Apoptosis induced by lipid-associated membrane proteins fromMycoplasma penetransis mediated by nuclear factor κB activation in mouse macrophage

2008 ◽  
Vol 54 (2) ◽  
pp. 150-158 ◽  
Author(s):  
Yanhua Zeng ◽  
Yimou Wu ◽  
Zhongliang Deng ◽  
Xiaoxing You ◽  
Cuiming Zhu ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Molecular Mechanisms ◽  
Specific Inhibitor ◽  
Annexin V ◽  
Fluorescein Isothiocyanate ◽  
Nuclear Factor Κb ◽  
Pyrrolidine Dithiocarbamate ◽  
Pathogenic Potential ◽  
Mouse Macrophages ◽  
Induced Apoptosis

Mycoplasma penetrans was shown to be involved in alteration of several eukaryotical cells functions and a causative agent in urogenital infectious diseases. Lipid-associated membrane proteins (LAMPs) may be responsible for the pathogenicity of some mycoplamas. In this study, we investigated whether M. penetrans LAMPs have pathogenic potential by inducing apoptosis in mouse macrophages. As analyzed by annexin-V – fluorescein isothiocyanate staining, significant early- and late-stage apoptosis was induced in M. penetrans LAMPs-challenged mouse macrophages. And agarose gel electrophoresis of the DNA of M. penetrans LAMPs-challenged cells revealed a ladder-like pattern of migration of DNA indicative of apoptosis. The possible molecular mechanisms responsible for the induction of apoptosis were also investigated by characterizing the activation of nuclear transcription factor κB (NFκB). NFκB was activated and translocated into the nucleus in mouse macrophages stimulated by M. penetrans LAMPs. The activation of NFκB and M. penetrans LAMPs-induced apoptosis in mouse macrophages was partially inhibited by the NFκB-specific inhibitor pyrrolidine dithiocarbamate. Thus, this study demonstrates that M. penetrans LAMPs may be an important etiological factor owing to their ability to induce apoptosis in mouse macrophages, which is probably mediated through the activation of NFκB.

scholarly journals Carfilzomib in Combination with Bortezomib Enhances Apoptotic Cell Death in B16-F1 Melanoma Cells

Biology ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 153
Author(s):  
Min Seung Lee ◽  
So Hyun Lim ◽  
Ah-Ran Yu ◽  
Chi Yeon Hwang ◽  
Insug Kang ◽  
...  
Keyword(s):  
Er Stress ◽  
Dose Reduction ◽  
Molecular Mechanisms ◽  
Apoptotic Cell ◽  
Proteasome Inhibitors ◽  
Annexin V ◽  
Fluorescein Isothiocyanate ◽  
Melanoma Cells ◽  
Eif2α Phosphorylation ◽  
Pharmacological Interactions

Proteasome inhibitors, such as bortezomib (BZ) and carfilzomib (CFZ), have been suggested as treatments for various cancers. To utilize BZ and/or CFZ as effective therapeutics for treating melanoma, we studied their molecular mechanisms using B16-F1 melanoma cells. Flow cytometry of Annexin V-fluorescein isothiocyanate-labeled cells indicated apoptosis induction by treatment with BZ and CFZ. Apoptosis was evidenced by the activation of various caspases, including caspase 3, 8, 9, and 12. Treatment with BZ and CFZ induced endoplasmic reticulum (ER) stress, as indicated by an increase in eIF2α phosphorylation and the expression of ER stress-associated proteins, including GRP78, ATF6α, ATF4, XBP1, and CCAAT/enhancer-binding protein homologous protein. The effects of CFZ on ER stress and apoptosis were lower than that of BZ. Nevertheless, CFZ and BZ synergistically induced ER stress and apoptosis in B16-F1 cells. Furthermore, the combinational pharmacological interactions of BZ and CFZ against the growth of B16-F1 melanoma cells were assessed by calculating the combination index and dose-reduction index with the CompuSyn software. We found that the combination of CFZ and BZ at submaximal concentrations could obtain dose reduction by exerting synergistic inhibitory effects on cell growth. Moreover, this drug combination reduced tumor growth in C57BL/6 syngeneic mice. Taken together, these results suggest that CFZ in combination with BZ may be a beneficial and potential strategy for melanoma treatment.

