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K Schulze-Osthoff

Publications and source records attributed to K Schulze-Osthoff.

At least 19 recordsLinked to original sources
PubMed2006

Apoptin is modified by SUMO conjugation and targeted to promyelocytic leukemia protein nuclear bodies.

Apoptin, a protein of the chicken anemia virus (CAV), represents a novel potential anticancer therapeutic, because it induces apoptotic death specifically in tumor but not normal cells. The cellular localization appears to be crucial for apoptin's selective toxicity. In normal cells apoptin remains in the cytoplasm, whereas in transformed cells it migrates into the nucleus and kills the cell. However, the manner by which apoptin is able to distinguish between tumor and normal cells is unknown. Here, we report for the first time that apoptin interacts directly with the promyelocytic leukemia protein (PML) in tumor cells and accumulates in PML nuclear bodies (NBs), which are involved in apoptosis induction and viral replication. We also demonstrate that apoptin is sumoylated and that a sumoylation-deficient apoptin mutant is no longer recruited to PML-NBs, but localizes in the nuclear matrix. This mutant fails to bind PML, but can still induce apoptosis as efficiently as wild-type apoptin. Moreover, apoptin kills also PML-/- cells and promyelocytic leukemia cells with defective PML expression. Our results therefore suggest that apoptin kills tumor cells independently of PML and sumoylation, however, the interaction of apoptin with PML and small ubiquitin-like modifier (SUMO) proteins might be relevant for CAV replication.

Capsid Proteins↗
PubMed2006

Staphylococcus aureus induces caspase-independent cell death in human peritoneal mesothelial cells.

Bacterial peritonitis remains a serious complication of peritoneal dialysis. Although Staphylococcus epidermidis is the most common pathogen involved, infections with Staphylococcus aureus lead to severe peritoneal damage and are often associated with a dramatic loss of mesothelial cells. Induction of cell death appears to be involved in peritoneal damage and mesothelial cell loss during bacterial infections. Using cultured human peritoneal mesothelial cells (HMCs), we investigated the ability of different S. epidermidis and S. aureus strains to damage the HMC monolayer and to trigger cell death. We show that only a subgroup of live S. aureus isolates, characterized by an invasive and alpha-hemolysin-producing phenotype, induces cell death. None of the tested S. epidermidis strains, which were not invasive or hemolytic, had a cytotoxic effect. After host cell invasion, S. aureus resided within phagocytic vacuoles, and HMCs were apparently able to degrade staphylococci. However, even after prolonged infection, a high percentage of S. aureus remained alive within HMCs and might be released after host cell death. Cell death induced by S. aureus was accompanied by apoptotic alterations, such as DNA fragmentation, but was independent of endogenous FasL and tumor necrosis factor-alpha death ligand expression. Moreover, caspases were not involved in S. aureus-induced mesothelial cell death. In conclusion, our data indicate that mesothelial cell death might represent a major mechanism of S. aureus-induced damage of the peritoneum during bacterial peritonitis.

Apoptosis↗
PubMed2006

CD95 ligand mediates T-cell receptor-induced apoptosis of a CD4+ CD8+ double positive thymic lymphoma.

Tumors in the thymus can be of different cellular origin. Among the most common tumors are thymoma and lymphoma, which are derived from transformed thymic epithelial cells and transformed lymphocytes, respectively. Thymic lymphoma and their response to apoptotic stimuli are poorly characterized. Here, we analyse apoptosis events in the thymic lymphoma cell line Thy278, which expresses cell surface antigens characteristic of immature double positive thymocytes. Upon T-cell receptor (TCR)/CD3 stimulation, Thy278 cells die by apoptosis, similar as primary thymocytes during negative selection. Caspases are crucial for deletion of both Thy278 cells and normal thymocytes. Moreover, we show that deletion of primary thymocytes and Thy278 cells upon CD3 stimulation is considerably impaired by neutralizing CD95L antibody. Thus, our results not only demonstrate that TCR-induced apoptosis is still functional in transformed thymocytes, but also suggest that Thy278 cells are a helpful model for the molecular analysis of negative selection.

Animals↗
PubMed2006

Apoptin-induced cell death is modulated by Bcl-2 family members and is Apaf-1 dependent.

