Biomedical subjects

H Lindner

Publications and source records attributed to H Lindner.

At least 19 recordsLinked to original sources

PubMed2005

Irreversible pan-ErbB tyrosine kinase inhibitor CI-1033 induces caspase-independent apoptosis in colorectal cancer DiFi cell line.

The epidermal growth factor receptor (EGFR) is overexpressed in the majority of colorectal carcinomas and represents a target for therapeutic interventions with signal transduction inhibitors. We investigated the ability of CI-1033 to induce apoptosis and inhibition of proliferation in the colorectal cancer cell lines DiFi and Caco-2, which both express high levels of EGFR. While in Caco-2 cells CI-1033 treatment at a concentration 0.1 microM for 72 hours demonstrated only antiproliferative (53.7 +/- 4.3%) but no pro-apoptotic effects, treatment of DiFi cells resulted in a reduced proliferation rate (31.4 +/- 3.1%) and in apoptosis (44.2 +/- 8.9%). In order to define proteins involved in the regulation of apoptosis, we aimed to determine differences in the proteome profile of both cell lines before and after treatment with CI-1033. Cellular proteins were analyzed by 2-D gel electrophoresis followed by computational image analysis and mass spectrometry. Our data show that DiFi cells differ from Caco-2 cells in nine significantly upregulated proteins, and their potential role in apoptosis is described. We demonstrate that induction of apoptosis was triggered via caspase-independent pathways. Overexpression of leukocyte elastase inhibitor (LEI) and translocation of cathepsin D to the cytosol accompanied by upregulation of other defined proteins resulted in Bax-independent AIF translocation from mitochondria into the nucleus and apoptosis. Definition of these proteins can pave the way for functional studies and contribute to a better understanding of the effects of CI-1033 and the pathways of caspase-independent cell death.

Amino Acid Chloromethyl Ketones↗

PubMed2004

[Investigation of color vision using a web-based color vision test].

BACKGROUND: Screening tests of visual functions using the Internet are theoretically possible. To use these tests as a screening test, they must deliver comparable results with conventional test procedures. A web-based color vision test was developed based on pseudoisochromatic color plates. METHOD: The web-based color vision test was developed according to the pseudoisochromatic color plates by Velhagen and Broschmann using the programming-languages HTML, Java, and Perl. Sixty-five voluntary subjects, including nine color-deficient subjects, were examined by luminescence color plates (via web-based color vision test) and pigment color plates (via book). The statistical analysis was performed by determining the correspondence and the 95%-confidence interval. RESULTS: The correspondence of the test results for all subjects was 0.98 and the 95%-confidence interval was within 0.91 and 0.99. The correspondence of the test results in the group of color-deficient subjects was 1.0 and because of the limited number the 95%-confidence interval was within 0.71 and 1.0. CONCLUSIONS: The web-based color vision test with luminescence color plates for color-efficient and color-deficient subjects delivers test results comparable to pigment color plates under standardized examination conditions. Further studies are needed to examine if the web-based color vision test can also be used as an Internet screening test.

Adult↗

PubMed2004

Visual function tests on the Internet--sense or nonsense?

BACKGROUND: The quantitative capability of the visual system can be tested using graphic presentations with defined size, form and color. For presentations, a chart projector or monitor can be used. Today, the number of visual function tests on the Internet is increasing constantly. Options and limitations of visual function tests using the METHODS: Internet and the authors' own test results are described. RESULTS: Several visual function tests, such as visual acuity tests, the Amsler-Grid, stereo and color vision tests, can already be given via Internet. The variability of the tests ranges from the simple presentation of graphic elements to the laboriously programmed interactive input by the user to specify the test result. Under standardized examination conditions, there was a very high correspondence between the results of the authors' own web-based color vision test and those of luminescence color test plates and conventional pigment color plates. CONCLUSIONS: However, the interpretation of the test results is difficult due to the absence of controls during the test as well as the heterogeneity of the hardware. In order to obtain comparable test results, differences in size and resolution as well as in brightness, contrast and color of computer monitors must be taken into consideration. Due to the deficits described in the tests, the value of visual function tests on the Internet is rather limited. Currently, the data of test distributers with respect to the test conditions are all still insufficient. Standards need to be defined for Internet-based visual function tests. However, visual function tests on the Internet can achieve test results comparable to those of conventional visual function tests under standardized examination conditions in clinical practice. Further studies are needed to check the accuracy of web-based screening examinations in ophthalmology.

