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- Haeger, M, et al.
(författare)
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Complement, neutrophil, and macrophage activation in women with severe preeclampsia and the syndrome of hemolysis, elevated liver enzymes, and low platelet count.
- 1992
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Ingår i: Obstetrics and gynecology. - 0029-7844. ; 79:1, s. 19-26
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Tidskriftsartikel (refereegranskat)abstract
- Activation of complement, neutrophils, and macrophages was studied in 14 women with severe preeclampsia, 11 of whom had the syndrome of hemolysis, elevated liver enzymes, and low platelet count; in 14 women with normal pregnancies; in seven normal pregnant women undergoing cesarean deliveries; and in 15 healthy nonpregnant women. Activation of complement, neutrophils, and macrophages was measured by plasma determinations of complement split products, polymorphonuclear (PMN) elastase, and neopterin, respectively. Women with severe preeclampsia had increased levels of C5a, terminal complement complex, PMN elastase, and neopterin at delivery and 1 day postpartum as compared with the normal pregnant group. One week postpartum, neopterin remained higher in preeclamptic women, whereas the complement components and PMN elastase had returned to normal. Cesarean delivery after normal pregnancy did not increase the levels of complement split products, PMN elastase (except for one value), or neopterin. The nonpregnant women had normal PMN elastase and neopterin levels. Accordingly, complement, neutrophils, and macrophages are activated in women with severe preeclampsia at delivery. The plasma levels of PMN elastase correlated positively to the formed terminal complement complexes in vivo. An in vitro study was performed to elucidate further the connection between complement and leukocyte activation. Recombinant C5a incubated in whole blood and in a neutrophil cell suspension gave a dose-dependent release of PMN elastase. Both the clinical and the in vitro results indicate that activation of the complement system may affect the function of neutrophils. This study supports the theory that the pathologic manifestations of severe preeclampsia may be explained by complement-induced release of biologically active substances from activated leukocytes.
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- Björquist, P, et al.
(författare)
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Plasminogen activator inhibitor type-1 interacts exclusively with the proteinase domain of tissue plasminogen activator.
- 1994
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Ingår i: Biochimica et Biophysica Acta. - 0006-3002 .- 1878-2434. ; 1209:2, s. 191-202
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Tidskriftsartikel (refereegranskat)abstract
- Two different techniques have been used to study the complex formation of recombinant human plasminogen activator inhibitor type-1, PAI-1, with either recombinant human two-chain tissue plasminogen activator, tc tPA (EC 3.4.21.68), or the tPA deletion variants tc K2P, containing the kringle 2 domain and the proteinase domain, and P, containing only the proteinase domain. The same value for Kon, 2.10(7) M-1s-1 for binding of PAI-1 was found for the three tPA forms by direct detection of the complex formation in real time by surface plasmon resonance, BIAcore, or indirectly by monitoring the time course of the inhibition of tPA using the chromogenic substrate N-methylsulfonyl-D-Phe-Gly-Arg-4-pNA-acetate. Apparently, no conformational change is involved in the rate-limiting step, since the kon value was found to be independent of the temperature from 20 to 35 degrees C. By the BIAcore technique, it was found that the complex between PAI-1 and tPA covalently coupled to the surface, was stable at 25 degrees C, since no dissociation was seen in buffer. However, extended treatment with 1 M NH4OH destroyed the complex with t 1/2 = 5 h. The same kon values and complex composition were found by measuring either the binding of tPA to PAI-1 captured on the monoclonal antibody MAI-11 or the binding of PAI-1 to tPA captured on the monoclonal antibody 2:2 B10. Quantification of the complex composition between PAI-1 captured on the monoclonal antibody MAI-11 with either tPA, K2P or P gave a one-to-one ratio with the fraction of active PAI-1, consistent with the results from SDS-PAGE and the specific activity of PAI-1. The complexes of the three tPA forms with PAI-1 captured on a large surface of MAI-11 dissociated more rapidly from MAI-11, with the same apparent koff, kdis, = 2.10(-3) s-1, compared with 0.7-10(-3) s-1 for the dissociation of PAI-1 alone. In consistance, the Kd, calculated from the direct determination of the kon and koff for the association of different form of PAI-1 to a small surface of MAI-11, was found to be higher for PAI-1 in complex with tPA than for free active PAI-1. Apparently, upon complex formation, a change is induced in PAI-1 at the binding epitope for MAI-11.(ABSTRACT TRUNCATED AT 400 WORDS)
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- Ekström, Per, et al.
(författare)
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A calmodulin inhibitor with high specificity, compound 48/80, inhibits axonal transport in frog nerves without disruption of axonal microtubules.
