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Biomedical subjects

C Weber

Publications and source records attributed to C Weber.

At least 109 records · Page 6Linked to original sources
PubMed2000

Can sterile and pyrogen-free on-line substitution fluid be routinely delivered? A multicentric study on the microbiological safety of on-line haemodiafiltration.

BACKGROUND: Microbial contamination is characterized not only by the presence of bacteria, but also by high concentrations of biologically active by-products. They are potentially able to cross ultrafiltration and dialysis membranes and stimulate immunocompetent blood cells to synthesize cytokines. In turn, cytokine induction causes acute symptoms and has been incriminated in the long-term complications of haemodialysis patients. Infusion of large volumes of substitution fluids following ultrafiltration of microbially contaminated dialysis fluids may place patients on on-line therapies at particular risk. METHODS: In this study we evaluated 30 machines with a two-stage ultrafiltration system in routine clinical haemodiafiltration settings in six centres for 6 months. Microbiological safety was assessed monthly and at the last use of the filters by determining microbial counts, endotoxin concentration and cytokine-inducing activity. RESULTS: No pyrogenic episodes were observed during the study period. Double-filtration of standard dialysis fluid (range, <1-895 cfu/ml, 0.0028-4.6822 IU/ml) resulted in sterile substitution fluids with endotoxin concentrations well below the Ph.Eur. standard for haemofiltration solutions (range, 0.0014-0.0281 vs 0.25 IU/ml). Moreover, they did not differ from commercial haemofiltration solutions and depyrogenated saline. Likewise, there was no difference in the cytokine-inducing activity between the solutions tested. The high microbiological quality of the ultrafiltered dialysis fluid, which was in the same range as substitution fluid, translates into both the absence of cytokine induction by dialyser back-transport and a redundant safety mode of the on-line system by a second filtration step. CONCLUSION: On-line HDF treatment can routinely be provided with ultra-pure dialysis fluids and sterile substitution fluids at pyrogen-free levels. The online preparation of substitution fluids thus can be considered microbiologically safe.

Bacteria↗
PubMed2000

Progression of cardiac allograft vascular disease as assessed by serial intravascular ultrasound: correlation to immunological and non-immunological risk factors.

OBJECTIVE: To characterise the severity and progression of cardiac allograft vascular disease (CAVD) in a large patient cohort, and to evaluate possible immunological and non-immunological risk factors for progression. DESIGN: A prospective observational study using intravascular ultrasound. SETTING: Two university hospitals. PATIENTS AND MAIN OUTCOME MEASURES: Changes in focal plaque, lumen, and total vessel area (worst site method) were assessed at baseline and after 12.1 (2.8) months (mean (SD)) of follow up in a cohort of 96 patients (79 male, 17 female; mean age 48.7 (9.6) years; time post-transplant 26.0 (32.4) months). RESULTS: Overall, the mean (SD) intimal index of worst sites increased by 6.7 (8.8)%. The increase in the first 12 months was 7.5 (9.4)%, v 5.9 (8.0)% after the first year (NS). Analysing immunological and non-immunological risk factors (age, underlying disease, sex, donor age, immunosuppression, cytomegalovirus, rejection episodes, cholesterol), low density lipoprotein (LDL) cholesterol was found to be the most important predictor of severe progression (as defined by an increase in intimal index of >/= 15% (p = 0.01). CONCLUSIONS: Progression of CAVD is characterised by a continuing increase in intimal hyperplasia, especially within the first year after heart transplantation. LDL cholesterol is an important predictor of major progression.

Adult↗
PubMed2000

Pyrrolidine dithiocarbamate-induced apoptosis depends on cell type, density, and the presence of Cu(2+) and Zn(2+).

