Bell, J, Besong, AA, Tipper, JL, Ingham, E, Wroblewski, BM, Stone, MH & Fisher, J 2000, 'Influence of gelatin and bovine serum lubricants on ultra-high molecular weight polyethylene wear debris generated in in vitro simulations', Proceedings of the Institution of Mechanical Engineers, Part H: Journal of Engineering in Medicine, vol. 214, no. 5, pp. 513-518.
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Ultra-high molecular weight polyethylene (UHMWPE) wear debris induced osteolysis has a major role in the late aseptic loosening and ultimate failure of total hip replacements (THR). Clinically relevant in vitro simulations of wear are essential to predict the osteolytic potential of bearing surfaces in artificial hip joints. Newborn calf or bovine serum has been accepted as a boundary lubricant for such in vitro tests, but its biological stability has been questioned. This study compared the wear factors, number of wear particles and levels of microbial contamination produced in bovine serum and a gelatin-based lubricant. The wear factors produced by the two lubricants were not significantly different, however the wear debris morphology produced was substantially different. The bovine serum became contaminated with micro-organisms within 28 h, whereas the protein-based lubricant remained uncontaminated. The results showed that bovine serum was not a stable boundary lubricant. They also showed that although the wear factors for the two solutions were not significantly different, the protein-based lubricant was not a suitable alternative to bovine serum because the wear debris produced was not clinically relevant.
Dr. Center, JR, Nguyen, TV, Sambrook, PN & Eisman, JA 2000, 'Hormonal and Biochemical Parameters and Osteoporotic Fractures in Elderly Men', Journal of Bone and Mineral Research, vol. 15, no. 7, pp. 1405-1411.
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Abstract Low testosterone has been associated with hip fracture in men in some studies. However, data on other hormonal parameters and fracture outcome in men is minimal. This study examined the association between free testosterone (free T) estradiol (E2), sex hormone-binding globulin (SHBG), 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), insulin-like growth factor I (IGF-I), and fracture in 437 elderly community-dwelling men. Age, height, weight, quadriceps strength, femoral neck bone mineral density (FN BMD), and fracture data (1989–1997) also were obtained. Fractures were classified as major (hip, pelvis, proximal tibia, multiple rib, vertebral, and proximal humerus) or minor (remaining distal upper and lower limb fractures). Fifty-four subjects had a fracture (24 major and 30 minor). There was no association between minor fractures and any hormonal parameter. Risk of major fracture was increased 2-fold for each SD increase in age, decrease in weight and height, and increase in SHBG, and risk of major fracture was increased 3-fold for each SD decrease in quadriceps strength, FN BMD, and 25(OH)D (univariate logistic regression). Independent predictors of major fracture were FN BMD, 2.7 (1.5–4.7; odds ratio [OR]) and 95% confidence interval [CI]); 25(OH)D, 2.8 (1.5–5.3); and SHBG, 1.7 (1.2–2.4). An abnormal value for three factors resulted in a 30-fold increase in risk but only affected 2% of the population. It is not immediately apparent how 25(OH)D and SHBG, largely independently of BMD, may contribute to fracture risk. They may be markers for biological age or health status not measured by methods that are more traditional and as such may be useful in identifying those at high risk of fracture.
Dr. Nguyen, TV & Eisman, JA 2000, 'Genetics of Fracture: Challenges and Opportunities', Journal of Bone and Mineral Research, vol. 15, no. 7, pp. 1253-1256.
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Heise, CE, O'Dowd, BF, Figueroa, DJ, Sawyer, N, Nguyen, T, Im, D-S, Stocco, R, Bellefeuille, JN, Abramovitz, M, Cheng, R, Williams, DL, Zeng, Z, Liu, Q, Ma, L, Clements, MK, Coulombe, N, Liu, Y, Austin, CP, George, SR, O'Neill, GP, Metters, KM, Lynch, KR & Evans, JF 2000, 'Characterization of the Human Cysteinyl Leukotriene 2 Receptor', Journal of Biological Chemistry, vol. 275, no. 39, pp. 30531-30536.
