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Biomedical subjects

M Weller

Publications and source records attributed to M Weller.

At least 199 records · Page 11Linked to original sources
PubMed1997

Adhesion of neutrophils to epidermal cells: prerequisites for and blocking by anti-CD11b antibodies.

Immigration of neutrophils into the epidermis is a hallmark of several so-called neutrophilic dermatoses. In this study we used a frozen section adhesion assay to study the impact of different activation procedures as well as of CD11 integrins on cell-cell adhesion between neutrophils and epidermal cells. Serial sections from punch biopsies of healthy donors were incubated with freshly prepared neutrophils for 40 min and adherent cells were counted after washing. We found that preactivation of the biopsies with interferon-gamma in vitro is a prerequisite for neutrophil binding. Activation of neutrophils with N-formyl-methionyl-leucyl-phenylalanine (fMLP), tumor necrosis factor-alpha (TNF-alpha) or platelet activating factor (PAF) then leads to a three- to sixfold increase in neutrophil adhesion. This enhanced binding could be blocked by anti-CD11b monoclonal antibodies, whereas anti-CD11a and anti-CD11c antibodies exerted no significant effect. At a temperature of 6 degrees C, 50% inhibition of neutrophil binding was observed. Complement factor C3 and fibrinogen, known ligands for CD11b/CD18, could not be detected on epidermal cells by immunohistochemistry. In summary, our study showed that enhanced binding of neutrophils to epidermal sections can be achieved by prestimulation of both the keratinocytes with interferon-gamma and the neutrophils with fMLP, TNF-alpha and PAF. This adhesion was furthermore revealed to be dependent on the function of the leucocyte integrin CD11b/CD18.

Antigen-Antibody Reactions↗
PubMed1997

Evidence for an active type of cell death with ultrastructural features distinct from apoptosis: the effects of 3-acetylpyridine neurotoxicity.

3-Acetylpyridine is a niacinamide antagonist with potent neurotoxic properties in vitro and in vivo. 3-Acetylpyridine neurotoxicity was associated with positive DNA end-labelling and displayed features of active cell death without the ultrastructural changes of apoptotic cell death. After systemic administration in rats (70 mg/kg), we detected labelled nuclei in the inferior olive using in situ DNA end-labelling. However, the conventional chromatin stain did not show chromatin condensation or fragmentation and electron microscopy studies failed to reveal features of apoptosis. Although areas of condensed chromatin were present in some nuclei, cytoplasmic damage with extensive organelle swelling was the most prominent finding. In vitro, 3-acetylpyridine (0.1-1 mM) induced degeneration of cerebellar granule neurons in a concentration- and time-dependent manner. The protein synthesis inhibitor cycloheximide (10 micrograms/ml) and the transcriptional inhibitor actinomycin D (10 microM) protected against 3-acetylpyridine toxicity. In contrast, neither the free radical scavenger alpha-phenyl-N-tertbutylnitron (100 microM), nor glutathione ethyl ester (10-100 microM), N-acetyl-cysteine (10-200 microM) or 3-aminobenzamide (0.1-4 mM), an inhibitor of poly(ADP-ribose) synthesis, were effective. 3-Acetylpyridine-induced neuronal death in vitro was associated with positive in situ DNA labelling. However, DNA fragmentation could not be demonstrated prior to neuronal cell loss and no DNA "laddering" was detected by DNA gel electrophoresis. Correspondingly, no apoptotic nuclei were revealed upon electron microscopy but organelle swelling and extensive vacuolization, changes similar to autophagocytosis. In conclusion, 3-acetylpyridine induces an active form of cell death that required de novo protein synthesis but is distinct from apoptosis. A loss of glutathione accompanies, but does not precede, cell death.

Animals↗
PubMed1997

A new look at the role of p53 in leukemia cell sensitivity to chemotherapy.

