PubMed Health⌕ Search

Biomedical subjects

G J Arnold

Publications and source records attributed to G J Arnold.

At least 19 recordsLinked to original sources
PubMed2006

Proteome research based on modern liquid chromatography--tandem mass spectrometry: separation, identification and quantification.

Recent developments of new generations of mass spectrometers and improvements in the field of chromatography have revolutionized protein analytics. Particularly the combination of liquid chromatography as a separation tool for proteins and peptides with tandem mass spectrometry as an identification tool referred to as LC-MS/MS has generated a powerful and broadly used technique in the field of proteomics. The resolution and sensitivity of state-of-the-art LC-MS/MS systems has reached dimensions allowing not only the analysis of individual proteins but also investigations on the level of complete proteomes. However, the enormous complexity and the extreme concentration range of proteins within typical eukaryotic proteomes are still the major challenge of this technique. This review gives an overview of modern LC-MS/MS based proteomics, describing state-of-the-art chromatography and modern mass spectrometry. Strategies to perform quantitative proteomics will be presented and capabilities as well as current limitations of this innovative methodology will be discussed.

Animals↗
PubMed2005

Functional genomics: tools for improving farm animal health and welfare.

The first genome sequence assemblies of farm animal species are now accessible through public domain databases, and further sequencing projects are in rapid progress. In addition, large collections of expressed sequences have been obtained, which will aid in constructing annotated transcript maps for many economically important species. Thus, the breeding of farm animals is entering the post-genome era. Functional genomics, defined as applying global experimental approaches to assess gene function, by using the information and reagents provided by structural genomics (i.e. mapping and sequencing), has become the focus of interest. Combining a holistic view of phenotypes at the molecular level with genetic marker data seems a particularly promising approach for improving health and welfare traits in farm animals. These traits are often difficult to define. They suffer from low heritabilities and a corresponding lack of genetic gain in conventional selection and breeding programmes. At the same time, genomic information from micro-organisms and parasites offers the potential for new vaccines and therapeutics. This review describes major functional genomics tools, lists genomic resources available for farm animals and discusses the prospects and challenges of functional genomics in improving the health and welfare of farm animals.

Animal Welfare↗
PubMed2003

Embryo-maternal communication in bovine - strategies for deciphering a complex cross-talk.

Early embryonic development, implantation and maintenance of a pregnancy are critically dependent on an intact embryo-maternal communication. So far, only few signals involved in this dialogue have been identified. In bovine and other ruminants, interferon tau is the predominant embryonic pregnancy recognition signal, exhibiting antiluteolytic activity. However, this is just one aspect of the complex process of embryo-maternal signalling, and a number of other systems are more likely to be involved. To gain a more comprehensive understanding of these important mechanisms, integrated projects involving specialists in embryology, reproductive biotechnology and functional genome research are necessary to perform a systematic analysis of interactions between pre-implantation stage embryos and oviduct or uterine epithelial cells, respectively. State-of-the-art transcriptomic and proteomic technologies will identify reciprocal signals between embryos and their maternal environment and the respective downstream reaction cascades. For in vivo studies, the use of monozygotic twins as recipient animals provides elegant model systems, thus eliminating genetic variability as a cause of differential gene expression. In addition, suitable systems for the co-culture of oviduct epithelial or endometrium cells with the respective embryonic stages need to be established for functional validation of candidate genes potentially involved in the dialogue between embryos and their maternal environment. The knowledge of these mechanisms should help to increase the pregnancy rate following embryo transfer and to avoid embryonic losses. Candidate genes involved in embryo-maternal communication will also be used to define new quality criteria for the selection of embryos for transfer to recipients. Another application is the supplementation of embryotrophic factors or components of embryo-maternal signalling in optimized formulations, such as bioartificial matrices. As a long-term goal, signalling mechanisms identified in bovine will also be functionally evaluated in other species, including the human.

Animals↗
PubMed2001

Insulin-like growth factor I (IGF-I) and long R(3)IGF-I differently affect development and messenger ribonucleic acid abundance for IGF-binding proteins and type I IGF receptors in in vitro produced bovine embryos.

