Van Der Ploeg Research Articles

Lower Extremity Muscle Strength in 6- to 8-Year-Old Children Using Hand-Held Dynamometry

The purposes of the study were to establish isometric torque reference values for healthy 6- to 8-year-old children for 6 lower extremity muscles and determine the effects of gender, age, height, weight, and physical activity upon strength. A hand-held dynamometer was used to obtain force/torque values for 154 children. Age-referenced force/torque and cutoff values are provided for each muscle group. Torque increased with age and height for all muscles and with weight for all muscles except knee extensors. No gender differences were found. Children who participated in 3 or more hours per week in organized sports were stronger in 4 of 6 muscles; the number of hours spent in active play did not affect torque. This study provides hand-held dynamometer strength reference values to enable clinicians to determine if clients of the same age, height, and weight have muscle weakness.

Protection against β-Amyloid-induced Apoptosis by Peptides Interacting with β-Amyloid

beta-Amyloid peptide produces apoptosis in neurons at micromolar concentrations, but the mechanism by which beta-amyloid exerts its toxic effect is unknown. The normal biological function of beta-amyloid is also unknown. We used phage display, co-precipitation, and mass spectrometry to examine the protein-protein interactions of beta-amyloid in normal rabbit brain in order to identify the biochemical receptors for beta-amyloid. beta-Amyloid was found to bind primarily to proteins involved in low density lipoprotein and cholesterol transport and metabolism, including sortilin, endoplasmic reticulum-Golgi intermediate compartment 2 (ERGIC2), ERGIC-53, steroid 5alpha-reductase, and apolipoprotein B. beta-Amyloid also bound to the C-reactive protein precursor, a protein involved in inflammation, and to 14-3-3, a protein that regulates glycogen synthase kinase-3beta, the kinase involved in tau phosphorylation. Of eight synthetic peptides identified as targets of beta-amyloid, three were found to be effective blockers of the toxic effect of beta-amyloid on cultured neuronal cells. These peptides bound to the hydrophobic region (residues 17-21) or to the nearby protein kinase C pseudo-phosphorylation site (residues 26-30) of beta-amyloid, suggesting that these may be the most critical regions for beta-amyloid effector action and for aggregation. Peptides or other small molecules that bind to this region may protect against beta-amyloid toxic effect by competitively blocking its ability to bind beta-amyloid effector proteins such as sortilin and 14-3-3.

Styles, Parables And Scripts: Diversity And Conformity In Australian And Finnish Agriculture

Two paradigms for understanding the acquisition of farmer identity, farming styles and cultural scripts, are compared using interview material with family owned and operated farms in Australia and Finland. The styles of farming approach of van der Ploeg has been considered in various Australian applications. However, it does not adequately address why farmers become farmers, or how farming styles and farmers’ ways of life are socially reproduced. Instead, these are explained by the concepts of farmers’ parables and cultural scripts, both a form of narrative. Parables are the stories farmers tell about hypothetical farmers and which act as a form of social control. Scripts, like parables, are the cultural models that farmers utilise in their stories. Scripts and parables provide an explanation for the preservation of traditional practice when discourses of globalisation and economic rationality advocate the abandoning of the farm.

Ivermectin Interaction with Transmembrane Helices Reveals Widespread Rearrangements during Opening of P2X Receptor Channels

P2X receptors are trimeric cation channels that open in response to binding of extracellular ATP. Each subunit contains a large extracellular ligand binding domain and two flanking transmembrane (TM) helices that form the pore, but the extent of gating motions of the TM helices is unclear. We probed these motions using ivermectin (IVM), a macrocyclic lactone that stabilizes the open state of P2X(4) receptor channels. We find that IVM partitions into lipid membranes and that transfer of the TM regions of P2X(4) receptors is sufficient to convey sensitivity to the lactone, suggesting that IVM interacts most favorably with the open conformation of the two TM helices at the protein-lipid interface. Scanning mutagenesis of the two TMs identifies residues that change environment between closed and open states, and substitutions at a subset of these positions weaken IVM binding. The emerging patterns point to widespread rearrangements of the TM helices during opening of P2X receptor channels.