A gene encoding the p53 val135 mutant, which assumes mutant conformation at 38.5 degrees C and wild-type conformation at 32.5 degrees C, was introduced into p53-deficient K562 myeloid leukemia cells. Forced expression of wild-type, but not mutant, p53 resulted in growth arrest, accumulation of p21 and Bax proteins, and delayed cell death. Wild-type p53 enhanced the cytotoxic effects of some drugs and attenuated those of others. Compared with wild-type p53, mutant p53 induced much stronger sensitization to drug cytotoxicity. This occurred in the absence of effects on cell cycle progression or activation of several p53 target genes. Although both mutant and wild-type p53 induced changes of immunophenotype, no specific pattern of differentiation was associated with enhanced chemosensitivity. Thus, (1) induction of growth arrest and activation of p53 target genes such as p21 and bax are linked to the wild-type conformation of p53; (2) p53 induces immunophenotypic changes of myeloid leukemia cells suggestive of multidirectional differentiation in a conformation-dependent manner; and (3) (so-called) mutant p53 induces chemosensitization in the absence of effects on cell cycle progression, activation of bax, p21, gadd45 and mdm-2, or a specific pattern of differentiation; and (4) chemosensitization mediated by wild-type p53 may be masked by transcription-dependent induction of growth arrest.

Animals↗
PubMed1997

Cooperative interception of neuronal apoptosis by BCL-2 and BAG-1 expression: prevention of caspase activation and reduced production of reactive oxygen species.

Neuronally differentiated PC12 cells undergo synchronous apoptosis when deprived of nerve growth factor (NGF). Here we show that NGF withdrawal induces actinomycin D- and cycloheximide-sensitive caspase (ICE-like) activity. The peptide inhibitor of caspase activity, N-acetyl-Asp-Glu-Val-Asp-aldehyde, was more potent than acetyl-Tyr-Val-Ala-Asp-chloromethyl ketone in preventing NGF withdrawal-induced apoptosis, suggesting an important role for caspase-3 (CPP32)-like proteases. We observed a peak of reactive oxygen species (ROS) 6 h after NGF withdrawal. ROS appear to be required for apoptosis, because cell death is prevented by the free radical spin trap, N-tert-butyl-alpha-phenylnitrone, and the antioxidant, N-acetylcysteine. ROS production was blocked by actinomycin D, cycloheximide, and caspase protease inhibitors, suggesting that ROS generation is downstream of new mRNA and protein synthesis and activation of caspases. Forced expression of either BCL-2 or the BCL-2-binding protein BAG-1 blocked NGF withdrawal-induced apoptosis, activation of caspases, and ROS generation, showing that they function upstream of caspases. Coexpression of BCL-2 and BAG-1 was more protective than expression of either protein alone.

Animals↗
PubMed1997

Plasminogen in proliferative vitreoretinal disorders.

OBJECTIVE: Intravitreal fibrin formation is a frequent observation after vitrectomy performed for a variety of vitreoretinal disorders including proliferative vitreoretinopathy (PVR), proliferative diabetic retinopathy (PDR), and endophthalmitis. Plasminogen activators (PA) have been used for the management of this postoperative complication. This approach requires the presence of plasminogen, the substrate for PA mediated fibrinolysis, in the vitreal cavity. METHODS: Quantification of plasminogen in the vitreous of 60 patients with PVR, PDR, and macular pucker was performed by streptokinase mediated activation using a chromogenic substrate. The presence of immunoreactive plasminogen was confirmed by immunoblot analysis of vitreal proteins and immunocytochemistry of surgically removed epiretinal membranes. RESULTS: Plasminogen levels were dramatically increased in the vitreous of PVR and PDR patients compared with macular pucker patients and normal controls. Staining for plasminogen in epiretinal membranes was confined to the extracellular matrix. Predominant staining of perivascular areas in PDR specimens indicated that breakdown of the blood-retinal barrier is an important source of intravitreal plasminogen in that condition. CONCLUSION: Plasminogen may play a role in traction membrane formation in PVR and PDR. Our biochemical analysis of presurgical vitreous indicates that there may be abundant substrate for PA mediated fibrinolysis in the vitreous cavity after vitrectomy.

Eye Diseases↗
PubMed1997

Ocular myasthenia gravis: response to long-term immunosuppressive treatment.