The insulin-like growth factor (IGF) system is a complex network, including ligands (IGF-I and -II), binding proteins (IGFBP-1 to -6), and receptors, of which the type I IGF receptor (IGF-I-R) is important for transmission of most biological effects of IGFs. As IGFs are secreted in large amounts by the female reproductive tract, it has been hypothesized that maternal IGFs may affect embryonic growth and differentiation in a fine-tuned manner, involving modulation of IGF effects by embryonic IGFBP and IGF-I-R expression. To address this point, we cultured in vitro produced bovine embryos in a chemically defined culture system in the presence (100 ng/ml) of recombinant human IGF-I, long R(3)IGF-I (LR(3)), or without IGF supplementation (control). The affinity of LR(3) to IGFBPs measured by competition assays and Western ligand blots is at least 3 orders of magnitude lower than that of IGF-I. LR(3) was most efficient in stimulating early embryonic cleavage, whereas further development was most potently supported by IGF-I. Total cell numbers of blastocysts were highest in the presence of LR(3) (105 +/- 4), followed by IGF-I (96 +/- 5), and the control group (91 +/- 3; P < 0.05). Differential cell staining of blastocysts revealed that these differences were mainly represented by trophectoderm cell numbers. Analysis of messenger RNA (mRNA) expression for IGFBPs and IGF-I-R was performed by RT-real-time PCR, using expression of the nonregulated housekeeping gene glyceraldehyde-3-phosphate dehydrogenase for normalization. Embryonic IGFBP-2 mRNA levels in the LR(3) treatment group were 1.7-fold (P < 0.001) and 2.8-fold (P < 0.001) higher than those in the IGF-I and control groups, respectively. IGFBP-5 mRNA levels were about 2-fold (P < 0.001) elevated in both IGF treatment groups, with slightly (P < 0.05) higher levels in IGF-I- than in LR(3)-treated embryos. Similarly, IGFBP-3 mRNA abundance was increased (P < 0.05) in embryos from the IGF-I vs. the LR(3) culture system. IGF-I-R mRNA levels were reduced by IGF-I (80% of control; P < 0.01), but increased by LR(3) (1.3-fold vs. control; P < 0.001). These data show that the affinity for IGFBPs of IGF peptides is relevant for their effects on preimplantation embryos and affects different parameters, i.e. development, cell numbers, and mRNA expression for components of the IGF system, in different directions.

Animals↗
PubMed2000

Human CLP36, a PDZ-domain and LIM-domain protein, binds to alpha-actinin-1 and associates with actin filaments and stress fibers in activated platelets and endothelial cells.

A 38-kd protein that associates with F-actin structures in activated platelets and endothelial cells was purified, cloned, and characterized. The protein contains an N-terminal PDZ motif, a large intervening sequence, and a C-terminal LIM domain and was identified as the human homolog of rat CLP36. The study showed that CLP36 associates with actin filaments and stress fibers that are formed during shape change and spreading of platelets and during migration and contraction of endothelial cells. CLP36 binds to alpha-actinin-1 as shown by coimmunoprecipitation, pull-down experiments, yeast 2-hybrid analysis, and blot overlay assays and colocalizes with alpha-actinin-1 along endothelial actin stress fibers. In contrast to alpha-actinin-1, CLP36 was absent from focal adhesions in both activated platelets and endothelial cells. The N-terminal part of CLP36 containing the PDZ domain and the intervening region, but not the LIM domain, targeted enhanced green fluorescent protein fusion proteins to stress fibers in endothelial cells. Yeast 2-hybrid analysis demonstrated that the intervening sequence, but not the PDZ or the LIM domain of CLP36, binds to the spectrinlike repeats 2 and 3 of alpha-actinin-1. The study further shows that CLP36 binds to alpha-actinin in resting platelets and translocates as a CLP36/alpha-actinin complex to the newly formed actin cytoskeleton in activated platelets. The results indicate that CLP36 binds via alpha-actinin-1 to actin filaments and stress fibers in activated human platelets and endothelial cells. The study suggests that CLP36 may direct alpha-actinin-1 to specific actin structures and at this position might modulate the function of alpha-actinin-1. (Blood. 2000;96:4236-4245)

Actin Cytoskeleton↗
PubMed1999

Avoiding inappropriate drug prescribing: fundamental principles for rational medication management.

Drug therapy is not only the most common form of medical therapy, but it can be one of the most effective, if used appropriately. Rapid pharmaceutical development poses a challenge for all prescribing professionals, especially for advanced practice nurses who have only recently acquired prescriptive privileges and who do not have the extensive background experience that guides more seasoned medication managers. For nurse practitioners, nurse midwives, nurse anesthetists, and clinical nurse specialists, excessive reliance on diagnosis-based drug protocols can result in the selection of inappropriate drugs or the prescription of inappropriate doses or durations of therapy. A rational system of drug decision making is necessary to avoid common errors in prescribing. By considering the fundamental principles of pharmacotherapy and by focusing on pathophysiologic objectives, detailed pharmacologic analysis, and a patient-specific plan of therapy, nurse prescribers will avoid errors in prescribing and enhance the safety and effectiveness of drug therapy.

Drug Monitoring↗
PubMed1999

Overexpression of insulin-like growth factor-binding protein-2 in transgenic mice reduces postnatal body weight gain.