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The contractile and inflammatory actions of the cysteinyl leukotrienes (CysLTs), LTC4, LTD4, and LTE4, are thought to be mediated through at least two distinct but related CysLT G protein-coupled receptors. The human CysLT1 receptor has been recently cloned and characterized. We describe here the cloning and characterization of the second cysteinyl leukotriene receptor, CysLT2, a 346-amino acid protein with 38% amino acid identity to the CysLT1 receptor. The recombinant human CysLT2 receptor was expressed in Xenopus oocytes and HEK293T cells and shown to couple to elevation of intracellular calcium when activated by LTC4, LTD4, or LTE4. Analyses of radiolabeled LTD4 binding to the recombinant CysLT2 receptor demonstrated high affinity binding and a rank order of potency for competition of LTC4 = LTD4 >> LTE4. In contrast to the dual CysLT1/CysLT2 antagonist, BAY u9773, the CysLT1 receptor-selective antagonists MK-571, montelukast (Singulair(TM)), zafirlukast (Accolate(TM)), and pranlukast (Onon(TM)) exhibited low potency in competition for LTD4 binding and as antagonists of CysLT2 receptor signaling. CysLT2 receptor mRNA was detected in lung macrophages and airway smooth muscle, cardiac Purkinje cells, adrenal medulla cells, peripheral blood leukocytes, and brain, and the receptor gene was mapped to chromosome 13q14, a region linked to atopic asthma.
HUTVÁGNER, G, MLYNÁROVÁ, L & NAP, J-P 2000, 'Detailed characterization of the posttranscriptional gene-silencing-related small RNA in a GUS gene-silenced tobacco', RNA, vol. 6, no. 10, pp. 1445-1454.
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Posttranscriptional gene-silencing phenomena such as cosuppression and RNA interference are associated with the occurrence of small, about 21-23 nt short RNA species homologous to the silenced gene. We here show that the small RNA present in silenced transgenic plants can easily be detected in total RNA isolated according to standard procedures. This will allow for the development of routine and early screenings for the presence of small RNA species and, therefore, gene silencing in transgenic plants. We further demonstrate that the small RNA fraction can be visualized with the SYBR Green II RNA stain, isolated from a gel, labeled and used as a specific probe. Using these approaches, we have fine-mapped the sequences of the GUS gene that are represented in the small RNA fraction of a GUS-silenced tobacco line containing an inverted repeat of the GUS gene. In this tobacco line, the silencing-associated small RNA is a mixture of fragments that cover the 3' two-thirds of the GUS coding region. The 5' coding and the 3' noncoding ends of the GUS mRNA are not represented in the small RNA fraction. The RNA fragments are not likely to be a primary synthesis product of an RNA-dependent RNA polymerase, but rather degradation products from nuclease activity. Surprisingly, RNA isolated from wild-type, untransformed plants showed the presence of a similar-sized small RNA pool. This might indicate the existence of small RNA species from putative endogenous genes that are down regulated by a similar posttranscriptional gene-silencing mechanism. The possibility of isolating and labeling the small RNA pool from wild-type plants will provide a way to identify and study such putative genes.
Jones, G & Nguyen, TV 2000, 'Associations Between Maternal Peak Bone Mass and Bone Mass in Prepubertal Male and Female Children', Journal of Bone and Mineral Research, vol. 15, no. 10, pp. 1998-2004.
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Abstract The aim of this study was to estimate heritability of bone density in premenopausal women, prepubertal male, and prepubertal female child pairs. We studied 291 pairs (mothers, mean age, 33 years, range 22–45 years; children, mean age, 7.92 years, range 7.32-8.92 years). Bone density and body composition were assessed by dual-energy X-ray absorptiometry. Height and weight were measured in both mother and child. Body size-adjusted heritability estimates for areal bone density (g/cm2) were all statistically significant (femoral neck, 59%; lumbar spine, 38%; total body, 41%) and were consistently and significantly higher in mother-daughter pairs (n = 105) as compared with mother-son pairs (n = 186). Heritability estimates for bone mineral apparent density (BMAD; g/cm3) were marginally lower but remained statistically significant at all sites (femoral neck, 51%; lumbar spine, 32%; total body, 38%). Maternal osteopenia was associated with significant reductions in bone mass at all sites in the children (femoral neck, 0.75 SD and p < 0.0001; lumbar spine, 0.61 SD and p < 0.0001; total body, 0.43 SD and p = 0.012). Mother-child bone areal bone density correlation coefficients and prediction of low bone mass in the child were greater (but this did not reach statistical significance) if the corresponding anatomical site in the mother was used for prediction with the exception of the total body. These data confirm that heritability of bone mass extends to prepubertal children and is gender- and possibly site-specific as well as under separate genetic control to growth. Furthermore, the strength of the mother-child association is such that bone density screening of mothers would make it possible to identify most prepubertal children at higher risk of osteoporosis in later life.
