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M C Vissers

Publications and source records attributed to M C Vissers.

At least 19 recordsLinked to original sources
PubMed2001

Regulation of apoptosis by vitamin C. Specific protection of the apoptotic machinery against exposure to chlorinated oxidants.

We have investigated the ability of intracellular vitamin C to protect human umbilical vein endothelial cells from exposure to hypochlorous acid (HOCl) and a range of derived chloramines. Ascorbate provided minimal protection against the cytotoxicity induced by these oxidants, as measured by propidium iodide uptake. In contrast, there was a marked effect on apoptosis, monitored by caspase-3 activation and phosphatidylserine exposure. Extended incubation of the cells with glycine chloramine or histamine chloramine completely blocked apoptosis initiated in the cells by serum withdrawal. This effect was significantly abrogated by ascorbate. Inhibition of apoptosis required the oxidant to be present for an extended period after serum withdrawal and occurred prior to caspase-3 activation. General protection of thiols by ascorbate was not responsible for the protection of apoptosis, because intracellular oxidation by HOCl or chloramines was not prevented in supplemented cells. The results suggest a new role for vitamin C in the regulation of apoptosis. We propose that, by protection of an oxidant-sensitive step in the initiation phase, ascorbate allows apoptosis to proceed in endothelial cells under sustained oxidative stress.

Antioxidants↗
PubMed2001

Hypochlorous acid stimulation of the mitogen-activated protein kinase pathway enhances cell survival.

We investigated the activation of three subfamilies of mitogen-activated protein kinases (MAP kinase), the extracellular regulated kinase (ERK1/2), p38, and c-Jun N-terminal kinase (JNK), by the myeloperoxidase-derived oxidant HOCl, in human umbilical vein endothelial cells (HUVEC) and human skin fibroblasts. Treatment of fibroblasts with 10-30 microM HOCl induced a dose-dependent increase in the tyrosine phosphorylation of several proteins. ERK1/2 was activated by exposure to sublethal concentrations of reagent HOCl or by HOCl generated by myeloperoxidase as shown by immune complex kinase assays. Maximum activation was seen at 20 microM and peak activation occurred within 10 min. Western blot analysis demonstrated activation of p38 with 30 microM HOCl, occurring at 15-30 min. No activation of JNK was detected in the concentration range investigated. These results show that HOCl is able to activate MAP kinases. Effective doses were considerably lower than with H2O2 and the lack of JNK activation contrasts with the activation frequently seen with H2O2. Exposure to HOCl caused a loss of viability in HUVEC that was markedly enhanced when ERK1/2 activation was inhibited by U0126. This suggests that the activation of ERK promotes cell survival in response to the oxidative challenge.

Cell Death↗
PubMed2001

Glutathione oxidation by hypochlorous acid in endothelial cells produces glutathione sulfonamide as a major product but not glutathione disulfide.

Treatment of cells with hypochlorous acid (HOCl) at sublethal doses causes a concentration-dependent loss in reduced glutathione (GSH) levels. We have investigated the products of the reaction of HOCl with GSH in human umbilical vein endothelial cells. Despite a complete loss of GSH, there were only very small increases in intracellular and extracellular glutathione disulfide and glutathione sulfonic acid after exposure to HOCl. (35)S labeling of the GSH pool showed only a minimal increase in protein-bound GSH, suggesting that S-thiolation was not a major contributor to HOCl-mediated loss of GSH in endothelial cells. Rather, the products of the reaction were mostly exported from cells and included a peak that co-eluted with the cyclic sulfonamide that is a product of the reaction of GSH with reagent HOCl. Evidence of this species in endothelial cell supernatants after HOCl treatment was also obtained using electrospray mass spectrometry. In conclusion, exposure to HOCl causes the irreversible loss of cellular GSH with the formation of novel products that are rapidly exported from the cell, and resynthesis of GSH will be required to restore levels. The loss of GSH would alter the redox state of the cell and compromise its defenses against further oxidative stress.

Cells, Cultured↗
PubMed2001

Fatty acid chlorohydrins and bromohydrins are cytotoxic to human endothelial cells.

Reaction of unsaturated lipids with the hypohalous acids (hypochlorous acid and hypobromous acid) results in the addition of the halide (X) across double bonds to form halohydrins (-CH2CH(OH)CH(X)CH2-). These modified lipids could be potentially destabilising to cell membranes due to their increased polarity. We have investigated the effect of pre-formed halohydrins on human umbilical vein endothelial cells (HUVEC) by incubating cultured cells with oleic acid micelles containing chlorohydrins or bromohydrins. Cell detachment and necrotic death were observed with increasing doses of halohydrins, whereas the cells were unaffected by equivalent doses of oleic acid. Bromohydrins caused more lysis than did chlorohydrins at equivalent doses. Complete lysis was seen with 200 microM fatty acid/chlorohydrin micelles and with 50 microM fatty acid/bromohydrin micelles. Chlorohydrin uptake was much less than the oleic acid control whereas bromohydrins were incorporated into the endothelial cells similarly to oleic acid. This difference or the bulkier nature of the bromohydrins could account for their increased toxicity. This study has demonstrated the potential toxicity of the halohydrins, and implications for their formation in inflammation are discussed.

