Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P08758 (annexin V)
 9,383 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Rad21 is one of the major cohesin subunits that holds sister chromatids together until anaphase, when proteolytic cleavage by separase, a caspase-like enzyme, allows chromosomal separation. We show that cleavage of human Rad21 (hRad21) also occurs during apoptosis induced by diverse stimuli. Induction of apoptosis in multiple human cell lines results in the early (4 h after insult) generation of 64- and 60-kDa carboxy-terminal hRad21 cleavage products. We biochemically mapped an apoptotic cleavage site at residue Asp-279 (D(279)) of hRad21. This apoptotic cleavage site is distinct from previously described mitotic cleavage sites. hRad21 is a nuclear protein; however, the cleaved 64-kDa carboxy-terminal product is translocated to the cytoplasm early in apoptosis before chromatin condensation and nuclear fragmentation. Overexpression of the 64-kDa cleavage product results in apoptosis in Molt4, MCF-7, and 293T cells, as determined by TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) and Annexin V staining, assaying of caspase-3 activity, and examination of nuclear morphology. Given the role of hRad21 in chromosome cohesion, the cleaved C-terminal product and its translocation to the cytoplasm may act as a nuclear signal for apoptosis. In summary, we show that cleavage of a cohesion protein and translocation of the C-terminal cleavage product to the cytoplasm are early events in the apoptotic pathway and cause amplification of the cell death signal in a positive-feedback manner.
Mol Cell Biol 2002 Dec
PMID:Linking sister chromatid cohesion and apoptosis: role of Rad21. 1241 29

Elevated DNA repair processes represent resistance mechanisms to the treatment of malignancies with alkylating agents. Recently, the cell cycle checkpoint abrogator, UCN-01, was reported to inhibit nucleotide excision repair in cell-free systems. We hypothesized that if UCN-01 was combined with DNA-damaging agents, UCN-01 might inhibit the damage repair processes, thereby enhancing cytotoxicity in quiescent cells. Here, we investigated the effect of UCN-01 on DNA repair and viability of quiescent normal lymphocytes and chronic lymphocytic leukemia lymphocytes treated with UV or the cyclophosphamide prodrug 4-hydroperoxycyclophosphamide (4-HC). DNA damage repair kinetics were determined as DNA single strand breaks by the alkaline single cell gel electrophoresis (comet) assay and by [3H]thymidine incorporation. Pretreatment with UCN-01 inhibited DNA repair initiated by UV or 4-HC in normal lymphocytes as well as chronic lymphocytic leukemia lymphocytes in a concentration-dependent manner at clinically relevant levels (50-300 nM). This inhibition was demonstrated by the decreases in incision capability, DNA resynthesis, and in rejoining, suggesting that UCN-01 inhibits the multiple sites of the repair processes. The higher UCN-01 concentration (300 nM) maximized the inhibitory effects and enhanced the UV- or 4-HC-induced cytotoxicity, as determined by annexin V binding or Hoechst 33342 staining. This enhancement was not obtained by the lower concentrations that incompletely inhibited the repair, suggesting the close association between the inhibition of the repair and the enhancement of the cytotoxicity. Our findings suggest that UCN-01 may be a good candidate for combination strategies of cancer treatment.
Mol Cancer Ther 2002 Feb
PMID:UCN-01 (7-hydroxystaurosporine) inhibits DNA repair and increases cytotoxicity in normal lymphocytes and chronic lymphocytic leukemia lymphocytes. 1246 24

The hydroxymethylglutaryl-CoA reductase inhibitor lovastatin is used widely to treat hypercholesterolemia and has been shown to have cell cycle-specific effects. In these studies, we have examined the effects of combining lovastatin and paclitaxel (Taxol), a microtubule-stabilizing agent, in the human leukemia K562 and HL-60 cell lines. Isobologram analysis of cytotoxicity assays revealed that there is a synergistic interaction between the two agents in both cell lines. Cell cycle analyses showed that lovastatin enhances paclitaxel-induced G2-M arrest in both cell lines. In addition, Annexin V apoptotic studies revealed that lovastatin enhances paclitaxel-induced apoptosis in HL-60 cells. Lovastatin did not affect levels of [3H]paclitaxel in cells. Whereas lovastatin induced an accumulation of unmodified Ras and caused an up-regulation of both RhoB and Rap1A, paclitaxel was found to have no effect on the isoprenylated proteins. Studies of the centromere-associated protein mitosin revealed that treatment with lovastatin and paclitaxel resulted in increased mitosin levels and that lovastatin altered the association of mitosin with condensed chromosomes. These findings provide insight into the mechanisms underlying the cell cycle effects of lovastatin and support the development of a novel therapeutic strategy directed toward altering deleterious cell proliferation.
Mol Cancer Ther 2001 Dec
PMID:Synergistic interaction of lovastatin and paclitaxel in human cancer cells. 1246 31

