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The insulin-like growth factor (IGF) axis is a multicomponent molecular network which has important biological functions in the development and maintenance of differentiated tissue function(s). One of the most important functions of the IGF axis is the control of skeletal tissue metabolism by the finely tuned regulation of the process of osteogenesis. To achieve this, the IGF axis controls the activity of several cell types-osteoprogenitor cells, osteoblasts, osteocytes and osteoclasts to achieve the co-ordinated development of appropriate hard tissue structure and associated matrix deposition. In addition, there is an increasing awareness that the IGF axis also plays a role in the process of odontogenesis (tooth formation). In this review, we highlight some of the key findings in both of these areas. A further understanding of the role of the IGF axis in hard tissue biology may contribute to tissue regeneration strategies in cases of skeletal tissue trauma. [ABSTRACT FROM AUTHOR]

Conformational transitions in a 4-way DNA junction when titrated with ionic solutions are studied using time-resolved fluorescence resonance energy transfer. Parameters characterising the transition in terms of critical ion concentration ( c 1/2) and the Hill coefficient for ion binding are obtained by fitting a simple two-state model using steady-state spectra. Data obtained from a fluorescence lifetime plate reader and analysed by fitting a single exponential to donor fluorescence lifetime decays are shown to be in good agreement with the parameters obtained from steady-state measurements. Fluorescence lifetimes, however, offer advantages, particularly in being independent of fluorophore concentration, output intensity, inhomogeneity in the excitation source and output wavelength. We demonstrate preliminary FRET-FLIM images of DNA junction solutions obtained using a picosecond gated CCD which are in agreement with results from a fluorescence lifetime plate reader. The results suggest that time-resolved FRET-FLIM is sensitive to subtle structural changes and may be useful in assays based on 4-way DNA junctions. [ABSTRACT FROM AUTHOR]

Objective:This paper describes baseline data on basal metabolic rate (BMR), thyroid hormone levels and body composition of middle-aged and older people participating in the ZENITH project and the correlation of thyroid hormone levels with zinc status.Design:A multicentre prospective intervention study employing a randomised double blind design.Setting:Clermont-Ferrand, Theix (France), Coleraine (Northern Ireland), Grenoble (France), Rome (Italy).Interventions:BMR has been measured on a subsample of 70 middle-aged volunteers (35 men and 35 women recruited in Clermont-Ferrand, France, aged 55–70 y) and 108 older volunteers (56 men and 52 women recruited in Rome, Italy, aged 70–85 y). Thyroid hormone levels were evaluated in the entire group of ZENITH volunteers (n=387). BMR was measured by indirect calorimetry. Fat-free mass (FFM) was derived by four skinfold thicknesses using Durnin and Womersley's equations. Concentrations of thyroid hormones (total T3 and T4) were measured using a competitive immunoassay with an enhanced chemiluminescence end point.Results:Italian older volunteers had a significantly lower FFM than middle-aged French volunteers (−7% P<0.01). A negative correlation between BMR and age (men, r= −0.64; women, r= −0.62; both P<0.0001) was observed: BMR was significantly (P<0.000001) lower in Italian elderly volunteers (4.03±0.46 kJ/min and 3.29±0.42 kJ/min for men and women, respectively) than in middle-aged French volunteers (4.84±0.45 kJ/min and 3.87±0.38 kJ/min for men and women, respectively), even after adjustment for FFM (−12%). No correlation has been observed between BMR and thyroid hormones both in French and Italian subjects. Total T4 (TT4) concentrations were lowest in middle-aged population (−10%, P<0.0001). A moderate negative correlation has been found with TT4 and red blood cell zinc (r=−0.12, P<0.02, slope −0.026).Conclusions:The results confirm an age-related decline in BMR not entirely explained by body composition or thyroid hormones differences.Sponsorship:The ZENITH study is supported by the European Commission ‘Quality of Life and Management of Living Resources’, Fifth Framework Programme, Contract no. QLK1-CT-2001-00168.European Journal of Clinical Nutrition (2005) 59, Suppl 2, S53–S57. doi:10.1038/sj.ejcn.1602299 [ABSTRACT FROM AUTHOR]

