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Biomedical subjects

L Kunz

Publications and source records attributed to L Kunz.

At least 19 recordsLinked to original sources
PubMed2006

Non-neuronal acetylcholine and choline acetyltransferase in oviductal epithelial cells of cyclic and pregnant pigs.

Certain female reproductive tissues are known to express the non-neuronal cholinergic system. Using different experimental approaches, we tested the hypothesis that acetylcholine (ACh) in the porcine oviduct may also be derived from non-neuronal structures. Immunohistochemistry was performed to detect acetylcholine synthesizing enzyme choline acetyltransferase (ChAT) in different segments of the oviduct of cyclic and pregnant sows. Immunohistochemical experiments revealed strong immunoexpression of ChAT in the entire oviductal epithelium at metoestrus. Thereby, a particular pronounced staining was found in the supranuclear region of almost all epithelial cells. Immunostaining of ChAT decreased markedly during dioestrus and prooestrus stages, respectively. At prooestrus, ChAT immunoreactivity was confined to ciliated cells. Furthermore, we found elevated level of staining intensity of ChAT in the pregnant oviduct at day 13. Using the same ChAT antibody for Western blot analyses, we detected immunoreactive bands of MW 69,000 and 46,000 mainly in ampulla, while MW 58,000 and 30,000 forms were present mainly in infundibulum and isthmus. Furthermore ACh was detected by HPLC and fluorimetric methods in oviductal epithelium. In conclusion, we show expression of ChAT in oviductal epithelial cells at different stages of the oestrus cycle and pregnancy, indicating that these cells can synthesize ACh in a cycle-dependent manner. These results suggest as yet unexplored roles of epithelial ACh in the oviduct.

Acetylcholine↗
PubMed2005

Divergent effects of the major mast cell products histamine, tryptase and TNF-alpha on human fibroblast behaviour.

Fibroblast proliferation is a key process in tissue remodeling and mast cells (MCs) are thought to play a crucial role. Having established that the three major MC products, tryptase, histamine and TNF-alpha (TNF) are normally present in human skin MCs, which are in close proximity to dermal fibroblasts, we studied their individual effects on cell cycle-controlled human dermal fibroblasts (HFFF2). These cells express receptors (H1, PAR2, TNFR1/2) for the major MC mediators, but only tryptase or a PAR2 agonist peptide stimulated proliferation and gene expression. TNF was antimitotic, and histamine, while elevating intracellular Ca2+ levels at high concentrations, did not affect proliferation. We conclude that MC products but also composition and numbers of respective receptors on fibroblasts are crucially responsible for fibroproliferative events.

Cell Cycle↗
PubMed2004

Tryptase inhibits motility of human spermatozoa mainly by activation of the mitogen-activated protein kinase pathway.

BACKGROUND: We previously localized protease-activated receptor 2 (PAR-2) on human spermatozoa and demonstrated that activation of PAR-2 by the mast cell (MC) product tryptase inhibits sperm motility. Importantly, tryptase-secreting MCs are encountered in the male and female genital tract, implying that MC-spermatozoa interactions may be as yet unrecognized factors affecting sperm fertilizing ability. In order to elucidate how tryptase via activation of PAR-2 acts in human spermatozoa, we studied intracellular signal transduction events. METHODS AND RESULTS: Impairment of sperm motility by tryptase was not dependent on the presence of extracellular Ca2+ and tryptase did not alter intracellular Ca2+ levels. Pre-incubation with pertussis toxin (PTX) failed to prevent tryptase effects on sperm motility. Western blot analyses revealed that tryptase increased phosphorylation of the mitogen-activated protein kinases (MAPK) ERK1/2, an effect which was blocked by the MAPK pathway inhibitor PD98059. Pre-treatment of spermatozoa with this inhibitor also blocked the inhibtion of sperm motility evoked by tryptase. CONCLUSIONS: These results indicate that tryptase acts via the ERK1/2 pathway to inhibit motility of human spermatozoa.

Calcium↗
PubMed2004

Correlation of real-time haemoglobin oxygen saturation monitoring during photodynamic therapy with microvascular effects and tissue necrosis in normal rat liver.

