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H Kolb

Publications and source records attributed to H Kolb.

At least 181 records · Page 10Linked to original sources
PubMed1993

Dihydrolipoic acid protects pancreatic islet cells from inflammatory attack.

In vitro models of pancreatic islet cell inflammation are the lysis of isolated islet cells by activated macrophages or by oxygen radicals released by the endothelial enzyme xanthine oxidase. Dihydrolipoic acid protected islet cells in both systems by different modes of action. Macrophage cytotoxicity towards islet cells, which is nitric-oxide-mediated, was suppressed by 2 h of preincubation of macrophages with lipoic acid. Similarly, 2 h of preincubation sufficed to protect islet cells against enzymatically produced oxygen radicals. Dihydrolipoic acid was found by chemiluminescence assay to scavenge directly such radicals. In macrophages dihydrolipoic acid suppressed the production of nitrite as a measure of nitric oxide release. These results suggest that dihydrolipoic acid is an anti-inflammatory agent which at the same time interferes with nitric oxide release from inflammatory macrophages and protects target cells from oxygen radical attack.

Animals↗
PubMed1993

Gangliosides protect from TNF alpha-induced apoptosis.

The modulation of tumor necrosis factor alpha-mediated cytotoxicity by gangliosides was analyzed. When cells of the TNF alpha-sensitive fibrosarcoma cell line L929 were incubated for 16 h with recombinant TNF alpha (156 or 312 pg/ml), they were lysed to 65.2% or 78.0%, respectively. The presence of a bovine brain ganglioside mixture (BBG, Cronassial) inhibited the TNF alpha-mediated lysis in a dose-dependent manner (IC50 approximately 1 mg/ml). Maximum inhibition was achieved with 2 mg/ml BBG (82.6% inhibition for 156 pg/ml TNF alpha and 88.5% inhibition for 312 pg/ml TNF alpha). In situ nick translation revealed that BBG (2 mg/ml) significantly reduces the number of cells with TNF alpha-induced DNA-strand breaks from 71.2% to 6.6% (P < 0.0001), indicating a protection against TNF alpha-mediated apoptosis. We conclude that BBG prevents the cytotoxic action of TNF alpha on tumor cells. The inhibitory effect may be due to an interference of gangliosides with intracellular signal transduction pathways, resulting in an inhibition of the activation of DNA-cleaving endonucleases.

Animals↗
PubMed1993

Protection of islet cells from inflammatory cell death in vitro.

Islet cells cocultured with activated macrophages are lysed within 15 h in vitro. We showed previously that nitric oxide generated by macrophages is a major mediator of islet cell death. We have now probed several pathways to interfere with the chain of events leading to islet cell death. Scavenging of extracellular oxygen radicals by superoxide dismutase and catalase did not improve islet cell survival. Scavenging of extra- and intracellular oxygen radicals by two potent substances, citiolone and dimethyl-thiourea, also did not reduce islet cell lysis, while a lipid-soluble scavenger, probucol, provided partial protection. These findings argue against a synergistic action of nitric oxide and oxygen radicals in islet cell toxicity. The inhibition of poly(ADP-ribose)polymerase by 3-aminobenzamide significantly improved islet cell survival. Selective inhibitors of cyclooxygenase, such as indomethacin or acetylsalicylic acid, did not improve islet cell survival. Full protection was seen in the presence of NDGA, an inhibitor of lipoxygenase, and partial suppression was caused by BW755c, an inhibitor of both lipoxygenase and cyclooxygenase. We conclude that inflammatory islet cell death caused by activated macrophages involves the activation of arachidonic acid metabolism and of poly(ADP-ribose)polymerase, but that scavenging of oxygen free radicals provides little protection from lysis.

Animals↗
PubMed1993

A method for selective intracellular labeling of immunostained neurons in turtle retina.

We describe a method for direct intracellular staining under visual control of immunolabeled neurons in the turtle retina. Substance P was the antiserum used. It labels two different sizes of ganglion cells in turtle retina. Intracellular labeling under visual control was achieved by iontophoresis of Lucifer yellow or Neurobiotin. The best immunolabeling of substance P-immunoreactive (SP-IR) ganglion cells occurred after either Triton X-100 or freeze-thaw techniques to get good penetration of the antisera. However, this inevitably resulted in leaky cells and inadequate morphology of the ganglion cells subsequently stained by Lucifer yellow and Neurobiotin. Most successful immunocytochemical labeling followed by intracellular labeling was achieved with light fixation (15 min in 4% paraformaldehyde) and long incubation time in the primary antiserum (4 days). Before intracellular labeling, dendritic tree shape, dendritic field size, and stratification of SP-IR ganglion cells were not sufficiently revealed for correct classification of these cells. After the selective intracellular staining described here, we were able to identify and characterize one of the populations of substance P-IR ganglion cells types as large-field, monostratified G20 ganglion cells.

