Biomedical subjects

C Becker

Publications and source records attributed to C Becker.

At least 235 records · Page 13Linked to original sources

PubMed1999

Tumour necrosis factor (TNF) production by T cell receptor-primed T lymphocytes is a target for low dose methotrexate in rheumatoid arthritis.

Methotrexate (MTX) is an effective immunosuppressive agent in various chronic inflammatory diseases such as rheumatoid arthritis (RA). However, its mechanisms of action are only partially understood. In this study, we assessed the effects of MTX on the differentiation of peripheral blood (PB) CD4+CD45RA 'naive' and CD4+CD45RO 'memory' T cells from healthy controls and patients with RA. Accordingly, purified T cells were primed and restimulated in vitro via the T cell receptor (TCR) in the presence of IL-2 to generate effector T cells secreting large amounts of Th1 and Th2 cytokines. We observed that low doses of MTX strongly suppress TNF and to a lesser extent interferon-gamma (IFN-gamma) production by T cells from both healthy donors and RA patients when present during T cell priming via the TCR. Similar data were obtained for TCR-primed synovial fluid mononuclear cells in RA. In contrast, production of IL-4 by TCR-primed CD45RA T cells was significantly increased upon MTX treatment. Interestingly, MTX did not enhance IL-4 production when present during restimulation of effector CD45RO T cells, although it still suppressed TNF production. The results indicate that MTX effects depend on the stage of T cell activation and identify TNF production by TCR-primed T lymphocytes as a target for low-dose MTX treatment in RA. These findings could explain the delayed clinical effects of MTX and may contribute to its potent anti-inflammatory and immunoregulatory properties.

Adult↗

PubMed1999

[Assessment of the effective dose for routine protocols in conventional CT, electron beam CT and coronary angiography].

PURPOSE: To compare the effective dose applied by sequential CT (SEQ), spiral CT (SCT), electron beam CT (EBT) and coronary angiography for investigations of the chest, abdomen and the heart. METHODS: The Alderson Phantom was used to compare the effective dose for all modalities. In addition, the effective dose for conventional CT (SEQ and SCT) was estimated with a mathematical phantom. RESULTS: For CT investigation of the chest and abdomen the dose was highest for the EBT (11 mSv and 25 mSv, respectively) and slightly lower for the SEQ (7.8 mSv and 21.5 mSv, respectively), whereas spiral CT required the least dose (5.3 mSv and 8.8 mSv, respectively). For coronary calcium screening (0.8 mSv) and EBT coronary angiography (1.7 mSv) the dose was lower than for coronary catheter angiography (3.3 mSv). For conventional CT the difference between the effective dose derived by the mathematical phantom and by the Alderson phantom was 2% to 20%. CONCLUSIONS: For investigations of the chest and abdomen the effective dose applied by SCT is significantly lower than that with EBT and SEQ. For investigation of the coronary arteries the effective dose applied by EBT is lower than that for coronary catheter angiography.

Body Burden↗

PubMed1999

2'Halo-ATP and -GTP analogues: rational phasing tools for protein crystallography.

The solution of the crystallographic macromolecular phase problem requires incorporation of heavy atoms into protein crystals. Several 2'-halogenated nucleotides have been reported as potential universal phasing tools for nucleotide binding proteins. However, only limited data are available dealing with the effect of 2'-substitution on recognition by the protein. We have determined equilibrium dissociation constants of 2'-halogenated ATP analogues for the ATP binding proteins UMP/CMP kinase and the molecular chaperone DnaK. Whereas the affinities to UMP/CMP kinase are of the same order of magnitude as for unsubstituted ATP, the affinities to DnaK are drastically decreased to undetectable levels. For 2'-halogenated GTP analogues, the kinetics of interaction were determined for the small GTPases p21ras(Y32W) (fluorescent mutant) and RabS. The rates of association were found to be within about one order of magnitude of those for the nonsubstituted nucleotides, whereas the rates of dissociation were accelerated by factors of approximately 100 (p21ras) or approximately 10(5) (Rab5), and the resulting equilibrium dissociation constants are in the nm or microM range, respectively. The data demonstrate that 2'halo-ATP and -GTP are substrates or ligands for all proteins tested except the chaperone DnaK. Due to the very high affinities of a large number of GTP binding proteins to guanine nucleotides, even a 10(5)-fold decrease in affinity as observed for Rab5 places the equilibrium dissociation constant in the microM range, so that they are still well suited for crystallization of the G-protein:nucleotide complex.

Adenosine Triphosphate↗

PubMed1999

Age-related changes and effect of exercise on the molecular composition of immature equine superficial digital flexor tendons.