Color Perception Tests↗

PubMed2003

[Influence of long term occupational exposure to solvents on colour vision].

BACKGROUND: The study was designed to determine the influence of chronic occupational exposition of organic solvent mixtures on colour vision of car painters. SUBJECTS AND METHODS: The 123 subjects (2 groups differing in organic solvents exposure and 1 control group) were examined using Ishihara-Panel, Lanthony Desaturated Panel D-15, Velhagen-Panel, and Tritan-Album. RESULTS: In the Velhagen-Panel 3% of the probands currently exposed to organic solvents, and 11% of formerly exposed probands developed a blue/yellow vision defect for the right eye. All control subjects perfectly finished this panel. In the Tritan-Album 3% of currently exposed subjects and 26% of formerly exposed painters expressed a blue/yellow vision defect for the right eye, but also 7% of controls showed anomalies. Similar results were found for both panels with the left eye. The CCI difference in the D-15 test was significant between all three groups. CONCLUSION: The impaired colour vision may also be an important indicator of neuro-ophthalmological effects after long-term occupational exposure to organic solvents.

Adult↗

PubMed2002

Polymer erosion in PLGA microparticles produced by phase separation method.

This article deals with polymer erosion in biodegradable microparticles produced using the phase separation method. Poly(lactic-co-glycolic acid) copolymers with different compositions and molecular weights were employed. The microparticles were stored in phosphate buffer for 6 months. The molecular weight of the polymers was determined by size exclusion chromatography, and the weight loss was monitored gravimetrically. No weight loss was measured in the first weeks, although the molecular weight decreased significantly already from the start. After a certain storage period which was found to be specific for the type of polymer, the weight of the microparticles decreased rapidly. The start of this weight loss occurred when the molecular weight of the polymer in the degrading microparticles reached a threshold of approximately 15,000. This critical molecular weight was found to be identical for all investigated polymers, i.e. it was independent of the initial molecular weight of the polymer and of the lactic-glycolic ratio.

Chromatography, Gel↗

PubMed2002

[Options and limits of visual function tests available on the internet].

BACKGROUND: The quantitative capability of the visual system can be tested using graphic presentations with defined size, form and colour. For presentations a chart projector or personal computer (PC) can be used. In the meantime the number of visual function tests on the internet is continually increasing. These tests are potentially available for over 400 million internet users with different hard- and software requirements. METHODS: From 38 on-line visual acuity tests and 19 on-line colour vision tests found by means of search engines on german language web pages, the offerer of the test, the type of chart used and the data for the test conditions, as well as hard- and software adjustment were registered. RESULTS. Various visual function tests, such as visual acuity tests, the Amsler grid, stereo and colour vision tests, can already be planned via internet. The variability of the tests ranges from a simple presentation of graphic elements to a laboriously programmed interactive input by the user to specify the test result. Most of the tests are presented by opticians and the optical industry. DISCUSSION: The analysed tests allow for a considerable variability of the test conditions, for example by missing data concerning the room lighting and lacking monitor calibration. Furthermore, problems of the web-based colour presentation were not considered or not considered sufficiently in most of the on-line colour tests. CONCLUSIONS: Because of the test deficits determined, the results of visual function tests on the internet are to be classified as doubtful. At present, most data of test offerers are not sufficient with respect to the test conditions. Standards should be defined for internet-based visual function tests and further studies should be followed to check the effectiveness of these tests for screening examinations.

Color Perception Tests↗

PubMed2002

Hyperphosphorylation of histone H2A.X and dephosphorylation of histone H1 subtypes in the course of apoptosis.