Peripheral Effect of α-Melanocyte-stimulating Hormone on Fatty Acid Oxidation in Skeletal Muscle

To study the peripheral effects of melanocortin on fuel homeostasis in skeletal muscle, we assessed palmitate oxidation and AMP kinase activity in alpha-melanocyte-stimulating hormone (alpha-MSH)-treated muscle cells. After alpha-MSH treatment, carnitine palmitoyltransferase-1 and fatty acid oxidation (FAO) increased in a dose-dependent manner. A strong melanocortin agonist, NDP-MSH, also stimulated FAO in primary culture muscle cells and C2C12 cells. However, [Glu6]alpha-MSH-ND, which has ample MC4R and MC3R agonistic activity, stimulated FAO only at high concentrations (10(-5) M). JKC-363, a selective MC4R antagonist, did not suppress alpha-MSH-induced FAO. Meanwhile, SHU9119, which has both antagonistic activity on MC3R and MC4R and agonistic activity on both MC1R and MC5R, increased the effect of alpha-MSH on FAO in both C2C12 and primary muscle cells. Small interference RNA against MC5R suppressed the alpha-MSH-induced FAO effectively. cAMP analogues mimicked the effect of alpha-MSH on FAO, and the effects of both alpha-MSH and cAMP analogue-mediated FAO were antagonized by a protein kinase A inhibitor (H89) and a cAMP antagonist ((Rp)-cAMP). Acetyl-CoA carboxylase activity was suppressed by alpha-MSH and cAMP analogues by phosphorylation through AMP-activated protein kinase activation in C2C12 cells. Taken together, these results suggest that alpha-MSH increases FAO in skeletal muscle, in which MC5R may play a major role. Furthermore, these results suggest that alpha-MSH-induced FAO involves cAMP-protein kinase A-mediated AMP-activated protein kinase activation.

Hypoallergens for Allergen-specific Immunotherapy by Directed Molecular Evolution of Mite Group 2 Allergens

Allergen-specific immunotherapy is the only treatment that provides long lasting relief of allergic symptoms. Currently, it is based on repeated administration of allergen extracts. To improve the safety and efficacy of allergen extract-based immunotherapy, application of hypoallergens, i.e. modified allergens with reduced IgE binding capacity but retained T-cell reactivity, has been proposed. It may, however, be difficult to predict how to modify an allergen to create a hypoallergen. Directed molecular evolution by DNA shuffling and screening provides a means by which to evolve proteins having novel or improved functional properties without knowledge of structure-function relationships of the target molecules. With the aim to generate hypoallergens we applied multigene DNA shuffling on three group 2 dust mite allergen genes, two isoforms of Lep d 2 and Gly d 2. DNA shuffling yielded a library of genes from which encoded shuffled allergens were expressed and screened. A positive selection was made for full-length, high-expressing clones, and screening for low binding to IgE from mite allergic patients was performed using an IgE bead-based binding assay. Nine selected shuffled allergens revealed 80-fold reduced to completely abolished IgE binding compared with the parental allergens in IgE binding competition experiments. Two hypoallergen candidates stimulated allergen-specific T-cell proliferation and cytokine production at comparable levels as the wild-type allergens in patient peripheral blood mononuclear cell cultures. The two candidates also induced blocking Lep d 2-specific IgG antibodies in immunized mice. We conclude that directed molecular evolution is a powerful approach to generate hypoallergens for potential use in allergen-specific immunotherapy.

Growth Hormone Corrects Proliferation and Transcription of Phosphoenolpyruvate Carboxykinase in Livers of Old Mice via Elimination of CCAAT/Enhancer-binding Protein α-Brm Complex

Growth hormone (GH), which is reduced with age, corrects the impaired proliferative capacity of livers of old animals. In this paper, we present a mechanism by which GH eliminates age-dependent negative control of proliferation and increases transcription of liver-specific genes in livers of old mice. The reduced proliferative capacities of the liver of old animals are associated with the CCAAT/enhancer-binding protein alpha (C/EBPalpha)-Brm complex, which inhibits E2F-dependent promoters. We found that a sequestration of C/EBPalpha into complexes with Brm leads to a weak interaction of C/EBPalpha with promoters of liver-specific genes, expression of which is reduced in old animals. Injection of either GH or the regulator of the amplitude of endogenous GH release, ghrelin, reduces the C/EBPalpha-Brm complex in livers of old mice, leading to a derepression of E2F targets, to increased interactions of C/EBPalpha with promoters of liver-specific genes, and to correction of their expression. GH-dependent elimination of the complex is mediated by the inhibition of cyclin D3-CDK4 activity and by elevation of a phosphatase, protein phosphatase 2A, which dephosphorylates C/EBPalpha and dissociates the complex.

