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Biomedical subjects

H Kolb

Publications and source records attributed to H Kolb.

At least 127 records · Page 7Linked to original sources
PubMed1996

Generation and characterization of inducible nitric oxide synthase deficient macrophage cell lines.

Stable inducible nitric oxide synthase deficient mouse macrophage cell lines were generated by the antisense technology. A 666 bp fragment of a mouse inducible nitric oxide synthase cDNA was cloned in antisense orientation into a mammalian expression vector behind the CMV promoter. This construct was transfected into J774.1A cells, a mouse macrophage cell line. The inducible nitric oxide synthase antisense lines showed up to 84% reduction of nitric oxide production in response to lipopolysaccharide stimulation and 66% reduction of nitric oxide production in response to interferon-gamma and a combination of interferon-gamma and lipopolysaccharide stimulation. The deficiency in inducible nitric oxide synthase expression had no impact on lipopolysaccharide induced tumor necrosis factor alpha and interleukin-1 secretion. The stable and specific inhibition of inducible nitric oxide synthase expression by antisense DNA vectors allows a direct analysis of contribution of inducible nitric oxide synthase activity to macrophage regulatory and immune defence functions.

Animals↗
PubMed1996

Meta-analysis of nicotinamide treatment in patients with recent-onset IDDM. The Nicotinamide Trialists.

OBJECTIVE: Nicotinamide, a vitamin of the B group, has in vitro actions capable of interfering with the pathogenetic process leading to IDDM. Since 1987, several studies have evaluated nicotinamide as a means of protecting beta-cells from end-stage destruction in insulin-treated patients with newly diagnosed IDDM. The aim of the study was to determine whether nicotinamide protects residual beta-cell function when given at IDDM diagnosis. RESEARCH DESIGN AND METHODS: We performed a meta-analysis of the integrated parameters of metabolic control (C-peptide, glycosylated hemoglobin, insulin dose) in 10 randomized (5 of which were placebo) controlled trials conducted in recent-onset IDDM patients for a total of 211 nicotinamide-treated patients. Data on the adverse effects of nicotinamide were also collected from an additional four trials to yield a grand total of 291 nicotinamide-receiving patients. RESULTS: One year after diagnosis, baseline C-peptide was significantly higher in nicotinamide-treated patients, compared with control patients (0.73 +/- 0.65 vs. 0.32 +/- 0.56 ng/ml, P < 0.005). This statistical difference remained also when the five placebo-controlled trials only were considered (P < 0.05). No differences were observed in the insulin dose required or glycosylated hemoglobin values between nicotinamide and control patients. Adverse effects were reported in few patients (transient elevation of transaminase, n = 2; skin rash, n = 2; recurrent hypoglycemia, n = 2). CONCLUSIONS: This combined analysis demonstrates a therapeutic effect of nicotinamide in preserving residual beta-cell function when given at IDDM diagnosis in addition to insulin. Since adverse effects were negligible, we suggest that prolonged use of nicotinamide after IDDM diagnosis should be tested to see whether residual beta-cell function can be preserved for longer periods.

Adolescent↗
PubMed1996

Histochemical demonstration of glycogen in neurons of the cat retina.

PURPOSE: To demonstrate histochemically the cellular distribution of particulate glycogen in the cat retina and to correlate it with glucose sensitivity of neuronal electrical activities. METHODS: Free-floating, ultrathin sections of cat eyes (without glucose challenges) were stained by the periodic acid-thiocarbohydrazide-silver proteinate procedure and examined by electron microscopy. RESULTS: Muller cells were filled uniformly with fine-grain glycogen throughout all retinal layers. Particle density was higher in Muller cells of the peripheral retina than in those of central retina. Astrocytes contained little, if any, particulate glycogen. Alpha and beta ganglion cells had a heavy content of clumped glycogen granules. Rod bipolar and A17 amacrine cells of the rod pathway were stained intensely with particulate glycogen. No glycogen was seen in photoreceptor cells, cone bipolar cells, and the majority of amacrine cells, including AII cells of the rod pathway. However, one type of cone bipolar-driven amacrine cell was intensely glycogen positive. Its ultrastructural morphology, stratification pattern, and synaptology suggest that is a wide-field, axon-bearing type called A22. CONCLUSIONS: Except for the cone bipolar-driven ON-OFF A22 amacrine cell, it appears that glycogen staining preferentially labels neurons of the rod pathway. These observations are compatible with the reported sensitivity of the rod-driven electroretinogram and optic nerve response to glucose in the cat retina.