Chromatin condensation paralleled by DNA fragmentation is one of the most important nuclear events occurring during apoptosis. Histone modifications, and in particular phosphorylation, have been suggested to affect chromatin function and structure during both cell cycle and cell death. We report here that phosphate incorporation into all H1 subtypes decreased rapidly after induction of apoptosis, evidently causing a strong reduction in phosphorylated forms of main H1 histone subtypes. H1 dephosphorylation is accompanied by chromatin condensation preceding the onset of typical chromatin oligonucleosomal fragmentation, whereas H2A.X hyperphosphorylation is strongly correlated to apoptotic chromatin fragmentation. Using various kinase inhibitors we were able to exclude some of the possible kinases which can be involved directly or indirectly in phosphorylation of histone H2A.X. Neither DNA-dependent protein kinase, protein kinase A, protein kinase G, nor the kinases driven by the mitogen-activated protein kinase (MAP) pathway appear to be responsible for H2A.X phosphorylation. The protein kinase C activator phorbol 12-myristate 13-acetate (PMA), however, markedly reduced the induction of apoptosis in TNFalpha-treated cells with a simultaneous change in the phosphorylation pattern of histone H2A.X. Hyperphosphorylation of H2A.X in apoptotic cells depends indirectly on activation of caspases and nuclear scaffold proteases as shown in zVAD-(OMe)-fmk- or zAPF-cmk-treated cells, whereas the dephosphorylation of H1 subtypes seems to be influenced solely by caspase inhibitors. Together, these results illustrate that H1 dephosphorylation and H2A.X hyperphosphorylation are necessary steps on the apoptotic pathway.

3T3 Cells↗

PubMed2001

Bladder cancer cells acquire competent mechanisms to escape Fas-mediated apoptosis and immune surveillance in the course of malignant transformation.

Mechanisms of resistance against Fas-mediated cell killing have been reported in different malignancies. However, the biological response of immune escape mechanisms might depend on malignant transformation of cancer cells. In this study we investigated different mechanisms of immune escape in 2 well-differentiated low-grade (RT4 and RT112) and 2 poorly differentiated high-grade (T24 and TCCSUP) bladder cancer cell lines. Fas, the receptor of Fas-ligand, is expressed and shedded by human transitional bladder carcinoma cell lines RT4, RT112, T24 and TCCSUP. Cytotoxicity and apoptosis assays demonstrate that in spite of the Fas expression, poorly differentiated T24 and TCCSUP cells are insensitive towards either recombinant Fas-ligand or agonistic apoptosis-inducing monoclonal antibody against Fas. In poorly differentiated T24 and TCCSUP cell lines we were able to detect marked Fas-ligand protein by flow cytometry and Western blot analysis. In grade 1 RT4 and RT112 cells only minor expression of Fas-ligand possibly because of proteinase action. Fas-ligand mRNA translation or post-translational processing seems to be regulated differentially in the cancer cell lines depending on malignant transformation. In co-culture experiments we show that poorly differentiated cells can induce apoptosis and cell death in Jurkat cells and activated peripheral blood mononuclear cells. This in vitro study suggests that bladder cancer cells can take advantage of different mechanisms of immune evasion and become more competent in avoiding immune surveillance during transformation to higher-grade malignant disease.

Antibodies, Monoclonal↗

PubMed2001

Age-dependent deamidation of asparagine residues in proteins.

Nonenzymatic deamidation of peptides and proteins represents an important degradation reaction occurring in vitro in the course of isolation or storage and in vivo during development and/or aging of cells. This review first presents a synopsis of the influence of structure on deamidation reaction proceeding via a five-membered succinimide intermediate, followed by an outline of procedures for separation and detection of deamidated forms. Selected examples for in vitro and in vivo deamidation are reviewed including the possible biological consequences of this protein degradation. Finally, the reaction of protein methyltransferase with L-isoaspartyl- and D-aspartyl residues and its possible role in protein repair is elucidated.

Aging↗

PubMed2001

Room temperature activates human blood platelets.

Temperatures ranging from room temperature (20 degrees C) to 42 degrees C are generally not considered to have an activating effect on platelets. However, this assumption is not supported by clinical phenomena that result in hemostatic failure related to hypothermia. In this study, we investigated the effect of temperatures between room temperature (20 degrees C) and 42 degrees C on human blood platelets and found that room temperature causes marked activation of platelets. Major changes in platelet morphology were seen at 20 degrees C compared to resting platelets at 37 degrees C. Platelet morphology was investigated with noninvasive live cell techniques (light microscopy and dynamic and static light scattering), as well as with transmission and scanning electron microscopy. The changes in platelet morphology correlated with the expression of the activation marker, activated glycoprotein (GP) IIb-IIIa, measured by flow cytometry. Twenty-five percent to 30% of platelets expressed activated GPIIb-IIIa after exposure to 20 degrees C for 10 minutes. In the presence of serotonin re-uptake inhibitors, the serotonin content of platelets at 20 degrees C was twice that of resting platelets. In comparison, moderate heat shock conditions (42 degrees C for 10 minutes) caused no signs of platelet activation as indicated by the absence of morphological alterations, no expression of activated GPIIb-IIIa, and no changes in serotonin content. These results show that room temperature by itself significantly activates platelets and has an effect on the platelet serotonin content. This may contribute to both the functional lesion associated with 22 degrees C storage of platelets for transfusion and the in vivo hemostatic failure after hypothermia.

