X-ray interference studies of crossbridge action in muscle contraction: evidence from muscles during steady shortening

During normal muscle shortening, the myosin heads must undergo many cycles of interaction with the actin filaments sliding past them. It is important to determine what range of configurations is found under these circumstances, and, in terms of the tilting lever arm model, what range of orientations the lever arms undergo. We have studied this using the X-ray interference technique described in the previous article, focusing mainly on the changes in the first order meridional reflection (M3) as compared to isometric. The change in ratio of the heights of the interference peaks indicates how far the mean lever arm angle has moved towards the end of the working stroke; the total intensity change depends on the angle change, on the number of heads now attached at any one time, and on the dispersion of lever arm angles. The latter provides a measure of the distance over which myosin heads remain attached to actin as they go through their working strokes. Surprisingly, the mean position of the attached heads moves only about 1 nm inwards (towards the center of the A-band) at low velocity shortening (around 0.9 T0): their dispersion changes very little. This shows that they must be detaching very early in the working stroke. However, at loads around 0.5 T0, the mean lever arm angle is about half way towards the end of the working stroke, and the dispersion of lever arm angles (with a uniform dispersion) is such as to distribute the heads throughout the whole of the working stroke. At higher velocities of shortening (at 0.3 T0), the mean position shifts further towards the end of the stroke, and the dispersion increases further. The details of the measurements, together with other data on muscle indicate that the force-generating mechanism within the myosin heads must have some unexpected properties.     

Interactions of protein kinase CK2 subunits

Several approaches have been used to study the interactions of the subunits of protein kinase CK2. The inactive mutant of CK2 that has Asp 156 mutated to Ala (CK2A156) is able to bind the CK2 subunit and to compete effectively in this binding with wild-type subunits and . The interaction between CK2A156 and CK2 was also demonstrated by transfection of epitope-tagged cDNA constructs into COS-7 cells. Immunoprecipitation of epitope-tagged CK2A156 coprecipitated the subunit and vice-versa. The assay of the CK2 activity of the extracts obtained from cells transiently transfected with these different subunits yielded some surprising results: The CK2 specific phosphorylating activity of these cells transfected with the inactive CK2A156 was considerably higher than the control cells transfected with vectors alone. Assays of the immunoprecipitated CK2A156 expressed in these cells, however, demonstrated that the mutant was indeed inactive. It can be concluded that transfection of the inactive CK2A156 affects the endogenous activity of CK2. Transfection experiments with CK2 and subunits and CK2A156 were also used to confirm the interaction of CK2 with the general CDK inhibitor p21WAF1/CIP1 co-transfected into these cells. Finally a search in the SwissProt databank for proteins with properties similar to those derived from the amino acid composition of CK2 indicated that CK2 is related to protein phosphatase 2A and to other phosphatases as well as to a subunit of some ion-transport ATPases.     

Germination and seedling survival in fens undergoing succession

We studied mechanisms of vegetation change in fens subject to succession from open water to floating mats and finally herbaceous rich-fens. Earlier research showed that these systems are characterized by transient seed banks. Our main question was whether seedlings of later successional fen stages are already present in earlier stages, remaining subordinate in the vegetation until conditions become suitable for them. If, however, conditions during succession change in a way that only a limited set of species can survive as seedlings during each of the successional stages, no seedling bank will exist. The transient character of the seeds would then imply that seeds will not germinate and will subsequently die and that seeds that have germinated in the “wrong” stage will not become established. We hypothesized that: (1) germination and seedling survival of fen species are significantly better in the successional fen stage for which these species are characteristic, (2) as a consequence no seedling banks occur in these fens. In a field experiment, seeds of five characteristic fen species in the standing vegetation of three successional fen stages i.e. raft fen, quaking fen and rich fen were sown in each of these stages in a turf pond in the Tienhoven area, The Netherlands. Germination and seedling survival were measured over two growing seasons together with environmental variables. Germination was higher in the “own” stage for all species groups as was survival for quaking fen species and rich fen species. For both these stages, percentage of germination and survival of four out of five characteristic species were significantly higher in the “own” stage. Germination and survival can be considered stage-dependent and it was concluded that seedling banks do not exist in these fens. Site-specific environmental variables act as a sieve and differentiate on species presence already during early life history stages. We found clues that the environmental sieve acts at the level of nutrient availability, tolerance for high sulphide concentration and light climate. Because of the transient seed bank and absence of a seedling bank in these fen wetlands, successful establishment of species necessitates a continuous dispersal of characteristic species until the environmental conditions permit establishment. This also implies that species of the whole successional sere should be present within dispersal distance.     

Human host genetic factors in mycobacterial and Salmonella infection: lessons from single gene disorders in IL-12/IL-23-dependent signaling that affect innate and adaptive immunity

Interleukin (IL)-12/IL-23 signal transduction-deficient individuals with genetic defects in IL12RB1 or IL12B often suffer from unusual mycobacterial and Salmonella infections. Here we discuss recent questions that have arisen from clinical observations that cast doubt on the necessity of IL-12/IL-23 signaling in controlling infections with intracellular bacteria. Alternative IL-12/IL-23-dependent, interferon-gamma-independent pathways of immunity to intracellular bacteria are also discussed.     