Alcohols↗
PubMed2000

Myeloperoxidase.

This review covers recent advances in the biology of myeloperoxidase. Mechanisms of posttranslational processing and how these fail in some of the common deficiency mutants are discussed. We also review the enzymology that points to myeloperoxidase having a number of physiologic substrates in addition to chloride and the evidence that it produces hypochlorous acid in the neutrophil phagosome in sufficient quantities to be bactericidal. Evidence is accumulating that myeloperoxidase-derived oxidants modify biologic macromolecules and cell-regulatory pathways and that they play a role in atherosclerosis. Investigation of disease incidence in relation to a polymorphism in the promoter region of the gene has produced interesting associations. These links with inflammatory diseases can now be pursued further using specific biomarkers of myeloperoxidase activity.

Humans↗
PubMed1999

Hypochlorous acid causes caspase activation and apoptosis or growth arrest in human endothelial cells.

We have investigated the effect of hypochlorous acid (HOCl) on cultured human umbilical-vein endothelial cells and shown that, whereas higher concentrations cause rapid necrosis, smaller amounts of this oxidant induce apoptosis or growth arrest. Exposure to 20-40 nmol of HOCl per 1.2x10(5) cells initiated apoptosis that was determined morphologically, by the identification of apoptotic nuclei with Hoechst 33342, and by detection of phosphatidylserine on the outer membrane. Degraded DNA was detected by flow cytometry. HOCl induced caspase activity; specific inhibition of caspases was shown to prevent apoptosis. No caspase activation could be detected with 50 nmol of HOCl per 1.2x10(5) cells, a dose that caused more extensive necrosis. Lower doses of HOCl, which did not cause cell death, resulted in a transient growth arrest that was apparent with as little as 5 nmol of HOCl per 1.2x10(5) cells. These results show that HOCl can modify cellular responses that are dependent on signal transduction pathways in a manner similar to that of other oxidants.

Apoptosis↗
PubMed1999

Loss of GSH and thiol enzymes in endothelial cells exposed to sublethal concentrations of hypochlorous acid.

We investigated the effect of sublethal concentrations of hypochlorous acid (HOCl) on intracellular thiol groups. Exposure of human umbilical vein endothelial cells to HOCl caused a decrease in cell viability, with concentrations of </=25 microM HOCl being sublethal. At these concentrations, we saw a loss of glutathione and total protein thiol groups. Of the thiol enzymes we investigated, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was particularly susceptible to inactivation, creatine kinase was moderately susceptible, and lactate dehydrogenase was unaffected by HOCl at the concentrations used. Similar results were obtained with HOCl generated over 30 min by myeloperoxidase. GAPDH activity could be regenerated on reincubation of cells in Hanks' balanced salt solution or reduction with dithiothreitol. In contrast, glutathione loss was not reversible, and further decreased with time. Cellular ATP levels decreased with sublethal HOCl concentrations and this appeared to be unrelated to the inactivation of GAPDH. Our results demonstrate that intracellular thiol groups differ in their reactivity with HOCl and suggest that HOCl may be able to regulate specific cellular functions.

Adenosine Triphosphate↗
PubMed1998

Comparison of human red cell lysis by hypochlorous and hypobromous acids: insights into the mechanism of lysis.

Human red blood cells are lysed by the neutrophil-derived oxidant hypochlorous acid (HOCl), although the mechanism of lysis is unknown. Hypobromous acid (HOBr), a similarly reactive oxidant, lysed red cells approx. 10-fold faster than HOCl. Therefore we compared the effects of these oxidants on thiols, membrane lipids and proteins to determine which reactions are associated with lysis. There was no difference in the loss of reduced glutathione or membrane thiols with either oxidant, but HOBr reacted more readily with membrane lipids and proteins. Bromohydrin derivatives of phospholipids and cholesterol were seen at approx. one-tenth the level of oxidant than chlorohydrins were. However, these products were detected only with high concentrations of HOCl or HOBr, which caused instant haemolysis. Membrane protein modification occurred at much lower doses of oxidant and was more closely correlated with lysis. SDS/PAGE analysis showed that band 3, the anion transport protein, was lost at the lowest dose of HOBr and at the higher concentrations of HOCl. Labelling the red cells with eosin 5-maleimide, a fluorescent label for band 3, suggested possible clustering of this protein in oxidant-exposed cells. There was also irreversible cross-linking of all the major membrane proteins; this reaction occurred more readily with HOBr. The results indicate that membrane protein modification is the reaction responsible for HOCl-mediated lysis. These effects, and particularly cross-link formation, might result in clustering of band 3 and other membrane and cytoskeletal proteins to form haemolytic pores.

