Thrombin Flux and Wall Shear Rate Regulate Fibrin Fiber Deposition State during Polymerization under Flow

Thrombin is released as a soluble enzyme from the surface of platelets and tissue-factor-bearing cells to trigger fibrin polymerization during thrombosis under flow conditions. Although isotropic fibrin polymerization under static conditions involves protofibril extension and lateral aggregation leading to a gel, factors regulating fiber growth are poorly quantified under hemodynamic flow due to the difficulty of setting thrombin fluxes. A membrane microfluidic device allowed combined control of both thrombin wall flux (10⁻¹³ to 10⁻¹¹ nmol/μm² s) and the wall shear rate (10-100 s⁻¹) of a flowing fibrinogen solution. At a thrombin flux of 10⁻¹² nmol/μm² s, both fibrin deposition and fiber thickness decreased as the wall shear rate increased from 10 to 100 s⁻¹. Direct measurement and transport-reaction simulations at 12 different thrombin flux-wall shear rate conditions demonstrated that two dimensionless numbers, the Peclet number (Pe) and the Damkohler number (Da), defined a state diagram to predict fibrin morphology. For Da < 10, we only observed thin films at all Pe. For 10 < Da < 900, we observed either mat fibers or gels, depending on the Pe. For Da > 900 and Pe < 100, we observed three-dimensional gels. These results indicate that increases in wall shear rate quench first lateral aggregation and then protofibril extension.     

Molecular and functional analysis of the ruv region of Escherichia coli K-12 reveals three genes involved in DNA repair and recombination

Summary Recombinant plasmids carrying ruvA, ruvB, or both were constructed and used to investigate the genetic defects in a collection of UV-sensitive ruv mutants. The results revealed that efficient survival of UV-irradiated cells depends on both ruvA and ruvB, and on a third gene, ruvC, located upstream of the ruvAB operon. Southern blotting analysis was used to locate insertions in ruv and to examine putative deletion mutants. Two Tn10 insertions were located to the region encoding ruvA. Since these insertions caused a deficiency in the activities of both ruvA and ruvB, we concluded that they must exert a polar effect on ruvB. Two putative ruv deletion mutants were shown to be the result of deletion-inversion events mediated during imprecise excision of Tn10. The relevant inversion breakpoints in these mutants were located to ruvA and ruvC.     

Purification and properties of the RuvA and RuvB proteins of Escherichia coli

Summary The RuvA and RuvB proteins of Escherichia coli play important roles in the post-replicational repair of damaged DNA, genetic recombination and cell division. In this paper, we describe the construction of over expression vectors for RuvA and RuvB and detail simple purification schemes for each protein. The purified 22 kDa RuvA polypeptide forms a tetrameric protein (M_r ca. 100000) as observed by gel filtration. The tetramer is stabilised by strong disulphide bridges that resist denaturation during SDS-PAGE (in the absence of boiling and -mercaptoethanol). In contrast, purified RuvB polypeptides (37 kDa) weakly associate to form a dimeric protein (M_r ca. 85000). At low protein concentrations, the RuvB dimer dissociates into monomers. The multimeric forms of each protein may be covalently linked by the bifunctional cross-linking reagent dimethyl suberimidate. Addition of purified RuvA and RuvB to a RecA-mediated recombination reaction was found to stimulate the rate of strand exchange leading to the rapid formation of heteroduplex DNA.     

Age-related changes in neuromuscular function of the quadriceps muscle in physically active adults

Substantial evidence exists for the age-related decline in maximal strength and strength development. Despite the importance of knee extensor strength for physical function and mobility in the elderly, studies focusing on the underlying neuromuscular mechanisms of the quadriceps muscle weakness are limited.The aim of this study was to investigate the contributions of age-related neural and muscular changes in the quadriceps muscle to decreases in isometric maximal voluntary torque (iMVT) and explosive voluntary strength. The interpolated twitch technique and normalized surface electromyography (EMG) signal during iMVT were analyzed to assess changes in neural drive to the muscles of 15 young and 15 elderly volunteers. The maximal rate of torque development as well as rate of torque development, impulse and neuromuscular activation in the early phase of contraction were determined. Spinal excitability was estimated using the H reflex technique. Changes at the muscle level were evaluated by analyzing the contractile properties and lean mass.The age-related decrease in iMVT was accompanied by a decline in voluntary activation and normalized surface EMG amplitude. Mechanical parameters of explosive voluntary strength were reduced while the corresponding muscle activation remained primarily unchanged. The spinal excitability of the vastus medialis was not different while M wave latency was longer. Contractile properties and lean mass were reduced.In conclusion, the age-related decline in iMVT of the quadriceps muscle might be due to a reduced neural drive and changes in skeletal muscle properties. The decrease in explosive voluntary strength seemed to be more affected by muscular than by neural changes.     