The mechanism leading to the high level of radiosensitivity of T lymphocytes has not yet been fully described. In our previous study, we demonstrated that human peripheral T lymphocytes revealed early apoptotic changes (annexin V-positive) and late apoptotic changes (propidium iodide-positive), at 13 and 24 h after irradiation of 5 Gy, respectively. Changes in mitochondrial membrane potential were observed at 10 h after irradiation of 5 Gy. Subsequently, mitochondrial cytochrome c release was confirmed. In order to elucidate the mechanism which occurs prior to the mitochondrial membrane potential changes, we examined in the present study the radiation dose and the timing of oxidative DNA damage induced in human peripheral T lymphocytes following 10 MV X-ray irradiation. As a result, the production of 8-oxoguanine, i.e., the product of oxidative DNA damage, was clearly identified starting at 10, 6, and 3 h, after 2, 5, and 20 Gy of irradiation, respectively. Therefore, we concluded that it remains necessary to evaluate the extent of radiation-induced oxidative DNA damage. Furthermore, it is important to analyze superoxide radical production and scavenging in terms of the variety of radiosensitivities found among various types of normal tissue cells and neoplastic cells.
Int J Mol Med 2003 Jan
PMID:Radiation-induced oxidative DNA damage, 8-oxoguanine, in human peripheral T cells. 1246 13

The objective of this study was to determine potential mechanisms of apoptotic activity of gemcitabine, a pyrimidine nucleoside analogue, in the MM1.S multiple myeloma (MM) cell line. A MM cell line that is sensitive to glucocorticoids (MM1.S) was used for this study. Immunoblotting analysis, cell cycle assays, and annexin V staining were performed to determine whether gemcitabine induced apoptosis in this model. Furthermore, we attempted to delineate the apoptotic pathway by measuring caspase-8 and -9 activity using fluorometric assays. Loss of mitochondrial membrane potential was measured by flow cytometry. Gemcitabine treatment caused apoptosis in MM cell lines as measured by an increase in DNA cleavage, an increase in annexin V binding, a decrease in the mitochondrial membrane potential, and activation of caspase activity. Furthermore, cleavage of the caspase substrate poly(ADP-ribose) polymerase and caspase-3 activation were documented as early as 8 h after treatment with gemcitabine. Caspase-8 and -9 were activated by gemcitabine treatment in this cell line, suggesting several mechanisms of action including death receptor pathway and mitochondrial damage. The addition of interleukin 6 to MM1.S cells treated with gemcitabine offered no protection against gemcitabine-induced cell death. Gemcitabine induced apoptosis in the MM1.S cell line, and its activity required caspase activation. There is a suggestion that mitochondrial integrity is being affected with gemcitabine in this system. Gemcitabine acts independently of interleukin 6, suggesting potential important therapeutic implications in MM patients.
Mol Cancer Ther 2002 Nov
PMID:Caspase activation is required for gemcitabine activity in multiple myeloma cell lines. 1247 3