scholarly journals Vulpinic Acid as a Natural Compound Inhibits the Proliferation of Metastatic Prostate Cancer Cell by Inducing Apoptosis

2021 ◽  
Author(s):  
Demet Cansaran Duman ◽  
Gamze Guney Eskiler ◽  
Betül Çolak ◽  
Elif Sozen Kucukkara
Keyword(s):  
Prostate Cancer ◽  
Metastatic Prostate Cancer ◽  
Molecular Mechanisms ◽  
Treatment Options ◽  
Annexin V ◽  
Current Treatment ◽  
Pcr Analysis ◽  
G1 Arrest ◽  
Advanced Analysis ◽  
Induced Apoptosis

Abstract Lichen secondary metabolites have drawn considerable attention in recent years due to limitations of current treatment options. Vulpinic acid (VA) obtained from Letharia vulpina lichen species exerts a remarkable cytotoxic effect on different cancer types. However, the therapeutic efficacy of VA in metastatic prostate cancer (mPC) cells has not been investigated. In the present study, we aimed to identify VA-mediated cytotoxicity in PC-3 mPC cells compared with control cells. After identification of the cytotoxic concentrations of VA, VA induced apoptosis was analyzed by Annexin V, cell cycle, acridine orange and propidium iodide staining and RT-PCR analysis. Our findings showed that VA significantly decreased the viability of PC-3 cells (p < 0.01) and caused a considerable early apoptotic effect through G0/G1 arrest, nuclear bleebing and the activation of particularly initiator caspases. Therefore, VA may be a potential treatment option for mPC patients. However, the underlying molecular mechanisms of VA-induced apoptosis with advanced analysis should be further performed.

scholarly journals Curcumin Enhanced Busulfan-Induced Apoptosis through Downregulating the Expression of Survivin in Leukemia Stem-Like KG1a Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-16 ◽  
Author(s):  
Guangyang Weng ◽  
Yingjian Zeng ◽  
Jingya Huang ◽  
Jiaxin Fan ◽  
Kunyuan Guo
Keyword(s):  
Specific Inhibitor ◽  
Annexin V ◽  
Chemotherapeutic Agents ◽  
Combination Group ◽  
Hematopoietic Stem ◽  
Leukemia Relapse ◽  
Conditioning Regimens ◽  
Underlying Mechanisms ◽  
Induced Apoptosis ◽  
Related Proteins

Leukemia relapse and nonrecurrence mortality (NRM) due to leukemia stem cells (LSCs) represent major problems following hematopoietic stem cell transplantation (HSCT). To eliminate LSCs, the sensitivity of LSCs to chemotherapeutic agents used in conditioning regimens should be enhanced. Curcumin (CUR) has received considerable attention as a result of its anticancer activity in leukemia and solid tumors. In this study, we investigated the cytotoxic effects and underlying mechanisms in leukemia stem-like KG1a cells exposed to busulfan (BUS) and CUR, either alone or in combination. KG1a cells exhibiting BUS-resistance demonstrated by MTT and annexin V/propidium iodide (PI) assays, compared with HL-60 cells. CUR induced cell growth inhibition and apoptosis in KG1a cells. Apoptosis of KG1a cells was significantly enhanced by treatment with CUR+BUS, compared with either agent alone. CUR synergistically enhanced the cytotoxic effect of BUS. Seven apoptosis-related proteins were modulated in CUR- and CUR+BUS-treated cells analyzed by proteins array analysis. Importantly, the antiapoptosis protein survivin was significantly downregulated, especially in combination group. Suppression of survivin with specific inhibitor YM155 significantly increased the susceptibility of KG1a cells to BUS. These results demonstrated that CUR could increase the sensitivity of leukemia stem-like KG1a cells to BUS by downregulating the expression of survivin.