Pyrrolidine dithiocarbamate (PDTC) has been found to induce or inhibit apoptosis in different cell types. Here we show that PDTC dose-dependently reduced the viability of rat smooth muscle cells (rSMC), human fibroblasts, and endothelial cells at low but not at high cell density. Endothelial cells were least sensitive, fibroblasts showed a medium sensitivity, and rSMC showed a high sensitivity to PDTC-mediated cell death. An early reduction in the mitochondrial membrane potential indicated a rapid onset of apoptosis in rSMC. Apoptosis was further confirmed by annexin V staining and DNA fragmentation analysis. Gel shift analysis demonstrated increased nuclear factor (NF)-kappaB activity in high-density rSMC compared with low-density cells. NF-kappaB has recently been shown to regulate the induction of anti-apoptotic proteins. Although PDTC is widely used as an inhibitor for NF-kappaB and a radical scavenger, our data show that PDTC rather enhanced NF-kappaB activity and, alone or in combination with menadione, induced oxygen radical generation. Notably, PDTC failed to reduce rSMC viability in medium without Cu(2+) or Zn(2+), and addition of Cu(2+) or Zn(2+) resulted in a dose-dependent increase in PDTC-induced cell death. Addition of both Cu(2+) and Zn(2+) showed synergistic effects. Our results indicate that the induction of apoptosis by PDTC requires Cu(2+) and Zn(2+) and is dependent on cell type and density. Such differential effects may have implications for studies of PDTC as an anti-atherosclerotic or immunomodulatory drug.

Animals↗
PubMed2000

Respiratory symptoms of obstructive lung disease in European crop farmers.

Crop farming as a risk factor for respiratory symptoms of obstructive lung disease was assessed. Random samples of crop farmers from four European countries were studied following a cross-sectional design. A questionnaire on respiratory symptoms and occupation was administered to determine prevalences, and the roles of the various crops as risk factors for respiratory symptoms were assessed through logistic regression modeling. The 4,793 crop farmers included in the study (response rate: 85.3%) reported the following respiratory symptoms: wheezing (14.9%), asthma (3.3%), nasal allergy (14.4%), chronic phlegm (12.4%), organic dust toxic syndrome (ODTS) (15.2%), and symptoms at work (22.0%). In the multivariate analysis, adjusting for age, sex, smoking, country, and exposure to other plants or livestock, flower growing was a risk factor for asthma (odds ratio [OR] 2.1, 95% confidence interval [CI] 1.1-3.9) and cultivating oil plants was associated with ODTS (OR 1.5, 95% CI 1.3-1.9), symptoms at work (OR 1.4, 95% CI 1.2- 1.7), and chronic phlegm (OR 1.3, 95% CI 1.1-1.6). Working inside greenhouses was a marginal risk factor for asthma (OR 2.1, 95% CI 0.9-4.5). We conclude that flower and oil plant production is associated with increased risk of respiratory symptoms in European crop farmers.

Adult↗
PubMed1999

Simple and efficient method for the detection of diethylenetriaminepentaacetic acid.

Diethylenetriaminepentaacetic acid (DTPA) is a commonly used chelating agent. Its antiviral, antibacterial and immunomodulatory effects are well documented. DTPA forms a highly stable complex with lead (II) with an increased absorption coefficient and a bathochromic shift of the absorption maximum compared to pure DTPA. Based on this complex a high-performance liquid chromatographic method for the quantitative detection of DTPA in biological fluids was developed. A calibration curve was prepared and linearity was shown in the concentration range between 10 mg l(-1) and 1000 mg l(-1) DTPA. The recovery in water and in human plasma showed the method to be suitable for routine use.

Adsorption↗
PubMed1999

A rapid method for the evaluation of compounds with mitochondria-protective properties.