Identification and Stage-specific Association with the Translational Apparatus of TbZFP3, a CCCH Protein That Promotes Trypanosome Life-cycle Development

The post-transcriptional control of gene expression is becoming increasingly important in the understanding of regulated events in eukaryotic cells. The parasitic kinetoplastids have a unique reliance on such processes, because their genome is organized into polycistronic transcription units in which adjacent genes are not coordinately regulated. Indeed, the number of RNA-binding proteins predicted to be encoded in the genome of kinetoplastids is unusually large, invoking the presence of unique RNA regulators dedicated to gene expression in these evolutionarily ancient organisms. Here, we report that a small CCCH zinc finger protein, TbZFP3, enhances development between life-cycle stages in Trypanosoma brucei. Moreover, we demonstrate that this protein interacts both with the translational machinery and with other small CCCH proteins previously implicated in trypanosome developmental control. Antibodies to this protein also co-immunoprecipitate EP procyclin mRNA and encode the major surface antigen of insect forms of T. brucei. Strikingly, although TbZFP3 is constitutively expressed, it exhibits developmentally regulated association with polyribosomes, and mutational analysis demonstrates that this association is essential for the expression of phenotype. TbZFP3 is therefore a novel regulator of developmental events in kinetoplastids that acts at the level of the post-transcriptional control of gene expression.

Book Reviews

Book reviewed in this article:Bioevolution: How Biotechnology is Changing Our World, by Michael FumentoCommunity and Forests: Where People Meet the Land, edited by Robert G. Lee and Donald R. FieldSocial Assessment: Theory, Process and Techbiques, 3rd ed., by C. Nicholas Taylor, C. Hobson Bryan and Colin G. GoodrichThe Concepts, Process and Methods of Social Impact Assessment: Rabel J. Burdge and Colleagues, by Rabel J. BurdgeWearing Cultural Styles in Japan: Concepts of Tradition and Modernity in Practice, edited by Christopher S. Thompson and John W. TraphaganThe Virtual Farmer: Past, Present and Future of the Dutch Peasantry, by Jan Douwe van der Ploeg

Altered Gastrointestinal and Metabolic Function in the GPR39-Obestatin Receptor–Knockout Mouse

The G-protein-coupled receptor GPR39 is a member of a family that includes the receptors for ghrelin and motilin. Recently the peptide obestatin was identified as a natural ligand for GPR39. The objective of this study was to gain insight into the biological function of the GPR39 receptor. GPR39(-/-) mice were generated and analyzed. Endogenous GPR39 expression was detected in the brain (septum-amygdala) and the gastrointestinal system (parietal cells, enterocytes, neurons, and pancreas). Gastric emptying of a solid meal (measured by the (14)C octanoic breath test) in GPR39(-/-) mice was accelerated significantly with a gastric half-emptying time of 49.5 +/- 2.2 minutes compared with 86.9 +/- 8.4 minutes in GPR39(+/+) mice. A more effective expulsion of distally located pellets (30%-75% of length) was observed in the colon of GPR39(-/-) mice. Four hours after pylorus ligation, the volume of gastric secretion was increased significantly (GPR39(-/-): 638 +/- 336 microL; GPR39(+/+): 225 +/- 170 microL), but gastric acid secretion was unchanged. The mature body weight and body fat composition of GPR39(-/-) mice was significantly higher compared with GPR39(+/+) mice, but this was not related to hyperphagia because 24-hour food intake did not differ between both genotypes. In contrast, deficiency of the GPR39 receptor led to reduced hyperphagia after fasting. The cholesterol levels were increased significantly in the GPR39(-/-) mice. Our data partially confirm and extend the described in vivo effects of obestatin and suggest that this peptide plays a functional role in the regulation of gastrointestinal and metabolic function through interaction with the GPR39 receptor.

