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Biomedical subjects

W Jacob

Publications and source records attributed to W Jacob.

At least 37 records · Page 2Linked to original sources
PubMed1996

Biotin- or digoxigenin-conjugated nucleotides bind to matrix vesicles in atherosclerotic plaques.

The present study analyzes the staining pattern of DNA in situ end-labeling techniques of human and rabbit atherosclerotic plaques. Both the terminal deoxynucleotidyl transferase end-labeling and the in situ nick translation technique detected, besides apoptotic nuclei, numerous round vesicles with diameters from 0.5 to 5 microns within the atherosclerotic plaques. These vesicles did not contain DNA but contained calcium. A pretreatment with EDTA or citric acid abolished the labeling of the vesicles but did not influence the detection of apoptotic nuclei. Ultrastructurally, the vesicles were of variable diameter and density, and their aspect was compatible with matrix vesicles, which are well known in epiphyses during bone formation. The larger vesicles contained cell organelles, and the small vesicles were very dense. X-ray microanalysis demonstrated high calcium and phosphorus levels within the most dense vesicles. Different stages of the process were present in the plaques. In this way we could demonstrate that cytoplasmic fragmentation of smooth muscle cells and subsequent formation of matrix vesicles are a frequent finding in atherosclerotic plaques. The association of apoptotic cell death and formation of matrix vesicles could be an interesting pathway in explaining calcification of atherosclerotic plaques. Both the terminal deoxynucleotidyl transferase end-labeling and the in situ nick translation technique detected simultaneously apoptotic nuclei and matrix vesicles if calcium is not removed from the sections.

Animals↗
PubMed1996

Applications of laser microprobe mass spectrometry in biology and medicine.

Laser microprobe mass spectrometry (LMMS) provides spot analysis with a lateral resolution of 1-5 microns. Focused laser ionisation, combined with mass spectrometry, yields information on elements, molecular identification of inorganic substances, and structural characterisation of organic molecules of typically less than 2-3 kDa. Quantification in complex heterogeneous systems, such as biological materials, is not possible at the present state of the art. The strength of the method lies in the qualitative information on the molecular composition of the analyte, not just element detection, with a lateral resolution at the light microscopy level. The applications of this technique in the field of biomedicine and biology are illustrated by examples that demonstrate the use of this qualitative information in practical test cases.

Fourier Analysis↗
PubMed1995

Lidocaine does not prevent the calcium paradox in rat hearts: a laser microprobe mass analysis (LAMMA) study.

The calcium paradox stands for the cell damage that occurs when isolated hearts are perfused with a Ca(2+)-free solution followed by perfusion with a Ca(2+)-containing solution. Although it is generally accepted that a massive Ca2+ influx during the Ca(2+)-repletion phase is responsible for the cell damage, there is no consensus about what makes the heart susceptible to the calcium paradox during the Ca(2+)-depletion phase. It has been suggested that the extent of the calcium paradox is primarily determined by accumulation of Na+ during Ca2+ depletion and a subsequent accumulation of Ca2+ via reverse Na(+)-Ca2+ exchange during Ca2+ repletion. According to another theory, weakening of intercalated disc junctions during Ca2+ depletion and contracture-mediated disruption of the cell membrane during Ca2+ repletion are responsible for the calcium paradox. In the present study we further investigated the possible role of Na+ in the development of the calcium paradox. During Ca2+ depletion, lidocaine was used to inhibit Na+ entry through the Na+ channels. Isolated rat hearts were perfused with Krebs Henseleit buffer (KH) containing 1.4 mM Ca2+ for 15 min, followed by 10 min of Ca(2+)-free perfusion and 10 min of reperfusion with Ca2+. In the treated group 0.1 mM lidocaine was present throughout the experiment. At the end of each experiment, Ca2+ cytochemistry was performed and the intracellular Ca2+ content was analyzed by laser microprobe mass analysis (LAMMA). The results show that during Ca2+ depletion, the intracellular Ca2+ content did not change significantly. Ca2+ repletion, however, gave rise to a full calcium paradox irrespective of the presence of lidocaine: massive cell damage and Ca2+ accumulation in the mitochondria. The results provide further evidence that intracellular Na+ accumulation during Ca2+ depletion is not involved in the occurrence of the calcium paradox.