OBJECTIVE: Ocular myasthenia gravis is a subtype of myasthenia gravis that causes relatively mild disability, but may convert into severe generalised muscle weakness. A universal management plan for ocular myasthenia gravis has not been established. This study was performed to determine the outcome of ocular myasthenia gravis with the currently available therapeutic options. METHODS: Retrospective analysis of 78 patients with ocular myasthenia gravis with a mean disease duration of 8.3 (range 0.5-58.3) years. RESULTS: In 54 patients (69%) symptoms and signs remained confined to the extraocular muscles during the observation period. The remaining 24 patients (31%) developed symptoms of generalised myasthenia gravis; 50% of them within two years, 75% within four years after onset. A somewhat reduced risk of generalisation was found in those with mild symptoms, normal repetitive nerve stimulation test, and low or absent antiacetylcholine receptor (AChR) antibodies at the time of diagnosis. Patients receiving immunosuppressive treatment (corticosteroids and/or azathioprine) rarely developed generalised myasthenia gravis (six of 50, 12%). Those without such treatment, usually due to uncertain diagnosis and late referral, converted into generalised myasthenia gravis significantly more often (18 of 28, 64%). CONCLUSIONS: The prognosis of ocular myasthenia gravis is good. A conventional scheme with short-term corticosteroids and long-term azathioprine seems adequate to achieve remission in most patients. The proportion of patients developing generalised myasthenia gravis was smaller in this population compared with previously published groups (usually 50%-70%). Early immunosuppressive treatment is at least partially responsible for this finding. Thymectomy (performed here in 12 patients with an abnormal chest CT) also correlated with a good outcome, but had no apparent advantage over medical treatment alone.

Adolescent↗
PubMed1997

Gluteal compartment syndrome due to rhabdomyolysis after heroin abuse.

We report a 30-year-old man who developed painful swelling of his right leg and complete sciatic nerve palsy after an i.v. injection of heroin. Excessive elevation of serum creatine phosphokinase indicated the presence of rhabdomyolysis. Fasciotomy of the gluteus maximus led to rapid and complete recovery from sciatic nerve palsy. Nontraumatic rhabdomyolysis may cause a gluteal compartment syndrome that requires immediate fasciotomy.

Adult↗
PubMed1997

Chemotherapy of human malignant glioma: prevention of efficacy by dexamethasone?

Steroids are commonly administered for the control of edema, mass effect, and side effects from therapy to patients with malignant glioma who are receiving radiotherapy and chemotherapy. Here, we report that therapeutic concentrations of dexamethasone (DEX) attenuate cytotoxicity and growth inhibition of human malignant glioma cells induced by exposure to several chemotherapeutics, including ACNU, VM-26, vincristine, cytarabine, methotrexate, and adriamycin. DEX-mediated cytoprotection is not linked to DEX effects on glioma cell proliferation. However, the cytoprotective effects of DEX appeared to be more prominent in cell lines with wild-type p53 status (n = 2) than in p53 mutant cell lines (n = 3). Further, DEX-mediated rescue from chemotherapy does not directly involve Bcl-2 family proteins since DEX failed to change the expression of Bcl-2 or Bax proteins and since bcl-2 gene transfer-mediated cytoprotection was not redundant with the effects of DEX. DEX thus appears to control a common, bcl-2-independent death pathway in glioma cells that is not limited to specific drug actions. Chemotherapy is usually given as an elective, adjuvant treatment to glioma patients in stable condition who can tolerate steroid withdrawal. To maximize therapeutic efficacy, steroids should be withdrawn from glioma patients prior to chemotherapy.

Antibiotics, Antineoplastic↗
PubMed1997

Hypericin-induced apoptosis of human malignant glioma cells is light-dependent, independent of bcl-2 expression, and does not require wild-type p53.

Hypericin and tamoxifen are experimental agents for the adjuvant chemotherapy of malignant glioma. We report that hypericin and tamoxifen induce apoptosis of 7 human malignant glioma cell lines in a concentration- and time-dependent manner. Illumination is essential for the cytotoxicity of hypericin but not tamoxifen. Apoptosis is unaffected by inhibitors of RNA and protein synthesis or free radical scavengers, does not require wild-type p53 activity, and occurs in glioma cells expressing high levels of bcl-2. There is no correlation between hypericin and tamoxifen-induced cytotoxicity and inhibition of protein kinase C (PKC). Ectopic expression of a murine bcl-2 transgene provides modest protection from tamoxifen but does not affect hypericin toxicity. Hypericin and tamoxifen do not modulate glioma cell killing induced by tumor necrosis factor-alpha (TNF-alpha) or CD95 ligand. Both drugs augment the acute cytotoxicity of various cancer chemotherapy drugs but fail to shift their EC50 values in modified colony formation assays. These data do not provide further supportive evidence how to enhance the limited efficacy of tamoxifen treatment for human malignant glioma. However, hypericin is a promising agent for the treatment of malignant glioma if local photodynamic activation of hypericin in the glioma tissue can be achieved.