Involvement of the sphingolipid ceramide in heat-shock-induced apoptosis of bovine oocytes

2011 ◽  
Vol 23 (7) ◽  
pp. 876 ◽  
Author(s):  
Dorit Kalo ◽  
Zvi Roth
Keyword(s):  
Heat Shock ◽  
De Novo ◽  
Specific Inhibitor ◽  
Annexin V ◽  
Developmental Competence ◽  
Control Group ◽  
Cleavage Rate ◽  
Oocyte Developmental Competence ◽  
Induced Apoptosis

Programmed cell death via the sphingomyelin pathway has been suggested to underlie heat-shock disturbance of oocyte developmental competence. A series of experiments were performed to characterise the role of the sphingolipid ceramide in heat-shock-induced apoptosis, and to determine whether ceramide formation can be regulated. Bovine cumulus–oocyte complexes (COCs) were aspirated from ovaries collected in the cold season (November–April), in vitro-matured, fertilised and cultured for 8 days. Exposure of COCs to heat shock (41°C) during maturation reduced cleavage rate and blastocyst formation relative to the control group (38.5°C). Annexin-V binding (V-FITC assay), which is associated with the early apoptotic event of membrane phosphatidylserine turnover, was higher in oocytes exposed to short-term versus long-term heat shock, suggesting that heat-shock-induced apoptosis involves membrane alterations. Similar to heat exposure, oocyte maturation with C2-ceramide had a dose-dependent deleterious effect on the first cleavages and subsequent embryonic development in association with increased annexin-V binding. Blocking endogenous ceramide generation with fumonisin B1, a specific inhibitor of dihydroceramide synthase (i.e. de novo formation), moderated, to some extent, the effects of heat shock on oocyte developmental competence, suggesting that ceramide plays an important role in heat-shock-induced apoptosis.

scholarly journals The Pyridone-Annelated Isoindigo (5‘-Cl) Induces Apoptosis, Dysregulation of Mitochondria and Formation of ROS in Leukemic HL-60 Cells

2015 ◽  
Vol 35 (5) ◽  
pp. 1958-1974 ◽  
Author(s):  
Ayman M. Saleh ◽  
Ahmad Aljada ◽  
Mustafa M. El-Abadelah ◽  
Salim S. Sabri ◽  
Jalal A. Zahra ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Molecular Mechanisms ◽  
Reactive Oxygen ◽  
Pyrrolidine Dithiocarbamate ◽  
Nuclear Staining ◽  
Dependent Manner ◽  
Oxygen Species ◽  
Mitochondrial Functions ◽  
Apoptotic Activity ◽  
Induced Apoptosis

Background/Aims: In our quest to develop an isoindigo with improved efficacy and bioavailability, we recently synthesized a series of novel substituted pyridone-annelated isoindigo and evaluated their antiproliferative effects. We identified the compound [(E)-1-(5'-Chloro-2'-oxoindolin-3'-ylidene)-6-ethyl-2,3,6,9-tetrahydro-2,9-dioxo-1H-pyrrolo[3,2-f] quinoline-8-carboxylic acid], abbreviated as 5'-Cl, which shows selective antiproliferative activities against various cancer cell lines mediated through apoptosis. Here we have investigated the molecular mechanisms underlying the apoptotic activity of 5'-Cl in the human promyelocytic leukemia HL-60 cells. Methods: We employed different methods to determine the apoptotic pathways triggered by 5'-Cl in HL-60 cells, using flow cytometry, nuclear staining, caspases activation, mitochondria functioning, generation of reactive oxygen species (ROS) and Western blotting techniques. Results: Low concentrations (1-8 µM) of 5'-Cl inhibited the growth of HL-60 cells in a dose and time-dependent manner. Cytotoxicity of this compound is found to be mediated by a caspase-dependent apoptosis. Also, there were indications of caspase independent apoptosis as z-VAD-FMK failed to fully rescue the cells from 5‘-Cl-induced apoptosis. In addition, the compound triggered generation of Reactive Oxygen Species (ROS), caused depolarization of the mitochondrial inner membrane, decreased the level of cellular ATP, modulated the expression and phosphorylation of Bcl-2 leading to loss of its association with Bax and increased the release of cytochrome c to the cytosol of treated cells. The effects of 5‘-Cl on mitochondria and apoptosis were substantially blocked in the presence of a combination between z-VAD-FMK and either of the ROS scavenger N-acetyl-L-cysteine (NAC) or pyrrolidine dithiocarbamate (PDTC). Conclusion: We demonstrated that the growth inhibitory effects of 5'-Cl in HL-60 cells involve multiple pathways of apoptosis and dysregulation of mitochondrial functions.