Adaptation, Ocular↗
PubMed1993

Islet cell DNA is a target of inflammatory attack by nitric oxide.

NO has been identified recently as the prime islet-toxic product of inflammatory macrophages. The adverse effects of IL-1 on isolated islets also have been reported to involve NO. We now show that exposure of an islet cell suspension to the NO donor nitroprusside or to activated macrophages leads to DNA strand breaks. Macrophages did not induce DNA damage in the presence of the NO synthase inhibitor NG-methyl-L-arginine. DNA strand breaks were demonstrated at the level of single cells by a modified nick-translation procedure and confirmed by analysis of DNA fragmentation by gel electrophoresis. DNA strand breaks occurred within 1 h and preceded islet cell lysis. DNA damage could not be prevented by inhibitors of endogenous endonucleases. We conclude that islet cell DNA is an early target of NO action.

Animals↗
PubMed1993

Oxygen radical production is increased in macrophages from diabetes prone BB rats.

Macrophages from autoimmune diabetes prone BB rats were found to produce radical oxygen intermediates (ROI) at an enhanced rate when compared to diabetes resistant BB or normal Wistar rats. The release of ROI was determined by chemiluminescence using in parallel luminol and lucigenin as detector molecules. In diabetes prone BB rats the spontaneous release of ROI was upregulated in macrophages from different compartments, i.e. peritoneum and spleen. Also, maximal output of ROI after activation of macrophages either in vivo by injection of Corynebacterium parvum or in vitro by LPS and IFN was highest for cells from diabetes prone BB rats. This macrophage abnormality was seen in animals prior to recognizable islet inflammation and also was present at the level of macrophages grown in vitro from precursor cells of diabetes prone BB rats. Hypersecretion of oxygen radicals may contribute to Beta cell loss and diabetes development in BB rats.

Animals↗
PubMed1992

Cyclosporin A protects pancreatic islet cells from nitric oxide-dependent macrophage cytotoxicity.

It has been shown earlier in an in-vitro model of inflammatory islet cell death that activated macrophages lyse islet cells via the release of nitric oxide. Here we report that cyclosporin A suppresses macrophage cytotoxicity. Control experiments showed that the immunosuppressive drug does not improve the defences of islet cells against nitric oxide but inhibits the release of nitric oxide from LPS-stimulated macrophages. This property of cyclosporin A may contribute to the preservation of beta cell function seen in cyclosporin A-treated patients with recent onset type I diabetes.

Animals↗
PubMed1992

Neurons of the human retina: a Golgi study.

Golgi techniques have been applied to post mortem specimens of human retina. Analysis was possible on 150 human retinas processed and viewed by light microscopy as wholemounts. Camera lucida drawings and photography were used to classify the impregnated neurons into 3 types of horizontal cell, 9 types of bipolar cell, 24 basic types of amacrine cell, a single type of interplexiform cell, and 18 types of ganglion cell. We have distinguished two types of midget bipolar cell: fmB (flat) and imB (invaginating). In central retina, both types are typically single-headed, each clearly contacting a single cone. Peripherally, they may be two- or even three-headed, obviously contacting more than one cone. Two types of small-field diffuse cone bipolars occurring as flat and invaginating varieties are found across the entire retina from fovea to far periphery. The single rod bipolar type appears about 1 mm from the fovea and increases in dendritic tree diameter from there into the far periphery. The putative "ON-center" blue cone bipolar and the giant bistratified bipolar first described by Mariani are also present in human retina and we add two previously undescribed bipolar cell types: a putative giant diffuse invaginating and a candidate "OFF-center" blue cone bipolar. Taking into account the variation of cell size with eccentricity at all points on the retina, we observed three distinct varieties of horizontal cell. The HI is the well known, long-axon-bearing cell of Polyak. HII is the more recently described multibranched, wavy-axoned horizontal cell. The third variety, HIII, introduced here, has been separated from the HI type on morphological criteria of having a larger, more asymmetrical dendritic field and in contacting 30% more cones than the HI at any point on the retina. Amacrine cells proved to be most diverse in morphology. Many of the amacrine cell types that have been described in cat retina (Kolb et al., '81: Vision Res. 21; 1081-1114) were seen in this study. Where there are no equivalent cells in cat, we have adopted the descriptive terminology used by Mariani in monkey retina. Thus eight varieties of small-field amacrines (under 100 microns dendritic trees), eight varieties of medium-field cells (100-500 microns dendritic span), and eight large-field varieties (over 500 microns dendritic trees) have been classified. Often a broadly described variety of amacrine cell can be subdivided into as many as three subtypes dependent on stratification levels of their dendrites in the inner plexiform layer.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Cytotoxic action of IL-1 beta against pancreatic islets is mediated via nitric oxide formation and is inhibited by NG-monomethyl-L-arginine.