To test the hypothesis that exercise at very young age may influence the eventual molecular composition (and hence the biomechanical properties) of tendon tissue in the horse, 43 Dutch Warmblood foals were allotted to 3 differently exercised groups (box-rest, box-rest with training and pasture exercise). Twenty-four superficial digital flexor tendons (SDFTs) were collected at age 5 months (8 from each exercise group) and the others were obtained at 11 months after an additional period of light exercise that was equal for all remaining foals and was intended to see if any induced changes would be reversible or not. Significant changes in DNA content (cellularity), hyaluronic acid (HA) and polysulphated glycosaminoglycans (PSGAGs) were found after the 5 month period of different exercise regimens. There was a tendency towards an exercise-related effect on hydroxylysine content and number of hydroxylysylpyridinoline (HP) crosslinks. Levels of Cartilage Oligomeric Matrix Protein (COMP), measured by homologous inhibition ELISA, showed significant differences at 5 months and were highest in foals kept at pasture and lowest in foals maintained in a box but given enforced exercise. At 11 months, the biochemical parameters of the tendons from the foals of the former box-rest and pasture groups became similar, indicating the capacity of the immature tendon to recover from a retarded development. However, the ratio of PSGAGs per unit of DNA of the former training group was significantly lower than those from the other groups, suggesting that the training regimen in this study had a lasting negative effect on the tenocytes resulting in a decrease of the production of PSGAGs. Therefore, inappropriate or excessive exercise may damage developing tendon, with limited recovery after normalising the exercise level. These possibly deleterious effects of a training regimen on tendon development may be important for the management of young would-be equine athletes.

Analysis of Variance↗

PubMed1999

Summary report of the TD-3 workshop: characterization of 83 antibodies against prostate-specific antigen.

Twelve research groups participated in the ISOBM TD-3 Workshop in which the reactivity and specificity of 83 antibodies against prostate-specific antigen (PSA) were investigated. Using a variety of techniques including cross-inhibition assays, Western blotting, BIAcore, immunoradiometric assays and immunohistochemistry, the antibodies were categorized into six major groups which formed the basis for mapping onto two- and three-dimensional (2-D and 3-D) models of PSA. The overall findings of the TD-3 Workshop are summarized in this report. In agreement with all participating groups, three main antigenic domains were identified: free PSA-specific epitopes located in or close to amino acids 86-91; discontinuous epitopes specific for PSA without human kallikrein (hK2) cross-reactivity located at or close to amino acids 158-163; and continuous or linear epitopes shared between PSA and hK2 located close to amino acids 3-11. In addition, several minor and partly overlapping domains were also identified. Clearly, the characterization of antibodies from this workshop and the location of their epitopes on the 3-D model of PSA illustrate the importance of selecting appropriate antibody pairs for use in immunoassays. It is hoped that these findings and the epitope nomenclature described in this TD-3 Workshop are used as a standard for future evaluation of anti-PSA antibodies.

Antibodies, Monoclonal↗

PubMed1999

Characterization of epitope structure for 53 monoclonal antibodies against prostate-specific antigen.

Prostate-specific antigen (PSA) is the most widely used marker of prostate cancer. Assays for PSA are based on anti-PSA antibodies, and the characterization and selection of these antibodies is important for determining their optimum performance. In our study, we characterized the reactivity of 53 antibodies, submitted to the ISOBM TD-3 PSA Workshop, using free PSA, PSA complexed to alpha1-antichymotrypsin (ACT) and purified ACT. Immunoblotting was performed after native agarose gel or reducing sodium dodecyl polyacrylamide gel electrophoresis. Immunoblotting after agarose gel electrophoresis revealed 10 antibodies that recognized only the free form of PSA, and 43 antibodies that detected both free PSA and PSA-ACT. Immunoblotting of reducing sodium dodecyl-polyacrylamide gels showed the linear or conformation-dependent nature of the epitopes. Two antibodies specific for free PSA and 18 antibodies that recognized both free PSA and PSA-ACT complex recognized linear epitopes. Moreover, 7 antibodies also detected fragmented forms of PSA.

Antibodies, Monoclonal↗

PubMed1999

Reperfusion pattern of the immature femoral head after critical ischemia: a microsphere study in pigs.