Landers, P, Kerr, KG, Rowbotham, TJ, Tipper, JL, Keig, PM, Ingham, E & Denton, M 2000, 'Survival and Growth of Burkholderia cepacia Within the Free-Living Amoeba Acanthamoeba polyphaga', European Journal of Clinical Microbiology & Infectious Diseases, vol. 19, no. 2, pp. 121-123.
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Lee, DK, Cheng, R, Nguyen, T, Fan, T, Kariyawasam, AP, Liu, Y, Osmond, DH, George, SR & O'Dowd, BF 2000, 'Characterization of Apelin, the Ligand for the APJ Receptor', Journal of Neurochemistry, vol. 74, no. 1, pp. 34-41.
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Abstract: The apelin peptide was recently discovered and demonstrated to be the endogenous ligand for the G protein‐coupled receptor, APJ. A search of the GenBank databases retrieved a rat expressed sequence tag partially encoding the preproapelin sequence. The GenBank search also revealed a human sequence on chromosome Xq25‐26.1, containing the gene encoding preproapelin. We have used the rat sequence to screen a rat brain cDNA library to obtain a cDNA encoding the full‐length open reading frame of rat preproapelin. This cDNA encoded a protein of 77 amino acids, sharing an identity of 82% with human preproapelin. Northern and in situ hybridization analyses revealed both human and rat apelin and APJ to be expressed in the brain and periphery. Both sequence and mRNA expression distribution analyses revealed similarities between apelin and angiotensin II, suggesting they that share related physiological roles. A synthetic apelin peptide was injected intravenously into male Wistar rats, resulting in immediate lowering of both systolic and diastolic blood pressure, which persisted for several minutes. Intraperitoneal apelin injections induced an increase in drinking behavior within the first 30 min after injection, with a return to baseline within 1 h.
Lee, DK, Lynch, KR, Nguyen, T, Im, D-S, Cheng, R, Saldivia, VR, Liu, Y, Liu, ISC, Heng, HHQ, Seeman, P, George, SR, O’Dowd, BF & Marchese, A 2000, 'Cloning and characterization of additional members of the G protein-coupled receptor family', Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression, vol. 1490, no. 3, pp. 311-323.
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A search of the expressed sequence tag (EST) database retrieved a human cDNA sequence which partially encoded a novel G protein-coupled receptor (GPCR) GPR26. A human genomic DNA fragment encoding a partial open reading frame (ORF) and a rat cDNA encoding the full length ORF of GPR26 were obtained by library screening. The rat GPR26 cDNA encoded a protein of 317 amino acids, most similar (albeit distantly related) to the serotonin 5- HT(5A) and gastrin releasing hormone BB2 receptors. GPR26 mRNA expression analysis revealed signals in the striatum, pons, cerebellum and cortex. HEK293 and Rh7777 cells transfected with GPR26 cDNA displayed high basal cAMP levels, slow growth rate of clonal populations and derangements of normal cell shape. We also used a sequence reported only in the patent literature encoding GPR57 (a.k.a. HNHCI32) to PCR amplify a DNA fragment which was used to screen a human genomic library. This resulted in the cloning of a genomic fragment containing a pseudogene, ψGPR57, with a 99.6% nucleotide identity to GPR57. Based on shared sequence identities, the receptor encoded by GPR57 was predicted to belong to a novel subfamily of GPCRs together with GPR58 (a.k.a. phBL5, reported only in the patent literature), putative neurotransmitter receptor (PNR) and a 5-HT4 pseudogene. Analysis of this subfamily revealed greatest identities (~56%) between the receptors encoded by GPR57 and GPR58, each with shared identities of ~40% with PNR. Furthermore, ψGPR57, GPR58, PNR and the 5-HT4 pseudogene were mapped in a cluster localized to chromosome 6q22-24. PNR and GPR58 were expressed in COS cells, however no specific binding was observed for various serotonin receptor-specific ligands. (C) 2000 Elsevier Science B.V.
Lee, S 2000, 'Oligomerization of Dopamine and Serotonin Receptors', Neuropsychopharmacology, vol. 23, no. 4, pp. S32-S40.
