsergebnis zwischen Ratte und Schaf zu erklären. Die LC/MS Methode von OZ78 für Schafplasma wurde für Rattenplasma angepasst und validiert. Zusätzlich konnte die OZ78 LC/MS Methode modifiziert, validiert und für MT04 Messung in Schaf und Ratten Plasma verwendet werden.
OZ78 und MT04 wurden in einer Dosis von 50 mg/kg Ratten verabreicht und nach verschiedenen Zeitpunkten wurden Samples gezogen. Studien mit MT04 betreffend Aktivität und Pharmakokinetik im Schaf sind noch ausstehend, sind aber in Planung.
Nach oraler Applikation von 50 mg/kg in Ratten, wurden Cmax Werte von MT04 und OZ78 49.8±9.1 und 70.1±19.1 µg/ml nach 2.7 h±1.2 und 1.6±1.6 h (Tmax) gemessen. Die berechnete AUC ergab 31’258.8±6’232.7 und 29’794.1±3’990.6 µg min/ml für MT04 und OZ78. Mittlere Halbwertszeiten (t1/2) für MT04 und OZ78 waren 6.4 ±5.7 h und 2.5±1.5 h.
Zuletzt wurde die Toxizität von OZ78 und MT04 mit einer Leberzelllinie (HegG2) bestimmt. Verschiedene Toxizitäts- und Viabilitäts Assays wurden mit OZ78 oder MT04 kombiniert mit verschiedenen Einsenlösungen (Hämin, Fe(II) und Fe(III)) angewandt um das Toxizitätsprofil besser zu verstehen. Reaktive Sauerstoffspezies (ROS) Produktion von MT04 und OZ78 wurden in Kombination mit diesen Eisenlösungen gemessen. OZ78 war nur mässig toxisch kombiniert mit Eisen, wobei eine signifikante Toxizität für MT04 in Kombination mit Eisen gefunden werden konnte. Ein hohes ROS Signal wurde für beide Substanzen vor allem in Kombination mit Haemin gemessen.
Zusammengefasst, OZ78 und MT04 sind vielversprechende „Lead“-Kandidaten in der Entwicklung eines Medikaments gegen Fasziola Infektionen. OZ78 und MT04 waren aktiv gegen juvenile und adulte Infektion in der Ratte. Das Therapieversagen in Schafen kann durch eine zu geringe Exposition der Saugwürmer im Schaf mit der Substanz erklärt werden. Eine andere Applikation im Schaf ist noch ausstehend. Bis jetzt lässt sich nicht sagen, wie toxisch die verwendeten Dosen sind, mehr Studien im Tier werden dafür benötigt. Die Aktivität der Substanzen scheint von der ROS Produktion abhängig zu sein
Darboux's Formula with Integral Remainder of Functions with Two Independent Variables
No full textIn the article, the noted Darboux’s formula of functions with single variable is generalized to that of functions of two independent variables with integral remainder, some important special cases of Darboux’s formula of functions with two variables are obtained, and some estimates of the integral remainders and Darboux’s expansion of the function ln(x + y) are given. These results generalize A. Sard’s formula in numerical integration
Correlation of Ang-2 levels with clinical and vascular parameters.
No full text<p>Ang-2 levels in dialysis individuals correlated positively with time on dialysis (A), serum urate levels (B), systolic blood pressure SDS (C) and cIMT (D). Independent variables are shown on the x-axis. Regression lines account for dialysis patients only. Dotted line in D indicates the value for cIMT in healthy age-matched controls. There was no correlation between Ang-2 and any clinical and vascular measures in pre-dialysis CKD patients.</p
Hemodynamic regulation of metalloendopeptidases EC3.4; 24.15 and EC3.4; 24.16: expression and function in the vascular endothelium
No full textHemodynamic forces, namely shear stress and cyclic strain, have been well characterised as modulators of vascular endothelial function, and have been assigned an important role in the maintainence of vascular tone, haemostasis, and regulation of vascular growth and health. They exert their influence in part by effecting changes in the production and release of vasoactive compounds by the endothelium, and by effecting changes in the levels and activity of various enzymes. Thimet oligopeptidase (EC3.4.24.15, EP24.15) and neurolysin (EC3.4.24.16, EP24.15) are closely related zinc metalloendopeptidases that have been shown to be expressed and active in the vascular endothelium. Their substrates include the vasoactive peptides bradykinin and angiotensin I, which have been identified as important regulators of both blood pressure and angiogenic processes. Other related peptidases, namely endothelin converting enzyme (ECE) and angiotensin converting enzyme (ACE), have been shown to be regulated by hemodynamic forces in the vascular endothelium. As such EP24.15 and EP24.16 are likely candidates for regulation by hemodynamic forces.
