Initiation of the extrinsic pathway of coagulation. Association of factor VIIa with a cell line expressing tissue factor

We have examined initial assembly of the extrinsic pathway of blood coagulation on cell surfaces with radiolabeled human factor VIIa and a human fetal lung cell line possessing abundant functional tissue factor activity. Binding of factor VIIa to these cells was observed and was time- and temperature-dependent. Binding of factor VIIa was quantitatively equivalent at 37 and 6 degrees C, although the kinetics of binding differed. The radiolabeled ligand bound by the cell was indistinguishable by sodium dodecyl sulfate-polyacrylamide gel analysis from the factor VIIa offered. Factor VIIa binding was influenced by calcium ions. The binding appears to involve at least two classes of calcium-dependent binding sites. Optimal binding occurred at 2 mM calcium for both classes of sites, and there was inhibition of binding to the high affinity sites at higher calcium. Association of factor VIIa was specific, saturable, had a Kd of 123 +/- 37 pm, and factor VIIa interacted with about 100,000 binding sites per cell. Once established, specific binding was rapidly reversible. Direct cellular binding of human factor X also was observed and was calcium, time- and temperature-dependent. Factor X binding was specific and saturable with half-maximal binding at 87.6 +/- 27.4 nM to 6.03 +/- 1.03 X 10(6) sites per cell. Specific high affinity binding of factor VIIa correlated with generation of factor Xa. A direct linear relationship was observed at low factor VIIa binding; however, at higher bound factor VIIa, the relationship was nonstoichiometric, i.e. less factor Xa was formed per mole of factor VIIa. Expression of specific binding sites for factors VIIa and X provides further substantiation for the molecular assembly hypothesized to initiate the extrinsic coagulation protease cascade on cells.     

Effect of brain antibodies on the lipid peroxidation process in the brain

The authors studies the effects of blood serum and IgG fraction from dogs immunized with brain and blood sera from patients with multiple sclerosis and schizophrenia on lipid peroxidation in rat brain homogenates. Measured the content of diene conjugates (DC) and malonic dialdehyde (MDA) in the rat brain after administering the IgG fraction. It was established that antioxidant activity of blood sera and IgG fraction from control animals and donors was significantly higher as compared to experimental. Administration of the IgG fraction brought about an increase in the content of DC and MDA in the brain of experimental animals. It is concluded that complement-dependent brain antibodies present in the blood serum of patients with schizophrenia and multiple sclerosis potentiate lipid peroxidation in the cerebral tissue and that the unsophisticated and informative method for antibody determination may be used in clinical practice.     

Factors affecting equal catchability in two swallowtail butterflies, Papilio polyxenes and P.glaucus

Abstract. 1. Recapture probabilities were analysed for individually marked black swallowtails, Papilio polyxenes F., and tiger swallowtails, P.glaucus L. (Lepidoptera: Papilionidae).
2. Recapture rates differed with sex and behaviour at time of capture for P.polyxenes , but not with age. For P.glaucus , only differences related to behaviour at time of capture were significant.
3. Black swallowtail males that were not physically restrained in identification had a recapture probability (73%) that was significantly higher than the 45% for males that were captured and handled. The higher rate was also exhibited by released, laboratory-reared males.
4. The capture effect was due to increased dispersal out of the areas and not to avoidance of capture or the capture site.     

Obligatory amino acids in primitive proteins

Conformational similarity among amino acid residues, a property derived by analysing (φ, ψ)-probability distributions of 20 proteinous amino acids from 38 different globular proteins, is used to arrive at a set of six ‘obligatory’ amino acids of primitive proteins. The amino acids Ser, Val, Leu, Asp, Gly and Pro have been argued to be ‘obligatory’ and to represent, conformationally, the remaining amino acids. The reasons for consideration of these six residues as ‘obligatory’ are discussed. Methods to check the validity of our proposition are suggested.     

The differential effect on two hybrid proteins of deletion mutations within the hydrophobic region of the Escherichia coli OmpA signal peptide

Oligonucleotide-directed site-specific mutagenesis was used to systematically shorten the hydrophobic region within the signal peptide of the Escherichia coli outer membrane protein OmpA. DNA encoding the wild type and mutant OmpA signal peptides were then fused in frame to DNA encoding the mature regions of Staphylococcus aureus nuclease A and TEM beta-lactamase. The ability of these signal peptides to direct processing of the resulting hybrid proteins was dependent on both their length and the protein to which they were fused. Deletion of two or more residues progressively slowed processing of pro-OmpA-nuclease. By contrast, pro-OmpA-beta-lactamase was less sensitive to the length of the hydrophobic region than to the nature of the deleted residue(s). Deletion of an Ala residue tended to reduce processing efficiency of pro-OmpA-beta-lactamase, while deletion of an Ile residue, together with the Ala residue, resulted in improvement. The loss of either 3 or 4 residues abolished processing of both hybrids. These data indicate that both the length as well as the identity of residues in the hydrophobic region are important. The relative importance of these two factors depends on the mature region of the protein being secreted.     

Phenotype modulation in primary cultures of arterial smooth-muscle cells. Dual effect of prostaglandin E1

The effects of prostaglandin E1 (PGE1) on the phenotypic state of enzymatically isolated arterial smooth-muscle cells in primary culture were studied by transmission electron microscopy, thymidine autoradiography, and cell counting. Early in culture (day 0-2), PGE1 stimulated conversion of the cells from contractile (less euchromatic nucleus and cytoplasm dominated by myofilament bundles) to synthetic state (more euchromatic nucleus and cytoplasm dominated by cisternae of rough endoplasmic reticulum and a large Golgi complex). The rate of entrance of the cells into DNA synthesis and mitosis was also increased at this time. Later on (day 3-6), when the majority of the cells had entered synthetic state, PGE1 inhibited DNA synthesis and cellular proliferation. These observations indicate that the effect of prostaglandins on arterial smooth muscle is dual in nature and dependent on the state of differentiation of the cells.