Several studies have suggested that high dietary fat intake, particularly essential fatty acids, is associated with pancreatic cancer development and growth. Our previous studies have demonstrated that blockade of either the 5-lipoxygenase (LOX) or 12-LOX pathway of arachidonic acid metabolism inhibited pancreatic cancer cell proliferation and induced apoptosis. This study investigated the underlying mechanisms for LOX inhibitor-induced apoptosis and the potential of LOX inhibitors as antipancreatic cancer agents using the athymic mice xenograft model. Apoptosis of pancreatic cancer cells induced by LOX inhibitors (including the nonselective LOX inhibitor nordihydroguaiaretic acid, the 5-LOX inhibitor Rev-5901, and the 12-LOX inhibitor baicalein) was confirmed by growth inhibition, annexin V binding, and terminal deoxynucleotidyl transferase-mediated nick end labeling assay in MiaPaCa-2 and AsPC-1 human pancreatic cancer cells. Expression of the antiapoptotic proteins Bcl-2 and Mcl-1 was significantly decreased after LOX inhibitor treatment while that of the proapoptotic protein bax was increased. LOX inhibitors also markedly induced the release of cytochrome c from mitochondria into the cytosol. Caspase-9, caspase-7, and caspase-3 but not caspase-8 were activated after treatment, concomitant with cleavage of the capase-3 substrate poly(ADP-ribose) polymerase. In vivo studies in the athymic mice xenograft model also confirmed the growth inhibitory effect and induction of apoptosis by these LOX inhibitors in pancreatic cancer. In conclusion, LOX inhibitors block pancreatic cancer cell proliferation and induce apoptosis through the mitochondrial pathway both in vivo and in vitro. LOX inhibitors are likely to be valuable for the treatment of human pancreatic cancer.
Mol Cancer Ther 2002 Sep
PMID:Lipoxygenase inhibitors attenuate growth of human pancreatic cancer xenografts and induce apoptosis through the mitochondrial pathway. 1248 14

Previously, we demonstrated that human peripheral T lymphocytes revealed early apoptotic changes (annexin V-positive) and late apoptotic changes (propidium iodide-positive), at 13 and 24 h, respectively, after irradiation of 5 Gy. Changes in mitochondrial membrane potential were observed at 10 h after irradiation of 5 Gy. Subsequently, mitochondrial cytochrome c-release was confirmed. In order to elucidate the mechanism which acts prior to the mitochondrial membrane potential changes, we examined in the previous study the radiation dose and the timing of oxidative DNA damage induced in human peripheral T lymphocytes following 10 MV X-ray irradiation. As a result, the production of 8-oxoguanine, i.e., the product of oxidative DNA damage, was clearly identified starting at 10, 6, and 3 h, after 2, 5, and 20 Gy of irradiation, respectively. Therefore, we examined in the present study reactive oxygen species (ROS) formation in T lymphocytes following 5 Gy of irradiation. Using a CCD camera system, we monitored fluorescence in T lymphocytes loaded with the succinimidyl ester of dichlorodihydrofluorescein diacetate (H2DCFDA), which is non-fluorescent until oxidized by ROS. We found that ROS formation occurred immediately after irradiation, continued for several hours, and resulted in oxidative DNA damage. Therefore, the origin of hyper-radiosensitivity of T lymphocytes seemed to be the high production of ROS in the mitochondrial DNA following irradiation.
Int J Mol Med 2003 Feb
PMID:Radiation-induced reactive oxygen species formation prior to oxidative DNA damage in human peripheral T cells. 1252 68

Inflammation and cell death are two important components of myocarditis. We evaluated the distribution of inflammation and apoptotic cell death in rats with autoimmune myocarditis using two radiotracers - technetium-99m Hynic-annexin V ((99m)Tc-annexin) as a marker of apoptotic cell death and carbon-14 deoxyglucose ((14)C-DG) as a marker of inflammation - in comparison with histologic findings. Three, 7 and 14 weeks after immunization with porcine cardiac myosin (acute, subacute, and chronic phases, respectively) (99m)Tc-annexin and (14)C-DG were injected. The uptake in the total heart was determined as the percentage of injected dose per gram (% ID/g) by tissue counting. Dual-tracer autoradiography with (99m)Tc-annexin and (14)C-DG was performed. The distribution of each of these agents was compared with the results of hematoxylin and eosin staining to identify areas of inflammation, and TUNEL staining to identify areas of apoptosis. Total cardiac uptake of (99m)Tc-annexin in the acute phase of myocarditis was significantly higher than that in normal rats (1.28%+/-0.30% vs 0.46%+/-0.01%; P<0.0001); it then decreased in the subacute phase and reached normal levels (0.56%+/-0.08% vs 0.60%+/-0.08%; P=NS). Total cardiac uptake of (14)C-DG in the acute phase of myocarditis was significantly higher than that in normal rats (2.78%+/-0.95% vs 1.02%+/-0.25%; P<0.0001); it then decreased in the subacute phase, but still remained higher than in controls (2.06%+/-0.52% vs 1.37%+/-0.46%; P<0.05). Using autoradiography and staining of tissue specimens, it was found that most histologic inflammatory foci corresponded to areas of high (14)C-DG uptake; some also corresponded to areas of high (99m)Tc-annexin uptake in the acute phase of myocarditis. (99m)Tc-annexin localization was strongly correlated with the number of TUNEL-positive cells (P<0.0001, r=0.83), but the uptake of (14)C-DG showed no relationship with it. There is a marked difference in the distribution of inflammation and apoptotic cell death in the myocardium of animals with immune myocarditis. These changes are mirrored by the localization of (14)C-DG and (99m)Tc-annexin. Sites of inflammation and zones of apoptotic cell death change over the course of immune myocarditis.
Eur J Nucl Med Mol Imaging 2003 Feb
PMID:99mTc-Hynic-annexin V imaging to evaluate inflammation and apoptosis in rats with autoimmune myocarditis. 1255 41