Photodynamic therapy (PDT) requires a photosensitising drug, light and oxygen. While it is known that the haemoglobin oxygen saturation (HbSat) can be altered by PDT, little has been done to correlate this with microvascular changes and the final biological effect. This report describes such studies on the normal liver of rats sensitised with aluminium disulphonated phthalocyanine. In total, 50 J of light at 670 nm, continuous or fractionated at 25 or 100 mW, was applied with a single laser fibre touching the liver surface. HbSat was monitored continuously 1.5-5.0 mm from the laser fibre using visible light reflectance spectroscopy (VLRS). Vascular shutdown was assessed by fluorescein angiography 2-40 min after light delivery. Necrosis was measured at post mortem 3 days after PDT. In all treatment groups at a 1.5 mm separation, HbSat fell to zero with little recovery after light delivery. At 2.5 mm, HbSat also decreased during light delivery, except with fractionated light, but then recovered. The greatest recovery of fluorescein perfusion after PDT was seen using 25 mW, suggesting an ischaemia/reperfusion injury. Necrosis was more extensive after low power and fractionated light than with 100 mW, continuous illumination. We conclude that VLRS is a useful technique for monitoring HbSat, although the correlation between HbSat, fluorescein exclusion and necrosis varied markedly with the light delivery regimen used.

Animals↗
PubMed2001

Photomodification of the electrical properties of the plasma membrane: a comparison between 6 different membrane-active photosensitizers.

The present study deals with photomodification of the electrical properties of the plasma membrane of an epithelial cell line (opossum kidney (OK) cells). The effect of photofrin II (previously investigated) is compared with that of 5 other membrane-active sensitizers: sulfonated Zn-phthalocyanine, merocyanine 540, rose bengal, methylene blue and protoporphyrin IX (an endogenous sensitizer induced by addition of its biosynthetic precursor 5-aminolaevulinic acid). The study was performed in order to investigate whether photomodification of the ion transport properties of the plasma membrane by membrane-active sensitizers is a general and early event in cellular photosensitization. The changes in the electrical properties were monitored by application of the whole-cell and the inside-out configuration of the patch-clamp technique. Illumination in the presence of the compounds (apart from merocyanine 540) gave rise to similar changes of the electrical properties of the membrane: depolarization of the membrane potential, inactivation of a large-conductance, Ca2+-dependent K+-channel (maxi-KCa), and a strong increase of the leak conductance of the membrane. This similarity indicates the general character of the functional photomodifications by membrane-active sensitizers previously reported for photofrin II.

Aminolevulinic Acid↗
PubMed2001

[Clinical pilot study of interstitial photodynamic therapy for treatment of advanced head and neck tumors].

BACKGROUND: Photodynamic therapy is a new treatment modality which uses a photochemical reaction to destroy tumour tissue. To date this treatment has only been applied effectively to the surface of small, superficial tumours due to the limitations imposed by light penetration. With the use of fibres introduced into the substance of the tumour, more bulky lesions can be treated successfully. STUDY: We describe the first clinical use of such a treatment in the management and, specifically, the amelioration of advanced head and neck cancer. We describe our results in 12 patients, in 11 of whom we were able to improve quality of life. In one patient there was no observable effect. The side-effects were minimal. DISCUSSION: We suggest that interstitial photodynamic therapy may be of use in the palliative care of patients with advanced disease and bulky tumours. It may also be of use in benign, bulky tumors, but this point requires further study.

Head and Neck Neoplasms↗
PubMed1999

Radiolytic and photodynamic modifications of ion transport through the plasma membrane of OK cells: a comparison.

PURPOSE: This study seeks to compare functional modifications of the plasma membrane induced either by water radiolysis (80 kV X-rays) or by visible light in the presence of the photosensitizer photofrin II. The two methods generate different kinds of free radicals and other reactive species such as singlet oxygen. Their effect on the electrical properties of the plasma membrane is investigated. METHOD: The patch-clamp technique (whole-cell and inside-out configuration) was applied to the plasma membrane of an epithelial cell line (opossum kidney (OK) cells). RESULTS: Generation of the reactive species gave rise to a decay of the membrane potential, to an inactivation of K+-channels, and to an increase of the leak conductance of the membrane. All three effects were strongly involved in photomodification of the membrane. Following X-ray exposure (doses up to 1000 Gy) on the other hand, the decay of the membrane conductance caused by inactivation of ion channels was predominant, while the increase of the leak conductance and the depolarization were comparatively minor responses. CONCLUSION: Photomodification of the plasma membrane by visible light in the presence of the membrane-active sensitizer photofrin II is of considerably greater consequence for the electrical properties of the membrane, compared with exposure to ionizing radiation of the membrane and its aqueous environment. The differences are thought to reflect the types of reactive species produced by the two methods and their site of generation.