Anthracenes↗
PubMed1997

Expression and upregulation of microtubule-associated protein 1B in cultured retinal pigment epithelial cells.

PURPOSE: During routine cell culture and under pathologic conditions, human retinal pigment epithelial (RPE) cells lose epithelial characteristics and change their morphology. In this study, changes in gene expression in RPE cells of different generations were evaluated by polymerase-chain-reaction-based differential display mRNA analysis (DD-RT-PCR). METHODS: Total RNA was prepared from freshly isolated and cultured human RPE cells of passages P0 and P3 and was subjected to DD-RT-PCR. One band with enhanced expression was excised, reamplified, and partially sequenced, using a modified dideoxy chain termination approach. Expression of the corresponding protein was ascertained by immunocytochemical analysis. RESULTS: Differential display RT-PCR showed enhanced expression of a specific RNA in P3 cells compared with that in P0 cells. Sequence alignment revealed 98% identity with the 3' end of the coding sequence of human microtubule-associated protein 1B (MAP1B). Confirmation of induced expression of MAP1B mRNA was obtained by PCR with specific primers and by immunocytochemical analysis in cultured RPE cells and in surgically removed epiretinal membranes from patients with proliferative vitreoretinopathy. No expression of MAP1B mRNA or protein was detected in freshly isolated RPE cells. CONCLUSIONS: Differential display RT-PCR in RPE cells with subsequent sequence analysis allows characterization of the maturation- and differentiation-dependent expression of previously undetected genes and gene products in cultured RPE cells.

Animals↗
PubMed1996

Potassium deprivation-induced apoptosis of cerebellar granule neurons: a sequential requirement for new mRNA and protein synthesis, ICE-like protease activity, and reactive oxygen species.

Potassium (K+) deprivation-induced apoptosis of cerebellar granule neurons requires new mRNA and protein synthesis. Using a fluorogenic substrate for interleukin-1beta converting enzyme (ICE), we show that K+ deprivation of cerebellar granule neurons induces cycloheximide-sensitive ICE-like protease activity. A peptide inhibitor of ICE-like protease activity, Ac-YVAD-chloromethylketone (Ac-YVAD-CMK), prevents K+ deprivation-induced apoptosis. Further, reactive oxygen species (ROS) are essential mediators of K+ deprivation-induced apoptosis of cerebellar granule neurons because neuronal death is also blocked by superoxide dismutase, N-acetyl-L-cysteine, and free radical spin traps. Using fluorescent assays, we show that ROS production after K+ deprivation is blocked by actinomycin D, cycloheximide, and Ac-YVAD-CMK, suggesting that ROS act downstream of gene transcription, mRNA translation, and ICE activation. Taken together, we show that new mRNA and protein synthesis, activation of ICE-like proteases, and ROS production are sequential events in K+ deprivation-induced apoptosis of cerebellar granule neurons.

Animals↗
PubMed1996

TGF-beta-induced apoptosis of cerebellar granule neurons is prevented by depolarization.

