Tissue factor-dependent autoactivation of human blood coagulation factor VII.

We recently showed that single-chain zymogen factor VII is converted to two-chain factor VIIa in an autocatalytic manner following complex formation with either cell-surface or solution-phase relipidated tissue factor apoprotein (Nakagaki, T., Foster, D. C., Berkner, K. L., and Kisiel, W. (1991) Biochemistry 30, 10819-10824). We have now performed a detailed kinetic analysis of the autoactivation of human plasma factor VII in the presence of relipidated recombinant tissue factor apoprotein and calcium. Incubation of factor VII with equimolar amounts of relipidated tissue factor apoprotein resulted in the formation of factor VIIa amidolytic activity coincident with the conversion of factor VII to factor VIIa. The time course for the generation of factor VIIa amidolytic activity in this system was sigmoidal, characterized by an initial lag phase followed by a rapid linear phase until activation was complete. The duration of the lag phase was decreased by the addition of exogenous recombinant factor VIIa. Relipidated tissue factor apoprotein was essential for factor VII autoactivation. No factor VII activation was observed following complex formation between factor VII and a recombinant soluble tissue factor apoprotein construct consisting of the aminoterminal extracellular domain in the presence or absence of phospholipids. Kinetic analyses revealed that factor VII activation in the presence of relipidated tissue factor apoprotein can be defined by a second-order reaction mechanism in which factor VII is activated by factor VIIa with an apparent second-order rate constant of 7.2 x 10(3) M-1 S-1. Benzamidine inhibited factor VII autoactivation with an apparent Ki of 1.8 mM, which is identical to the apparent Ki for the inhibition of factor VIIa amidolytic activity by this active site competitive inhibitor. Our data are consistent with a factor VII autoactivation mechanism in which trace amounts of factor VIIa rapidly activate tissue factor-bound factor VII by limited proteolysis.     

A mathematical model for adaptive transport network in path finding by true slime mold

We describe here a mathematical model of the adaptive dynamics of a transport network of the true slime mold Physarum polycephalum, an amoeboid organism that exhibits path-finding behavior in a maze. This organism possesses a network of tubular elements, by means of which nutrients and signals circulate through the plasmodium. When the organism is put in a maze, the network changes its shape to connect two exits by the shortest path. This process of path-finding is attributed to an underlying physiological mechanism: a tube thickens as the flux through it increases. The experimental evidence for this is, however, only qualitative. We constructed a mathematical model of the general form of the tube dynamics. Our model contains a key parameter corresponding to the extent of the feedback regulation between the thickness of a tube and the flux through it. We demonstrate the dependence of the behavior of the model on this parameter.     

Anti-cytokine autoantibodies are ubiquitous in healthy individuals

Anti-cytokine autoantibodies in healthy individuals have been widely reported but the occurrence is variable and inconstant. We hypothesized that cytokine-binding in vivo may explain their variable and infrequent detection. Therefore, we focused on the detection of the cytokine-autoantibody complexes and found that anti-cytokine autoantibody to IL-2, IL-8, tumor necrosis factor-alpha, vascular endothelial growth factor and granulocyte-colony stimulating factor were present in all 15 individuals evaluated, while those to IL-3, osteopontin and macrophage-colony stimulating factor were not detected in anyone. Autoantibodies against IL-4, IL-6, IL-10, and interferon-gamma were variously detected. Thus, we discovered that anti-cytokine autoantibodies to multiple cytokines are ubiquitous in healthy individuals.     

Traffic optimization in railroad networks using an algorithm mimicking an amoeba-like organism, Physarum plasmodium

Traffic optimization of railroad networks was considered using an algorithm that was biologically inspired by an amoeba-like organism, plasmodium of the true slime mold, Physarum polycephalum. The organism developed a transportation network consisting of a tubular structure to transport protoplasm. It was reported that plasmodium can find the shortest path interconnecting multiple food sites during an adaptation process (Nakagaki et al., 2001. Biophys. Chem. 92, 47-52). By mimicking the adaptation process a path finding algorithm was developed by Tero et al. (2007). In this paper, the algorithm is newly modified for applications of traffic distribution optimization in transportation networks of infrastructure such as railroads under the constraint that the network topology is given. Application of the algorithm to a railroad in metropolitan Tokyo, Japan is demonstrated. The results are evaluated using three performance functions related to cost, traveling efficiency, and network weakness. The traffic distribution suggests that the modified Physarum algorithm balances the performances under a certain parameter range, indicating a biological process.     