Animals↗
PubMed1995

Bronchoscopic insufflation of room air for the treatment of lobar atelectasis in mechanically ventilated patients.

Segmental and lobar pulmonary atelectasis is a common occurrence in mechanically ventilated patients. Standard therapy for atelectasis relies on positive pressure ventilation, positive and expiratory pressure (PEEP), tracheobronchial toilet and regular chest physiotherapy. Various adjuncts to physiotherapy such as bronchoscopic clearance of secretions have not proved to be of additional benefit. Bronchoscopic clearance of secretions followed by insufflation of room air at 30 cm H2O into the atelectatic segment was employed on ten occasions in mechanically ventilated patients. Rapid re-expansion of the collapsed segment or lobe occurred in seven out of the ten treatments.

APACHE↗
PubMed1995

The effect of ischemia and reperfusion on mitochondrial contact sites in isolated rat hearts.

Contact sites may be described as energy channels between the mitochondria and the cytosol, created by fusion of the inner and the outer mitochondrial membranes, and their number depends highly on the energy state of the cell. The aim of the present study was to examine the early changes of ischemia and reperfusion on the number of mitochondrial contact sites. Therefore isolated rat hearts were subjected to short periods of ischemia followed by reperfusion. The left ventricular pressure (LVP), the contractility (dP/dtmax) and the heart rate were measured. The number of contact sites was morphometrically evaluated. As the flow was stopped, LVP, dP/dtmax and HR declined rapidly and became undetectable after 2 min of ischemia. The number of contact sites fell to a minimum after 10 min of ischemia after an initial increase (1 min of ischemia). A 15 min ischemic period resulted in a high number of contact sites which decreased again after 20 min of ischemia. Reperfusion after 2 min of ischemia caused an immediate functional recovery and a high presence of contact sites. After 15 min of reperfusion, all values returned to control values. Reperfusion after 10 min of ischemia resulted in a slow recovery of the number of contact sites and after 15 min of ischemia the number of contact sites remained low upon reperfusion. We may conclude that mitochondria lose the ability to form contact sites after more than 15 min of ischemia and this might be a first indication of irreversible injury.

Animals↗
PubMed1994

The effect of calcium on mitochondrial contact sites: a study on isolated rat hearts.

Mitochondrial contact sites are dynamic structures created by fusion of the inner and outer mitochondrial membranes. Stimulation of the metabolism results in an increase of the number of contact sites. Functionally, it is shown that mitochondrial creatine kinase (Mi-CK) is active in contact sites and therefore, Mi-CK cytochemistry was performed (using a tetrazolium salt) to improve the visibility of the contact sites. As calcium is involved as an intracellular messenger of hormonal stimulation, the effect of increasing extracellular calcium concentrations on the number of contact sites was investigated. Therefore, isolated rat hearts were perfused with Krebs-Henseleit buffers differing in their calcium content. During the perfusions the heart function was evaluated and at the end of each experiment, the hearts were processed for Mi CK cytochemistry and the number of contact sites was expressed as the ratio of surface densities contact sites to mitochondrial membranes (Ss). At 2.2 mM calcium perfusion, the physiological parameters and the Ss reached a maximum. This was in contrast to the 0.6 and the 3.6 mM of calcium perfusions whereby both the physiological values and the Ss were decreased. Treatment with noradrenaline in vivo, as was done in previous studies or perfusion with 2.2 mM of calcium ends up with similar values for Ss. From these results, it could be suggested that there might be a link between calcium, heart function and the formation of Mi CK active contact sites.

Animals↗
PubMed1994

Ultrastructural localisation of carnitine acetyltransferase activity in mitochondria of rat myocardium.