Apoptin, a chicken anemia virus-derived protein, selectively induces apoptosis in transformed but not in normal cells, thus making it a promising candidate as a novel anticancer therapeutic. The mechanism of apoptin-induced apoptosis is largely unknown. Here, we report that contrary to previous assumptions, Bcl-2 and Bcl-xL inhibit apoptin-induced cell death in several tumor cell lines. In contrast, deficiency of Bax conferred resistance, whereas Bax expression sensitized cells to apoptin-induced death. Cell death induction by apoptin was associated with cytochrome c release from mitochondria as well as with caspase-3 and -7 activation. Benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone, a broad spectrum caspase inhibitor, was highly protective against apoptin-induced cell death. Apoptosis induced by apoptin required Apaf-1, as immortalized Apaf-1-deficient fibroblasts as well as tumor cells devoid of Apaf-1 were strongly protected. Thus, our data indicate that apoptin-induced apoptosis is not only Bcl-2- and caspase dependent, but also engages an Apaf-1 apoptosome-mediated mitochondrial death pathway.

Amino Acid Chloromethyl Ketones↗
PubMed2006

Induction of p21CIP/WAF-1 and G2 arrest by ionizing irradiation impedes caspase-3-mediated apoptosis in human carcinoma cells.

There is an ongoing controversy regarding the relevance of apoptosis induction by ionizing irradiation as compared with other end points including transient or permanent cell cycle arrest of damaged cells. Here, we show that such permanent cell cycle arrest and apoptosis represent two sides of the same coin. MCF-7 cells fail to express procaspase-3, which results in resistance to apoptosis induced by anticancer drugs. Conversely, restoration of procaspase-3 sensitizes MCF-7 cells to chemotherapeutics including epirubicine, etoposide and taxol. In contrast, irradiation does not trigger apoptotic cell death but results in prolonged arrest in the G2 phase of the cell division cycle regardless of procaspase-3 expression. This suggested that the propensity of MCF-7 cells to arrest at the G2 checkpoint results in resistance to apoptosis upon gamma-irradiation. This G2 arrest was associated with upregulation of p21CIP/WAF-1. Inhibition of DNA-damage-induced stress kinases and p21CIP/WAF-1 expression by caffeine abrogated G2 arrest and induced apoptosis of the irradiated cells in a caspase-3-dependent manner. Inhibition of cell cycle progression by adenoviral expression of the cyclin dependent kinase inhibitor p21CIP/WAF-1 prevented apoptosis upon caffeine treatment indicating that cell cycle progression, that is, G2-release, is required for induction of apoptosis. Likewise, cells homozygously deleted for p21CIP/WAF-1 (HCT116 p21-/-) display enhanced irradiation-induced apoptosis via a caspase-3-dependent mechanism. These data indicate that the disruption of G2 checkpoint control overcomes cell cycle arrest and resistance to gamma-irradiation-induced cell death. Thus, DNA damage may trigger a permanent G2 arrest as an initial inactivation step of tumor cells where the phenomenon of apoptosis is hidden unless cell cycle arrest is overcome. The efficient induction of apoptosis upon G2 release thereby depends on the propensity to activate the key executioner caspase-3. This finding is of crucial importance for the understanding of molecular steps underlying the efficacy of ionizing radiation to delete tumor cells.

Antineoplastic Agents↗
PubMed2006

The tyrosine kinase Lck is a positive regulator of the mitochondrial apoptosis pathway by controlling Bak expression.

Tyrosine kinases of the Src family have been implicated in key biological processes. Here, we provide evidence that p56(Lck), a lymphoid-specific Src kinase, is involved in the activation of the mitochondrial apoptosis pathway. Lck-deficient T cells were completely resistant to anticancer drugs. In contrast, apoptosis sensitivity to death receptors was not altered, indicating a specific interference of Lck with the mitochondrial pathway. Re-expression of Lck restored sensitivity to drug-induced apoptosis and triggered mitochondrial cytochrome c release and caspase activation. Further analysis identified that the sensitization by Lck was independent of classical mediators of T-cell signaling, but essentially involved the Bcl-2 protein Bak. Expression of Bak was completely absent in Lck-deficient cells, while re-expression of Lck transcriptionally triggered Bak expression and conferred sensitivity to apoptosis, associated with a proapoptotic conformational change of Bak. Furthermore, in vitro the truncated fragment of Bid specifically activated Bak and cytochrome c release only from mitochondria of Lck-expressing cells. These results do not only demonstrate a sentinel role of Lck in drug resistance but also delineate a hitherto unknown pathway of Src kinases in regulation of Bcl-2 proteins.