Animals↗
PubMed1995

The organization of the turtle inner retina. I. ON- and OFF-center pathways.

Intracellular recordings and dye injections of Lucifer yellow, horseradish peroxidase, or Neurobiotin were made in bipolar, amacrine, and ganglion cells of the Pseudemys turtle retina. By using a standard light-stimulation protocol in a sample of 375 labeled neurons, we were able to identify morphological and physiological characteristics of 11 types of bipolar cell, 37 types of amacrine cell, and 24 types of ganglion cell. To make sense of these data, we have chosen to group the 72 essentially different neuron types into traditional, functionally significant pathways. In this paper we look at the neuronal types in the inner plexiform layer (IPL) in terms of their contribution to generalized luminosity responses such as sustained ON- or OFF-center and transient ON-OFF ganglion cells; in the companion paper (J. Ammermüller, J.F. Muller, and H. Kolb, 1995, J. Comp. Neurol. 358:35-62) we look at them in terms of their involvement in color opponency and directional selectivity. A functional organization of the turtle IPL into OFF sublaminae (strata 1 and 2) and ON sublaminae (strata 3, 4, and 5), as has been described for other vertebrate retinas, was quite clear for two varieties of OFF-center bipolar cells (B4 and B5) and for all four types of sustained ON-center bipolar cell (B1, B2, B6, and B7). Thus, we found no sustained ON-center bipolar cell terminating in strata 1 and 2. We did, however, see three varieties of sustained OFF-center bipolar cells (B3, B9, and B10) having axon terminals in strata 3-5 (the ON sublamina) in addition to their terminations in stratum 1 or 2 (the OFF sublamina). Monostratified sustained ON- and OFF-center amacrine and ganglion cells rigidly obeyed the border of ON and OFF sublaminae. However, multistratified and diffuse sustained amacrine and ganglion cells could be either ON-center or OFF-center, and they did not strictly obey the border: such ON-center cells always had processes in one of the ON sublaminae (strata 3-5), and the equivalent OFF-center cells always had processes in one of the OFF sublaminae (strata 1 and 2). Monostratified transient amacrine and ganglion cells were concentrated in the middle of the IPL (around stratum 3), whereas bi-, tri-, or multistratified transient amacrine or ganglion cells always had processes in both the ON and the OFF sublaminae.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1995

The organization of the turtle inner retina. II. Analysis of color-coded and directionally selective cells.

Color coding and directional selectivity (DS) of retinal neurons were studied in the Pseudemys turtle by using similar intracellular recording and staining techniques as in the preceding paper (J. Ammermüller and H. Kolb, 1995, J. Comp. Neuronal. 358:1-34). Color-coded responses were elicited by red (621 or 694 nm), green (525 or 514 nm), and blue (455 nm) light flashes. In addition to red/green and yellow/blue types of chromaticity horizontal cells, in our sample of 305 identified cells we found that 17% of bipolar cells, 6.5% of amacrine cells, and 18% of ganglion cells exhibit color-coded responses. DS responses were found in 37% of the tested ganglion cells and 41% of the tested amacrine cells. Two morphologically identified bipolar cell types, B10 and B11, were red-ON/blue-OFF and red-OFF/green, blue-ON, respectively. Of five identified amacrine cell types, three were red-OFF/blue-ON center (A1, A3, A23b), one was red-OFF/green-ON center (A32), and one (A33) was double color-opponent of red-ON/blue-OFF center:red-OFF/blue-ON surround. Five ganglion cell types had variously color-coded centers (G14 and G24) or surrounds (G3 and G18), including one type, G6, that was double color-opponent (red-OFF/green-ON center:red-ON/green-OFF surround). Responses to colors were found primarily in sustained responses of bipolar and ganglion cells. However, in amacrine cells, transient components of the response also showed color dependence. Red-OFF-center responses were found in ganglion cells that were in a position to make connections at the strata 2/3 border with the red-OFF bipolar cell (B11); red-ON-center responses occurred in ganglion cells with branches in stratum 4 of the IPL where the red-ON-center bipolar (B10) ended. Blue-ON-center signals appeared to be processed mainly in strata 1-2/3, and blue-OFF-center signals in strata 3-5 of the IPL, with contributions of amacrine cells and bipolar cells. Labeled DS amacrine cells could be identified as A9, A20, and A22, and ganglion cells as G19, G20, and G24. The latter type (G24) showed DS and color coding. All response types (ON-center, OFF-center, ON-OFF) were encountered. DS amacrine cells were monostratified near the middle of the IPL, whereas DS ganglion cells were mono-, bi-, and multistratified, although all DS ganglion cells had one feature in common: they had dendrites in stratum 1 of the IPL.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1995