The acetyl CoA/CoA ratio is an important regulating factor of beta-oxidation in mitochondria and hence of energy production in the myocardium. Carnitine acetyltransferase provides one of the control mechanisms for this ratio during changing energy demand in the heart muscle, possibly by buffering the CoA and carnitine concentration for sustained beta-oxidation. In search for a possible correlation between the activity of this enzyme and ultrastructural changes in heart mitochondria, carnitine acetyltransferase was cytochemically localised in rat myocardium, brought into different metabolic states. In this work we confirm previous observations, namely the formation of contact sites between inner and outer mitochondrial membranes upon catecholaminergic stimulation of the myocardium. It is further shown that this contact site formation might be a prerequisite for carnitine acetyltransferase to demonstrate enzymatic activity and hence control of beta-oxidation in myocardial mitochondria.

Animals↗
PubMed1994

Co-storage in large 'dense-core' vesicles of dopamine and cholecystokinin in rat striatum.

The subcellular localization of cholecystokinin in the striatum--an area where a high density of cholecystokinin containing terminals has been demonstrated--was studied using biochemical techniques. Cholecystokinin containing vesicles were partially purified using iso-osmotic Ficoll gradients. As judged from their size and their buoyant density in isopycnic gradients, cholecystokinin containing vesicles represent large 'dense-core' vesicles. Negative staining and subsequent immunolabelling for synaptophysin at the electron microscopical level, showed labelled vesicles of 50-70 nm. binding of dihydrotetrabenazine was detected in the cholecystokinin containing fractions. The results suggest that dopamine is co-stored with cholecystokinin in large dense vesicles in rat striatum.

Animals↗
PubMed1993

Study of intracellular deposition of the anti-leprosy drug clofazimine in mouse spleen using laser microprobe mass analysis.

Laser microprobe mass analysis (LAMMA) was used to study the composition of the brick-red crystalline material which had accumulated in the spleen of mice that had received the anti-leprosy drug Clofazimine in their diet for several months. The crystalline deposits light-microscopically resembled pure Clofazimine crystals. The presence of the drug in the crystals was indicated by LAMMA by the appearance of the chloride mass peaks in the negative mass spectra. More specific information was obtained from the positive mass spectra. A mass signal for the protonated molecule was present.

Animals↗
PubMed1993

Erythema chronicum migrans: an electron-microscopic study.

Spirochaetal organisms are found in skin specimens obtained by biopsy from a erythema chronicum migrans lesion. The histological picture shows a logical localization of a lymphohistiocytic cell infiltrate: deep dermal in the central papule and superficial in the erythematous border. The electron-microscopic characteristics of the micro-organisms in these specimens (regular waving appearance with coils every 0.8-1.2 microns, oblique striation of periplasmatic fibrils, cross-section of 0.3 micron, membranes) correspond to the spiral-like structure of Borreliae isolated from blood. The Borreliae seem to move freely through the matrix of the dermis and are not phagocytized. This brings on the typical clinical picture of the centrifugally spreading erythematous band.

Animals↗
PubMed1991

Dehiscence of the facial canal: developmental aspects.

In a series of human fetuses, the course of the facial canal in the temporal bone was investigated by the use of light and scanning electron microscopy. The normal development of the facial canal was correlated to clinical aspects of facial nerve dehiscences. Our observations demonstrate a more complex way of facial canal development not limited to the 'simple' ossification of the otic capsule. Endochondral ossification of the otic capsule does not virtually change the shape of the primitive facial sulcus. The fibrous layers surrounding the facial nerve seem to be responsible for the final architecture of the facial canal and not the otic capsule ossification by itself. The time sequence of their histological development is equally important and permitted us to distinguish three phases in facial canal development. The role of disturbances in epigenetic control for the initiation of dehiscences is discussed.

Cartilage↗
PubMed1990

Contact site between inner and outer mitochondrial membrane: a dynamic microcompartment for creatine kinase activity.

Creatine kinase is involved in the integration of high-energy metabolism in various tissues. In this study the tissue-specific distribution of the mitochondrial isoform was investigated, both by electrophoresis of rat tissue extracts, and by ultrastructural localisation of creatine kinase activity. Furthermore, the influence of uncoupling of oxidative phosphorylation on mitochondrial creatine kinase activity associated with intermembrane contacts was investigated by enzyme cytochemistry and morphometric analysis. The results of the cytochemical survey indicate that contact sites are a prerequisite for creatine kinase to demonstrate enzymatic activity. Moreover, the extent of creatine kinase active membrane contacts depends on the metabolic state of the mitochondrion, as shown for heart mitochondria in vivo and in vitro, before and after treatment with dinitrophenol.