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Until recently, it has largely been assumed that G proteincoupled receptors (GPCRs) function as monomeric entities. However, over the past few years, we and others havedocumented that GPCRs can form dimers and oligomers, leading to a re-evaluation of the mechanisms thought tomediate GPCR function. Despite the growing number ofinvestigations into dimerization, little is known about thestructural basis of receptor-receptor interactions and thefunctional consequences of dimer formation. Here, wepresent a brief review of some insights we have gained intothe dimerization of dopamine and serotonin receptors. Wehave demonstrated that agonist-regulated trafficking isidentical for receptor monomers and dimers, however, agonist treatment appears to stabilise the receptor oligomers. An investigation of the structural assemblybetween receptors involved in dimerization showed thatthere are several sites of interaction including hydrophobictransmembrane domain interactions and intermoleculardisulphide bonds. We have also examined receptor hetero-oligomerization and demonstrated the potential for novelfunctions as a result of these associations. Finally, as aresult of these observations, we have been able to presentevidence that GPCRs function as oligomers in the cell. © 2000, American College of Neuropsychopharmacology.
Lee, SP, O'Dowd, BF, Ng, GYK, Varghese, G, Akil, H, Mansour, A, Nguyen, T & George, SR 2000, 'Inhibition of Cell Surface Expression by Mutant Receptors Demonstrates that D2 Dopamine Receptors Exist as Oligomers in the Cell', Molecular Pharmacology, vol. 58, no. 1, pp. 120-128.
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Numerous mutant G protein-coupled receptors with diminished or no function have been described that are naturally occurring or that are the product of gene manipulation. It has largely been assumed that receptor mutants do not affect the function of the wild-type receptor; however, the occurrence of G protein-coupled receptor dimerization suggests the possibility that an intermolecular interaction between mutant and wild-type receptors can occur. We have shown previously that the D2 dopamine receptor (D2DR) exists as dimers in cell lines and brain tissue. In this study, we demonstrated that mutant D2DR can modulate the function of the wild-type D2DR. While attempting to elucidate the structure of the D2DR dimer, we demonstrated that nonfunctional D2DR substitution and truncation mutants antagonized wild-type D2DR function. Furthermore, from analyses of this interaction between the receptor mutants and the D2DR, using photoaffinity labeling, we provide evidence that the D2DR is oligomeric in the cell.
Mcnie, CM, Barton, DC, Ingham, E, Tipper, JL, Fisher, J & Stone, MH 2000, 'The prediction of polyethylene wear rate and debris morphology produced by microscopic asperities on femoral heads', Journal of Materials Science: Materials in Medicine, vol. 11, no. 3, pp. 163-174.
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Nguyen, TV, Blangero, J & Eisman, JA 2000, 'Genetic Epidemiological Approaches to the Search for Osteoporosis Genes', Journal of Bone and Mineral Research, vol. 15, no. 3, pp. 392-401.
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Abstract Important progress has been made in the identification of specific environmental factors and estimation of hereditary components in bone density, quantitative ultrasound (QUS), and bone turnover indices. By contrast, the search for specific genes that regulate bone mass has progressed rather slowly, and the results are more difficult to interpret and reproduce. This article reviews the genetics of osteoporosis and problems plaguing genetic research. It is argued that the search for genes involved in the expression of osteoporotic phenotypes should be based on linkage studies in relatively homogeneous populations. Strategies for increasing the power of studies, such as making use of information from extended pedigrees and multivariate analysis, are discussed. With the advent of a comprehensive human genetic linkage map, a complete identification of genes for osteoporosis appears feasible. Understanding the genetic mechanisms and their interactions with environmental factors should allow more focused and cost-effective osteoporosis prevention and treatment strategies.
Nguyen, TV, Center, JR & Eisman, JA 2000, 'Association between breast cancer and bone mineral density: the Dubbo Osteoporosis Epidemiology Study', Maturitas, vol. 36, no. 1, pp. 27-34.
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Nguyen, TV, Pocock, NA & Eisman, JA 2000, 'Interpretation of Bone Mineral Density Measurement and Its Change', Journal of Clinical Densitometry, vol. 3, no. 2, pp. 107-119.
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O'Dowd, BF, Lee, DK, Huang, W, Nguyen, T, Cheng, R, Liu, Y, Wang, B, Gershengorn, MC & George, SR 2000, 'TRH-R2 exhibits similar binding and acute signaling but distinct regulation and anatomic distribution compared with TRH-R1.', Mol Endocrinol, vol. 14, no. 1, pp. 183-193.