Mitochondrial dysfunction has been implicated in a number of neurodegenerative diseases, such as ischemia and Parkinson's disease. We present here a method that allows the rapid quantification of interventions, aimed at inhibiting the effect of mitochondrial membrane potential uncouplers, based on the ratioing properties of the fluorescent probe 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1), by using currently available 96-well fluorescent plate readers. A method is presented for evaluation of cross-talk between the two excitation/emission channels. Further characterization of the probe shows that the effect of plasma membrane potential changes on JC-1 fluorescence ratio are negligible, but that the signal is very sensitive to pH. One of the most exciting applications is the possibility to perform end-point measurements, thanks to the ratioing properties of the probe. The system is tested in different culture types with different mitochondrial uncouplers. As an example of a quantitative evaluation, we show that flunarizine is able to inhibit, dose-dependently, FCCP mediated JC-1 signal increase. The procedure is simple and allows for the fast screening of mitochondria-protective compounds.

Animals↗
PubMed1999

Effect of potent and selective inhibitors of the Grb2 SH2 domain on cell motility.

Cell motility has been correlated both with oncogenic invasiveness and metastatic potential. The development of selective inhibitors of motility has thus great potential importance. Grb2 is a SH2/SH3 domain-containing adaptor protein that links growth factor receptor tyrosine kinases to the Ras signaling pathway. We have developed specific small molecule inhibitors of the Grb2 SH2 domain as potential leads for drug discovery. Synthesis of the inhibitors and their effects on growth factor-induced growth in cells have been reported previously. In the current study, we establish that these inhibitors inhibit hepatocyte growth factor/scatter factor-induced A431 and Madin-Darby canine kidney cell motility and various cell motility-related events, including epidermal growth factor-induced ruffling of A431 cells and epidermal growth factor-induced translocation of the small GTPase Rac in these cells. We demonstrate for the first time a direct role for Grb2 in cell motility and indicate a new avenue for cancer therapeutics.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Monocyte arrest and transmigration on inflamed endothelium in shear flow is inhibited by adenovirus-mediated gene transfer of IkappaB-alpha.

Mobilization of nuclear factor-kappaB (NF-kappaB) activates transcription of genes encoding endothelial adhesion molecules and chemokines that contribute to monocyte infiltration critical in atherogenesis. Inhibition of NF-kappaB has been achieved by pharmacological and genetic approaches; however, monocyte interactions with activated endothelium in shear flow following gene transfer of the NF-kappaB inhibitor IkappaB-alpha have not been studied. We found that overexpression of IkappaB-alpha in endothelial cells using a recombinant adenovirus prevented tumor necrosis factor-alpha (TNF-alpha)-induced degradation of IkappaB-alpha and suppressed the upregulation of vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin mRNA and surface protein expression and the upregulation of transcripts for the chemokines monocyte chemoattractant protein 1 (MCP-1) and growth-related activity-alpha (GRO-alpha) by TNF-alpha. This was associated with a reduction in endothelial MCP-1 secretion and GRO-alpha immobilization. Adhesion assays under physiological shear flow conditions showed that firm arrest, spreading, and transmigration of monocytes on TNF-alpha-activated endothelium was markedly inhibited by IkappaB-alpha overexpression. Inhibition with monoclonal antibodies and peptide antagonists inferred that this was due to reduced expression of Ig integrin ligand as well as of chemokines specifically involved in these events. In contrast, rolling of monocytes was increased by IkappaB-alpha transfer and was partly mediated by P-selectin; however, it appeared to be unaffected by the inhibition of E-selectin induction. Thus, our data provide novel evidence that selective modulation of NF-kappaB by adenoviral transfer of IkappaB-alpha impairs the expression of multiple endothelial gene products required for subsequent monocyte arrest and emigration in shear flow and thus for monocyte infiltration in atherosclerotic plaques.

Adenoviridae↗
PubMed1999

Nuclear factor-kappa B regulates induction of apoptosis and inhibitor of apoptosis protein-1 expression in vascular smooth muscle cells.