Antineoplastic Agents↗
PubMed2006

Unique and overlapping substrate specificities of caspase-8 and caspase-10.

Although caspase-8 has an established role as an initiator of death receptor-mediated apoptosis, the function of its closest homolog, caspase-10, is almost completely unknown. To gain a closer insight into the physiological function of caspase-10, we compared the cleavage of known caspase-8 substrates by both initiator caspases. We demonstrate that caspase-10 and -8 have overlapping cleavage preferences for several substrates such as the kinases RIP and PAK2. Interestingly, in other substrates, such as the Bcl-2 protein Bid, we found additional and distinct cleavage sites for both caspases, which might have important consequences for mitochondrial targeting and propagation of the death signal. Caspase-8 and -10 also caused different interchain cleavage patterns of their enzyme precursors. Together, these results suggest that caspase-8 and -10, despite having overlapping functions, also have selective substrate cleavage specificities and might thereby exert nonredundant roles in apoptosis signaling.

Amino Acid Sequence↗
PubMed2005

Apoptosis-based therapies and drug targets.

The pathogenesis of many diseases is most closely connected with aberrantly regulated apoptotic cell death. The past 15 years have witnessed an explosion in the basic knowledge of mechanisms that regulate apoptosis and the mediators that either trigger or inhibit cell death. Consequently, great interest has emerged in devising therapeutic strategies for modulating the key molecules of life-and-death decisions. Numerous novel approaches are currently being followed employing gene therapy and antisense strategies, recombinant biologics or classical organic and combinatorial chemistry in order to target specific apoptotic regulators. Although drug development is still in its infancy, several therapeutics have progressed to clinical testing or have even been approved in record time. This review outlines the recent advances in the field of apoptosis-based therapies and explores some highlights of a very active field of drug development.

Animals↗
PubMed2004

Transcription factor AP-2alpha triggers apoptosis in cardiac myocytes.

Idiopathic-dilated cardiomyopathy (IDC) is a common primary myocardial disease of unknown etiology associated with apoptosis, cardiac dilatation, progressive heart failure and increased mortality. An elevation of the transcription factor activator protein 2alpha (AP-2alpha) is involved in vertebrate embryonic development and oncogenesis. Here, we show that AP-2alpha protein is expressed in the human heart and increased in human failing myocardium with IDC. Adenovirus-mediated overexpression of human AP-2alpha triggered apoptosis and increased mRNA levels of Bcl-2 family members Bax and Bcl-x in rat cardiomyocytes. Immunohistological analysis of human myocardium revealed an increased percentage of AP-2alpha-positive nuclei in IDC and, interestingly, a colocalization of AP-2alpha-positive but not -negative cells with a caspase-cleaved fragment of poly(ADP-ribose)polymerase. We suggest AP-2alpha as a novel cardiac regulator implicated in the activation of apoptosis in IDC.

Adenoviridae↗
PubMed2003

Apoptosis in hepatitis C virus infection.

Infection with hepatitis C virus (HCV) is characterized by inflammatory liver damage and a long viral persistence associated with an increased risk of developing hepatocellular carcinoma. Both in liver damage and in oncogenesis a disturbance of apoptosis has been implicated, although the underlying mechanisms in these apparently opposite processes are incompletely understood. HCV-triggered liver injury is mediated mainly by host immune mechanisms and eventually by direct cytopathic effects of HCV. Recent data shows that caspase activation, either triggered by death ligands, other cytokines, granzyme B or HCV proteins, is considerably upregulated in HCV-infected liver. Interestingly, caspase activation appears to correlate closely with the inflammatory response. Data about the role of single HCV proteins, either in cultured cells or transgenic animals models, however, are contradictory, as both pro- and anti-apoptotic effects have been observed. Nevertheless, apoptosis induction upon HCV infection may critically contribute to liver damage, while inhibition of apoptosis may result in HCV persistence and development of hepatocellular carcinoma.

Animals↗
PubMed2003

Many cuts to ruin: a comprehensive update of caspase substrates.

