Protein phosphatase 2A holoenzyme and its subunits from Medicago sativa

We detected an about 200 kDa holoenzyme of protein phosphatase 2A (PP2A) in the crude extract of Medicago sativa microcallus cells by gel permeation chromatography. By polymerase chain reaction (PCR) we isolated two M. sativa cDNA fragments corresponding to the catalytic (C) subunit, and one each coding for the A and the B regulatory subunits of PP2A. The C subunit sequences were different from that published previously, indicating the existence of at least three different isoforms in M. sativa. Using the PCR fragments as probes, we obtained two distinct full-length clones for both the A and B subunits from an alfalfa cDNA library. Our results demonstrate that the components of the PP2A holoenzyme, namely the catalytic and regulatory subunits, are present in alfalfa in several isoforms and that their sequences are highly similar to their plant, yeast and animal counterparts. The distinct regulatory subunit genes are constitutively expressed during the cell cycle. Interestingly, two A-B subunit pairs had parallel mRNA steady-state levels in different plant tissues suggesting that not all of the possible isoform combinations are present in all tissues. The expression of the MsPP2A B subunit form was induced by abscisic acid indicating a specific function for this protein in the stress response.     

Biomanipulation as an Application of Food-Chain Theory: Constraints, Synthesis, and Recommendations for Temperate Lakes

The aim of this review is to identify problems, find general patterns, and extract recommendations for successful biomanipulation. An important conclusion is that the pelagic food chain from fish to algae may not be the only process affected by a biomanipulation. Instead, this process should be viewed as the “trigger” for secondary processes, such as establishment of submerged macrophytes, reduced internal loading of nutrients, and reduced resuspension of particles from the sediment. However, fish reduction also leads to a high recruitment of young-of-the-year (YOY) fish, which feed extensively on zooplankton. This expansion of YOY the first years after fish reduction is probably a major reason for less successful biomanipulations. Recent, large-scale biomanipulations have made it possible to update earlier recommendations regarding when, where, and how biomanipulation should be performed. More applicable recommendations include (1) the reduction in the biomass of planktivorous fish should be 75% or more; (2) the fish reduction should be performed efficiently and rapidly (within 1–3 years); (3) efforts should be made to reduce the number of benthic feeding fish; (4) the recruitment of YOY fish should be reduced; (5) the conditions for establishment of submerged macrophytes should be improved; and (6) the external input of nutrients (phosphorus and nitrogen) should be reduced as much as possible before the biomanipulation. Recent biomanipulations have shown that, correctly performed, the method also achieves results in large, relatively deep and eutrophic lakes, at least in a 5-year perspective. Although repeated measures may be necessary, the general conclusion is that biomanipulation is not only possible, but also a relatively inexpensive and attractive method for management of eutrophic lakes, and in particular as a follow-up measure to reduced nutrient load. Received 14 April 1998; accepted 31 August 1998     

Supression of hemin-mediated oxidation of low-density lipoprotein and subsequent endothelial reactions by hydrogen sulfide (H2S)

Heme-mediated oxidative modification of low-density lipoprotein (LDL) plays a crucial role in early atherogenesis. It has been shown that hydrogen sulfide (H₂S) produced by vascular smooth muscle cells is present in plasma at a concentration of about 50 µmol/L. H₂S is a strong reductant which can react with reactive oxygen species like superoxide anion and hydrogen peroxide. The current study investigated the effect of H₂S on hemin-mediated oxidation of LDL and oxidized LDL (oxLDL)-induced endothelial reactions. H₂S dose dependently delayed the accumulation of lipid peroxidation products—conjugated dienes, lipid hydroperoxides (LOOH), and thiobarbituric acid reactive substances—during hemin-mediated oxidation. Moreover, H₂S decreased the LOOH content of both oxidized LDL and lipid extracts derived from soft atherosclerotic plaque, which was accompanied by reduced cytotoxicity. OxLDL-mediated induction of the oxidative stress responsive gene, heme oxygenase-1, was also abolished by H₂S. Finally we have shown that H₂S can directly protect endothelium against hydrogen peroxide and oxLDL-mediated endothelial cytotoxicity. These results demonstrate novel functions of H₂S in preventing hemin-mediated oxidative modification of LDL, and consequent deleterious effects, suggesting a possible antiatherogenic action of H₂S.     