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TRH (thyroliberin) is a tripeptide (pGlu-His-ProNH2) that signals via G protein-coupled receptors. Until recently, only a single receptor for TRH was known (TRH-R1), but two groups identified a second receptor, TRH-R2. We independently discovered TRH-R2. Using an extensive set of TRH analogs, we found no differences in TRH-R1 and TRH-R2 binding or in acute stimulation of signaling. TRH-R2 was more rapidly internalized upon binding TRH and exhibited a greater level of TRH-induced down-regulation than TRH-R1. During prolonged exposure to TRH, cells expressing TRH-R2 exhibited a lower level of gene induction than cells expressing TRH-R1. TRH-R2 receptor mRNA was present in very discrete nuclei and regions of rat brain. A major mRNA transcript for TRH-R2 was seen in the cerebral cortex, pons, thalamus, hypothalamus, and midbrain with faint bands found in the striatum and pituitary. The extensive distribution of TRH-R2 in the brain suggests that it mediates many of the known functions of TRH that are not transduced by TRH-R1. The variations in agonist-induced internalization and down-regulation/desensitization, and anatomic distribution of TRH-R2 compared with TRH-R1, suggest important functional differences between the two receptors.
Randell, AG, Nguyen, TV, Bhalerao, N, Silverman, SL, Sambrook, PN & Eisman, JA 2000, 'Deterioration in Quality of Life Following Hip Fracture: A Prospective Study', Osteoporosis International, vol. 11, no. 5, pp. 460-466.
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Thompson, MD, Gonzalez, N, Nguyen, T, Comings, DE, George, SR & O'Dowd, BF 2000, 'Serotonin transporter gene polymorphisms in alcohol dependence', Alcohol, vol. 22, no. 2, pp. 61-67.
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The serotonin transporter (5-HTT) gene is a candidate gene in alcohol dependence because serotonin reuptake inhibitors (SRIs) can alleviate alcohol withdrawal. Studies of the 5-HTT gene in alcohol dependence have not resulted in a consensus. Recent studies have examined the transcriptionally active promoter polymorphism, a 44-bp deletion resulting in short (S) or long (L) alleles. In this study, 131 alcohol-dependent patients of Northern and Western European descent were genotyped. Seventy of these patients were diagnosed with alcohol dependence without comorbid disorders. Sixty-one patients were diagnosed with alcohol dependence comorbid with Tourette syndrome (alcoholic-TS). We found an excess of the S allele in alcohol-dependent patients (47%) compared with 125 ethnically matched controls (39%). A similar trend was found in 150 ethnically matched TS patients without alcohol dependence comorbidity (51%). However, the statistical significance of this trend in the data was not present after Bonferroni correction. The data presented suggests a trend toward increased frequency of the S promoter allele in alcohol-dependent, alcoholic-TS and TS patients. (C) 2000 Elsevier Science Inc.
Tipper, JL, Ingham, E, Hailey, JL, Besong, AA, Fisher, J, Wroblewski, BM & Stone, MH 2000, 'Quantitative analysis of polyethylene wear debris, wear rate and head damage in retrieved Charnley hip prostheses', Journal of Materials Science: Materials in Medicine, vol. 11, no. 2, pp. 117-124.
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Submicrometer- and micrometer-sized ultra-high molecular weight polyethylene (UHMWPE) wear particles have been associated with osteolysis and failure of total artificial joints. Previous studies have isolated predominantly submicrometer-sized particles at the expense of larger particles ( > 10 μm). This study aimed to isolate and characterize quantitatively all sizes of UHMWPE wear particles generated in 18 Charnley hip prostheses. In addition, to analyze the wear debris with respect to the total volumetric wear of the cup and damage to the femoral head. Particle size distributions ranged from 0.1 to - > 1000 μm. A significant proportion (3-82%) of the mass of the wear debris isolated was > 10 μm. The mode of the frequency distribution of the particles was in the range 0.1-0.5 μm for all patients. However, analysis of the mass of wear debris as a function of its size allowed differentiation of the wear debris from different patients. Femoral head damage was associated with high volumetric wear and increased numbers of biologically active submicrometer-sized particles. (C) 2000 Kluwer Academic Publishers.
Tran, T 2000, 'Risk factors for postcesarean surgical site infection', Obstetrics & Gynecology, vol. 95, no. 3, pp. 367-371.
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