Bodies That Matter

Recent press coverage of the release of two new studies about breastfeeding and violence towards women highlighted for me the importance of Irma van der Ploeg's book. In May 2004 many UK newspapers led their front pages with the news that 'bottle fed babies face higher risk of heart death' (Boseley, 2004). The reports, and the studies upon which they were based, talked about 'breastfeeding' as if it were a practice devoid of any fleshy body, or, at least, the practice of a body that can easily be regulated for the greater good of public health. Missing from the discussion was any reference to 'mothers' and their embodied experiences of breastfeeding: experiences that can often be painful, messy and exhausting. This absence is perhaps one reason why it can seem so easy for policy to have been changed in the UK recently, with the recommended age at which babies should be fed purely on breast milk being increased from four to six months. One week following the headlines about the importance of breastfeed ing, the Sunday edition of the same newspaper carried an article headed 'Increase in Attacks on Pregnant Women' (Revill & Hobson, 2004). This article outlined research reporting midwives' beliefs that they are seeing an increase in attacks on pregnant women by partners who have become angry that they no longer 'have ownership' over the woman's body. The research highlighted how the problem of violence against women has been largely ignored by government and health professionals, even though pregnancy and childbirth can escalate an existing violent relationship and violence can be a response to infant feeding (including breastand bottle feeding) . Irma van der Ploeg's book deals with just such an absenting or deleting of women's bodies and experiences. She focuses on practices that impinge

Abstracts for Poster Presentation

Abstracts for Poster Presentation

Mapping of Binding Site III in the Leptin Receptor and Modeling of a Hexameric Leptin·Leptin Receptor Complex

The leptin.leptin receptor (LR) system shows strong similarities to the long chain cytokine interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF) cytokine.cytokine receptor systems. The IL-6 family cytokines interact with their receptors through three different binding sites (I-III). We demonstrated previously that leptin has similar binding sites I-III and mapped the interactions between binding site II and cytokine receptor homology domain II (CRH2) (Peelman, F., Van Beneden, K., Zabeau, L., Iserentant, H., Ulrichts, P., Defeau, D., Verhee, A., Catteeuw, D., Elewaut, D., and Tavernier, J. (2004) J. Biol. Chem. 279, 41038-41046). In this study, we built homology models for the CRH1 and Ig-like domains of the LR. The Ig-like domain shows a large conserved surface patch in the beta-sheet formed by beta-strands 3, 6, and 7. Mutations in this patch almost completely abolished the leptin-induced STAT3-dependent reporter activity. We propose that a conserved cluster of residues Leu370, Ala407, Tyr409, His417, and His418 forms the center of binding site III of the LR. We built a hexameric leptin.LR complex model based on the hexameric IL-6 complex. In this model, a conserved hydrophobic protuberance of Val36, Thr37, Phe41, and Phe43 in the A-B loop of leptin fits perfectly in the CRH2 domain, corresponding to the IL-6 alpha-receptor, and forms the center of binding site I. The 2:4 hexameric leptin.LR complex offers a rational explanation for mutagenesis studies and residue conservation.

Wild-type Presenilin 1 Protects against Alzheimer Disease Mutation-induced Amyloid Pathology

Mutations in presenilin 1 (PS1) lead to dominant inheritance of early onset familial Alzheimer disease (FAD). These mutations are known to alter the gamma-secretase cleavage of the amyloid precursor protein, resulting in increased ratio of Abeta42/Abeta40 and accelerated amyloid plaque pathology in transgenic mouse models. To investigate the factors that drive the Abeta42/Abeta40 ratio and amyloid pathogenesis and to investigate the possible interactions between wild-type and FAD mutant PS1, which are co-expressed in transgenic animals, we expressed the PS1 M146V knock-in allele either on wild-type PS1 (PS1M146V/+) or PS1 null (PS1M146V/-) background and crossed these alleles with the Tg2576 APP transgenic mice. Introduction of the PS1 M146V mutation on Tg2576 background resulted in earlier onset of plaque pathology. Surprisingly, removing the wild-type PS1 in the presence of the PS1 M146V mutation (PS1M146V/-) greatly exacerbated the amyloid burden; and this was attributed to a reduction of gamma-secretase activity rather than an increase in Abeta42. Our findings establish a protective role of the wild-type PS1 against the FAD mutation-induced amyloid pathology through a partial loss-of-function mechanism.