Molecular Characterization of Melanoma Cases in Denmark Suspected of Genetic Predisposition

Both environmental and host factors influence risk of cutaneous melanoma (CM), and worldwide, the incidence varies depending on constitutional determinants of skin type and pigmentation, latitude, and patterns of sun exposure. We performed genetic analysis of CDKN2A, CDK4, BAP1, MC1R, and MITFp.E318K in Danish high-risk melanoma cases and found CDKN2A germline mutations in 11.3% of CM families with three or more affected individuals, including four previously undescribed mutations. Rare mutations were also seen in CDK4 and BAP1, while MC1R variants were common, occurring at more than twice the frequency compared to Danish controls. The MITF p.E318K variant similarly occurred at an approximately three-fold higher frequency in melanoma cases than controls. To conclude, we propose that mutation screening of CDKN2A and CDK4 in Denmark should predominantly be performed in families with at least 3 cases of CM. In addition, we recommend that testing of BAP1 should not be conducted routinely in CM families but should be reserved for families with CM and uveal melanoma, or mesothelioma.     

Yield‐biodiversity trade‐off in patchy fields of Miscanthus × giganteus

Increasing crop productivity to meet rising demands for food and energy, but doing so in an environmentally sustainable manner, is one of the greatest challenges for agriculture to date. In Ireland, Miscanthus × giganteus has the potential to become a major feedstock for bioenergy production, but the economic feasibility of its cultivation depends on high yields. Miscanthus fields can have a large number of gaps in crop cover, adversely impacting yield and hence economic viability. Predominantly positive effects of Miscanthus on biodiversity reported from previous research might be attributable to high crop patchiness, particularly during the establishment phase. The aim of this research was to assess crop patchiness on a field scale and to analyse the relationship between Miscanthus yield and species richness and abundance of selected taxa of farmland wildlife. For 14 Miscanthus fields at the end of their establishment phase (4–5 years after planting), which had been planted either on improved grassland (MG) or tilled arable land (MT), we determined patchiness of the crop cover, percentage light penetration (LP) to the lower canopy, Miscanthus shoot density and height, vascular plants and epigeic arthropods. Plant species richness and noncrop vegetation cover in Miscanthus fields increased with increasing patchiness, due to higher levels of LP to the lower canopy. The species richness of ground beetles and the activity density of spiders followed the increase in vegetation cover. Plant species richness and activity density of spiders on both MT and MG fields, as well as vegetation cover and activity density of ground beetles on MG fields, were negatively associated with Miscanthus yield. In conclusion, positive effects of Miscanthus on biodiversity can diminish with increasing productivity. This matter needs to be considered when assessing the relative ecological impacts of developing biomass crops in comparison with other land use.     

Left ventricular assist device for end-stage heart failure: results of the first LVAD destination program in the Netherlands

PurposeMechanical circulatory support with a continuous-flow left ventricular assist device (LVAD) may be a valuable treatment in end-stage heart failure patients for an extended period of time. The purpose of this study was to evaluate the safety and efficacy of implantation of a continuous-flow LVAD in end-stage heart failure patients within the first destination program in the Netherlands.MethodsA third-generation LVAD was implanted in 16 heart failure patients (age 61 ± 8; 81 % male; left ventricular ejection fraction 20 ± 6 %) as destination therapy. All patients were ineligible for heart transplant. At baseline, 3 and 6 months, New York Heart Association (NYHA) functional class, quality-of-life and exercise capacity were assessed. Clinical adverse events were registered.ResultsSurvival at 30 days and 6 months was 88 and 75 %, respectively. In the postoperative phase, 6 (38 %) patients required continuous veno-venous haemofiltration for renal failure and 2 (13 %) patients required extracorporeal membrane oxygenation because of severe right ventricular failure. During follow-up, NYHA functional class and quality-of-life improved from 3.7 ± 0.1 to 2.3 ± 0.1 and 57 ± 5 to 23 ± 3 at 6 months (P < 0.001), respectively. The 6 min walking distance improved from 168 ± 42 m to 291 ± 29 m at 6 months (P = 0.001).ConclusionContinuous-flow LVAD therapy is a promising treatment for patients with end-stage heart failure ineligible for heart transplant.     