In vitro assays for chromosome aberrations (i.e., in vitro micronucleus and in vitro metaphase analysis tests) frequently produce false-positive or exaggerated-positive results. Our previous work suggested that apoptosis interferes with these tests, producing misleading results. These previous studies were conducted by performing the in vitro micronucleus test in CTLL-2 cells and a CTLL-2 cell derivative stably transfected with the apoptosis inhibitor gene bcl2. In the present study, these previous observations were extended by examining micronucleus induction with a larger number of compounds in both CTLL-2 and CTLL-2 bcl2 cells and measuring apoptosis with annexin V-FITC. Both cell lines were treated with different classes of compounds that were anticipated to be exclusively apoptosis inducers, or compounds known to be clastogens or aneugens, some of which were anticipated to be both genotoxic and apoptotic. We were able to confirm that compounds that are only apoptogenic induced micronuclei in CTLL-2 but not CTLL-2 bcl2 cells, indicating that the positive responses are due to apoptosis in CTLL-2 cells. Some genotoxins (clastogens and aneugens) did not produce apoptosis by the annexin V assay and gave similar responses in CTLL-2 and CTLL-2 bcl2 cells. Finally, higher responses were induced in CTLL-2 cells compared to CTLL-2 bcl2 cells that were treated with aneugens or clastogens that were also apoptosis inducers, suggesting that the greater response in CTLL-2 cells is a consequence of both genotoxicity and apoptosis. Finally, it was demonstrated that just eliminating CTLL-2 cells having three or more micronuclei from scoring was not adequate for correctly evaluating agents that only produce apoptosis. The results indicate that coupling the in vitro micronucleus test in both CTLL-2 and CTLL-2 bcl2 cells with the measurement of apoptosis is able to distinguish the genotoxic effects of a test compound from its apoptotic potential and is able to avoid interference from apoptosis in the in vitro micronucleus test. These observations may provide the basis for a useful genotoxicity assay.
Environ Mol Mutagen 2003
PMID:Using CTLL-2 and CTLL-2 bcl2 cells to avoid interference by apoptosis in the in vitro micronucleus test. 1255 88

Apoptosis may limit the utility of cell-based therapies for articular cartilage disorders. We tested the hypothesis that chondrocyte apoptosis can be reduced by optimizing the conditions employed to expand chondrocyte numbers in culture. Chondrocyte apoptosis was examined in monolayer and suspension culture, in the presence of fetal bovine serum (FBS) or autologous serum, and for culture periods of 2 or 4 days. The effect of these variables was assessed by measuring cell viability, Annexin V labeling and mitochondrial membrane potential. After 2 days of culture, the greatest increase in viable cell number (3.7-fold) occurred in monolayer cultures with autologous serum. After 4 days of culture, the greatest increase in cell number (9.0-fold) occurred in monolayer cultures supplemented with FBS. By Annexin V staining, the proportion of cells undergoing apoptosis after 2 days was not affected by the type of serum used or by culture in monolayer versus suspension. After 4 days of culture, the proportion of apoptotic cells was significantly reduced (35% to 13%, p<0.02) in suspension cultures with autologous serum. Apoptosis assessed by loss of mitochondrial membrane potential was decreased in the presence of autologous serum. These data suggest that suspension culture with autologous serum is useful in simultaneously maintaining cell proliferation and minimizing apoptosis in cultured human articular chondrocytes.
Int J Mol Med 2003 Mar
PMID:Role of apoptosis inhibition in various chondrocyte culture systems. 1257 30


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