Constitutive Dimerization of Glycoprotein VI (GPVI) in Resting Platelets Is Essential for Binding to Collagen and Activation in Flowing Blood

The platelet collagen receptor glycoprotein VI (GPVI) has been suggested to function as a dimer, with increased affinity for collagen. Dissociation constants (K(d)) obtained by measuring recombinant GPVI binding to collagenous substrates showed that GPVI dimers bind with high affinity to tandem GPO (Gly-Pro-Hyp) sequences in collagen, whereas the markedly lower affinity of the monomer for all substrates implies that it is not the collagen-binding form of GPVI. Dimer binding required a high density of immobilized triple-helical (GPO)(10)-containing peptide, suggesting that the dimer binds multiple, discrete peptide helices. Differential inhibition of dimer binding by dimer-specific antibodies, m-Fab-F and 204-11 Fab, suggests that m-Fab-F binds at the collagen-binding site of the dimer, and 204-11 Fab binds to a discrete site. Flow cytometric quantitation indicated that GPVI dimers account for ～29% of total GPVI in resting platelets, whereas activation by either collagen-related peptide or thrombin increases the number of dimers to ～39 and ～44%, respectively. m-Fab-F inhibits both GPVI-dependent static platelet adhesion to collagen and thrombus formation on collagen under low and high shear, indicating that pre-existing dimeric GPVI is required for the initial interaction with collagen because affinity of the monomer is too low to support binding and that interaction through the dimer is essential for platelet activation. These GPVI dimers in resting circulating platelets will enable them to bind injury-exposed subendothelial collagen to initiate platelet activation. The GPVI-specific agonist collagen-related peptide or thrombin further increases the number of dimers, thereby providing a feedback mechanism for reinforcing binding to collagen and platelet activation.     

Dynamic organization of ATP and birefringent fibrils during free locomotion and galvanotaxis in the plasmodium of Physarum polycephalum

Directed migration by a cell is a good phenomenon for studying intracellular coordination. Dynamic organization of both ATP and birefringent fibrils throughout the cell was studied in the multinuclear ameboid cell of the Physarum plasmodium during free locomotion and galvanotaxis. In a directionally migrating plasmodium, waves of ATP as well as thickness oscillations propagated from just inside the advancing front to the rear, and ATP concentration was high at the front on the average. In a DC electric field, locomotion was inhibited more strongly, ATP concentration decreased more, and birefringent fibrils were formed more abundantly at the anodal than at the cathodal side. Inside the cell there were a few undulations in the distributions of ATP and birefringent fibrils. In short, birefringent fibrils become abundant where ATP concentration decreases. The possible mechanism of the coordination in the directed migration and the implications of the scaling law are discussed.     

Partial characterization of Fusobacterium necrophorum protease

Partial characterization of Fusobacterium necrophorum protease was investigated. The protease was partially purified by gel filtration with Toyopearl HW 55. The final preparation was inactivated completely by heating at 60 degrees C for 30 min and inhibited by ascorbic acid, sodium thioglycollate and p-hydromercuribenzoate. Antibody response to the protease was demonstrated in mice receiving 10(4) CFU of F. necrophorum.     

Organization of collagen bundles during tendon healing in rats treated with L-NAME

The Achilles tendon can support high tension forces and may experience lesions. The damaged tissue does not regenerate completely, with the organization and mechanical properties of the repaired tendon being inferior to those of a healthy tendon. Nitric oxide (NO) plays an important role in wound repair. We have examined the structural reorganization and repair in Achilles tendon after injury in rats treated with the NO synthase inhibitor L-NAME. The right Achilles tendon of male Wistar rats was partially transected. One group of rats was treated with L-NAME (~300 mg/kg per day, given in drinking water) for 4 days prior to tendon sectioning and throughout the post-operative period. Control rats received water without L-NAME. The tendons were excised at 7, 14, and 21 days post-injury and used to quantify hydroxyproline and for mechanical tests. Tendons were also processed for histomorphological analysis by polarized light microscopy, which showed that the collagen fibers were disorganized by day 7 in non-treated and L-NAME-treated rats. In non-treated rats, the organization of the extracellular matrix was more homogeneous by days 14 and 21 compared with day 7, although this homogeneity was less than that in normal tendon. In contrast, in injured tendons from L-NAME-treated rats, the collagen fibers were still disorganized on day 21. Tendons from treated rats had more hydroxyproline but lower mechanical properties compared with those from non-treated rats. Thus, NO modulates tendon healing, with a reduction in NO biosynthesis delaying reorganization of the extracellular matrix, especially collagen. T.C.T. and W.R.N were supported by studentships from CAPES, and S.H. was supported by a research fellowship from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).     

Fully decentralized control of a soft-bodied robot inspired by true slime mold

Animals exhibit astoundingly adaptive and supple locomotion under real world constraints. In order to endow robots with similar capabilities, we must implement many degrees of freedom, equivalent to animals, into the robots’ bodies. For taming many degrees of freedom, the concept of autonomous decentralized control plays a pivotal role. However a systematic way of designing such autonomous decentralized control system is still missing. Aiming at understanding the principles that underlie animals’ locomotion, we have focused on a true slime mold, a primitive living organism, and extracted a design scheme for autonomous decentralized control system. In order to validate this design scheme, this article presents a soft-bodied amoeboid robot inspired by the true slime mold. Significant features of this robot are twofold: (1) the robot has a truly soft and deformable body stemming from real-time tunable springs and protoplasm, the former is used for an outer skin of the body and the latter is to satisfy the law of conservation of mass; and (2) fully decentralized control using coupled oscillators with completely local sensory feedback mechanism is realized by exploiting the long-distance physical interaction between the body parts stemming from the law of conservation of protoplasmic mass. Simulation results show that this robot exhibits highly supple and adaptive locomotion without relying on any hierarchical structure. The results obtained are expected to shed new light on design methodology for autonomous decentralized control system.