Apoptosis is important in normal development as well as in diseases such as atherosclerosis. However, the regulation of apoptosis is still not completely understood. We now show that the transcription factor nuclear factor-kappaB (NF-kappaB) controls the induction of apoptosis in human and rat vascular smooth muscle cells (SMCs). SMCs in high-density culture exhibited a high NF-kappaB activity and were insensitive to induction of apoptosis. Inhibition of NF-kappaB by adenovirus-mediated overexpression of its inhibitor IkappaBalpha caused a marked increase in cell death at low but not high cell density. Elevating endogenous IkappaBalpha levels by inhibiting its degradation with proteasomal inhibitors resulted in induction of apoptosis in low-density SMCs, as detected by increased binding of annexin V, reduced mitochondrial membrane potential, and increased hypodiploid DNA. In high-density cultures, protection against apoptosis was associated with the expression of inhibitor of apoptosis protein-1 (IAP-1). Transfer of IkappaBalpha reduced human IAP-1 mRNA levels, which suggested that IAP-1 is transcriptionally regulated by NF-kappaB. This was confirmed through identification of a motif with NF-kappaB-like binding activity in the human IAP-1 promoter region. Moreover, antisense inhibition of IAP-1 sensitized high-density SMCs to the induction of cell death. Together, our data imply that SMCs at high density are protected by an antiapoptotic mechanism that involves increased expression of NF-kappaB and IAP-1. Interference with pathways that control the susceptibility to programmed cell death may be helpful in the treatment of diseases where dysregulation of apoptosis is involved, eg, atherosclerosis and restenosis.

Animals↗
PubMed1999

Molecular and functional analysis of a conserved CTL epitope in HIV-1 p24 recognized from a long-term nonprogressor: constraints on immune escape associated with targeting a sequence essential for viral replication.

It has been hypothesized that sequence variation within CTL epitopes leading to immune escape plays a role in the progression of HIV-1 infection. Only very limited data exist that address the influence of biologic characteristics of CTL epitopes on the emergence of immune escape variants and the efficiency of suppression HIV-1 by CTL. In this report, we studied the effects of HIV-1 CTL epitope sequence variation on HIV-1 replication. The highly conserved HLA-B14-restricted CTL epitope DRFYKTLRAE in HIV-1 p24 was examined, which had been defined as the immunodominant CTL epitope in a long-term nonprogressing individual. We generated a set of viral mutants on an HX10 background differing by a single conservative or nonconservative amino acid substitution at each of the P1 to P9 amino acid residues of the epitope. All of the nonconservative amino acid substitutions abolished viral infectivity and only 5 of 10 conservative changes yielded replication-competent virus. Recognition of these epitope sequence variants by CTL was tested using synthetic peptides. All mutations that abrogated CTL recognition strongly impaired viral replication, and all replication-competent viral variants were recognized by CTL, although some variants with a lower efficiency. Our data indicate that this CTL epitope is located within a viral sequence essential for viral replication. Targeting CTL epitopes within functionally important regions of the HIV-1 genome could limit the chance of immune evasion.

Animals↗
PubMed1999

alpha-tocopheryl succinate-induced apoptosis in Jurkat T cells involves caspase-3 activation, and both lysosomal and mitochondrial destabilisation.

Alpha-Tocopheryl succinate (alpha-TOS), but not a-tocopherol, triggered apoptosis in Jurkat T cells. Apoptosis was induced by alpha-TOS in a time- and concentration-dependent mode, and signs of apoptosis were visible at concentrations of alpha-TOS as low as 30 microM, and within 3-5 h after addition of the ester. Employing a specific fluorogenic substrate, caspase-3 was found to be activated rapidly in response to alpha-TOS at 50 microM. We also found that Jurkat T cells challenged with alpha-TOS, when exposed to the lysosomotropic weak base acridine orange, showed decreased lysosomal uptake of the dye. This is suggestive of the involvement of lysosomal destabilisation in apoptosis of the cells. Apoptosis of Jurkat T cells induced with alpha-TOS also involved a drop in the mitochondrial membrane potential, although this phenomenon occurred after the initiation of lysosomal rupture. All apoptotic features observed with alpha-TOS were very similar to those found when cross-linking of the Fas receptor triggered apoptosis. These findings are consistent with the recent idea that vitamin E can contribute to elimination of malignant cells by the induction of apoptosis, and can be of (patho)physiological significance.