Overexpression of heat shock protein 27 (HSP27) increases gemcitabine sensitivity in pancreatic cancer cells through S‐phase arrest and apoptosis

We previously established a role for HSP27 as a predictive marker for therapeutic response towards gemcitabine in pancreatic cancer. Here, we investigate the underlying mechanisms of HSP27‐mediated gemcitabine sensitivity. Utilizing a pancreatic cancer cell model with stable HSP27 overexpression, cell cycle arrest and apoptosis induction were analysed by flow cytometry, nuclear staining, immunoblotting and mitochondrial staining. Drug sensitivity studies were performed by proliferation assays. Hyperthermia was simulated using mild heat shock at 41.8°C. Upon gemcitabine treatment, HSP27‐overexpressing cells displayed an early S‐phase arrest subsequently followed by a strongly increased sub‐G1 fraction. Apoptosis was characterized by PARP‐, CASPASE 3‐, CASPASE 8‐, CASPASE 9‐ and BIM‐ activation along with a mitochondrial membrane potential loss. It was reversible through chemical caspase inhibition. Importantly, gemcitabine sensitivity and PARP cleavage were also elicited by heat shock‐induced HSP27 overexpression, although to a smaller extent, in a panel of pancreatic cancer cell lines. Finally, HSP27‐overexpressing pancreatic cancer cells displayed an increased sensitivity also towards death receptor‐targeting agents, suggesting another pro‐apoptotic role of HSP27 along the extrinsic apoptosis pathway. Taken together, in contrast to the well‐established anti‐apoptotic properties of HSP27 in cancer, our study reveals novel pro‐apoptotic functions of HSP27—mediated through both the intrinsic and the extrinsic apoptotic pathways—at least in pancreatic cancer cells. HSP27 could represent a predictive marker of therapeutic response towards specific drug classes in pancreatic cancer and provides a novel molecular rationale for current clinical trials applying the combination of gemcitabine with regional hyperthermia in pancreatic cancer patients.     

Can intensively farmed arable land be favourable for birds during migration? The case of the Eurasian golden plover Pluvialis apricaria

Today's intensive farming practices are known to have affected farmland biodiversity negatively in many different ways. As far as birds are concerned, they are known to have suffered during both summer and winter. Relatively little is known about the effects on birds during migration. We studied the stopover ecology of the Eurasian golden plover Pluvialis apricaria, a species listed in EU Birds Directive, in intensively farmed arable land in southernmost Sweden in the autumns of 2003–2007. We used key ecological variables (length of stay, fat deposition and moult) as fitness proxies to evaluate how the birds manage in this habitat. Eurasian golden plovers were present in large numbers mainly on arable fields from early August to November and radio‐tagged birds were found to stay in the area for up to three months. Adult birds carried out a substantial part of their flight feather moult during their stay. Body mass increased only somewhat during moult, but from the last stages of moult and onwards fuel loads corresponding to 24% above lean body mass (LBM) were accumulated at a rate of 0.5% of LBM per day, before the birds departed. Juveniles arrived later, from mid Sep., and had a similar pattern of fuel deposition. The fact that the birds choose to stay for long periods, moult in the area, and manage to store substantial fuel loads strongly suggests that Eurasian golden plovers do well in this intensively farmed arable land.