In this regard we have investigated the effect of cyclic strain on the expression and activty of EP24.15 and EP24.16 in cultured bovine aortic endothelial cells (BAECs). We have shown that exposure to cyclic strain significantly increases the mRNA expression as well as both the cellular and secreted activity of both enzymes. We have demonstrated that up-regulation of both enzymes is dependent on Gi-protein mediated signalling, although with varying Gia/G(3y subunit specificity for either enzyme. Using immunocytochemistry, we have also demonstrated a strain-dependent increase in EP24.15 protein expression within the nucleus and cytoplasm in parallel with an increase in membrane associated EP 24.15
The effects of strain on the ability of BAECs in culture to cleave both Ang I, and BK in an EC24.15/EC24.16 specific manner was also studied. We observed that exposure to cyclic strain induces a significant increase in the EP24.15 specific hydrolysis of both exogenously added BK and Ang I.
The potential of the observed effects of cyclic strain on EP24.15 to effect changes in endothelial cell function were also examined. Use of the dual EP24.15/EP24.16 inhibitor, cFP-AAF-pAB, and the EP24.15 specific antisense, FLIP, was seen to significantly attenuate cyclic strain-induced endothelial cell tubule formation and migration. We also found that the effects of FLIP transfection on cyclic strain-induced endothelial cell tubule formation could be largely reversed by addition of exogenous Ang-(l-7). Taken together these results suggest that strain-induced endothelial cell angiogenesis and migration putatively involves EP24.15 cleavage of vasoactive peptide substrates
Proliferative synergy of ANG II and EGF in porcine aortic vascular smooth muscle cells
No full text To test growth effects of angiotensin II (ANG II) in porcine vascular smooth muscle cells (VSMC) and potential ANG II synergy with epidermal growth factor (EGF), we exposed subconfluent, near-quiescent porcine aortic VSMC to ANG II, EGF, or ANG II + EGF (each 10(-9) M) in Dulbecco's modified Eagle's-Ham's F-12 medium with insulin + 0.4% fetal calf serum (FCS) selected for minimal ANG II-degrading capacity. Cell number and DNA and protein synthesis (by [3H]-thymidine and [35S]methionine incorporation, respectively) were determined serially over 1-6 days. ANG II alone induced an early 20% increase and then a plateau in cell number over the 0.4% FCS control (P < 0.01; n = 8), thus without sustained increase in proliferation rate. Yet ANG II alone did not increase fractional DNA or protein synthesis (each as cpm/10(3) cells) and, by flow cytometry, reduced S phase fraction without increase in cell size. EGF alone induced brisk DNA synthesis yet minimal cell division over days 0-4, thus late-cycle arrest. ANG II + EGF, despite no increase in fractional DNA or protein synthesis rates over EGF alone, induced significant indomethacin-resistant dose-dependent (P < 0.001) increase in cell proliferation rate over EGF alone with a median effective dose of 5 x 10(-10) M ANG II, thus proliferative synergy. We propose that 1) ANG II induces a subpopulation of cells arrested in or beyond S phase to proceed through mitosis but does not influence G1 traversal or S phase entry and 2) ANG II + EGF achieve proliferative synergy by complementary actions at sequential cell cycle loci, with EGF supporting progression from G0/G1 to S phase and ANG II inducing completion of mitosis by cells already in or beyond S phase ("late-cycle completion"). </jats:p
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