Substance P-immunoreactive neurons in the human retina.

Substance P (SP) is a neuropeptide that acts as a neurotransmitter or a neuromodulator in the retina. The aim of this study was to identify the type(s) and the distribution of the SP-immunoreactive (SP-IR) cells in the human retina. We have used an antiserum to SP to immunostain neurons in postmortem human retinae. Immunostained retinae were processed with the avidin-biotin complex (ABC) to visualize the cells either whole mounted in glycerol or embedded in plastic. Some retinae were also sectioned at 20 microns in order to obtain radial views of stained cells. SP-IR amacrine cells stain intensely and appear to be of a single type in the human retina. They are large-field cells with large cell bodies (16 microns diameter) lying in normal or displaced positions on either side of the inner plexiform layer (IPL). Their sturdy, spiny, and appendage-bearing dendrites stratify in stratum 3 (S3) of the IPL, where many overlapping, fine dendrites intermingle to form a plexus of stained processes. Either cell bodies or primary dendrites emit an "axon-like" process that, typically, divides into two long, fine processes, which run in opposite directions for hundreds of micrometers in S5 and S3 before disappearing as distinct entities in the stained plexus in S3. Long, fine dendrites also pass from the dendritic plexus to run in S5 and down to the nerve fiber layer to end as large varicosities at blood vessel walls. In addition, fine processes are emitted from the dendritic plexus that runs in S1, and some pass up to the outer plexiform layer (OPL) to run therein for short distances. The SP-IR amacrine cell has many similarities to the thorny, type 2 amacrine cells described from Golgi studies. In addition to the SP-IR amacrine cells, a presumed ganglion cell type is faintly immunoreactive. Its 20-22 microns cell body gives rise to a radiate, sparsely branched, wide-spreading dendritic tree running in S3. Its dendrites and cell body become enveloped by the more intensely SP-IR processes and boutons from the SP-IR amacrine cell type. The SP-IR ganglion cell type most resembles G21 from a Golgi study.

Cell Count↗
PubMed1995

Suppression of nitric oxide toxicity in islet cells by alpha-tocopherol.

We show here that preincubation of pancreatic islet cells with alpha-tocopherol significantly improves their resistance to toxic doses of nitric oxide (NO). No protection was afforded by other antioxidants such as vitamin C or glutathione-monoethyl ester. The pathway of NO induced islet cell death involves DNA damage and excessive activation of poly(ADP-ribose)polymerase leading to irreversible depletion of intracellular NAD+. alpha-Tocopherol was found to interfere at early steps of this pathway, by preventing the occurrence of DNA strand breaks. This indicates that alpha-tocopherol directly interacts with NO or its reactive intermediates. We conclude that alpha-tocopherol is not only part of the cellular defence system against oxygen radicals but also protects eukaryotic cells from NO toxicity.

Animals↗
PubMed1995

Inactivation of the poly(ADP-ribose) polymerase gene affects oxygen radical and nitric oxide toxicity in islet cells.