Apoptosis↗
PubMed1999

Differential immobilization and hierarchical involvement of chemokines in monocyte arrest and transmigration on inflamed endothelium in shear flow.

Monocyte extravasation into areas of inflammation involves sequential interactions of multiple adhesion molecules. However, differential contribution of chemokines produced by cytokine-stimulated endothelium and their receptors to leukocyte attachment and transmigration under flow conditions remains to be elucidated. The activation of endothelial cells with TNF-alpha up-regulated mRNA and protein expression of the CXC chemokine GRO-alpha and the CC chemokine monocyte chemotactic protein (MCP)-1, which act through the receptors CXCR2 and CCR2 expressed on monocytes, respectively. Whereas GRO-alpha was immobilized to endothelial cells via heparan sulfate proteoglycans, MCP-1 was secreted in a soluble form. Consequently, inhibition experiments with blocking peptide analogues and monoclonal antibodies revealed that GRO-alpha and its receptor CXCR2 but not MCP-1 and its receptors substantially contributed to conversion of rolling into firm, shear-resistant arrest of monocytes to TNF-alpha-stimulated endothelium in physiological flow. In contrast, MCP-1 and CCR2 but not GRO-alpha and CXCR2 mediated spreading, shape change and subsequent transendothelial migration, which was evident in flow but rarely in stasis and may thus require the establishment of a diffusible MCP-1 gradient. Differential patterns of presentation may determine a functional specialization and hierarchy of chemokines and their receptors in sequential steps of monocyte emigration on inflamed endothelium and shear flow.

Antigen Presentation↗
PubMed1999

Downregulation by tumor necrosis factor-alpha of monocyte CCR2 expression and monocyte chemotactic protein-1-induced transendothelial migration is antagonized by oxidized low-density lipoprotein: a potential mechanism of monocyte retention in atherosclerotic lesions.

The subintimal infiltration with monocytes is crucially involved in the development of complex atherosclerotic plaques. Monocyte chemotactic protein-1 (MCP-1) and its receptor CCR2 are important for monocyte extravasation and formation of atherosclerotic lesions. However, mechanisms of monocyte persistence in atherosclerotic plaques remain to be elucidated. Flow cytometric analysis revealed that monocytoid Mono Mac 6 cells that had transmigrated endothelium towards a MCP-1 gradient expressed higher levels of CCR2 than the non-migratory fraction, while input cells were intermediate, suggesting that high CCR2 levels are essential for transendothelial chemotaxis. Pretreatment of Mono Mac 6 cells or isolated human blood monocytes with the inflammatory cytokine tumor necrosis factor (TNF)-alpha dose- and time-dependently reduced MCP-1-induced transendothelial chemotaxis, which was inhibited by the CCR2 receptor antagonist 9-76 analog. This was paralleled by a decrease in CCR2 surface protein and mRNA expression. as assessed by flow cytometry and reverse transcription-polymerase chain reaction, inferring that inhibition of monocyte transmigration was due to downregulation of CCR2 to levels insufficient for chemotaxis. In contrast, treatment of monocytes with oxidized low-density protein (oxLDL) containing oxidized lipids, such as cholesteryl linoleate 13-hydroxide. but not with LDL, increased CCR2 protein and mRNA expression. Notably, oxLDL counteracted the TNF-alpha-mediated downregulation of CCR2 and CCR2-dependent transendothelial chemotaxis. Macrophage-colony-stimulating factor hardly affected CCR2 expression and function, suggesting that differentiation was not responsible for effects on CCR2. In conclusion, TNF-alpha impairs MCP-1-induced transendothelial migration of monocytes by downregulating CCR2 which appears critical for migration. Exposure to oxLDL antagonized the effects of TNF-alpha, and may thus contribute to monocyte retention and perpetuation of a chronic inflammatory reaction in unstable atherosclerotic lesions.