The topographic reperfusion pattern of the femoral head after critical ischemia has not yet been investigated. We determined the blood flow of the porcine hip regions with the femoral head epiphysis divided into 24 subregions by the tracer microsphere technique. Blood flow was measured under steady-state conditions, at the end of a 6-hour increase in intracapsular hip joint pressure to 250 mm Hg, and 4 hours after release of the joint tamponade. Total femoral head epiphyseal blood flow decreased with ischemia and regained steady-state perfusion after tamponade. The reperfusion pattern of the femoral head epiphysis appeared identical with that of the steady state before ischemia. However, 2 of the 11 experimental epiphyses remained ischemic in the reperfusion phase. We conclude that hip joint tamponade above the arterial pressure level for 6 hours causes global ischemia in the femoral head epiphysis in the immature pig, without regional differences in reperfusion, and that reperfusion occurs at a level like that of the steady state before ischemia.

Age Factors↗

PubMed1999

Constitutive and inducible in vivo protein-DNA interactions at the tumor necrosis factor-alpha promoter in primary human T lymphocytes.

Tumor necrosis factor-alpha (TNF-alpha) is a key cytokine of lymphocytes with major regulatory functions in immunomodulation, chronic inflammation, and septic shock. However, only limited information on TNF promoter regulation in vivo in primary lymphocytes is available. To determine and compare protein-DNA interactions at the native TNF locus in primary lymphocytes, we analyzed the human TNF-alpha promoter by ligation-mediated polymerase chain reaction (LM-PCR) techniques. Accordingly, primary CD4+ T lymphocytes from peripheral blood were cultured in the presence of various stimuli and analyzed by LM-PCR. Inducible in vivo protein-DNA interactions at the TNF promoter were detected between -120 and -70 bp of the human TNF promoter relative to the transcriptional start site. This area includes binding sites for transcription factors such as ETS-1, NFAT, ATF-2/c-jun, SP-1/Egr-1, and NF-kappaB. In contrast, no protein-DNA interactions were observed at various binding sites with reported regulatory function in tumor cell lines such as the k2 element, the NFAT site at -160, the AP1 site at -50, and the SP1 site at -65. Additional mutagenesis and transfection studies demonstrated that NF-kappaB and CREB/AP-1 are important regulators of inducible TNF promoter activity in primary human T lymphocytes. These results provide novel insights into the complex regulation of TNF gene transcription in primary T lymphocytes in vivo by constitutive and inducible protein-DNA interactions that appear to be at least partially different compared to previously characterized tumor cell lines.

Base Sequence↗

PubMed1998

Does an asparaginyl-specific cysteine endopeptidase trigger phaseolin degradation in cotyledons of kidney bean seedlings?

An asparaginyl-specific cysteine endopeptidase which was named 'legumain-like proteinase' (LLP) and has an apparent molecular mass of 38.1 kDa was isolated from cotyledons of kidney bean (Phaseolus vulgaris L.) seedlings and partially characterized. It is, to our knowledge, the first known proteinase which in vitro extensively degrades native phaseolin, the major storage globulin of this grain legume. Phaseolin that in vitro had been partially degraded by LLP (Pvitro) and phaseolin that was isolated after partial in vivo breakdown 6 days after the start of seed imbibition (Pvivo) showed similar fragment patterns on SDS/polyacrylamide gels. The fragments had identical cleavage sites in Pvitro and Pvivo as determined by partial amino acid sequencing. In both types of partially degraded phaseolin, these cleavage sites have asparagine in the P1 position. Two of the cleavage sites are located in the beta-barrel domain of the C-terminal module and only one cleavage site was found in the beta-barrel domain of the N-terminal module according to the consensus structural model of phaseolin subunits. These results suggest that very likely LLP could in vivo be responsible for the initiation of phaseolin proteolysis. Two different legumain-specific clones named cp6b and p21b were isolated from a cDNA library of germinated bean cotyledons. Cp6b encodes LLP, while p21b encodes a VPE-like enzyme. Southern-blot analysis revealed a single gene copy for Pv-VPE and, presumably, at least two gene copies for LLP in the kidney bean genome. Northern-blot analysis indicated that mRNAs for both clones appear de novo during seed germination. However, the developmental patterns of the transcript levels corresponding to the two clones differed significantly. The temporal pattern of phaseolin degradation and of LLP polypeptide levels agreed well with the suggestion that LLP plays a key role in the mobilization of phaseolin during and after kidney bean germination.

Amino Acid Sequence↗

PubMed1998

[Accidental falls and fall-related injuries in the elderly. Diagnosis and intervention].

Falls in the elderly should be seen as treatable symptoms of impending or already present loss of mobility and therefore often loss of independence. The most important European and American epidemiological data are presented, and the diagnostic strategies and appropriate interventions discussed. The latter represent the most effective measures for minimizing functional impairment in the elderly that are currently available. The aim of the present paper is to demonstrate the feasibility of such measures and their value in the prevention of loss of mobility in the elderly.

Accidental Falls↗

PubMed1998