Blood Platelets↗

PubMed2001

[Illumination conditions of visually impaired people under private domestic circumstances - clinical study on 91 patients].

BACKGROUND: The illuminance and quality of illumination are important factors for visually impaired people to improve contrast sensitivity and reduce glare sensitivity at domestic reading places. The ability of visually impaired people to read can often be improved by simple strategies. At the onset of multiple ophthalmic diseases an optimum of lighting is often the tool of choice to restore reading capability. PATIENTS AND METHODS: 91 visually impaired people were included in this study. 76 were equipped with optical reading tools and 15 used a television reading aid. The illumination conditions of reading places were investigated. This included measurements of illuminance and information about the light source and localization, direction of light and modelling. The understanding of the importance of optimal illumination was judged by a graded scale from 1 - 5. RESULTS: Twenty-two of the 76 visually impaired people equipped with optimal reading tools were using an extra reading place. All others used livingroom or kitchen tables as their preferred reading place. 10 % of the reading places had a illuminance of above 5000 lx, whereas 63 % had less than 500 lx. Incandescent lamps dominated as artificial light sources. In 60 % of all reading places ceiling light sources were predominant. Only 40 % of the patients used an additional reading place illumination. For television reading aids the ceiling light caused a decrease of contrast sensitivity in 1 % - 7 %. If questioned, 90 % of the patients were highly sensitive to problems of good illumination of their reading places. This is, as our study revealed, in contrast to the conditions under which most of the visually impaired people read. CONCLUSIONS: Domestic reading places revealed considerable deficits concerning the illumination. Most visually impaired people do not realize this problem. Ophthalmologists, opticians, low-vision-, orientation- and mobility-trainers are recommended for rehabilitation.

Aged↗

PubMed2000

Reverse signaling through transmembrane TNF confers resistance to lipopolysaccharide in human monocytes and macrophages.

We have previously reported that the CD14+ monocytic subpopulation of human PBMC induces programmed cell death (apoptosis) in cocultured endothelial cells (EC) when stimulated by bacterial endotoxin (LPS). Apoptosis is mediated by two routes, first via transmembrane TNF-alpha (mTNF) expressed on PBMC and, in addition, by TNF-independent soluble factors that trigger apoptosis in EC. Neutralizing anti-TNF mAb completely blocked coculture-mediated apoptosis, despite the fact that there should have been additional soluble cell death factors. This led to the hypothesis that a reverse signal is transmitted from the TNF receptor on EC to monocytes (MO) via mTNF that prevents the production of soluble apoptotic factors. Here we have tested this hypothesis. The results support the idea of a bidirectional cross-talk between MO and EC. Peripheral blood MO, MO-derived macrophages (MPhi), or the monocytic cell line Mono Mac 6 were preincubated with human microvascular EC that constitutively express TNF receptor type I (TNF-R1) and subsequently stimulated with LPS. Cell-free supernatants of these preparations no longer induced EC apoptosis. The preincubation of MO/MPhi with TNF-reactive agents, such as mAb and soluble receptors, also blocked the production of death factors, providing further evidence for reverse signaling via mTNF. Finally, we show that reverse signaling through mTNF mediated LPS resistance in MO/MPhi as indicated by the down-regulation of LPS-induced soluble TNF and IL-6 as well as IL-1 and IL-10.