Anion Exchange Protein 1, Erythrocyte↗
PubMed1998

Presenting treatment protocols with Web technology.

This paper describes a Web-based version of a protocol information system (ProtoVIEW) with which a wide range of diagnostic or therapeutic protocols can be retrieved and viewed. The Web-based version contains all functionalities of the non web-based version plus several new functionalities. The web version contains an X-ray viewer and a great deal of interactivity such as validation of electronic patient data forms. The most important additional function is the context sensitive protocol support which may lead to improved protocol adherence. Finally, the web-based version can be accessed from any working place since patient data and protocols are stored centrally.

Clinical Protocols↗
PubMed1997

Hypochlorous acid disrupts the adhesive properties of subendothelial matrix.

We have investigated whether the cell adhesion-promoting properties of the subendothelial matrix are affected by exposure to neutrophil-derived oxidants. Native subendothelial matrix was exposed to increasing doses of H2O2 in the presence of myeloperoxidase and Cl- or to reagent hypochlorous acid (HOCl). Increasing doses of either oxidant system resulted in progressive loss in the adhesive properties of the matrix, and phase contrast microscopy showed that the cells failed to attach to and spread on the oxidant-treated surface. When cells were replated on the treated matrix in the presence of 20% serum, they did attach, but showed abnormal spreading and morphology in longer-term culture. In a modified ELISA system, binding of antibodies specific to fibronectin, thrombospondin and laminin was also disrupted by prior exposure of the matrix to HOCl. Of these components, the cell-binding region of fibronectin was most affected by HOCl, thrombospondin and laminin were less sensitive, and the collagen-binding region of fibronectin was the most resistant. SDS-PAGE of 35S-labelled subendothelial matrix proteins indicated that there was no major irreversible crosslink formation or fragmentation after exposure to HOCl or the myeloperoxidase system, although formation of disulfides is quite likely.

Cell Adhesion↗
PubMed1997

Introduction of a computerised protocol in clinical practice: is there anything to gain?

OBJECTIVES: To assess the potential benefit of a protocol for the diagnostic work-up and management of patients with obstructive jaundice, by comparing its recommendations with the policies actually followed in patients and to compare local expertise with diagnostic and therapeutic procedures with that described in published reports. DESIGN: A retrospective analysis of patients' records. SETTING: University hospital, The Netherlands. SUBJECTS: 49 consecutive patients who presented to the departments of internal medicine and surgery between June 1990 and June 1992 with serum alkaline phosphatase activities > 125 mumol/L, and serum bilirubin concentrations > 17 mumol/L. MAIN OUTCOME MEASURES: The proportions of diagnostic and therapeutic decisions that deviated from the recommendations, and the success rates of diagnostic and therapeutic procedures. RESULTS: In patients with bile duct stones the treatment strategies did not deviate from those recommended in the protocol. In patients with cancer 38 (30%) of the 128 diagnostic decisions and 4 (11%) of the 37 therapeutic decisions deviated from the protocol. Success rates of all diagnostic investigations were comparable with those reported, and success rates of endoscopic biliary drainage tended to be lower than those reported. CONCLUSIONS: The introduction of a protocol for the diagnostic work-up of patients with obstructive jaundice may reduce unnecessary investigations and diagnostic omissions by half. Because local expertise of some of the procedures seems to be significantly less than reported elsewhere it may be necessary to modify the protocol to better fit local circumstances.

Aged↗
PubMed1996

Impact of a protocol processing system (ProtoVIEW) on clinical behaviour of residents and treatment.

A protocol processing system (ProtoVIEW), containing therapeutic trauma protocols, was used in the Accident and Emergency (A and E) department for a period of 7 months to investigate the impact of automated protocols on firstly, medical decision making of physicians and secondly, on quality of treatments eventually received by the patients. A randomized controlled trial showed that mandatory use of the system led to a more uniform working strategy while fracture treatment only seemed to improve in a subgroup of patient for whom residents established a correct diagnosis.

Adolescent↗
PubMed1996

Effects of a supportive protocol processing system (ProtoVIEW) on clinical behaviour of residents in the accident and emergency department.