Apoptotic cell death is executed by the caspase-mediated cleavage of various vital proteins. Elucidating the consequences of this endoproteolytic cleavage is crucial for our understanding of cell death and other biological processes. Many caspase substrates are just cleaved as bystanders, because they happen to contain a caspase cleavage site in their sequence. Several targets, however, have a discrete function in propagation of the cell death process. Many structural and regulatory proteins are inactivated by caspases, while other substrates can be activated. In most cases, the consequences of this gain-of-function are poorly understood. Caspase substrates can regulate the key morphological changes in apoptosis. Several caspase substrates also act as transducers and amplifiers that determine the apoptotic threshold and cell fate. This review summarizes the known caspase substrates comprising a bewildering list of more than 280 different proteins. We highlight some recent aspects inferred by the cleavage of certain proteins in apoptosis. We also discuss emerging themes of caspase cleavage in other forms of cell death and, in particular, in apparently unrelated processes, such as cell cycle regulation and cellular differentiation.

Animals↗
PubMed2003

Staphylococcus aureus alpha-toxin-induced cell death: predominant necrosis despite apoptotic caspase activation.

Recent data suggest that alpha-toxin, the major hemolysin of Staphylococcus aureus, induces cell death via the classical apoptotic pathway. Here we demonstrate, however, that although zVAD-fmk or overexpression of Bcl-2 completely abrogated caspase activation and internucleosomal DNA fragmentation, they did not significantly affect alpha-toxin-induced death of Jurkat T or MCF-7 breast carcinoma cells. Caspase inhibition had also no effect on alpha-toxin-induced lactate dehydrogenase release and ATP depletion. Furthermore, whereas early assessment of apoptosis induction by CD95 resulted solely in the generation of cells positive for active caspases that were, however, not yet permeable for propidium iodide, a substantial proportion of alpha-toxin-treated cells were positive for both active caspases and PI. Finally, electron microscopy demonstrated that even in the presence of active caspases, alpha-toxin-treated cells displayed a necrotic morphology characterized by cell swelling and cytoplasmic vacuolation. Together, our data suggest that alpha-toxin-induced cell death proceeds even in the presence of activated caspases, at least partially, in a caspase-independent, necrotic-like manner.

Amino Acid Chloromethyl Ketones↗
PubMed2001

alpha-Toxin is a mediator of Staphylococcus aureus-induced cell death and activates caspases via the intrinsic death pathway independently of death receptor signaling.

Infections with Staphylococcus aureus, a common inducer of septic and toxic shock, often result in tissue damage and death of various cell types. Although S. aureus was suggested to induce apoptosis, the underlying signal transduction pathways remained elusive. We show that caspase activation and DNA fragmentation were induced not only when Jurkat T cells were infected with intact bacteria, but also after treatment with supernatants of various S. aureus strains. We also demonstrate that S. aureus-induced cell death and caspase activation were mediated by alpha-toxin, a major cytotoxin of S. aureus, since both events were abrogated by two different anti-alpha-toxin antibodies and could not be induced with supernatants of an alpha-toxin-deficient S. aureus strain. Furthermore, alpha-toxin-induced caspase activation in CD95-resistant Jurkat sublines lacking CD95, Fas-activated death domain, or caspase-8 but not in cells stably expressing the antiapoptotic protein Bcl-2. Together with our finding that alpha-toxin induces cytochrome c release in intact cells and, interestingly, also from isolated mitochondria in a Bcl-2-controlled manner, our results demonstrate that S. aureus alpha-toxin triggers caspase activation via the intrinsic death pathway independently of death receptors. Hence, our findings clearly define a signaling pathway used in S. aureus-induced cytotoxicity and may provide a molecular rationale for future therapeutic interventions in bacterial infections.

Adaptor Proteins, Signal Transducing↗
PubMed2001

The role of ceramide in receptor- and stress-induced apoptosis studied in acidic ceramidase-deficient Farber disease cells.

The activation of sphingomyelinases leading to the generation of ceramide has been implicated in various apoptotic pathways. However, the role of ceramide as an essential death mediator remains highly controversial. In the present study, we investigated the functional relevance of ceramide in a genetic model by using primary cells from a Farber disease patient. These cells accumulate ceramide as the result of an inherited deficiency of acidic ceramidase. We demonstrate that Farber disease lymphocytes and fibroblasts underwent apoptosis induced by various stress stimuli, including staurosporine, anticancer drugs and gamma-irradiation, equally as normal control cells. In addition, caspase activation by these proapoptotic agents occurred rather similarly in Farber disease and control fibroblasts. Interestingly, Farber disease lymphoid cells underwent apoptosis induced by the CD95 death receptor more rapidly than control cells. Our data therefore suggest that ceramide does not play an essential role as a second messenger in stress-induced apoptosis. However, in accordance with a role in lipid-rich microdomains, ceramide by altering membrane composition may function as an amplifier in CD95-mediated apoptosis.