OBJECTIVE:: To determine copper absorption from copper containing foods labelled either intrinsically or extrinsically with a highly enriched Cu-65 stable isotope label. DESIGN:: A longitudinal cross-over study. SETTING:: The study was conducted at the Institute of Food Research, Human Nutrition Unit, Norwich, UK. SUBJECTS:: Subjects were recruited locally via advertisements placed around the Norwich Research Park. A total of 10 volunteers (nine female, one male) took part in the study, but not all volunteers completed each of the test meals. INTERVENTIONS:: A highly enriched Cu-65 stable isotope label was administered to volunteers in the form of a reference dose or in breakfast test meals consisting of red wine, soya beans, mushrooms or sunflower seeds. Faecal monitoring and mass spectrometry techniques were used to estimate the relative quantities of copper absorbed from the different test meals. RESULTS:: True copper absorption from the reference dose (54%) was similar to extrinsically labelled red wine (49%) and intrinsically labelled sunflower seeds (52%), but significantly higher than extrinsically labelled mushrooms (35%), intrinsically (29%) and extrinsically (15%) labelled soya beans and extrinsically labelled sunflower seed (32%) test meals. CONCLUSIONS:: The use of Cu-65 extrinsic labels in copper absorption studies requires validation according to the food being examined; intrinsic and extrinsic labelling produced significantly different results for sunflower seeds. SPONSORSHIP:: Funding from the European Commission and the Biotechnology and Biological Sciences Research Council.European Journal of Clinical Nutrition (2005) 59, 363-368. doi:10.1038/sj.ejcn.1602082 Published online 3 November 2004 [ABSTRACT FROM AUTHOR]

Hexadecane-in-water emulsion droplets were formed in a homogeniser in the presence of a mixture of an anionic surfactant (sodium dodecyl sulfate, SDS) and nonionic surfactants of various chain lengths [nonylphenol ethoxylate (C9φEN, N=100, 40 and 30) or an alcohol ethoxylate (Brij35)]. The dynamic mobility of the oil droplets was then measured using a flow-through version of an AcoustoSizer. Large changes were observed in the dynamic mobility of the particles formed with the mixed surfactants compared to particles formed with SDS alone. O'Brien's “gel layer” model was employed to interpret the data. The characteristics of the adsorbed layer appeared to be similar whether the nonionic surfactant was adsorbed concurrently with the SDS as the emulsion formed or was merely added afterwards to the emulsion established. The particle size, the charge and the molar fraction of SDS had virtually no effect. The layers formed with the nonionic surfactants decreased in thickness with decreasing molecular weight as expected. Passage through the homogeniser itself had no effect on the properties of the largest nonionic surfactant and, hence, on the adsorption layer formed with it. [ABSTRACT FROM AUTHOR]

Using monolayer cultures of costal chondrocytes established from four week old Clun Forest lambs, we have demonstrated that, under serum free conditions the cells release three IGFBPs (32, 29 and 21 kDa) into the medium. The most abundant of these-the 32 kDa BP-was shown to be IGFBP-2 by Western blotting. Furthermore we demonstrate that the levels of IGFBP 2 in conditioned medium are acutely increased (6, 12 and 24 h time points) following treatment of cells with bovine GH (1-100 ng/ml). In a parallel set of experiments, using ovine fibroblasts (derived from dermis) we show that IGFBPs of Mr 32, 29 and 21 kDa are also secreted by this cell type. However the relative abundance of these BPs differed from that seen in the chondrocyte cultures, with the 21 kDa species now the most abundant. In addition, prolonged exposure of autoradiographs indicated that fibroblasts secreted a higher Mr IGFBP (most probably ovine BP-3) that was not detected in any of our chondrocyte cultures. Most significant however was the demonstration that bGH did not dramatically affect the levels of IGFBPs in fibroblast cell cultures. We conclude that GH stimulates BP-2 production from chondrocytes and this is a cell-type specific effect in as much as it is not replicated in cultures of dermal fibroblasts. [ABSTRACT FROM AUTHOR]

Using a solution phase assay we have demonstrated that sheep adipose tissue explants secrete insulin-like growth factor binding proteins (IGFBPs) when cultured in serum-free medium over a 24 h period. Further, we demonstrate that secretion of IGFBP(s) is inhibited (up to 50%) by incubation of the cultures in the presence of 10M dexamethasone. This inhibitory effects is overcome when insulin (10 ng/ml) and ovine growth hormone (100 ng/ml) are incubated together (but not separately) with glucocorticoid. Further characterisation of this IGF binding activity by high performance size exclusion chromatography and Western ligand blot analysis indicated that under our culture conditions sheep adipose tissue explants secrete one predominant 21 kDa IGFBP and it is this BP which is hormonally regulated as described above. We discuss our results in the context of endocrine/paracrine/autocrine control of adipose tissue metabolism and differentiation. [ABSTRACT FROM AUTHOR]