Animals↗
PubMed1998

Photofrin II sensitized modifications of ion transport across the plasma membrane of an epithelial cell line: I. Electrical measurements at the whole-cell level.

Photofrin II is a photosensitizer frequently applied in photodynamic therapy. Light-induced tumor cell inactivation observed in the presence of this substance has been suggested to start with modifications at the level of cellular membranes. In the present study electrophysiological techniques are applied in order to investigate the action of photofrin II on functional properties of the plasma membrane of opossum kidney (OK) cells (as an epithelial model system) and of fibroblasts. Illumination of the cells in the presence of photofrin II (or Zn-phthalocyanine) leads to comparatively fast depolarization of the membrane potential. It is caused by a strong change of the membrane conductance which proceeds in two phases. Both phases contribute to a loss of ion selectivity of the plasma membrane between K+ and Na+. In the first phase, specific pathways for K+, which determine the resting potential under physiological conditions, are inactivated. The second phase is distinguished by a marked increase of a nonselective conductance. The increase of the latter - after light-induced initiation - continues in the dark. The conclusions are derived from light-induced, time-dependent changes of the membrane conductance and of the shape of the current-voltage relationship detected under different experimental conditions.

Animals↗
PubMed1998

Photofrin II sensitized modifications of ion transport across the plasma membrane of an epithelial cell line: II. Analysis at the level of membrane patches.

In the first part of this study, photofrin II sensitized membrane modifications of OK-cells were investigated at the level of macroscopic membrane currents. In this second part, the inside-out configuration of the patch-clamp technique is applied to analyze the phenomena at the microscopic level. It is shown that the characteristic single channel fluctuations of the electric current disappear after the start of illumination of membrane patches in the presence of photofrin II. This holds for all three types of ion channels investigated: the large-conductance Ca2+-dependent K+ channel (maxi-KCa), a K+ channel of small conductance (sK), and a stretch-activated nonselective cation channel (SA-cat). Part of the experiments show a transient activation of the channels (indicated by an increase of the probability in the open-channel state) before the channels are converted into a closed nonconductive state. Inactivation of all three channel types proceeds by a continuous reduction of their open probability, while the single channel conductance values are not affected. The process of photodynamically induced channel inactivation is followed by a pronounced increase of the leak conductance of the plasma membrane. The latter process - after light-induced initiation - is found to continue in the dark. The ionic pathways underlying the leak conductance also allow permeation of Ca2+ ions. The resulting Ca2+-flux may contribute to the photodynamically induced increase of the intracellular Ca2+ concentration observed in various cell lines.

Animals↗
PubMed1997

Photodynamic membrane damage at the level of single ion channels.

Illumination of cellular membranes by visible light in the presence of appropriate photosensitizers is known to inactivate specific ionic pathways and to increase the unspecific leak conductance of the membranes. While previous studies have concentrated on the macroscopic ionic currents, the present study separates the two phenomena at the microscopic level. Using opossum kidney (OK) cells as epithelial model system and photofrin II as sensitizer, the patch-clamp technique in inside-out configuration has been applied to show the inactivation of single ion channels immediately after start of illumination and the subsequent strong increase of the leak conductance. Inactivation is shown for two kinds of channels: the large-conductance Ca2+-dependent K+ channel (maxi-K(Ca)) and the stretch-activated nonselective cation channel (SA-cat).

Animals↗
PubMed1995

Photodynamic and radiolytic inactivation of ion channels formed by gramicidin A: oxidation and fragmentation.