Antibodies, Monoclonal↗

PubMed2000

Lipoprotein-associated alpha-tocopheryl-succinate inhibits cell growth and induces apoptosis in human MCF-7 and HBL-100 breast cancer cells.

alpha-Tocopheryl succinate (alpha-TS) is a potent inhibitor of tumor cell proliferation. The goal of the present study was to investigate whether and to what extent alpha-TS associates with plasma lipoproteins and if alpha-TS-enriched lipoproteins inhibit breast cancer cell growth in a manner comparable to the free drug. In vitro enrichment of human plasma revealed that alpha-TS readily associated with the main lipoprotein classes, findings confirmed in vivo in mice. At the highest alpha-TS concentrations, lipoproteins carrying 50000 (VLDL), 5000 (LDL) and 700 (HDL) alpha-TS molecules per lipoprotein particle were generated. alpha-TS enrichment generated lipoprotein particles with slightly decreased density and increased particle radius. To study whether the level of LDL-receptor (LDL-R) expression affects alpha-TS uptake from apoB/E containing lipoprotein particles human breast cancer cells with low (MCF-7) and normal (HBL-100) LDL-R expression were used. The uptake of free, VLDL- and (apoE-free) HDL(3)-associated alpha-TS was nearly identical for both cell lines. In contrast, uptake of LDL-associated alpha-TS by HBL-100 cells (normal LDL-R expression) was about twice as high as compared to MCF-7 cells (low LDL-R expression). VLDL and LDL-associated alpha-TS inhibited proliferation most effectively at the highest concentration of alpha-TS used (100% inhibition of MCF-7 growth with 20 microg/ml of lipoprotein-associated alpha-TS). However, also alpha-TS-free VLDL and LDL inhibited HBL-100 cell proliferation up to 55%. In both cell lines, alpha-TS-enriched HDL(3) inhibited cell growth by 40-60%. Incubation of both cell lines in the presence of free or lipoprotein-associated alpha-TS resulted in DNA fragmentation indicative of apoptosis. Collectively, the present findings demonstrate that: (1) alpha-TS readily associates with lipoproteins in vitro and in vivo; (2) the lipoprotein-enrichment efficacy was dependent on the particle size and/or the triglyceride content of the lipoprotein; (3) uptake of LDL-associated alpha-TS was apparently dependent on the level of LDL-R expression; and (4) lipoproteins were efficient alpha-TS carriers inducing reduced cell proliferation rates and apoptosis in human breast cancer cells as observed for the free drug.

Animals↗

PubMed2000

The distribution of aminoacylase I among mammalian species and localization of the enzyme in porcine kidney.

Aminoacylase I (Acy-1, EC 3.5.1.14) is found in many mammalian tissues, with highest activities occurring in kidney. The enzyme hydrolyzes a variety of N-acylated amino acids; however, the physiological role and the exact cellular localization of Acy-1 are still a matter of debate. The comparison of Acy-1 activities in kidney and liver homogenates of 11 mammalian species showed that the enzyme is most abundant in true herbivores such as sheep and cattle as well as in omnivores, while activities were very low in both rodents and the cat. Acy-1 activity was not detected in livers of dogs of five different breeds. Using in situ hybridization of porcine kidney sections with DIG-labeled RNA probes, Acy-1 mRNA was shown to be evenly distributed throughout the tubular system, while glomeruli and the interstitium were free of stain. During subcellular fractionation, porcine Acy-1 behaved like a typical cytosolic enzyme. Commonly, Acy-1 is thought to catalyze hydrolytic reactions, i.e., the formation of free amino acids from acylated derivatives. Based on the present results and literature data, we propose a novel hypothesis, i.e., that Acy-1 catalyzes the synthesis (rather than the hydrolysis) of hippurate that is formed as a detoxification product of aromatic compounds.

Amidohydrolases↗

PubMed2000

Naive monocytes can trigger transendothelial migration of peripheral blood cells through the induction of endothelial tumour necrosis factor-alpha.