Pharmacological therapy of osteoporosis reduces bone loss and risk of fracture in patients. Modulation of bone mineral density cannot explain all effects. Other aspects of bone quality affecting fragility and ways to monitor them need to be better understood. Keratinous tissue acts as surrogate marker for bone protein deterioration caused by oestrogen deficiency in rats. Ovariectomised rats were treated with alendronate (ALN), parathyroid hormone (PTH) or estrogen (E2). MicroCT assessed macro structural changes. Raman spectroscopy assessed biochemical changes. Micro CT confirmed that all treatments prevented ovariectomy-induced macro structural bone loss in rats. PTH induced macro structural changes unrelated to ovariectomy. Raman analysis revealed ALN and PTH partially protect against molecular level changes to bone collagen (80% protection) and mineral (50% protection) phases. E2 failed to prevent biochemical change. The treatments induced alterations unassociated with the ovariectomy; increased beta sheet with E2, globular alpha helices with PTH and fibrous alpha helices with both ALN and PTH. ALN is closest to maintaining physiological status of the animals, while PTH (comparable protective effect) induces side effects. E2 is unable to prevent molecular level changes associated with ovariectomy. Raman spectroscopy can act as predictive tool for monitoring pharmacological therapy of osteoporosis in rodents. Keratinous tissue is a useful surrogate marker for the protein related impact of these therapies.The results demonstrate utility of surrogates where a clear systemic causation connects the surrogate to the target tissue. It demonstrates the need to assess broader biomolecular impact of interventions to examine side effects. Osteoporotic treatments exhibit substantial differences in biochemical impact.Alendronate preserved the bone tissue in the state closest to the sham group.Parathyroid hormone prevents ovariectomy changes, induces different changes.Estrogen preserves tissue macro structure, but unable to prevent biochemical changes.Systemic conditions affect structural proteins in both bone and claw. [ABSTRACT FROM AUTHOR]

Herbage analysis offers a definitive means of determining the N, P, K and S status of perennial ryegrass swards. Unfortunately, the results of such analyses can be difficult to interpret, simply because theminimum or 'critical' concentration of a nutrient in plant tissue for optimum growth, varies both with crop age and with changes in the concentrations of other nutrients. The Diagnosis and Recommendation Integrated System (DRIS) could help to improve the reliability of such interpretations. Diagnoses made using DRIS are based on relative rather than on absolute concentrations of nutrients in plant tissue, and as such should be comparatively independent of crop age.The aim of this study was to establish and test DRIS methodology for high-yieldingperennial ryegrass swards. Because of prohibitive costs, setting up a whole new series of field experiments to evaluate DRIS model parameters for perennial ryegrass was out of the question. Instead, the diagnostic norms and associated coefficients of variation for the model were evaluated using data from a single (large) multi-factorial glasshouse experiment.Of the nutrient ratios selected to form the diagnostic norms, K/N and S/N had the clearest physiological rationale, whereas those involving Ca and Mg in combination with N, P, K and S appeared to have little physiological basis. It was reasoned, though, that because Ca and Mg uptake by plants are largely passive processes (ultimately governed by plant growth), the DRIS indices for these nutrients, together reflected the degree to which growth may be limited by non-nutritional (environmental) factors relative to nutritional ones. Both indices were combined to form a single reference (Ri)index. Without such an internal reference, plant growth could be limited by multiple nutrient deficiencies, and yet N, P, K and S indicesmight all be close to, or equal to zero (i.e. the optimum), simply because the absolute concentrations of each nutrient (while low) had been [ABSTRACT FROM AUTHOR]

A DRIS model for perennial ryegrass, based on data collected from a glasshouse experiment, was used to diagnose the nutrient sufficiency status of perennial ryegrass swards growing in field situations. Initially, DRIS overestimated the N and P status of these swards and underestimated their S status. However, by calibrating the model outputs on the basis of actual sward dry matter yield responses to specific fertiliser inputs, correction factors were evaluated, and the nutrientindices modified to reflect the nutritional status of swards in field situations.Modified DRIS diagnoses of the N, P, K and S status of swards were compared with those made using the critical value approach. DRIS proved to be as reliable as the critical value approach at diagnosing N deficiency (both approaches having reliability scorings of 90%), but was superior to the latter at diagnosing P, K and S deficiencies, having reliability scorings of 100% (P), 90% (K) and 70% (S), compared with scorings of only 0% (P), 80% (K) and 30% (S), for the critical value approach. [ABSTRACT FROM AUTHOR]