IL-1 beta has been previously shown to act as a cytotoxic agent in islets. Here we show by electron microscopy of alginate encapsulated islets, that islet cell lysis is induced by culturing islets for 24 or 48 h in the presence of IL-1 beta. The extent of lysis depends on the IL-1 beta concentration and is slightly enhanced by the addition of TNF-alpha. Cells can be protected from lysis by NG-monomethyl-L-arginine. Lysis is paralleled by an increase in nitrite concentration in culture supernatants of whole islets but not in supernatants of isolated endocrine cells. The results indicate that IL-1 beta toxicity occurs via inducing in non-endocrine islet cells the synthesis and release of nitric oxide, which has been shown earlier to be highly toxic for islet cells.

Animals↗
PubMed1992

Localization of GABA, glycine, glutamate and tyrosine hydroxylase in the human retina.

A light microscope study using postembedding immunocytochemistry techniques to demonstrate the common neurotransmitter candidates gamma-aminobutyric acid (GABA), glycine, glutamate, and tyrosine hydroxylase for dopamine has been done on human retina. By using an antiserum to GABA, we found GABA-immunoreactivity (GABA-IR) to be primarily in amacrine cells lying in the inner nuclear layer (INL) or displaced to the ganglion cell layer (GCL). A few stained cells in the INL, which are probably interplexiform cells, were observed to project thin processes towards the outer plexiform layer (OPL). There were heavily stained bands of immunoreactivity in strata 1, 3 and 5 of the inner plexiform layer (IPL). An occasional ganglion cell was also GABA-IR. By using an antiserum to glycine, stained cells were observed at all levels of the INL. Most of these were amacrines, but a few bipolar cells were also glycine-IR. Displaced amacrine cells and large-bodied cells, which are probably ganglion cells, stained in the GCL. The bipolar cells that stained appeared to include both diffuse and midget varieties. The AII amacrine cell of the rod pathway was clearly stained in our material but at a lower intensity than two other amacrine cell types tentatively identified as A8 and A3 or A4. Again, there was stratified staining in the IPL, with strata 2 and 4 being most immunoreactive. An antiserum to glutamate revealed that most of the neurons of the vertical pathways in the human retina were glutamate-IR. Rod and cone photoreceptor synaptic endings labeled as did the majority of bipolar and ganglion cells. The rod photoreceptor stained more heavily than the cone photoreceptor in our material. While both midget and diffuse cone bipolar cell types were clearly glutamate-IR, rod bipolars were not noticeably stained. The most strongly staining glutamate-IR processes of the IPL lay in the outer half, in sublamina a. The antiserum to tyrosine hydroxylase (TOH) revealed two different amacrine cell types. Strongly immunoreactive cells (TOH1) had their cell bodies in the INL and their dendrites ramified in a dense plexus in stratum 1 of the IPL. Fine processes arising from their cell bodies or from the stratum 1 plexus passed through the INL to reach the OPL but did not produce long-ranging ramifications therein. The less immunoreactive amacrines (TOH2) lay in the INL, the center of the IPL or the GCL and emitted thick dendrites that were monostratified in stratum 3 of the IPL.

Aged↗
PubMed1992

Immunocytochemical staining with antibodies against protein kinase C and its isozymes in the turtle retina.

An LM immunocytochemical study has investigated the patterns of staining in turtle retina with monoclonal antibodies to the alpha, beta and gamma isozymes of protein kinase C. The protein kinase C-gamma antibody reveals cells in the ganglion cell layer, occasional amacrine cells and faint banding in strata 2 and 4 of the inner plexiform layer. The protein kinase C-beta antibody stains primarily amacrine cells that have dendrites running in strata 2, in 4 close to the 3/4 border and on the 4/5 border of the inner plexiform layer. Protein kinase C-alpha immunoreactivity is seen in a population of bipolar cells. The latter are characterized by stained axon terminals in strata 3 and 4 of the inner plexiform layer. A type of amacrine cell, different from those seen with the other antibodies, is also immunoreactive to protein kinase C-alpha. EM immunocytochemistry (using a polyclonal antibody) reveals protein kinase C immunoreactivity in photoreceptor cells, bipolar cells, amacrine cells and ganglion cells. In photoreceptors protein kinase C immunoreactivity occurs as patchy staining associated with vesicles and the plasmalemma in pedicles and telodendria. Some varieties of bipolar cell display protein kinase C reaction product throughout the entire cell. Their dendrites contact photoreceptor pedicles at wide-cleft basal junctions and ribbon and non-ribbon related narrow cleft junctions. A few lateral elements per cone or rod pedicle are always protein kinase C-immunoreactive. Amacrine and ganglion cells typically show small clumps of protein kinase C immunoreactivity around vesicles and close to the postsynaptic membranes. Synaptic boutons of some varieties of amacrine cell stain more uniformly. Protein kinase C-immunoreactive bipolar cells are most commonly presynaptic in stratum 4 of the inner plexiform layer, while protein kinase C-immunoreactive amacrine cells are both pre- and postsynaptic throughout strata 1, 2, 3 and 4. Stratum 5 appears to be almost devoid of protein kinase C-immunoreactive neural profiles.