In this manuscript we describe a potentially new mechanism by which unstimulated human monocytes activate endothelial cells (EC) through the secondary induction of endothelial tumour necrosis factor alpha (TNF-alpha). Serum free supernatants (SN) of peripheral blood mononuclear cells (PBMC) strongly induce the expression of intercellular adhesion molecule 1 (ICAM-1, CD54), vascular cell adhesion molecule 1 (VCAM-1, CD106), and endothelial-leukocyte adhesion molecule 1 (ELAM-1, CD62E) on human EC 24 and 4 h post treatment, respectively. Further characterization of the responsible subpopulation revealed the CD14+ monocytes and a monocytic cell line (MM6) to produce an endothelial activating factor (EAF). The EAF also triggers an adhesion and a transendothelial migration (TEM) of peripheral blood cells. Using neutralization with an anti TNF-alpha MoAb MAK195, EAF is not identical with TNF-alpha, but induces the expression of endothelial TNF-alpha, since MAK195 blocked TEM only when coincubated with EC, not with monocytes. Furthermore, intracellular TNF-alpha was significantly upregulated in EC after treatment with SN-MM6. Another evidence for a secondary autocrine mechanism was provided by culturing the EC with a conditioned medium of SN-MM6 treated EC. This conditioned medium induces an adhesion molecule expression and TEM in a similar way to SN-MM6 and can completely be inactivated by anti TNF-alpha. Taken together, these data may have an impact for, e.g. transplantational settings that donor monocytes may trigger an inflammatory response in the absence of further activation signals by eliciting an endogenous TNF-alpha response in the host.

Cell Adhesion↗

PubMed2000

Mutational analysis of two PWW sequence motifs in human aminoacylase 1.

Human and porcine aminoacylase 1 (Acy1) contain two peculiar sequence motifs located near the interface between the two major domains of the Acy1 subunit. Each motif consists of the sequence PWW preceded by four strongly polar residues. In order to examine the significance of these sequences for Acy1 stability and/or catalysis, we used site-directed mutagenesis of human Acy1 to replace the tryptophan residues in either motif with alanines. Both mutants showed unchanged zinc binding and normal substrate affinity. Modification in PWW motif 1 (residues 192 -194) and motif 2 (residues 321 - 323) resulted in markedly reduced specific activity in the first case and diminished stability in either mutant. Fluorescence quenching measurements showed that all four tryptophans of the PWW motifs are solvent-accessible. We conclude that PWW motif 1 maintains the native conformation of the active site by creating the proper spatial relationship between dimerization domains and catalytic domains, while motif PWW2 is necessary for the stability of the catalytic domain.

Amidohydrolases↗

PubMed1999

The essential transfer protein TraM binds to DNA as a tetramer.

The TraM proteins encoded by F-like plasmids are sequence specific DNA binding proteins that are essential for conjugative DNA transfer. We investigated the quarternary structure and the DNA binding properties of the TraM wild-type protein of the resistance plasmid R1 and two mutant forms thereof. Size-exclusion chromatography and differential scanning calorimetry showed that purified TraM protein (amino acids 2-127) forms stable tetramers in solution. A truncated version of the protein termed TraMM26 (amino acids 2-56) forms dimers. Thus, the dimerization and tetramerization domains can be assigned to the N-terminal and C-terminal domains of TraM, respectively. Further analyses using chemical cross-linking and light scattering corroborated the preferentially tetrameric nature of the protein but also suggest that TraM has a tendency to form higher aggregates. Band-shift and fluorescence spectroscopy investigations of TraM-DNA complexes revealed that the TraM protein is also tetrameric when bound to its minimal DNA binding site. The deduced binding constant in the range of 10(8) M(-1) demonstrated a very strong binding of TraM to its preferred DNA sequence. Secondary structure analysis based on CD measurements showed that TraM is mainly alpha-helical with a significant increase in alpha-helicity (48 to 58%) upon DNA-binding, indicating an induced fit mechanism.

Bacterial Proteins↗

PubMed1999

The N-terminally acetylated form of mammalian histone H1(o), but not that of avian histone H5, increases with age.

We report here on the HPCE separation of two chicken H5 histones, which do not show the heterogeneity (Gln/Arg) at residue 15 first found by Greenaway and Murray [Greenaway and Murray (1971) Nat. New Biol. 229, 233-238]. The two subfractions obtained were identified using reversed-phase HPLC, hydrophilic interaction HPLC, Edman degradation, and MALDI-MS analysis. We found that the two H5 subcomponents differ only by an acetylated (designated H5a) and an unacetylated N-terminus (H5b). In contrast to the N-terminally acetylated form of rat kidney histone H1(o), which increased by about 40% with aging of the animal, the corresponding form of chicken H5 did not: the ratio N-terminally acetylated: unacetylated remained constant (30:70) when histone H5 was extracted from erythrocytes of newly hatched chickens and from adult chickens, respectively. The HPCE technique used in this investigation represents a quick and convenient method for analyzing N-terminally acetylated proteins in the presence of unacetylated forms.

Acetylation↗