Equimolar Production of Amyloid β-Protein and Amyloid Precursor Protein Intracellular Domain from β-Carboxyl-terminal Fragment by γ-Secretase

We showed previously that cells expressing wild-type (WT) beta-amyloid precursor protein (APP) or coexpressing WTAPP and WT presenilin (PS) 1/2 produced APP intracellular domains (AICD) 49-99 and 50-99, with the latter predominating. On the other hand, the cells expressing mutant (MT) APP or coexpressing WTAPP and MTPS1/2 produced a greater proportion of AICD-(49-99) than AICD-(50-99). In addition, the expression of amyloid beta-protein (Abeta) 49 in cells resulted in predominant production of Abeta40 and that of Abeta48 leads to preferential production of Abeta42. These observations suggest that epsilon-cleavage and gamma-cleavage are interrelated. To determine the stoichiometry between Abeta and AICD, we have established a 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonic acid-solubilized gamma-secretase assay system that exhibits high specific activity. By using this assay system, we have shown that equal amounts of Abeta and AICD are produced from beta-carboxyl-terminal fragment (C99) by gamma-secretase, irrespective of WT or MTAPP and PS1/2. Although various Abeta species, including Abeta40, Abeta42, Abeta43, Abeta45, Abeta48, and Abeta49, are generated, only two species of AICD, AICD-(49-99) and AICD-(50-99), are detected. We also have found that M233T MTPS1 produced only one species of AICD, AICD-(49-99), and only one for its counterpart, Abeta48, in contrast to WT and other MTPS1s. These strongly suggest that epsilon-cleavage is the primary event, and the produced Abeta48 and Abeta49 rapidly undergo gamma-cleavage, resulting in generation of various Abeta species.

Caspase-3 Cleaves and Inactivates the Glutamate Transporter EAAT2

EAAT2 is a high affinity, Na+-dependent glutamate transporter with predominant astroglial localization. It accounts for the clearance of the bulk of glutamate released at central nervous system synapses and therefore has a crucial role in shaping glutamatergic neurotransmission and limiting excitotoxicity. Caspase-3 activation and impairment in expression and activity of EAAT2 are two distinct molecular mechanisms occurring in human amyotrophic lateral sclerosis (ALS) and in the transgenic rodent model of the disease. Excitotoxicity caused by down-regulation of EAAT2 is thought to be a contributing factor to motor neuron death in ALS. In this study, we report the novel evidence that caspase-3 cleaves EAAT2 at a unique site located in the cytosolic C-terminal domain of the transporter, a finding that links excitotoxicity and activation of caspase-3 as converging mechanisms in the pathogenesis of ALS. Caspase-3 cleavage of EAAT2 leads to a drastic and selective inhibition of this transporter. Heterologous expression of mutant SOD1 proteins linked to the familial form of ALS leads to inhibition of EAAT2 through a mechanism that largely involves activation of caspase-3 and cleavage of the transporter. In addition, we found evidence in spinal cord homogenates of mutant SOD1 ALS mice of a truncated form of EAAT2, likely deriving from caspase-3-mediated proteolytic cleavage, which appeared concurrently to the loss of EAAT2 immunoreactivity and to increased expression of activated caspase-3. Taken together, our findings suggest that caspase-3 cleavage of EAAT2 is one mechanism responsible for the impairment of glutamate uptake in mutant SOD1-linked ALS.