Animals↗
PubMed1992

Oxygen radicals generated by the enzyme xanthine oxidase lyse rat pancreatic islet cells in vitro.

The endothelium-associated enzyme xanthine oxidase is known to generate reactive oxygen intermediates which may damage the surrounding tissue. We investigated whether reactive oxygen intermediates released by xanthine oxidase exert a toxic effect on isolated rat islet cells. The xanthine oxidase (25 mU/ml)/hypoxanthine (0.5 mmol/l) system released reactive oxygen intermediates in vitro as detected by luminol in a chemiluminescence analysing system. The addition of nicotinamide inhibited the release of reactive oxygen intermediates in a dose-dependent manner (50% inhibition at 20 mmol/l). Exposure of islet cells to enzyme generated reactive oxygen intermediates caused lysis of 39% of the cells within 15 h. Monitoring the mitochondrial function of islet cells by the conversion of tetrazolium bromide to its formazan product revealed a significant reduction of the respiratory activity down to 51% of that of the controls by 30 min after the initiation of the xanthine oxidase reaction. Mitochondrial dysfunction preceded plasma membrane damage. The addition of nicotinamide, a radical scavenger and inhibitor of the DNA repair enzyme poly(ADP-ribose) synthetase protected the islet cells from lysis and partially preserved their mitochondrial activity in the presence of reactive oxygen intermediates. We conclude that activation of the endothelial enzyme xanthine oxidase, known to be induced by mediators of immune cells or by episodes of ischaemia and reperfusion causes islet cell damage with subsequent cell death in early phases of pancreatic islet cell destruction.

Animals↗
PubMed1992

Ganglioside therapy of type I diabetes: enhancement of hyperglycemia in the low dose streptozotocin model.

The therapeutic potential of bovine brain gangliosides on the development of insulin deficient diabetes was analysed. Daily ganglioside administration (50 mg/kg body weight) caused a more pronounced rise of blood glucose levels (p less than 0.05) in low dose streptozotocin treated mice, a model of human type I diabetes. Hyperglycemia induced by the injection of a single high dose of streptozotocin was slightly increased by ganglioside administration (not significant). The previously reported protective effect of bovine brain gangliosides on the development of diabetes in NOD mice was thus not found in a second mouse model.

Animals↗
PubMed1992

Toxicity of chemically generated nitric oxide towards pancreatic islet cells can be prevented by nicotinamide.

Previous studies have indicated that nitric oxide is involved in the lysis of pancreatic islet cells by inflammatory macrophages. Here we show that the incubation of islet cells with chemical NO-donors leads to cell lysis in a concentration and time dependent way. Islet cell death could be prevented by nicotinamide and 3-aminobenzamide, which are known to inhibit ADP-ribosylation, while several scavengers of oxygen radicals, N-acetylcysteine, dihydrolipoic acid, dimethylthiourea and citiolone, provided no protection.

Acetylcysteine↗
PubMed1992

Neurons immunoreactive to choline acetyltransferase in the turtle retina.

Light microscopic immunocytochemistry using anti-choline acetyltransferase (ChAT) was performed to stain putative cholinergic amacrine cells in turtle retina. ChAT-immunoreactive somata lie in the inner nuclear (INL) and ganglion cell (GCL) layers. Three types of amacrine cells were found according to the location of their somata and their dendritic stratification pattern in the inner plexiform layer (IPL). Type I amacrines lie in the row of cells closest to the INL/IPL limits and they branch along the s1/s2 border of the IPL. Type II amacrines are displaced to the GCL and they ramify along the s3/s4 border of the IPL. Type III amacrines lie in the middle of the INL, 2-3 rows away from the IPL limits and their dendrites appear to be bi- or tri-stratified in s1 and s3-s4 of the IPL. The turtle ChAT-IR amacrines are thus similar to the types described in chicken retina. A regular, non-random mosaic formed by stained type II amacrine cells was observed in the GCL. Their density in mid-central retina was 750 cells/mm2, tapering off to 393 cells/mm2 in peripheral retina. Our study indicates that a pair of cholinergic amacrine cell types in turtle retina is arranged in mirror-image symmetry contributing to sublamina "a" and sublamina "b" of the IPL, like in other vertebrate retinas.

Animals↗