Antineoplastic Agents↗
PubMed2001

Rapid extracellular release of cytochrome c is specific for apoptosis and marks cell death in vivo.

Diverse death stimuli including anticancer drugs trigger apoptosis by inducing the translocation of cytochrome c from the outer mitochondrial compartment into the cytosol. Once released, cytochrome c cooperates with apoptotic protease-activating factor-1 and deoxyadenosine triphosphate in caspase-9 activation and initiation of the apoptotic protease cascade. The results of this study show that on death induction by chemotherapeutic drugs, staurosporine and triggering of the death receptor CD95, cytochrome c not only translocates into the cytosol, but furthermore can be abundantly detected in the extracellular medium. The cytochrome c release from the cell is a rapid and apoptosis-specific process that occurred within 1 hour after induction of apoptosis, but not during necrosis. Interestingly, elevated cytochrome c levels were observed in sera from patients with hematologic malignancies. In the course of cancer chemotherapy, the serum levels of cytochrome c in the majority of the patients grew rapidly as a result of increased cell death. These data suggest that monitoring of cytochrome c in the serum of patients with tumors might serve as a useful clinical marker for the detection of the onset of apoptosis and cell turnover in vivo.

Adult↗
PubMed2001

Homocysteine inhibits tumor necrosis factor-induced activation of endothelium via modulation of nuclear factor-kappa b activity.

Homocystinuria is a metabolic disorder associated with an increased incidence of vascular disease. Here, we analyzed the effects of homocysteine on endothelial cell activation that is a prerequisite for the recruitment of leukocytes to sites of evolving atherosclerotic plaques. Exposure of human umbilical vein endothelial cells to homocysteine alone did not modulate expression of the adhesion molecules E-selectin, intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, and the chemokines monocyte chemotactic protein-1 and interleukin-8. In contrast, tumor necrosis factor (TNF)-induced upregulation of these molecules was almost completely inhibited by homocysteine, but not by related thiol amino acids. Using electrophoretic mobility shift and reporter gene assays, the inhibitory effect of homocysteine could be attributed to inhibition of DNA binding and transcriptional activity of NF-kappa B. TNF-induced phosphorylation and degradation of I kappa B-alpha, however, were not affected. Neither was NF-kappa B-independent activation of endothelial cells by interferon-gamma influenced by homocysteine. In summary, our data indicate that homocysteine alters the response to injury of endothelial cells which may have fundamental impacts on mechanisms of leukocyte recruitment to sites of inflammation. Our findings might refer to a novel pathway by which homocysteine is involved in vascular disorders associated with homocystinuria.

Cells, Cultured↗
PubMed2001

Ionizing radiation but not anticancer drugs causes cell cycle arrest and failure to activate the mitochondrial death pathway in MCF-7 breast carcinoma cells.

There is considerable evidence that ionizing radiation (IR) and chemotherapeutic drugs mediate apoptosis through the intrinsic death pathway via the release of mitochondrial cytochrome c and activation of caspases -9 and -3. Here we show that MCF-7 cells that lack caspase-3 undergo a caspase-dependent apoptotic cell death in the absence of DNA fragmentation and alpha-fodrin cleavage following treatment with etoposide or doxorubicin, but not after exposure to IR. Re-expression of caspase-3 restored DNA fragmentation and alpha-fodrin cleavage following drug treatment, but it did not alter the radiation-resistant phenotype of these cells. In contrast to the anticancer drugs, IR failed to induce the intrinsic death pathway in MCF-7/casp-3 cells, an event readily observed in IR-induced apoptosis of HeLa cells. Although IR-induced DNA double-strand breaks were repaired with similar efficiencies in all cell lines, cell cycle analyses revealed a persistent G2/M arrest in the two MCF-7 cell lines, but not in HeLa cells. Together, our data demonstrate that caspase-3 is required for DNA fragmentation and alpha-fodrin cleavage in drug-induced apoptosis and that the intrinsic death pathway is fully functional in MCF-7 cells. Furthermore, they show that the radiation-resistant phenotype of MCF-7 cells is not due to the lack of caspase-3, but is caused by the failure of IR to activate the intrinsic death pathway. We propose (1) different signaling pathways are induced by anticancer drugs and IR, and (2) IR-induced G2/M arrest prevents the generation of an apoptotic signal required for the activation of the intrinsic death pathway.

Antineoplastic Agents↗