Single, high-dose 17ß-estradiol therapy has anti-apoptotic effect and induces cerebral plasticity following transient forebrain ischemia in gerbils (Short communication).

Although much is known about the protective effect of acute estrogen therapy in cerebral ischemia, relatively little is known about the importance of apoptosis and cerebral plasticity in this mechanism. In this work 10 min global cerebral ischemia was produced by transient bilateral carotid occlusion in 4-month-old ovariectomized female gerbils. In every of our experimental group (sham for ischemia group, ischemia group and ischemia + a high, single dose 17ß-estradiol pre-treatment group) apoptotic (bcl-Xl, bax) and cerebral plasticity (GAP-43, synapsin-I, nestin) hippocampal genes' expression was measured four days after surgery. Expression of the anti-apoptotic bcl-Xl (p<0.01) and the cerebral plasticity marker synapsin-I and nestin (p<0.01) increased with acute estrogen pretreatment in ischemic animals. No change, however, in bax or GAP-43 expression was detected in estrogen treated animals compared to ischemic gerbils. These results suggest that acute estrogen therapy has anti-apoptotic effect and increases cerebral plasticity, which play an important role in cytoprotection or cerebroprotection.

Hypercapnia stimulates prostaglandin E(2) but not prostaglandin I(2) release in endothelial cells cultured from microvessels of human fetal brain.

Hypercapnia-induced cerebral vasodilation involves prostanoids, in newborns. The source of these prostanoids, however, is not yet determined. In the present study we address the hypothesis that microvascular endothelial cells of human fetal cerebrum increase the synthesis of dilator prostanoids in response to high pCO(2). Cells were isolated from a 22-week-old human fetus. Indication of induced abortion was 46 XY-t(3,10) 3q-25 chromosome abnormality. Normocapnia or hypercapnia was performed during normoxic and normothermic conditions in the medium of the cell culture. After normocapnic or hypercapnic stimuli, the amounts of released prostaglandin E(2) and 6-keto-prostaglandin F(1alpha) (the stable metabolite of prostaglandin I(2)) were measured by radioimmunoassay. Endothelial cells cultured from human fetal brain microvessels express PGE(2) and 6-keto-PGF(1alpha) in different degrees. Hypercapnic stimulus induced a significant increase of PGE(2), while expression of 6-keto-PGF(1alpha) was not augmented by the same stimulus. PGE(2) of endothelial origin, therefore, could be a factor in the mediation of the hypercapnia-induced vasodilation in human fetuses.

Streptokinase does not activate the complement system.

Streptokinase is an extensively used thrombolytic agent. However, different preparations cause severe hypotension during therapy, partially related to the complement cascade activation. In four ischaemic stroke patients treated with Streptase, an increased level of soluble terminal complement complex (SC5b-9) was measured. In the sera of normal subjects, the increase in SC5b-9 induced by Streptase, Kabikinase and Calbiochem streptokinases was highly significant (P < 0.005). Sigma streptokinase did not activate the complement system. Sigma streptokinase analyzed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed a homogeneous band. The other three preparations were contaminated with albumin and other proteins. Based on our in vivo and in vitro data, we conclude that complement activation is related to contamination of different streptokinase products rather than the streptokinase itself.

Expression of membrane-bound and soluble cell adhesion molecules by human brain microvessel endothelial cells.

Expression of membrane-bound (mb) and soluble (s) forms of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) induced by tumor necrosis factor-alpha (TNF-alpha) has been measured by enzyme-linked immunosorbent assay in cultured human brain microvessel endothelial cells. Both the mb and the s forms of VCAM-1 and ICAM-1 were upregulated by TNF-alpha; however, the stimulation of the s forms was delayed in time. When piracetam, a neuroprotective drug, was added to the tissue culture medium simultaneously with TNF-alpha, the expression of mbVCAM-1 and ICAM-1 was lowered. Differential upregulation of mb and s forms of adhesion molecules and a novel effect of piracetam have been demonstrated in human brain microvessel endothelial cell cultures.

Endothelial cells cultured from human brain microvessels produce complement proteins factor H, factor B, C1 inhibitor, and C4.

The inflammatory mediators, cytokines and complement proteins are believed to regulate the sequential events during the development of lesions secondary to ischaemia and reperfusion. The endothelial cell monolayer of the brain microvasculature is the critical interface between the blood-borne mediators and brain tissue. The involvement of these cells in complement production and regulation has not been well documented. In the present study, expression of complement proteins (C1 inhibitor, factor H, factor B, C4) by cultured endothelial cells obtained from human brain microvessels has been characterized. Interferon gamma upregulates the production of all the complement factors studied. Serine proteases, plasmin and miniplasmin induce the expression of C4, decrease the level of ELISA detectable C1 inhibitor, and do not affect the production of factors H and B. These data indicate that complement proteins are expressed locally by the brain microvessels, and may modulate the inflammatory responses of brain tissue.

Regulation of endothelin release from human brain microvessel endothelial cells.

