Pharmacokinetics and Pharmacodynamics of an Oral Formulation of Apixaban in Horses After Oral and Intravenous Administration

Horses with inflammatory and infectious disorders are often treated with injectable heparin anticoagulants to prevent thrombotic complications. In humans, a new class of direct oral acting anticoagulants (DOAC) appear as effective as heparin, while eliminating the need for daily injections. Our study in horses evaluated apixaban, a newly approved DOAC for human thromboprophylaxis targeting activated factor X (Xa). Our goals were to: (1) Determine pharmacokinetics and pharmacodynamics of apixaban after oral (PO) and intravenous (IV) administration in horses; (2) Detect any inhibitory effects of apixaban on ex vivo Equid herpesvirus type 1 (EHV-1)-induced platelet activation, and (3) Compare an anti-Xa bioactivity assay with ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) for measuring apixaban concentrations. In a blinded placebo-controlled cross-over study, five horses received a single dose (0.2 mg/kg) of apixaban or placebo PO or IV. Blood was collected before and at 3 (IV) or 15 (PO) min, 30 and 45 min, and 1, 2, 3, 4, 6, 8, and 24 h after dosing for measuring apixaban UPLC-MS concentrations and anti-Xa activity. Pharmacodynamic response was measured in a dilute prothrombin time (dPT) assay. Flow cytometric EHV-1-induced platelet P-selectin expression and clinical pathologic safety testing were performed at baseline, 2 and 24 h and baseline and 24 h, respectively. We found no detectable apixaban in plasma PO administration. After IV administration, plasma apixaban levels followed a two-compartment model, with concentrations peaking at 3 min and decreasing to undetectable levels by 8 h. The elimination half-life was 1.3 ± 0.2 h, with high protein binding (92–99%). The dPT showed no relationship to apixaban UPLC-MS concentration and apixaban did not inhibit EHV-1-induced platelet activation after IV dosing. Apixaban anti-Xa activity showed excellent correlation to UPLC-MS (r2 = 0.9997). Our results demonstrate that apixaban has no apparent clinical utility as an anticoagulant for horses due to poor oral availability

Physiology and pathophysiology of vascular signaling controlled by guanosine 3',5'-cyclic monophosphate-dependent protein kinase.

Recent medical advances suggest that the cellular natriuretic peptide/cGMP and NO/cGMP effector systems represent important signal transduction pathways especially in the cardiovascular system. These pathways also appear to be very interesting targets for the possible prevention of cardiovascular diseases. Exciting candidates for prevention include cGMP-dependent signaling networks initiated by natriuretic peptides (NP) and nitric oxide (NO) which are currently explored for their diagnostic and therapeutic potential. cGMP signaling contributes to the function and interaction of several vascular cell types, and its dysfunction is involved in the progression of major cardiovascular diseases such as atherosclerosis, hypertension and diabetic complications. This review will take a focussed look at key elements of the cGMP signaling cascade in vascular tissue. Recent advances in our knowledge of cGMP-dependent protein kinases (cGK, also known as PKG), the potential for assessing the functional status of cGMP signaling and the possible cross talk with insulin signaling will be reviewed

Immature platelets in COVID-19

Platelets play a critical role in immune response. Coronavirus disease 2019 (COVID-19) patients with a severe course often show pathological coagulation parameters including thrombocytopenia, and at the same time the proportion of immature platelets increases. In this study, the platelet count and the immature platelet fraction (IPF) of hospitalized patients with different oxygenation requirements was investigated daily over a course of 40 days. In addition, the platelet function of COVID-19 patients was analyzed. It was found that the number of platelets in patients with the most severe course (intubation and extracorporeal membrane oxygenation (ECMO)) was significantly lower (111.5 ∙ 106 /mL) than in the other groups (mild (no intubation, no ECMO): 203.5 ∙ 106 /mL, p < .0001, moderate (intubation, no ECMO): 208.0 ∙ 106 /mL, p < .0001). IPF tended to be elevated (10.9%). Platelet function was reduced. Differentiation by outcome revealed that the deceased patients had a highly significant lower platelet count and higher IPF (97.3 ∙ 106 /mL, p < .0001, 12.2%, p = .0003)