Insulin-like growth factor (IGF)-binding protein-2 (IGFBP-2) has been shown to inhibit IGF-dependent cell proliferation in a number of in vitro studies. However, no in vivo model of IGFBP-2 overexpression has been established so far. Therefore, we have generated transgenic mice, in which expression of a mouse IGFBP-2 complementary DNA is controlled by the cytomegalovirus (CMV) promoter. In two independent transgenic strains, transgene expression was highest in pancreas and stomach, followed by skeletal muscle, heart, colon, spleen, adipose tissue, brain, and kidney. Within the pancreas, IGFBP-2 expression was found in the islets but not in the exocrine part. Serum IGFBP-2 levels of CMV-IGFBP-2 transgenic mice were about 3-fold (P < 0.05) increased, compared with controls, whereas serum levels of IGF-I and IGF-II were unaffected by IGFBP-2 overexpression. Fasted serum glucose and fasted insulin levels were slightly reduced in transgenic mice, compared with controls. Postprandial serum glucose insulin levels were not affected by the genotype. At days later than 23, body weights of transgenic mice were significantly (P < 0.05) reduced in both sexes, compared with nontransgenic littermates. This reduction in body weight was mainly attributable to significantly (P < 0.05) lower carcass weights of CMV-IGFBP-2 transgenic vs. control mice. In contrast, absolute organ weights were not (or only as a tendency) reduced, except for the weight of the spleen, which was significantly (P < 0.05) lower in male transgenic than in control mice. Our data suggest that IGFBP-2 represents a negative regulator of postnatal growth in mice, potentially by reducing the bioavailability of IGF-I.

Animals↗
PubMed1998

Clinical recognition of adverse drug reactions: obstacles and opportunities for the nursing profession.

Knowing that adverse drug reactions (ADRs) can occur is only part of the challenge facing nurses today. Clinical recognition of ADRs when they occur is an equally challenging component of pharmacotherapeutics. By understanding the obstacles to their clinical recognition, nurses can design strategies that will engender renewed enthusiasm and vigilance for these complications of drug therapy. These strategies will afford the nursing profession with enhanced opportunities to assume a leadership role in the recognition and intervention of ADRs.

Adverse Drug Reaction Reporting Systems↗
PubMed1997

A comprehensive approach to evaluating nipple discharge.

While emphasis is justifiably placed on the importance of detecting breast masses during breast examination, the equally important need for accurate assessment of nipple discharge is often overlooked. Inspection and palpation for nipple discharge should be part of every breast examination. When detected, a critical analysis should be made of every nipple discharge, with the ultimate objective being differentiation between benign and malignant discharges. All nipple discharges can be defined by the physical characteristics of laterality, spontaneity, color, consistency, number of ducts involved, and duration. By correlating these characteristics with certain historical features (e.g., age, pregnancy, trauma, drugs), accurate determination of etiology can be made. A four-step approach is presented that offers a logical method for making the essential correlations. The sequence of the four relevant questions proposed gives the examiner a logical basis from which to assign clinical importance to each discharge. An algorithm is outlined that correlates diagnostic considerations with therapeutic actions.

Breast↗
PubMed1996

Multiplex reverse transcription polymerase chain reaction combined with temperature gradient gel electrophoresis as a tool for the normalized quantitation of intrinsic factor mRNA.

For the quantitation of intrinsic factor (IF) mRNA, an assay based on competitive reverse transcription and subsequent polymerase chain reaction (RT-PCR) combined with temperature gradient gel electrophoresis (TGGE) was established and validated with respect to precision and accuracy. IF-specific mRNA segments ("targets") were coamplified with known amounts of homologous "standard" RNA molecules, which differed from the targets by one base substitution. Following amplification, TGGE heteroduplex analysis proved to be a powerful method facilitating the efficient separation of these nearly identical target and standard DNA products. The measured absolute copy numbers of IF mRNA were put into relation to the constitutively expressed mRNA specific for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), quantified simultaneously by competitive multiplex RT-PCR. The resulting normalized IF mRNA expression rate in terms of n copies of IF mRNA/copy of GAPDH mRNA is independent of the mRNA heterogeneity and the abundance of specific transcripts within the RNA population of interest. Therefore, normalization relative to the housekeeping gene GAPDH provides a widely applicable value for comparative studies of gene expression on the level of mRNA. Here, a normalized IF mRNA expression rate of three copies per GAPDH mRNA molecule was measured in human stomach mucosa.

Base Sequence↗
PubMed1988

Functional dissection of 5' and 3' extragenic control regions of human tRNA(Val) genes reveals two different regulatory effects.