After approval by the Local Ethical Committee, brain microvessel endothelial cells from human cadavers were isolated by enzymatic digestion and gradient centrifugation. Basal levels of endothelin-1 (ET) in the supernatant increased over time (3 h, 18.3 +/- 4.3 pg/ml; 6 h, 31.3 +/- 1.1 pg/ml; 24 h, 88.0 +/- 5.7 pg/ml; 48 h, 86.3 +/- 11.2 pg/ml, mean +/- SD). Tumor necrosis factor-alpha (TNF-alpha) (270 U/ml) increased ET concentration dose-dependently: 3 h, 190 +/- 70%; 24 h, 217 +/- 39%; 48 h, 207 +/- 5%; TNF-alpha at 210 U/ml: 3 h, 137%; 24 h, 170%; 48 h, 212% (values are relative changes from control, run in parallel to the stimulated wells). Interleukin-1 alpha (IL-1 alpha) (38.8 U/ml) also increased ET dose-dependently: (3 h, 129%; 24 h, 161%; 48 h, 212%; IL-1 alpha 1.4 U/ml: 3 h, 116%; 24 h, 122%; 48 h, 180%). Lipoprotein (a) (Lp(a)) had a dual effect on ET, increasing ET in the first 3 h but reducing it by the end of the 48-h observation period. This effect was not dose-dependent in the concentration range tested: Lp(a) 450 micrograms/ml; 3 h, 188%; 24 h, 91%; 48 h, 85%; Lp(a) 360 micrograms/ml: 3 h, 180%; 24 h, 94%; 48 h, 52%). Lp(a) reduced the stimulatory effect of cytokines on ET release. Maximal values at 48 h were TNF-alpha 207%, TNF-alpha + Lp(a) 91%, IL-1 alpha 212%, IL-1 alpha + Lp(a) 64%. In HPLC analysis, the total ET-like immunoreactivity co-eluted with the synthetic human ET standard. A cell culture of human brain microvessel endothelial cells was established. TNF-alpha and IL-1 alpha increased ET secretion, whereas Lp(a) had a dual effect. When given together, Lp(a) reduced the effect of cytokines on ETs.

Human brain microvessel endothelial cell culture as a model system to study vascular factors of ischemic brain.

Cerebral ischemia is caused by reduced blood supply at the microcirculatory level. In the microvessels, the main elements of the reperfusion injury following brain ischemia are the transformation of endothelial cell-surface from anticoagulant to procoagulant property, leukocyte adhesion, sludge or clot formation. There is a paucity of information on how hemostatic factors, cytokines, lipoprotein(a) (Lp(a)) and endothelin-1 (ET-1), being responsible for ischemic/reperfusion injury, interact with human brain microvessel endothelium (HBEC). There are no data furthermore about the expression of complement proteins of HBEC influenced by cytokines or fibrinolytic factors. Previously we established optimal conditions for culturing HBEC. Cell contraction induced by thrombin, plasmin, miniplasmin was recorded. The reassembly of F-actin was observed after thrombin treatment. ICAM-1 upregulation was measured following TNF-alpha, IL-1-alpha and thrombin incubation. Plasmin and miniplasmin downregulated the ICAM-1 in our cell culture system. Lp(a) modulated the thromboresistant cell-surface by reduction of t-PA and u-PA, but PAI-1 remained unchanged. Lp(a) modulated the ET-1 production by early increasing and late decreasing, in a bimodal manner. The increased secretion of ET-1 by cytokines (TNF-alpha, IL-1-alpha) was reduced in the presence of Lp(a). Gradual increase of complement proteins (factor H, factor B, C4) was induced by cytokines. Plasmin and miniplasmin augmented a rapid increase of C4. Some factors of complex relationship between regulators and modulators of endothelial adhesion molecules have been demonstrated in a human cell culture system prepared from brain microvessel endothelium. A unified concept of sequential events of ischemia/reperfusion in the brain has not yet developed.

Quality of plasma separated from (buffy coat + plasma). An alternative way of blood processing.

A CPD/ADSOL triple-bag system was used to produce plasma, buffy coat and resuspended erythrocytes. These components could be produced in a quadruple-bag system when working according to the conventional technique. In the experimental technique, buffy coat and plasma are transferred together into the satellite bag and are separated from each other only after the second centrifugation. The plasma complement system is not activated and factor IXa is not generated when applying the experimental technique. The quality of plasma meets the international requirements. The blood component processing technique using a triple-bag system is less expensive compared to the quadruple-bag one.

[Poly-electrolyte fractionated porcine factor VIII in the treatment of hemophilia A]. / Polielektrolit-frakcionált porcin VIII. faktor egy esetben való alkalmazásáról gátlótestes haemophiliában.

In approximately 10 to 15 percent of congenital hemophilia A patients circulating antibodies to factor VIII appear in the blood that poses a serious problem in their treatment. A number of methods and preparations are used in the clinical practice to overcome this problem. Authors report their favourable clinical experience with the administration of polyelectrolyte-fractionated porcine factor VIII. concentrate in a hemophiliac child for the first time in Hungary and a brief review of the clinical methods in use in the management of factor VIII. inhibitors.