Funktionsanalysen der AAA Peroxine Pex1p und Pex6p aus Saccharomyces cerevisiae

Zum Beginn dieser Arbeit waren die beiden AAA-Proteine der Hefe S cerevisiae, Pex1p und Pex6p als Peroxine identifiziert. Ihre intrazelluläre Lokalisation wird unterschiedlich diskutiert. Dementsprechend wurde in der Arbeit die intrazellulären Lokalisation von Pex1p und Pex6p untersucht. Um einen Einblick in die Funktion der AAA-Peroxine zu gewinnen, wurden in Anlehnung an Arbeiten mit NSF, die Auswirkung von Mutationen in den WalkerA bzw. WalkerB-Motiven der ersten bzw. zweiten Domäne beider AAA-Peroxine untersucht. Bei der Suche nach weitere Interaktionspartner der AAA-Peroxine, konnte mit Pex15p auch ein zweiter Bindungspartner (neben der bekannten Interaktion der beiden Peroxine miteinander) unter den Peroxinen von S cerevisiae für Pex6p gefunden werden. Die Interaktionspartner wurden dann in die Untersuchungen bezüglich Lokalisation und Funktion mit einbezogen werden

Kopplung von IL-2 an Tumorzellen

Ziel der vorliegenden Doktorarbeit war es, zu untersuchen, ob rekombinantes humanes Interleukin-2 mittels heterobifunktioneller Linker an Tumorzellen gebunden werden kann. Dazu sollte rekombinantes Interleukin-2 sowohl kovalent über Thioether- bzw. Disulfidbindung als auch durch Adhäsion über das Lektin ConcanavalinA an Zellen gekoppelt werden. In Vorversuchen diente FITC als Modellsubstanz für IL-2. Die Ergebnisse zeigen, daß eine Kopplung von IL-2 mit verschiedenen Linkern an Myelomzellen möglich ist. Außerdem sprechen die Ergebnisse der Untersuchungen mittels FACScan für eine höhere Kopplungsrate bei Verwendung von kovalenten Linkern. Die stetige Abnahme von IL-2 im CTLL-Test deutet auf eine kontinuierliche Abgabe von IL-2 durch die gekoppelten Zellen hin. Die nachgewiesenen Trimere und Tetramere des IL-2 und die kontinuierliche Abgabe von IL-2 können einen möglichen neuen Therapieansatz von Tumorerkrankungen im Rahmen der ASI-Therapie darstellen

Strain-dependent interactions of Streptococcus gallolyticus subsp. gallolyticus with human blood cells

Abstract Background Streptococcus gallolyticus subsp. gallolyticus (S. gallolyticus) is the causative pathogen in up to 20% of streptococcal-induced infective endocarditis (IE) cases. However, the underlying mechanisms of pathogenesis in S. gallolyticus have not yet been solved. Pathogens causing IE need to employ virulent strategies to initiate and establish infections, such as escape the bloodstream, invade the host-cell, and persist intracellularly. In this study, we examined the induction of inflammation by different S. gallolyticus strains in relation to their survival in whole blood and cell culture models as well as their ability to induce platelet aggregation. Phagocytosis of these bacteria by macrophages, followed by intracellular survival, was also quantified. Methods In whole blood and THP-1 cell culture assays bacterial growth kinetics was determined by plating, followed by colony counting. Induction of interleukin (IL)-6 expression in whole blood of three healthy volunteers, caused by different strains, was quantified by ELISA. Gene expression of cytokines (IL1B, IL6 and IL8) was quantified by real-time PCR after stimulating THP-1 monocytes with bacteria. Induction of platelet aggregation was analyzed by light transmission aggregometry using the BORN method. A macrophage model was used to analyze phagocytosis of strains and their survival in macrophages within 48 h. Results Strains promoted IL-6 secretion in a time-dependent fashion. For example, DSM16831 induced IL-6 secretion in whole blood earlier than other isolates, and was eliminated in the whole blood of one volunteer, whereas UCN34 could grow. Platelet aggregation depended on the different isolates used and on the individual platelet donor. Two strains (AC1181 and 010672/01) induced cytokine gene expression in THP-1 monocytes only marginally, compared to other strains. The phagocytosis rate of S. gallolyticus isolates differed significantly, and the isolates UCN34 and BAA-2069 could persist for a considerable time in the phagocytes. Conclusion The strain-dependent differences of S. gallolyticus isolates, observed during interaction with human blood cells, support the hypotheses that divergences in individual virulence factors determine a distinct pathogenicity of the isolates. These data constitute an additional step towards the elucidation of mechanisms in the complex, multifactorial pathogenesis of this IE pathogen