Influence of simulated ischemia on apoptosis induction by oxidative stress in adult cardiomyocytes of rats

2000 ◽  
Vol 278 (1) ◽  
pp. H94-H99 ◽  
Author(s):  
J. Inserte ◽  
G. Taimor ◽  
B. Hofstaetter ◽  
D. Garcia-Dorado ◽  
H. M. Piper
Keyword(s):  
Oxidative Stress ◽  
Ischemia Reperfusion ◽  
Annexin V ◽  
Fluorescein Isothiocyanate ◽  
Adult Rats ◽  
Partial Protection ◽  
Adult Cardiomyocytes ◽  
Propidium Iodide Staining ◽  
Induction Of Apoptosis ◽  
Induced Apoptosis

Oxidative stress may cause apoptosis of cardiomyocytes in ischemic-reperfused myocardium. We investigated whether ischemia-reperfusion modifies the susceptibility of cardiomyocyte induction of apoptosis by oxidative stress. Ischemia was simulated by incubating isolated cardiomyocytes from adult rats in an anoxic, glucose-free medium, pH 6.4, for 3 h. Annexin V-fluorescein isothiocyanate/propidium iodide staining and the detection of DNA laddering were used as apoptotic markers. H2O2(7.5 μmol/l) induced apoptosis in 20.1 ± 1.8% of cells under normoxic conditions but only 14.4 ± 1.6% ( n = 6, P < 0.05) after ischemia-reoxygenation. This partial protection of ischemic-reoxygenated cells was observed despite a reduction in their cellular glutathione content, from 11.4 ± 1.9 in normoxic controls to 2.9 ± 0.8 nmol/mg protein ( n = 3, P < 0.05). Elevation of end-ischemic glutathione contents by pretreatment with 1 mmol/l N-acetylcysteine entirely protected ischemic-reoxygenated cells against induction of apoptosis by H2O2. In conclusion, ischemia-reperfusion can protect cardiomyocytes against induction of apoptosis by exogenous oxidative stress. This endogenous protective effect is most clearly demonstrated when control and postischemic cardiomyocytes are compared at similar glutathione levels.

scholarly journals Sheng Mai San protects H9C2 cells against hyperglycemia-induced apoptosis

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Bing Pang ◽  
Li-Wei Shi ◽  
Li-juan Du ◽  
Yun-Chu Li ◽  
Mei-Zhen Zhang ◽  
...  
Keyword(s):  
Cell Viability ◽  
Molecular Mechanisms ◽  
Annexin V ◽  
Signalling Pathway ◽  
H9c2 Cell ◽  
Protective Effects ◽  
H9c2 Cells ◽  
P53 Expression ◽  
Fasl Gene ◽  
Induced Apoptosis