The regulation of programmed cell death in the developing nervous system involves target-derived survival factors, afferent synaptic activity, and hormone- and cytokine-dependent signaling. Cultured immature cerebellar granule neurons die by apoptosis within several days in vitro unless maintained in depolarizing (high) concentrations of potassium (25 mM K+). Here we report that transforming growth factors (TGF)-beta1, -beta2, and -beta3 accelerate apoptosis of these neurons when maintained in physiological (low) K+ medium (5mM K+) as assessed by measures of viability, quantitative DNA fragmentation, and nuclear morphology. TGF-beta-induced apoptosis of these neurons is not blocked by CNTF and LIF, cytokines that enhance neuronal survival when applied alone, or by IGF-I, which prevents apoptosis upon potassium withdrawal. In contrast, neurons that differentiate in high K+ medium for several days in vitro acquire resistance to TGF-beta-mediated cell death. Granule neurons maintained in either low or high K+ medium produce latent, but not bioactive, TGF-beta1 and -beta2. Because neutralizing TGF-beta antibodies fail to augment survival of low K+ neurons, the cerebellar neurons are apparently unable to activate latent TGF-beta. Thus, apoptosis of low K+ neurons is not attributable to endogenous production of TGF-beta. Taken together, our data suggest that TGF-beta may limit the expansion of postmitotic neuronal precursor populations by promoting their apoptosis but may support survival of those neurons that have maturated, differentiated, and established supportive synaptic connectivity.

Animals↗
PubMed1996

Maturation-dependent modulation of apoptosis in cultured cerebellar granule neurons by cytokines and neurotrophins.

Immature cerebellar granule neurons die by apoptosis within 1 week in vitro unless maintained in depolarizing (high) concentrations of potassium (25 mM K+). Neurons allowed to survive and differentiate in high K+ medium for several days in vitro are still induced to undergo apoptosis when switched back to physiological (low) concentrations of K+ (5 mM). Here we have investigated the effects of various cytokines and growth factors in these two well-defined paradigms of neuronal apoptosis. Tumour necrosis factor-alpha, leukaemia inhibitory factor, ciliary neurotrophic factor, interleukin-10 and interleukin-13 delayed apoptosis and prolonged survival of cerebellar granule neurons maintained in low K+ medium. The effect observed required continuous exposure of the cultures to the cytokines and appeared not to involve modulation of Bcl-2 protein expression. Brain-derived neurotrophic factor accelerated neuronal death in low K+ medium. In contrast, when apoptosis of the neurons was precipitated by switching mature high K+ neurons to low K+ medium, neither tumour necrosis factor-alpha, leukaemia inhibitory factor, ciliary neurotrophic factor, interleukin-10 nor interleukin-13 prevented apoptosis. When testing the cytokines and growth factors for their capacity to alter N-methyl-D-aspartate receptor-mediated excitotoxicity of differentiated cerebellar granule neurons, no significant effect was observed. These data appear to define a maturation-dependent modulation of cerebellar granule cell survival by cytokines and neurotrophic factors that are expressed in a developmental pattern in the mammalian brain.

Animals↗
PubMed1996

Apoptosis in human alveolar macrophages is induced by endotoxin and is modulated by cytokines.

A central factor in the pathogenesis of inflammatory and fibrotic lung disease (adult respiratory distress syndrome, sarcoidosis, idiopathic pulmonary fibrosis) is the locally elevated number of alveolar macrophages (AM). An elevation in the production rate of AM, chemoattraction and differentiation of monocytes, or a diminution in the death rate might be underlying mechanisms. The aim of the present study was to investigate the modulatory role of endotoxin and cytokines on the death rate of human AM. Lipopolysaccharide (LPS) treatment resulted in a 4-fold increase (7.6 to 30.2%) of AM death. AM death was apoptotic as assessed by in situ DNA end labeling (ISDE), transmission electron microscopy, DNA gel electrophoresis, fluorometry of fragmented DNA, and an ELISA specific for histone-associated DNA fragments. Among the different bacterial cell wall components tested, LPS was the only inducer of apoptosis in human AM. None of the tested cytokines (interleukin-1 beta [IL-1 beta], IL-4, IL-6, IL-10, tumor necrosis factor-alpha [TNF-alpha], transforming growth factor-beta 2 [TGF-beta 2], interferon-gamma [IFN-gamma], macrophage colony-stimulating factor [M-CSF], granulocyte colony-stimulating factor [G-CSF], and granulocyte-macrophage colony-stimulating factor [GM-CSF]) was capable of enhancing the spontaneous rate of apoptosis. However, LPS-induced apoptosis was significantly enhanced by the macrophage-activating cytokine IFN-gamma, and reduced by the macrophage-deactivating cytokines IL-4, IL-10, and TGF-beta.

Apoptosis↗