Functional determination of plasminogen activator in arginine-stabilized plasma.

Reliable data on plasminogen activator (PA) activities in blood of patients receiving fibrinolytic treatment are lacking. This is due to the continuing in vitro action of PA after blood withdrawal. We have elaborated a new simple stabilization technique for plasma involving the addition of arginine in final concentrations greater than 500 mM. In this study, new assays for PA in stabilized plasma are developed. The assay was performed with substrate plasma, that is, pooled normal plasma, preoxidized with chloramine-T; oxidant amount and oxidation time were optimized. The chloramine consumption by plasma was assayed with a KJ-assay (absorbance increase at 405 nm by addition of 200 microL 4 M KJ to 25 microL oxidized plasma). The substrate plasma concentration in the PA assay and the PA acting time was optimized. The inhibition of PA by the cations Na(+), K(+), Mg(2+), and Ca(2+) was evaluated. The optimized PA assay consists of incubation of 10 microL arginine-stabilized plasma with 10 microL 1.5 M arginine, pH 8.7 and 10 microL 100 mM CT in PBS. After 30 minutes (37 degrees C), 175 microL 15 mM CT oxidized EDTA plasma are added. After 40 minutes (37 degrees C), 75 microL Stop-CS Reagent is added and DeltaA at 405 nm was determined, giving PA + plasmin activity in plasma. A control value (basal plasmin activity) consists of the addition of Stop-CS Reagent before 175 microL oxidized EDTA plasma. To obtain plasmatic PA activity, the control value has to be subtracted from the PA main value. The assay is matrix-independent and linear up to 1250 IU/mL t-PA, 790 U/mL reteplase, or 199 IU/mL u-PA (37 nM). With arginine stabilization of plasma and the described determination of plasminogen activator activity in arginine-stabilized plasma, it is feasible to determine the activity of plasminogen activators in blood of patients receiving fibrinolytic treatment without artefactual in vitro changes of the samples.     

BDNF overexpression induces differential increases among subsets of sympathetic innervation in murine back skin

Besides their recognized dependence on nerve growth factor (NGF) during development, the dependence of mature sympathetic ganglion neurons on other neurotrophins is still unclear. Here, we have investigated the sympathetic innervation of back skin in mice overexpressing brain‐derived neurotrophic factor (BDNF) under the alpha‐myosin heavy‐chain promoter, as well as in BDNF knockout (–/–) mice. Compared with wild‐type controls, the dorsal skin of BDNF overexpressing mice displayed a significantly enhanced number of adrenergic, tyrosine hydroxylase‐immunoreactive (IR) nerve fibres, while cholinergic or peptidergic sensory nerve fibres appeared unaltered. The adrenergic hyperinnervation in dorsal skin of BDNF overexpressing mice was most pronounced in the arrector pili muscle of hair follicles, while no increase of tyrosine hydroxylase‐or neuropeptide Y‐IR fibres associated with subcutaneous blood vessels was found. Instead, back skin of BDNF knockout (–/–) mice contained significantly fewer tyrosine hydroxylase‐IR dermal nerve fibres than wild‐type animals. This suggests that BDNF plays an important role in the control of different subsets of adrenergic innervation in murine back skin, and indicates that paravertebral sympathetic ganglia display a previously unrecognized differential BDNF‐dependence in vivo.     

Dysgammaglobulinämic Typ I

Zusammenfassung Es wird ein Fall von Dysgammaglobulinämie Typ I (Einteilung nach Rosen und Janeway [20]) mitgeteilt. Der 7jährige Knabe zeigte die typischen Zeichen eines Antikörpermangelsyndroms seit seinem ersten Lebensjahr. Die Acetatfolienelektrophorese war unauffällig, trotzdem fehlten bei dem Patienten IgG und IgA vollständig. Erst die immunelektrophoretische Analyse deckte den Defekt auf: Eine im ₂-Bereich nachweisbare Doppellinie entsprach zwei verschiedenen Typen von IgM-Molekülen, nämlich IgM-kappa und IgM-lambda, wie durch entsprechende Erschöpfung des Antiserums mit Bence-Jones-Proteinen vom kappa- und/ oder lambda-Typ gezeigt werden konnte. Die analytische Ultrazentrifugation und die Gelfiltration über Sephadex G 200 bestätigte das Vorliegen einer polyklonalen Makroglobulinämie, der Serum-IgM-Gehalt betrug etwa 880 mg/100 ml (Ergebnis der Ultrazentrifugation). Das exzessiv vermehrte IgM erschien inert, es konnten keine Isoagglutinine nachgewiesen werden.     

In Vitro Effects of Fentanyl, Methohexital, and Thiopental on Brain Endothelial Permeability

Background: The use of anesthetics can lead to changes of the permeability of the blood-brain barrier (BBB). To eliminate those factors, such as varying hemodynamic effects that are associated with anesthesia, an in vitro model of the BBB consisting of brain microvascular endothelial cells (BMEC) was used to study the direct effects of the opiate, fentanyl, and the barbiturates methohexital and thiopental, which are widely used in the clinical setting, on the permeability of confluent monolayers.

Methods: BMEC isolated from porcine brains were grown to confluence on collagen-coated polycarbonate membranes, which were placed into 24 well dishes, thus forming a two-compartment chamber. The permeability of the BMEC monolayer to ions--determined by measurements of the transendothelial resistance (TER)--the passage of sucrose, Evans Blue albumin (EBA), and alpha-aminolsobutyric acid (AIB) across the BMEC monolayer were assessed in the presence and absence of fentanyl (25-100 ng/ml), methohexital (10-50 micro gram/ml), and thiopental (25-100 micro gram/ml).

Results: The permeability of cultured BMEC to the tracers used increased significantly after exposure of the monolayer to arabinose and after removal of calcium ions. Fentanyl, methohexital, and thiopental did not change the permeability of the cell monolayer to ions, sucrose, albumin, and AIB. Only thiopental at the concentration of 100 micro gram/ml increased the flux of AIB.     

Betablocking drugs in essential hypertension: transdermal bupranolol compared with oral metoprolol.

In the present study the antihypertensive efficacy and tolerability of transdermal bupranolol (30 mg once-daily) was compared with oral metoprolol (100 mg once-daily). Blood pressure measurements were performed in the office, at home, and with ambulatory 24-h blood pressure devices. Systemic and local side-effects, as well as compliance and acceptance, were evaluated every two weeks. The treatment period lasted eight weeks. The results showed a significant decrease in blood pressure under the bupranolol transdermal therapeutic system in the office, at home, and with 24-h blood pressure measurements day- (08h00-20h00) and night-time (20h00-08h00). Under oral metoprolol there was a significant blood pressure decrease in the office, at home, and in the mean daytime values of the 24-h blood pressure measurements. The night-time values, however, demonstrated only a slight decrease in blood pressure, being significant only for diastolic values. Systemic side-effects were comparable in both groups. 69% of the patients had local side-effects at the patch side (erythema, papulous exanthema, pruritus). Six patients dropped out because of localized urticarial exanthema (five patients treated with transdermal bupranolol, one patient treated with oral metoprolol). In comparison to the oral form, twice as many patients had admitted to have been non-compliant with the patches (13 versus 7 patients). At the end of the study, 24 out of 32 patients preferred to be treated with capsules.