Activation of the nuclear enzyme poly(ADP-ribose) polymerase (PARP) is an early response of cells exposed to DNA-damaging compounds such as nitric oxide (NO) or reactive oxygen intermediates (ROI). Excessive poly-(ADP-ribose) formation by PARP has been assumed to deplete cellular NAD+ pools and to induce the death of several cell types, including the loss of insulin-producing islet cells in type I diabetes. In the present study we used cells from mice with a disrupted and thus inactivated PARP gene to provide direct evidence for a causal relationship between PARP activation, NAD+ depletion, and cell death. We found that mutant islet cells do not show NAD+ depletion after exposure to DNA-damaging radicals and are more resistant to the toxicity of both NO and ROI. These findings directly prove that PARP activation is responsible for most of the loss of NAD+ following such treatment. The ADP-ribosylation inhibitor 3-aminobenzamide partially protected islet cells with intact PARP gene but not mutant cells from lysis following either NO or ROI treatment. Hence the protective action of 3-aminobenzamide must be due to inhibition of PARP and does not result from its other pharmacological properties such as oxygen radical scavenging. Finally, the use of mutant cells an alternative pathway of cell death was discovered which does not require PARP activation and NAD+ depletion. In conclusion, the data prove the causal relationship of PARP activation and subsequent islet cell death and demonstrate the existence of an alternative pathway of cell death independent of PARP activation and NAD+ depletion.

Animals↗
PubMed1995

Insulitis and islet-cell antibody formation in rats with experimentally reduced beta-cell mass.

We studied the effect of severe reduction of beta-cell mass by 90% pancreatectomy on the immune tolerance to the endocrine pancreas. Four months after subtotal pancreatectomy all LEW.Han rats had developed mononuclear infiltration of islets and 9 of 14 rats were positive for islet-cell antibodies. Electron microscopy revealed lymphocytic invasion of endocrine tissue, lysis of beta cells and phagocytotic macrophages. None of these changes were seen 2 weeks after 90% pancreatectomy or 4 months after 10% pancreatectomy. Weekly substitution of islet antigens in the form of a homogenate of 100 islets into 90% pancreatectomized LEW.Han rats almost completely prevented the development of insulitis and autoantibodies. The dependence of insulitis on T cells was shown when 90% pancreatectomy in LEW.rnu rats (i.e., the congenic athymic nude strain), did not result in islet infiltration. The exocrine tissue remained normal in all experimental groups. During the observation period insulitis was not associated with overt diabetes but was accompanied by substantial enlargement of islets and of beta-cell mass, as shown by morphometry. Suppression of islet inflammation by injection of islet antigens abolished beta-cell regeneration, despite continuing metabolic stress in rats with 90% pancreatectomy. The findings indicate induction of islet autoimmunity in response to 90% but not to 10% pancreatectomy. We conclude that severe reduction of the islet-antigen mass allows the development of T-cell-dependent islet autoimmunity which indicates a loss of immune tolerance. In addition, the data suggest the existence of islet-antigen autoreactive immune cells in rats not genetically predisposed to autoimmune diabetes. Finally, we conclude that selective beta-cell regeneration occurs in association with insulitis.

Animals↗
PubMed1995

Levels of the terminal complement complex, C3a-desArg and C1-inhibitor in adult patients with capillary leak syndrome following bone marrow transplantation.