Animals↗
PubMed1990

Early bone formation in the human fetal otic capsule. A methodological approach.

The present study was concentrated on the use of energy-dispersive X-ray analysis and backscattered electron imaging as practical tools for advanced autopsy of human fetuses when diagnostic evaluation of ear pathology is required. These methods were used to revisit the primary calcification front of the fetal otic capsule between 18 and 36 weeks' gestational age. Energy-dispersive X-ray analysis indicates an equal Ca/P ratio in all the three layers of the otic capsule. These results are discussed in view of calcium homeostasis and inner ear function.

Calcification, Physiologic↗
PubMed1989

Ultrastructural localisation of creatine kinase activity in the contact sites between inner and outer mitochondrial membranes of rat myocardium.

The mitochondrial isoenzyme of creatine kinase, together with the ADP/ATP translocase, most probably belongs to a functional multi-enzyme complex located on the inner mitochondrial membrane. The outer membrane is a necessary constituent of this microcompartment. On the other hand, electron microscopic visualisation demonstrated the formation of contact sites between inner and outer mitochondrial membranes as a reaction to variations of the energy metabolism. In search for a possible correlation between these biochemical and morphological phenomena, rat myocardia were brought into the required energy state by stimulation through catecholaminergic mechanisms or adjusted perfusion with amytal. Subsequently, creatine kinase was cytochemically localised. Creatine kinase activity is demonstrated in membrane contacts between inner and outer mitochondrial membranes. The extent of contact sites and creatine kinase activity depends on the metabolic state as shown by morphometric analysis of the surface density of cytochemical reaction product. This surface density diminishes drastically after inhibiting the metabolic activity with amytal. It is concluded that these contact sites are dynamic micro-environments in which the active site of creatine kinase, oxidative phosphorylation and ADP/ATP transport interact during basal and stimulated metabolism.

Amobarbital↗
PubMed1989

Early ossification within the human fetal otic capsule: morphological and microanalytical findings.

Besides the use of conventional techniques such as light and polarization microscopy, the present paper proposes the combined use of transmission electron microscopy, secondary and backscattered electron imaging, energy dispersive X-ray analysis and computed tomography for the diagnostic evaluation of ear pathology in the human fetus. These methods were used to revisit the primary calcification front of the fetal otic capsule between 16 and 23 weeks gestational age. Ultramicroscopic evaluation demonstrates similar fetal bone formation to that found in other bones of the human fetus. The formation of the endosteal and periosteal layers is a typical example of early intra-membranous ossification. The enchondral layer is made up of fibrillar bone, laid down around the calcified cartilage remnants. Microchemical analysis indicates a significantly higher Ca/P ratio in the endochondral layer with respect to the endosteum and periosteum. The consequences of a lower Ca/P ratio in the endosteal layer are discussed in view of calcium homeostasis and inner ear function.

Ear Cartilage↗
PubMed1988

In-depth limitation of the Lamma 500 for the in situ localization of organic compounds in biological embedded tissue samples.

The objective of this study was exploration of the potential, offered by the laser microprobe mass spectrometer (LAMMA), for the in situ localization of organic targets in embedded tissues by means of structurally relevant ions. A series of model systems was designed to evaluate stepwise the analytical problems involved. A preliminary screening pointed to the position of the target in comparison to the actual sample surface as a determining parameter. Refined simulations were carried out with sandwich samples, consisting of an epon carrier (thickness 1 micron), coated with microcrystalline targets and covered with a layer between 5 and 50 nm of different materials (epon, carbon, formvar). In addition to the stimulated conversion of molecular into fragment ions, the presence of a barrier leads to a drastic loss of sensitivity (20-50 x) and unacceptable degradation of the mass spectrometric quality (resolution, calibration).

Gas Chromatography-Mass Spectrometry↗