<Emphasis Type="Italic">In vitro</Emphasis> plantlet production of the endangered <Emphasis Type="Italic">Pinguicula vulgaris</Emphasis>

This study describes the development of a micropropagation protocol for Pinguicula vulgaris using cultures initiated from in vitro produced seedlings. P. vulgaris is a carnivorous plant with a northern, disjunctly circumpolar distribution and specific habitat requirements, and is hence becoming increasingly rare. Shoot proliferation was significantly influenced by Murashige and Skoog (MS) macronutrient concentration, showing higher proliferation rates in 1/4MS, but was not affected by the addition of 0.1 mg/L 6-benzyladenine (BA) or zeatin (Zea). The best medium for propagating P. vulgaris was plant growth regulator (PGR) free ¼MS. An average of 7.62 new shoots per initial explant could be obtained after 8 weeks of culture, of which over 79% produced roots during proliferation. Moreover, rooting percentages of 100% were obtained for the initial explants in all the tested media, including media without PGRs. The plantlets were successfully acclimatized to ex vitro conditions, exhibiting normal development.     

Polar bear (Ursus maritimus) cub mortality at a den site in northern Alaska

In March 2009, we documented the death of one member of a triplet polar bear (Ursus maritimus) litter at its den site in the southern Beaufort Sea (SBS) of Alaska. We used a self-contained video camera unit to document activity between den emergence and departure. All three cubs showed low activity levels relative to other cubs observed, and one died within one week of den emergence. Necropsy confirmed that the dead cub had a low body weight and was malnourished. Capture later confirmed that the two surviving cubs were also undersized. Polar bear cub survival is influenced by many factors including litter size and sea ice conditions. Triplet litters are often smaller and suffer higher mortality rates than singletons and twins. This cub was not only a triplet but also born following 2 years of record minimum sea ice extent, both of which may have played a role in this cub’s demise.     

Growth and nutrition of birch seedlings in relation to potassium supply rate

Summary Growth of hydroponically cultivated birch seedlings (Betula pendula Roth.) at sub- and supra-optimum potassium supply rates was investigated. Potassium was supplied either as a relative addition rate (r _k = 5, 10, 15 and 20% increase day^-1) or as fixed concentrations (0.2, 3, 6, 12 and 15 mM) in the culture solution. After an acclimation period the growth rate of the seedlings in the suboptimum treatments reached values close to the treatment variable, the relative rate of K-addition. Deficiency symptoms, in the form of chlorosis and necroses along the leaf margins, developed initially in all suboptimum treatments, but very few new symptoms appeared once the seedlings had reached the phase of steady-state nutrition and growth. At supra-optimum K-supply levels, i.e. at 0.2–15 mM K in the culture solution, no symptoms of deficiency or toxicity developed, and the relative growth rate of the seedlings remained maximum. The relative growth rate of the seedlings was linearly related to the plant K-status for K contents ranging from 0.2 to 1.0% of dry weight (DW). At higher internal K-concentrations, 1.0–3.0% DW, no further increase in relative growth rate was achieved. A shortage of K resulted in a decrease in the net assimilation rate. This effect was counterbalanced by the absence of shift in he leaf weight ratio as well as by the production of relatively thin leaves. The fraction of dry matter allocated to roots decreased in K-limited plants, as did the leaf contents of soluble carbohydrates and starch.     

Meta-performance evaluation of sustainability indicators

There are many different kinds of frameworks for evaluating environmental and sustainability performance at the organizational level (profit or not-for-profit, private or public), sectoral level (e.g. industry, transport, agriculture and tourism), and local, regional or country levels. Despite the diversity of methods and tools to measure sustainable development, indicators are one of the approaches most used. However, these tools do not usually include evaluation of the performance measurement instrument itself. The main objective of this research is to develop a conceptual framework to design and assess the effectiveness of the sustainability indicators themselves. To put the proposed tool into practice, a set of key good-practice factors and meta-performance evaluation indicators is proposed for adoption in a national case study—the national sustainable development indicators system, SIDS Portugal, and the usefulness of this methodology is demonstrated. This approach aims to evaluate how appropriate a set of sustainability indicators is and allow an evaluation of overall performance-monitoring activities and results. Stakeholder involvement is an essential component of the proposed framework. The tool developed could support continuous improvement in the performance of ongoing sustainability indicator initiatives, allowing greater guidance, objectivity and transparency in sustainability assessment processes.     

Angiogenic Properties of Human Dental Pulp Stem Cells

Angiogenesis, the formation of capillaries from pre-existing blood vessels, is a key process in tissue engineering. If blood supply cannot be established rapidly, there is insufficient oxygen and nutrient transport and necrosis of the implanted tissue will occur. Recent studies indicate that the human dental pulp contains precursor cells, named dental pulp stem cells (hDPSC) that show self-renewal and multilineage differentiation capacity. Since these cells can be easily isolated, cultured and cryopreserved, they represent an attractive stem cell source for tissue engineering. Until now, only little is known about the angiogenic abilities and mechanisms of the hDPSC. In this study, the angiogenic profile of both cell lysates and conditioned medium of hDPSC was determined by means of an antibody array. Numerous pro-and anti-angiogenic factors such as vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1), plasminogen activator inhibitor-1 (PAI-1) and endostatin were found both at the mRNA and protein level. hDPSC had no influence on the proliferation of the human microvascular endothelial cells (HMEC-1), but were able to significantly induce HMEC-1 migration in vitro. Addition of the PI3K-inhibitor {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 and the MEK-inhibitor U0126 to the HMEC-1 inhibited this effect, suggesting that both Akt and ERK pathways are involved in hDPSC-mediated HMEC-1 migration. Antibodies against VEGF also abolished the chemotactic actions of hDPSC. Furthermore, in the chicken chorioallantoic membrane (CAM) assay, hDPSC were able to significantly induce blood vessel formation. In conclusion, hDPSC have the ability to induce angiogenesis, meaning that this stem cell population has a great clinical potential, not only for tissue engineering but also for the treatment of chronic wounds, stroke and myocardial infarctions.