A randomized two period crossover trial was performed at the Accident and Emergency (A & E) department of the University Hospital in Nijmegen (The Netherlands). We assessed what the impact was of (mandatory) consultation of a protocol for the management of isolated traumas on treatment decisions of residents. All eight surgical residents who regularly worked in the A & E department participated in the trial. All patients who entered the A & E department between October 13, 1992 and June 9, 1993, of age 16 years or older with an isolated fracture without concomitant lesions were admitted to the study. During the experimental periods, the management protocol was available on computer (using ProtoVIEW) and during the control periods on paper. Main measurements were treatment adjustments made by residents (after consulting different information sources), and their opinion about ProtoVIEW as an information source assessed by means of a questionnaire. When protocol consultation was mandatory, residents changed their treatments almost four times more often towards the protocol than during the control periods (P = 0.01 Chi-square test). Most residents found ProtoVIEW easy to use, liked it as a useful training source while half of them said they would use the system in daily clinical practice. We conclude that mandatory protocol consultation using ProtoVIEW influenced protocol adherence positively.

Attitude↗
PubMed1995

Oxidation of intracellular glutathione after exposure of human red blood cells to hypochlorous acid.

Exposure of human red blood cells to low doses of hypochlorous acid (HOCl) resulted in the loss of intracellular GSH. Oxidation occurred less than 2 min after the addition of HOCl, and required approx. 2.5 mol of HOCl per mol of GSH lost. Loss of GSH preceded oxidation of membrane thiols, the formation of chloramines and haemoglobin oxidation. The susceptibility of intracellular GSH to oxidation by HOCl was two-thirds that of GSH in cell lysates. These results indicate that HOCl can penetrate the red cell membrane, which provides little barrier protection for cytoplasmic components, and that GSH oxidation by HOCl may be a highly selective process. Virtually all of the GSH lost was converted into GSSG. If glucose was added to the medium, most of the GSH oxidized by low doses of HOCl was rapidly regenerated. At higher doses, recovery was less efficient. However, when HOCl was added as a slow infusion rather than in a single bolus, there was increased recovery at higher doses. This indicates that in metabolically active cells regeneration is rapid and GSH may protect cell components from damage by HOCl. HOCl-induced lysis was only slightly delayed by adding glucose to the medium, indicating that lytic injury is not ameliorated by GSH.

Cell Membrane Permeability↗
PubMed1995

Consultation behaviour of residents supported with a protocol processing system (ProtoVIEW) at the emergency ward.

We evaluated the consultation behaviour of residents using a protocol processing system in routine clinical practice. A total of 125 consecutive patients, of age 16 years or older with an isolated fracture without concomitant lesions, were treated with computer support between 13 October 1992 and 9 June 1993. All eight surgical residents who worked at the emergency ward of the University Hospital in Nijmegen, The Netherlands participated. The mean consultancy time, method of information retrieval, number of correct protocols found, number of windows retrieved and attitude towards ProtoVIEW as a useful information source were estimated. Main results are: a mean consultancy time of 1.5 min per case, residents browsed through the protocol information more often than using keyword search. The correct protocols were found in 98% of the cases while on average a minimum number of text-browse windows was retrieved. Residents were positive about the way protocols were presented and about the information supplied by ProtoVIEW. From this study we may conclude that ProtoVIEW consultation is hardly time consuming, and easy to use. Since keyword search was hardly used, expanding the number of synonyms may stimulate searching by keyword more often.

Adolescent↗
PubMed1995

Development, implementation and a first evaluation of a protocol processing system (ProtoVIEW).

In this contribution the protocol information system ProtoVIEW is presented. The system provides the necessary information to physicians about diagnostic procedures and therapies. It is implemented as a stand alone system. The design criteria are discussed and the results of a first evaluation presented. It appears that interns can easily find the required information with the help of the system. The time that they need for accessing the relevant information is relatively short (about 1 min). The users expressed the opinion that the system is easy to use and does support them in the management of their patients. On the basis of this evaluation and evaluations reported elsewhere it is concluded (a) that stand-alone protocol systems can support daily patient management in a positive way and (b) that the design criteria for a protocol information system as presented in this paper are useful for prospective protocol information system developers.

Attitude of Health Personnel↗
PubMed1994

A single assay for measuring the rates of phagocytosis and bacterial killing by neutrophils.

We have developed a method that enables the rates of phagocytosis and killing of bacteria by neutrophils to be measured in a single assay. Neutrophils were incubated with bacteria, and at specific intervals were separated from uningested bacteria by low speed centrifugation. Rates of phagocytosis and killing were calculated from the decrease in number of extracellular bacteria and change in the number of intracellular bacteria. Both phagocytosis and killing were shown to follow first-order kinetics, and rate constants were calculated without having to separate the assay into two phases. In contrast to two-step methods, our method measures killing from the moment the neutrophils start ingesting the bacteria, and also eliminates the need to halt neutrophil activity temporarily and restart the assay after the extracellular bacteria have been removed. We obtained reproducible results for the phagocytosis and killing of Staphylococcus aureus (t1/2 = 9 min and 6 min respectively) and Escherichia coli (t1/2 = 10 min and 2 min respectively). We also were able to detect a 56% impairment in the rate of killing of S. aureus by neutrophils from an individual with a low level of myeloperoxidase.

Cell Separation↗