Abstract Background Sheng Mai San (SMS) has been proven to exhibit cardio-protective effects. This study aimed to explore the molecular mechanisms of SMS on hyperglycaemia (HG)-induced apoptosis in H9C2 cells. Methods HG-induced H9C2 cells were established as the experimental model, and then treated with SMS at 25, 50, and 100 μg/mL. H9C2 cell viability and apoptosis were quantified using MTT and Annexin V-FITC assays, respectively. Furthermore, Bcl-2/Bax signalling pathway protein expression and Fas and FasL gene expression levels were quantified using western blotting and RT-PCR, respectively. Results SMS treatments at 25, 50, 100 μg/mL significantly improved H9C2 cell viability and inhibited H9C2 cell apoptosis (p < 0.05). Compared to the HG group, SMS treatment at 25, 50, and 100 μg/mL significantly downregulated p53 and Bax expression and upregulated Bcl-2 expression (p < 0.05). Moreover, SMS treatment at 100 μg/mL significantly downregulated Fas and FasL expression level (p < 0.05) when compared to the HG group. Conclusion SMS protects H9C2 cells from HG-induced apoptosis probably by downregulating p53 expression and upregulating the Bcl-2/Bax ratio. It may also be associated with the inhibition of the Fas/FasL signalling pathway.

Motexafin Gadolinium Induces Apoptosis in Lymphoid Cell Lines and Demonstrates Enhanced Biological Activity with Akt Kinase Inhibitors.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3406-3406
Author(s):  
Louie Naumovski ◽  
Jason Ramos ◽  
Jun Chen ◽  
Mint Sirisawad ◽  
David Lucas ◽  
...  
Keyword(s):  
Cell Death ◽  
Chronic Lymphocytic Leukemia ◽  
Cell Lines ◽  
Specific Inhibitor ◽  
Annexin V ◽  
Lymphocytic Leukemia ◽  
Jurkat Cells ◽  
Motexafin Gadolinium ◽  
Akt Phosphorylation ◽  
Induced Apoptosis

Abstract Motexafin gadolinium (MGd, Xcytrin®) is a tumor-selective redox mediator that catalytically oxidizes intracellular reducing metabolites and produces reactive oxygen species (ROS). In this report, we demonstrate that MGd induces apoptosis or growth inhibition in several hematopoietic tumor-derived cell lines and tumor cells from patients with chronic lymphocytic leukemia. Lymphoma (HF-1, Ramos, DHL-4, DB, Hut78 and Raji) and leukemia (Jurkat, HL-60) cell lines were cultured in RPMI 1640 media with 10% heat inactivated fetal bovine serum with or without 50 uM MGd. MGd inhibited the growth of 6 of the cell lines (HF-1, Ramos, HL-60, DHL-4, Jurkat and DB) and was cytotoxic to HF-1. ROS were implicated in MGd-induced cell death since their presence was detected by dichlorofluorescein diacetate staining and peroxiredoxin oxidation in MGd treated HF-1 cells that undergo apoptosis, but not in Jurkat cells that do not undergo MGd-induced apoptosis. MGd triggered an apoptotic pathway in HF-1 cells as demonstrated by loss of mitochondrial membrane potential, release of cytochrome c from mitochondria, activation of caspases, cleavage of PARP and annexin-V binding. MGd also induced cell death and activated caspases in vitro in primary chronic lymphocytic leukemia cells. Protein lysates from cultured cell lines (HF-1, Ramos) were subjected to immunoblot analysis to determine caspase cleavage patterns, and the phosphorylation status of Akt, a kinase that regulates survival pathways. In MGd treated HF-1, phospho-Akt protein levels initially increased 2–3 fold between 30 min and 1 hr (n=4) and then decreased to 40–50% of control levels by 24–48 hrs (n=4). The drop in phospho-Akt protein coincided with an increase in apoptotic cell death as indicated by morphology, staining with Annexin-V and activation of caspases. Addition of a specific inhibitor of Akt phosphorylation (SH-5) reduced Akt phosphorylation in MGd treated HF-1 cells by 90% and enhanced the cytotoxic effect of MGd. In Ramos cells, which do not undergo apoptosis when treated with MGd, co-treatment with MGd and SH-5 decreased phospho Akt levels by only 15% and did not result in cytotoxicity. These data point to a potential role for Akt in MGd-induced apoptosis and suggest that MGd activity may be enhanced by inhibition of Akt. These data show that the pro-apoptotic effects of MGd involve caspase activation and provide a rationale to evaluate MGd in the treatment of lymphoma and leukemia patients.

scholarly journals Mink Cell Focus-Forming Murine Leukemia Virus Killing of Mink Cells Involves Apoptosis and Superinfection