Arteriosclerosis↗
PubMed1999

Coenzyme Q10 does not prevent oral dyskinesias induced by long-term haloperidol treatment of rats.

Tardive dyskinesia (TD) is a debilitating side effect of long-term treatment with neuroleptics with an unclear pathophysiologic basis. It has been proposed that TD may be a result of neuroleptic-induced oxidative stress. To investigate this hypothesis, we studied if neuroleptic-induced oral dyskinesias in rats, a putative analogue to human TD, could be prevented by the antioxidant coenzyme Q10 (CoQ10). Rats received 16 weeks of treatment with haloperidol decanoate (HAL) IM alone or together with orally administered CoQ10, and the behavior was recorded during and after treatment. HAL significantly increased the level of oral dyskinesias, and the increase persisted for 12 weeks after drug withdrawal. Cotreatment with CoQ10 did not attenuate the development of HAL-induced oral dyskinesia. Despite adequate absorption of orally administered CoQ10, shown by the increased serum levels of CoQ10, no increase of either CoQ10 or coenzyme Q9 was detected in the brain. These results suggest that cotreatment with CoQ10 does not inhibit the development of HAL-induced oral dyskinesias in rats, and that further studies seem to be needed in order to clarify the pharmacokinetics of CoQ10 in rats.

Animals↗
PubMed1999

Psychophysical measures of sensitization to tonic heat discriminate chronic pain patients.

Sensitization to continued nociceptive stimulation is supposed to be involved in the development of chronic pain at several levels of the CNS, but experimental studies investigating the perceptual dynamics of sensitization in humans are rare, and the diagnostic validity of experimental pain models is not known. The present study used a tonic heat paradigm to assess early sensitization (15-100 s) to experimental pain in 30 chronic pain patients (15 musculoskeletal/back pain, 15 headache) and 23 healthy controls. Change in pain sensation during prolonged stimulation was measured by a dual sensitization method which combines subjective ratings and behavioural responses in an indirect psychophysical protocol protected against response bias. Phasic and tonic pain thresholds were measured for control purposes. The degree of sensitization was linearly related to stimulus temperature, and groups differed significantly in this 'sensitization gradient': chronic pain patients sensitized earlier and stronger than healthy subjects, musculoskeletal pain patients showed the strongest effect. Pain thresholds were lowered in headache patients only. Discriminant analysis demonstrated good sensitivity and specificity of individual sensitization measures for distinguishing pain syndromes, particularly in combination with pain thresholds. The results are in accordance with current models of spinal plasticity contributing to pathological pain states. They argue for the diagnostic value of psychophysical measures of sensitization.

Adolescent↗
PubMed1999

Opioid-induced emesis among hospitalized nonsurgical patients: effect on pain and quality of life.

There is very little information in the medical literature regarding opioid-induced emesis and its relationship to patient outcomes. Two-hundred and six nonsurgical patients in a 400-bed teaching hospital with minimal known risks of disease-associated emesis were interviewed to examine emesis and associated outcomes following the administration of opioids for acute pain management. The mean age, weight, and height of the study group were 54.4 (+/- 19.6) years, 175.8 (+/- 45.7) pounds, and 67.1 (+/- 4.4) inches, respectively. Seventy-three (35.4%) patients experienced nausea; 28 (13.6%) patients vomited; and 15 (7.3%) patients retched following the opioid therapy. These symptoms were mild and discomforting for relatively short periods of time. The patients' ability to concentrate and eat was affected by the incidence of nausea/vomiting. The intensity, duration, and severity of nausea were positively associated with the magnitude of the functional limitations. The symptoms also influenced patients' ratings of various hospital satisfaction measures. In conclusion, emesis due to opioids represents a notable burden on nonsurgical patients. Successful therapies that prevent opioid-induced emesis are likely to positively influence patient outcomes by reducing adverse effects, improving functional outcomes, and enhancing quality of life.

Adolescent↗