Capillary leak syndrome (CLS) is a severe complication after bone marrow transplantation (BMT). To investigate whether there is a pathogenetic role of the complement system, we monitored the levels of the terminal complement complex C5b-9 (TCC) and C3a-desArg as indicators of an activation of the complement system and the inhibitor of the classical pathway of the complement cascade, C1 inhibitor (C1-INH), in 48 bone marrow transplant recipients from 1 week before to 5 weeks after transplantation. Capillary leak syndrome developed in 7 out of 48 patients between days 1 and 12 after BMT. Complement activation as indicated by TCC levels was more pronounced in patients with CLS (n = 7) from day -8 to +28 (p < 0.05; day -1) and the elevation of TCC levels lasted longer in CLS patients (peak day 21) than in patients without this complication (peak day 7). Mean C3a-desArg levels were highest in patients with CLS reaching a peak at day 7. During the early posttransplant period a significant elevation of C1-INH levels (p < 0.01 and p < 0.05 respectively) compared with baseline levels (day -8) was found in patients with and without CLS, which was more pronounced in those patients with CLS (p < 0.05). Although we could not observe an absolute C1-INH deficiency as compared to healthy individuals our data support the presence of a relative deficiency of the inhibitor which might explain the reported beneficial effects of C1-INH substitution in BMT related CLS.

Adult↗
PubMed1995

Three-dimensional reconstruction and surface rendering of the five different spectral types of cone pedicle in the turtle retina.

Pseudo-3-dimensional models and surface renderings of a small group of neighboring cone pedicles in the turtle retina, in which the 5 different spectral types are represented, have been made from scanned serial electron micrographs using a Macintosh personal computer, Autocad and Studio Pro software. The resultant computer generated images clarify the morphological differences between the spectral types and show how they are related to each other via telodendrial connections and different levels of ending in the neuropil of the outer plexiform layer (OPL). The double cone pedicles end highest in the neuropil and the single red and green pedicles lie slightly lower into the OPL with the single blue and ultraviolent (UV)-sensitive cone pedicles coming in from obliquely angled axons to assume positions between the others much more vitread in the OPL. Telodendria interconnect the double and single red and green cones but not the blue and UV-sensitive cones.

Animals↗
PubMed1995

Regeneration of beta-cells in response to islet inflammation.

Subtotal pancreatectomy (90%) in Lewis rats induces chronic islet inflammation and tissue damage in the remaining pancreas 4 months after surgery. Concomitantly, significant enlargement of the islets of Langerhans was observed (90% pancreatectomy: islet/pancreas area: 25.6 +/- 9.6 x 10(-3), beta-cell/pancreas area: 12.4 +/- 4.4 x 10(-3); n = 4; controls without pancreatectomy: islet/pancreas area: 5.5 +/- 1.7 x 10(-3), beta-cell/pancreas area: 4.6 +/- 1.5 x 10(-3); n = 4; p < 0.05, respectively). Islet growth is mainly due to an increase in beta-cell mass. Beta-cell regeneration was not caused by the surgical manipulations or by metabolic stress. The former was ruled out by performing 10% pancreatectomy which did not cause islet enlargement after 4 months (islet/pancreas area: 13.6 +/- 11.3 x 10(-3), beta-cell/pancreas area: 7.1 +/- 2.0 x 10(-3); n = 3). An influence of metabolic stress was excluded by continuous substitution of syngenic islet antigens, which inhibits insulitis. In the absence of islet inflammation, despite persistent metabolic stress, beta-cell regeneration did not occur (islet/pancreas area: 7.0 +/- 5.5 x 10(-3), beta-cell/pancreas area: 5.5 +/- 4.1 x 10(-3); n = 4). Continuous treatment of animals after 90% pancreatectomy by insulin implants (1.5 U/day) avoided insulitis and beta-cell growth (islet/pancreas area: 9.2 +/- 1.1 x 10(-3), beta-cell/pancreas area: 6.8 +/- 1.0 x 10(-3), n = 3): All enlarged islets observed 4 months after 90% pancreatectomy without further treatment were infiltrated. Thus, beta-cell growth appears to be a response to insulitis. The stimulus for beta-cell growth could result from tissue damage caused by infiltrating cells or from cytokines secreted by the infiltrating cells, or both.

Animals↗
PubMed1995

Heat shock induces resistance in rat pancreatic islet cells against nitric oxide, oxygen radicals and streptozotocin toxicity in vitro.