2001 ◽  
Vol 75 (13) ◽  
pp. 6007-6015 ◽  
Author(s):  
Fayth K. Yoshimura ◽  
Tao Wang ◽  
Suparna Nanua
Keyword(s):  
Murine Leukemia Virus ◽  
Leukemia Virus ◽  
Annexin V ◽  
Fluorescein Isothiocyanate ◽  
Viral Dna ◽  
Infected Cells ◽  
Mink Cell ◽  
Induced Apoptosis ◽  
Murine Leukemia

ABSTRACT Induction of apoptosis by different types of pathogenic retroviruses is an important step in disease development. We have observed that infection of thymic lymphocytes by the mink cell focus-forming murine leukemia virus (MCF MLV) during the preleukemic period resulted in an enhancement of apoptosis of these cells. To further study the ability of MCF MLVs to induce apoptosis and the role of this process in viral pathogenesis, we have developed an in vitro system of virus-induced apoptosis. MCF13 MLV infection of mink epithelial cells resulted in the production of cytopathic foci. In contrast, infection of mink cells with the 4070A amphotropic MLV did not produce any cytopathic effects. Staining of MCF13 MLV-infected cells with propidium iodide and annexin V-fluorescein isothiocyanate indicated that virus-induced cell death was due to apoptosis. At 6 days postinfection, the percentage of apoptotic MCF13 MLV-infected cells was 27% compared with 2 to 3% for mock- or amphotropic MLV-infected cells, representing a 9- to 14-fold difference. Assays for caspase-3 activation confirmed the detection by flow cytometry of apoptosis of MCF13 MLV-infected cells. Large amounts of unintegrated linear viral DNA were detectable by Southern blot analysis during the acute phase of infection, which indicated that MCF13 MLV is able to superinfect mink cells. Unintegrated viral DNA of only the linear form was detectable in thymic lymphocytes isolated from MCF13 MLV-inoculated mice during the preleukemic period. These results indicated that the ability of MCF13 MLV to induce apoptosis is correlated with its ability to superinfect cells and that this occurs as an early step in thymic lymphoma development.

scholarly journals Regulation of the Activity of p38 Mitogen-Activated Protein Kinase by Akt in Cancer and Adenoviral Protein E1A-Mediated Sensitization to Apoptosis

2003 ◽  
Vol 23 (19) ◽  
pp. 6836-6848 ◽  
Author(s):  
Yong Liao ◽  
Mien-Chie Hung
Keyword(s):  
Molecular Mechanisms ◽  
Specific Inhibitor ◽  
Human Tumor ◽  
Tumor Stage ◽  
Mitogen Activated Protein Kinase ◽  
Necrosis Factor Alpha ◽  
Akt Phosphorylation ◽  
Tumor Tissues ◽  
Mitogen Activated Protein ◽  
Induced Apoptosis

ABSTRACT The adenoviral early region 1A (E1A) protein mediates sensitization to different stimulus-induced apoptosis, such as tumor necrosis factor alpha, UV and gamma irradiation, and different categories of anticancer drugs. However, the molecular mechanisms underlying E1A-mediated sensitization to apoptosis are still not completely defined. Here, we show that E1A-mediated sensitization to apoptosis by the inactivation of a key survival factor Akt and the activation of a pro-apoptotic factor p38. Also, inactivation of Akt by either a specific inhibitor or a genetic knockout of Akt1 results in p38 activation, possibly through the release of the activity of p38 upstream kinases, including ASK1 and MEKK3. In addition, we showed that p38 phosphorylation is downregulated and Akt phosphorylation is upregulated in multiple human tumor tissues, and this correlates with tumor stage in human breast cancer. A deletion mutation of a conserved domain of E1A, which is required for E1A-induced downregulation of Akt activity, disrupts E1A-mediated upregulation of p38 activity and also eliminates E1A-mediated chemosensitization. Thus, activation of p38 and inactivation of Akt may have general implications for tumor suppression and sensitization to apoptosis.

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