Moths in fragments: insights into the biology and ecology of the Australian endangered golden sun moth Synemon plana (Lepidoptera: Castniidae) in natural temperate and exotic grassland remnants

The conservation and management of endangered species requires an adequate understanding of their biology and ecology. Although there has been an increasing appreciation in Australia of the need for greater efforts to conserve insects, there is only limited information available that can be used to underpin conservation efforts. The endangered golden sun moth, Synemon plana (Lepidoptera: Castniidae) is a flagship species endemic to natural temperate grassland in south-eastern Australia. Most populations of this species are at considerable risk from habitat loss, weed invasion and inadequate management. Despite the considerable knowledge that exists about the species biology and ecology, efforts to improve the species conservation status are hampered because there are still critical gaps in our understanding of the species’ natural history. In particular, the ecology of the larvae is not known. Our study examined the abundance, population structure and reproductive biology of the moths in a broad sample of both natural temperate and exotic grassland remnants in and near Canberra in the Australian Capital Territory (ACT) in south-eastern Australia. The results fill critical gaps in the knowledge needed to achieve effective conservation management. From our findings, it is clear that the species inhabits grasslands dominated by a mixture of native wallaby grasses (Rytidosperma spp. (formerly Austrodanthonia)) and spear grasses (Austrostipa spp.). In contrast to earlier suggestions that S. plana is entirely confined to natural temperate grassland, mature and immature life stages of the species were also present in grasslands comprised entirely of the exotic Chilean needlegrass (Nassella neesiana). Most of the S. plana populations surveyed in the ACT were characterised by low relative abundance with only very few large populations being recorded. The conservation of exotic grasslands as substitute habitat for S. plana is discussed and suggestions regarding future monitoring and research of the species are provided.     

Very rapid phosphorylation kinetics suggest a unique role for Lhcb2 during state transitions in Arabidopsis

Light‐harvesting complex II (LHCII) contains three highly homologous chlorophyll‐a/b‐binding proteins (Lhcb1, Lhcb2 and Lhcb3), which can be assembled into both homo‐ and heterotrimers. Lhcb1 and Lhcb2 are reversibly phosphorylated by the action of STN7 kinase and PPH1/TAP38 phosphatase in the so‐called state‐transition process. We have developed antibodies that are specific for the phosphorylated forms of Lhcb1 and Lhcb2. We found that Lhcb2 is more rapidly phosphorylated than Lhcb1: 10 sec of ‘state 2 light’ results in Lhcb2 phosphorylation to 30% of the maximum level. Phosphorylated and non‐phosphorylated forms of the proteins showed no difference in electrophoretic mobility and dephosphorylation kinetics did not differ between the two proteins. In state 2, most of the phosphorylated forms of Lhcb1 and Lhcb2 were present in super‐ and mega‐complexes that comprised both photosystem (PS)I and PSII, and the state 2‐specific PSI–LHCII complex was highly enriched in the phosphorylated forms of Lhcb2. Our results imply distinct and specific roles for Lhcb1 and Lhcb2 in the regulation of photosynthetic light harvesting.     

Postnatal remains of the foetal membranes affect growth and survival of wild European rabbit pups

During a long-term study on reproduction of the European rabbit (Oryctolagus cuniculus L.) that was carried out on a population living under semi-natural conditions, occasionally single pups per litter were discovered being stuck in remnants of the foetal membranes. Here we describe this so far unknown phenomenon, quantify the occurrence, and investigate the consequences for the individual growth rate and for survival. Furthermore, we tested if maternal social rank and age were correlated with the occurrence of this phenomenon. Pups comprising the described adherence of the foetal membranes had lower growth rates than their unaffected litter siblings, and usually died during the nestling period or short after emergence above ground. Our findings point out that there was an individual predisposition of the mother for having affected litters. However, an impact of maternal social rank or age on the occurrence of this feature in their offspring was not supported by our data. Nevertheless, a pathological background also has to be considered as a possible cause.