When cultures of pancreatic islet cells are exposed to the nitric oxide donor sodium nitroprusside, to enzymatically generated reactive oxygen intermediates or to streptozotocin cell lysis occurs after 4-12 h. We report here that a heat shock at 43 degrees C for 90 min reduces cell lysis from nitric oxide (0.45 mM sodium nitroprusside) by 70%, from reactive oxygen intermediates (12 mU xanthine oxidase and 0.05 mM hypoxanthine) by 80% and from streptozotocin (1.5 mM) by 90%. Heat shock induced resistance was observed immediately after termination of the 90 min culture at 43 degrees C and correlated with enhanced expression of hsp70. The occurrence of DNA strand breaks, a major early consequence of nitric oxide, reactive oxygen intermediates, or streptozotocin action, was not suppressed by heat shock treatment. However, the depletion of NAD+, the major cause of radical induced islet cell death, was suppressed after heat shock (P < 0.01). We conclude that pancreatic islet cells can rapidly activate defence mechanisms against nitric oxide, reactive oxygen intermediates and streptozotocin by culture at 43 degrees C. Islet cell survival is due to the prevention of lethal NAD+ depletion during DNA repair, probably by slowing down poly(ADP-ribose)polymerase activation.

Adenosine Diphosphate Ribose↗
PubMed1995

Substance P: a neurotransmitter of amacrine and ganglion cells in the vertebrate retina.

A short history and summary of the occurrence of substance P in the vertebrate body is presented. Substance P is now generally accepted to be a neurotransmitter and can be visualized by immunocytochemistry to occur in various nerve cells in the CNS. In the retina, substance P-immunoreactivity (SP-IR) occurs in amacrine cell populations in all the species so far studied. In some vertebrates retinas SP is also apparent in one or more ganglion cell types. Anatomical investigations have revealed the morphology and connectivity of SP-IR amacrine cells: they branch in several strata of the inner plexiform layer receiving input from bipolar and amacrine cells and making synapses upon bipolar and ganglion cells. Most commonly SP-IR amacrines emit axon-like process that pass to both the outer plexiform layer and the ganglion cell and nerve fiber layers. These processes often end upon the retinal vasculature. SP-IR ganglion cells have been described in turtle, rabbit and human retinas. In turtle, intracellular dye injection has revealed the morphology of one type of SP-IR ganglion cell as being a large-field monostratified cell with a branches in the outer stratum of the inner plexiform layer. It may correspond to a "Dogiel cell" type. Intracellular investigation of SP-IR amacrine cells in turtle reveal their physiological responses to be ON-OFF in nature with some color-coding characteristics. In general SP acts as an excitatory neurotransmitter raising the spontaneous activity level of ganglion cell responses. The SP-IR ganglion cell is an OFF-center unit in the turtle retina and may be driven in the center of its receptive field by luminosity bipolar cells and in its surround by amacrine cells with color-opponent properties.

Animals↗
PubMed1995

Skeletal growth of transgenic mice with elevated levels of circulating insulin-like growth factor-II.

Insulin-like growth factor-II (IGF-II) is a major factor produced by skeletal tissues. To evaluate endocrine effects of IGF-II on bone growth, we measured skeletal dimensions of 12-week-old transgenic mice harbouring fusion genes where a human IGF-II cDNA is transcriptionally controlled by rat phospheonolpyruvate carboxykinase (PEPCK) promoter sequences. Transgene expression in liver, kidney and intestine resulted in circulating IGF-II levels in transgenic mice which were 2-3-fold higher than in controls. Serum IGF-I concentrations of transgenic mice were lower than in controls. Body weight was not influenced by the expression of the IGF-II transgene. Only 1 out of 5 measurements taken from the radius was significantly affected by the presence of the transgene, while in 60 measurements taken from eight other bones there was no difference between transgenic mice and controls. Furthermore, serum levels of calcium and phosphate as well as alkaline phosphatase activity were not significantly altered in PEPCK-IGF-II transgenic mice. Our findings demonstrate that moderately increased levels of circulating IGF-II do not cause major changes in skeletal growth and turnover in mice. This may be due to a lack of activity of circulating IGF-II on bone growth or to physiological consequences of elevated IGF-II, like a reduction of circulating IGF-I or an increase in IGF binding proteins.

Animals↗