Two natural human tRNA(Val) genes, pHtV1 and pHtV3, differ in their transcription efficiency by an order of magnitude. The extragenic control regions (ECRs) responsible for this effect were compared with respect to the kinetics and thermodynamics of transcription complex formation. The 5' ECR of pHtV1 acts by increasing both the rate of stable complex formation and the equilibrium constant of association between tDNA and at least one transcription factor present in the stable complex. The stability of the preinitiation complexes is not affected by ECRs. For the formation of a stable preinitiation complex, we suggest a two-step mechanism, comprising (i) the ECR-controlled association of at least one transcription factor (TFIIIC) with the tDNA, and (ii) an ECR-independent conformational change of this tDNA-protein complex. The function of 3' ECRs could be discriminated from the 5' ECR-mediated effects by transcriptional analysis of two chimeric constructs derived from pHtV1 and pHtV3. Surprisingly, the pHtV1 3' ECR causes an eight-fold increase of transcription efficiency, although it has only minor influence on stable preinitiation complex formation. Instead, this ECR stimulates transcription by promoting the transition of the preinitiation complex into an activity synthesizing transcription complex. This novel function of a 3' ECR contributes an additional regulatory level for tRNA gene expression.

Base Sequence↗
PubMed1987

A variant gene and a pseudogene for human 5S RNA are transcriptionally active in vitro.

Screening of a human genomic DNA library with ribosomal 5S RNA yielded a variant 5S rRNA gene (pH5S1) and a pseudogene lacking the first 9 bp and the last 33 bp (pH5S2). Sequence analysis revealed that both genes contain several mutations in their coding region as compared to human 5S rRNA; however, their intragenic promoters are highly conserved. Both genes are transcribed in a homologous HeLa cell S100 extract. pH5S1 gives rise to a 5S-sized product, whereas the two pH5S2-derived RNAs are about 220 and 240 nucleotides long. pH5S1 is transcribed more efficiently than pH5S2; however, its ability to form a stable preinitiation complex is impaired.

Base Sequence↗
PubMed1987

Unrelated leader sequences can efficiently promote human tRNA gene transcription.

The 5'-leader sequence of a human tRNA gene encoding the major tRNA(IACVa 1) species was replaced by several unrelated sequences of human and bacterial origin. Transcription in a HeLa cell extract revealed an extragenic control region (ECR) between positions -51 and -16. Competition assays demonstrate that the wild-type ECR acts as a positive modulator of transcription factor binding. The amount of active transcription complex formed is shown to be dependent on the ECR, whereas the stability of transcription complexes formed under the control of wild-type and mutant ECRs seems not to be affected. One bacterial DNA provided transcription controlling properties indistinguishable from those of the natural human leader sequence. The poor homology between these two sequences indicates that ECRs of human tRNA genes do not consist of highly conserved boxes like intragenic control regions, but of fairly individual DNA elements.

Base Sequence↗
PubMed1986

The human tRNAVal gene family: organization, nucleotide sequences and homologous transcription of three single-copy genes.

At least 13 independent tRNAVal gene loci were detected in the human genome. Three of these genes were isolated and shown to occur only once in the haploid genome. No further functional tRNA genes are located on the isolated clones. Two tRNAVal genes encode the known major and minor tRNAVal isoacceptors, the third may be a pseudogene because a corresponding tRNAVal is not yet known. Comparison of extragenic sequences did not reveal significant homologies, indicating the separation of these genes early in vertebrate evolution. An Alu-type repeat was found in two of the clones within several hundred bp distance from the tDNA. All three genes are transcriptionally active in a HeLa nuclear extract. We show here for the first time that homologous in vitro transcription of mammalian tRNA genes strongly depends on extragenic control regions: interestingly, as a consequence of different flanking regions, the transcription efficiencies vary by an order of magnitude among the genes for the major and the minor tRNAVal and thus reflect the concentrations of these tRNAs in vivo.

Base Sequence↗
PubMed1985

Serotyping of Campylobacter jejuni from an outbreak of enteritis implicating chicken.

An outbreak of campylobacter enteritis involving 7 of 17 people over a period of 5 days followed a dinner at a restaurant. A chicken casserole dish was implicated with a food-specific attack rate of 58%. Campylobacter jejuni Penner serotype 18/21/29, resistant to metronidazole, was isolated from 3 of 4 symptomatic patients and from three raw fresh chicken samples closely associated with the implicated chicken. Numbers of C. jejuni in the chicken ranged from 5.3 X 10(1) to 7.5 X 10(2) colony forming units per square centimeter of surface area. This is the first outbreak of campylobacter enteritis reported in Australia in which C. jejuni has been isolated from both human and food sources and the isolates serologically confirmed as identical.

Animals↗