Ion channels formed by the peptide gramicidin A in planar lipid membranes have been reported to react very sensitively upon irradiation of the membrane by ionizing radiation (radiolysis), by UV light (photolysis), or by visible light in the presence of appropriate photosensitizers (photodynamic inactivation). In all three cases the effect is due to the presence of the four tryptophan residues of the pentadecapeptide. Modifications of these amino acids--due to an interaction with free radicals formed upon water radiolysis or due to light absorption--have been found to reduce the membrane conductance by many orders of magnitude. The present study was intended to correlate functional changes, observed at the level of single ion channels, with changes of the molecular structure identified by mass spectrometry. About 98% of the inactivated channels showed a single-channel conductance of virtually zero, while about 2% of the channels present before irradiation are converted to a state of reduced conductance (and reduced lifetime). On the structural level, irradiation in the presence of the photosensitizer Rose Bengal was found to produce oxidation and fragmentation of the peptide at the positions of the tryptophan residues. Our results provide evidence that the main effect of radiolysis, or of photodynamic treatment, is the cleavage of the peptide backbone leading to immediate closure of an open ion channel.

Amino Acid Sequence↗
PubMed1993

Staphylococcal alpha-toxin kills human keratinocytes by permeabilizing the plasma membrane for monovalent ions.

Incubation of human keratinocytes with nanomolar concentrations of Staphylococcus aureus alpha-toxin leads to irreversible depletion of cellular ATP. The toxin forms hexamers in the target cell membranes, and rapid transmembrane flux of K+, Na+, and 86Rb+ is observed. Unexpectedly, pores formed in keratinocytes through application of low but lethal doses of alpha-toxin appeared to be considerably smaller than those formed in erythrocyte membranes. They permitted neither rapid influx of Ca2+ or propidium iodide, nor efflux of carboxyfluorescein. Larger pores allowing flux of all three markers did form when the toxin was applied at high concentrations. Flux of monovalent ions and reduction in cellular ATP levels evoked by low toxin doses correlated temporally with a fall in oxygen consumption, which was interpreted to reflect breakdown of mitochondrial respiration. The lethal event could not be thwarted by manipulating the extracellular K+ or Ca2+ concentrations. Realization that alpha-toxin may form very small pores in nucleated cells is important for future research on cellular toxin effects and membrane repair processes.

Bacterial Toxins↗
PubMed1989

Interrelationship among morphology, metabolism, and proliferation of tumor cells in monolayer and spheroid culture.

The proliferative activity of tumor cells is positively correlated with the cellular respiration rate. Thus, a decrease in the proliferative fraction during monolayer growth of EMT6 cells is associated with a decrease in the oxygen consumption rate both per cell and per cellular volume. These changes are paralleled by variations in the morphology of the tumor cells. The cellular volume, the cytoplasmic volume, the number and volume of mitochondria is decreased, whereas the volume of the nucleus remains relatively constant during the transition of cells from the exponential to the plateau growth phase. Similar observations have been made in outer, highly proliferative and inner non-proliferating cells in EMT6 spheroids. However, the local oxygen consumption in this three dimensional, tissue-like assemblage of cells is obviously modified by the cellular packing density leading to a rather uniform oxygen uptake in the proliferating and non-proliferating cell areas.

Animals↗
PubMed1980

[Relationships between interruption abortion, and premature birth and low birth weight (author's transl)].

Polydimensional retrospective studies were conducted into correlations between interruption of pregnancy, abortion, and premature birth of infants weighing less than 2,501 g, on the one hand, and the general phenomenon of low birth weight, on the other. Reference was made to parity. The studies were undertaken over a period between 1969 and 1977. The birth total investigated was 17,076, stillbirths and twins excluded.-The impact of all factors involved as well as the statistical significance were tested and verified by calculation of information measure 2-I, followed by an approximative chi 2-test. The results suggested that the parameters studied were of somewhat staggered impact upon low birth weight. Premature birth, interruption of pregnancy, and abortion was the order of impact at all parity levels studied.

Abortion, Induced↗
PubMed1979

[Polydimensional studies into frequency of "infants of low birth weight", with reference to age, parity, and occupation (author's transl)].

Retrospective polydimensional studies were conducted into correlations between age, parity, and occupation (social status) of gravidae, on the one hand, and deliveries of "infants of low birth weight", on the other. The reference group amounted to 16.729 newborns, twins and stillbirth excluded, with 5.85 per cent of those newborns with low birth weights up to 2,500 g. Included in the above figure are all newborns delivered in the Regional Hospital of Schwerin, between 1969 and 1977.--Any single factor quoted, age, parity, and occupation, was found to affect birth weight, effects being by the following order: parity, age, occupation.

Adult↗