- 1991
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Ingår i: Acta physiologica Scandinavica. - 0001-6772. ; 142:2, s. 181-9
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Tidskriftsartikel (refereegranskat)abstract
- The calmodulin inhibitor compound 48/80 has previously been shown to arrest axonal transport in vitro in the regenerating frog sciatic nerve. The inhibition was limited to the outgrowth region of nerves, which had been allowed to regenerate in vivo for 6 days after a crush lesion, before they were incubated with or without drugs in vitro overnight. The effects of compound 48/80 on the regenerating nerve were further investigated. A concentration of compound 48/80 (50 micrograms ml-1), which effectively inhibits axonal transport, did not cause observable changes of the microtubules of regenerating axons in the outgrowth region as judged by electron microscopy. Furthermore, it was shown that also a lower concentration (25 micrograms ml-1) inhibited axonal transport. As a measure of possible metabolic effects, the level of ATP was assessed in the regenerating nerve after exposure to compound 48/80. Compound 48/80 at 25 micrograms ml-1 did not change the level of ATP in the nerve. The assembly of bovine brain microtubule proteins in a cell-free system was unaffected by 25 micrograms ml-1 of compound 48/80 and slightly inhibited by 50 micrograms ml-1. At higher concentrations (greater than 100 micrograms ml-1) assembly of microtubules appeared stimulated, and microtubule spirals as well as closely aligned microtubules could be seen. These effects appeared to be unrelated to the transport effects. The present results indicate that compound 48/80 arrests axonal transport via mechanisms other than destruction of axonal microtubules or interference with the energy metabolism. It is possible that these mechanisms involve inhibition of calmodulin-regulated events essential to the transport.
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- Frick, Inga-Maria, et al.
(författare)
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Convergent evolution among immunoglobulin G-binding bacterial proteins
- 1992
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Ingår i: Proceedings of the National Academy of Sciences. - 1091-6490. ; 89:18, s. 8532-8536
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Tidskriftsartikel (refereegranskat)abstract
- Protein G, a bacterial cell-wall protein with high affinity for the constant region of IgG (IgGFc) antibodies, contains homologous repeats responsible for the interaction with IgGFc. A synthetic peptide corresponding to an 11-amino acid-long sequence in the COOH-terminal region of the repeats was found to bind to IgGFc and block the interaction with protein G. Moreover, two other IgGFc-binding bacterial proteins (proteins A and H), which do not contain any sequences homologous to the peptide, were also inhibited in their interactions with IgGFc by the peptide. Finally, a decapeptide based on a sequence in IgGFc blocked the binding of all three proteins to IgGFc. This unusually clear example of convergent evolution emphasizes the complexity of protein-protein interactions and suggests that bacterial surface-protein interaction with host protein adds selective advantages to the microorganism.
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- Heimgartner, U, et al.
(författare)
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Purification and Partial Characterization of Milk Clotting Proteases from Flowers of Cynara cardunculus
- 1990
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Ingår i: Phytochemistry. - : Elsevier BV. - 0031-9422 .- 1873-3700. ; 29:5, s. 1405-1410
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Tidskriftsartikel (refereegranskat)abstract
- Three proteases (cynarases 1, 2 and 3) with milk-clotting activity have been purified from dried flowers of Cynara cardunculus. The proteases are each composed of one large and one small subunit. The native Mr of the dimeric proteins is 49 000. The three proteases are glycoproteins containing N-linked high mannose type glycans. Cynarase 3 shows the highest proteolytic and milk-clotting activity. All three enzymes express maximum activity at pH 5.1. Inhibitor studies indicate that the cynarases are of the aspartic acid type. Antibodies raised against the large subunit of cynarase 3 cross-reacts with the large subunits of the other two cynarases after destruction of the glycan structure by periodate oxidation.
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- Nakao, Naoyuki, et al.
(författare)
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Lazaroids improve the survival of grafted rat embryonic dopamine neurons
- 1994
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - 0027-8424. ; 91:26, s. 12-12408
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Tidskriftsartikel (refereegranskat)abstract
- In rodent models of Parkinson disease in which transplants of dissociated rodent and human embryonic mesencephalic tissue, rich in dopamine neurons, have been studied, only 5-20% of the dopamine neurons survive the implantation procedure. We have investigated the effects of inhibiting free radical generation with two lazaroids, U-74389G and U-83836E, on the survival of embryonic rat dopamine neurons. U-74389G is a 21-aminosteroid, and U-83836E combines the piperazinyl pyrimidine portion of 21-aminosteroids with the antioxidant ring of alpha-tocopherol. In an initial study, we found that the lazaroids markedly prolonged the period after tissue dissociation that an embryonic mesencephalic cell suspension exhibits high cell viability in vitro, as assessed by using a dye exclusion method. In a second series of experiments, addition of lazaroids to dissociated mesencephalic graft tissue increased the yield of surviving rat dopamine neurons 2.6-fold after implantation in the dopamine-denervated rat striatum. The improved survival correlated with an earlier onset of graft-induced functional effects in the amphetamine-induced rotation test. Thus, inhibition of free radical generation can significantly increase the yield of grafted embryonic dopamine neurons. Addition of lazaroids to the graft preparation is a relatively simple modification of the transplantation protocol and could readily be applied in a clinical setting. Moreover, since iron-dependent lipid peroxidation has been suggested to play a role in the death of nigral dopamine neurons in Parkinson disease and lazaroids are particularly potent inhibitors of such processes, the findings may have implications for the pathogenesis of this disease.
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