Guide to reform of higher education: a European perspective

Although there are exceptions, most European universities and institutions of higher education find it difficult to compete with the best universities in the Anglo-Saxon world. Despite the Bologna Agreement and the ambitions of the Lisbon Agenda, European universities are in need of fundamental reforms. We look at structural reforms of higher education and propose more effective use of public subsidies, more efficient modes of financing institutions of higher education, more diversity, competition, and transparency, larger private contributions and more equity. In the process we discuss the nature and governance of an institution of higher education, selection, hierarchy in higher education, grade-inflation, fair competition, private and social returns to education, income-contingent loans, equity, and transparency. We sum up with seven recommendations for reform of higher education in Europe. — Bas Jacobs and Frederick van der Ploeg

The Social and Intellectual Construction of Farming Styles: Testing Dutch Ideas in Australian Agriculture

AbstractVan der Ploeg's ‘styles of farming’ approach attempts to explain the social nature of diversity in agriculture. For over 10 years we have attempted to apply this concept to a range of farming situations in Australia. This article reviews the issues posed by Vanclay et al. in 1998 and provides insight into what have been the developments in the concept of farming styles. We utilise the results of three studies in Australia: a study of farming styles in broadacre cropping and grazing farmers in the Riverina region of New South Wales; a study of grape growers in the Sunraysia district of Victoria and a separate study of grape growers across south‐eastern Australia. While this research was intended to validate the existence of styles and to provide evidence of the usefulness of the concept and its application in rural industries, we were unable to replicate or validate van der Ploeg's original conceptualisation and methodology. We conclude that farming styles are more of an intellectual construction than a social construction. Nevertheless, we believe that the concept of farming styles is a useful heuristic and a new framework for understanding farming styles is presented.

Multiple Elements Jointly Determine Inhibitor Selectivity of Cyclic Nucleotide Phosphodiesterases 4 and 7

Phosphodiesterase (PDE) inhibitors have been widely studied as therapeutics for treatment of human diseases. However, the mechanism by which each PDE family recognizes selectively a category of inhibitors remains a puzzle. Here we report the crystal structure of PDE7A1 catalytic domain in complex with non-selective inhibitor 3-isobutyl-1-methylxanthine and kinetic analysis on the mutants of PDE7A1 and PDE4D2. Our studies suggest at least three elements play critical roles in inhibitor selectivity: 1) the conformation and position of an invariant glutamine, 2) the natures of scaffolding residues, and 3) residues that alter shape and size of the binding pocket. Kinetic analysis shows that single PDE7 to PDE4 mutations increase the sensitivity of PDE7 to PDE4 inhibitors but are not sufficient to render the engineered enzymes comparable with the wild types. The triple S373Y/S377T/I412S mutation of PDE7A1 produces a PDE4-like enzyme, implying that multiple elements must work together to determine inhibitor selectivity.

Phosphorylation of GRK1 and GRK7 by cAMP-dependent Protein Kinase Attenuates Their Enzymatic Activities

Phosphorylation of G protein-coupled receptors is a critical step in the rapid termination of G protein signaling. In rod cells of the vertebrate retina, phosphorylation of rhodopsin is mediated by GRK1. In cone cells, either GRK1, GRK7, or both, depending on the species, are speculated to initiate signal termination by phosphorylating the cone opsins. To compare the biochemical properties of GRK1 and GRK7, we measured the K(m) and V(max) of these kinases for ATP and rhodopsin, a model substrate. The results demonstrated that these kinases share similar kinetic properties. We also determined that cAMP-dependent protein kinase (PKA) phosphorylates GRK1 at Ser(21) and GRK7 at Ser(23) and Ser(36) in vitro. These sites are also phosphorylated when FLAG-tagged GRK1 and GRK7 are expressed in HEK-293 cells treated with forskolin to stimulate the endogenous production of cAMP and activation of PKA. Rod outer segments isolated from bovine retina phosphorylated the FLAG-tagged GRKs in the presence of dibutyryl-cAMP, suggesting that GRK1 and GRK7 are physiologically relevant substrates. Although both GRKs also contain putative phosphorylation sites for PKC and Ca(2+)/calmodulin-dependent protein kinase II, neither kinase phosphorylated GRK1 or GRK7. Phosphorylation of GRK1 and GRK7 by PKA reduces the ability of GRK1 and GRK7 to phosphorylate rhodopsin in vitro. Since exposure to light causes a decrease in cAMP levels in rod cells, we propose that phosphorylation of GRK1 and GRK7 by PKA occurs in the dark, when cAMP levels in photoreceptor cells are elevated, and represents a novel mechanism for regulating the activities of these kinases.