Senescence, apoptosis or autophagy? When a damaged cell must decide its path--a mini-review.

PubMed

Vicencio, José Miguel; Galluzzi, Lorenzo; Tajeddine, Nicolas; Ortiz, Carla; Criollo, Alfredo; Tasdemir, Ezgi; Morselli, Eugenia; Ben Younes, Amena; Maiuri, Maria Chiara; Lavandero, Sergio; Kroemer, Guido

2008-01-01

Many features of aging result from the incapacity of cells to adapt to stress conditions. When damage accumulates irreversibly, mitotic cells from renewable tissues rely on either of two mechanisms to avoid replication. They can permanently arrest the cell cycle (cellular senescence) or trigger cell death programs. Apoptosis (self-killing) is the best-described form of programmed cell death, but autophagy (self-eating), which is a lysosomal degradation pathway essential for homeostasis, reportedly contributes to cell death as well. Unlike mitotic cells, postmitotic cells like neurons or cardiomyocytes cannot become senescent since they are already terminally differentiated. The fate of these cells entirely depends on their ability to cope with stress. Autophagy then operates as a major homeostatic mechanism to eliminate damaged organelles, long-lived or aberrant proteins and superfluous portions of the cytoplasm. In this mini-review, we briefly summarize the molecular networks that allow damaged cells either to adapt to stress or to engage in programmed-cell-death pathways. (c) 2008 S. Karger AG, Basel.

Tight regulation between cell survival and programmed cell death in GBM stem-like cells by EGFR/GSK3b/PP2A signaling.

PubMed

Gürsel, Demirkan B; Banu, Matei A; Berry, Nicholas; Marongiu, Roberta; Burkhardt, Jan-Karl; Kobylarz, Keith; Kaplitt, Michael G; Rafii, Shahin; Boockvar, John A

2015-01-01

Malignant gliomas represent one of the most aggressive forms of cancer, displaying high mortality rates and limited treatment options. Specific subpopulations of cells residing in the tumor niche with stem-like characteristics have been postulated to initiate and maintain neoplasticity while resisting conventional therapies. The study presented here aims to define the role of glycogen synthase kinase 3 beta (GSK3b) in patient-derived glioblastoma (GBM) stem-like cell (GSC) proliferation, apoptosis and invasion. To evaluate the potential role of GSK3b in GBM, protein profiles from 68 GBM patients and 20 normal brain samples were analyzed for EGFR-mediated PI3kinase/Akt and GSK3b signaling molecules including protein phosphatase 2A (PP2A). To better understand the function of GSK3b in GBM, GSCs were isolated from GBM patient samples. Blocking GSK3b phosphorylation at Serine 9 attenuated cell proliferation while concomitantly stimulating apoptosis through activation of Caspase-3 in patient-derived GSCs. Increasing GSK3b protein content resulted in the inhibition of cell proliferation, colony formation and stimulated programmed cell death. Depleting GSK3b in GSCs down regulated PP2A. Furthermore, knocking down PP2A or blocking its activity by okadaic acid inactivated GSK3b by increasing GSK3b phosphorylation at Serine 9. Our data suggests that GSK3b may function as a regulator of apoptosis and tumorigenesis in GSCs. Therapeutic approaches targeting GSK3b in glioblastoma stem-like cells may be a useful addition to our current therapeutic armamentarium.

Myeloblastic leukemia cells conditionally blocked by myc-estrogen receptor chimeric transgenes for terminal differentiation coupled to growth arrest and apoptosis.

PubMed

Selvakumaran, M; Liebermann, D; Hoffman-Liebermann, B

1993-05-01

Conditional mutants of the myeloblastic leukemic M1 cell line, expressing the chimeric mycer transgene, have been established. It is shown that M1 mycer cells, like M1, undergo terminal differentiation coupled to growth arrest and programmed cell death (apoptosis) after treatment with the physiologic differentiation inducer interleukin-6. However, when beta-estradiol is included in the culture medium, M1 mycer cells respond to differentiation inducers like M1 myc cell lines, where the differentiation program is blocked at an intermediate stage. By manipulating the function of the mycer transgene product, it is shown that there is a 10-hour window during myeloid differentiation, from 30 to 40 hours after the addition of the differentiation inducer, when the terminal differentiation program switches from being dependent on c-myc suppression to becoming c-myc suppression independent, where activation of c-myc has no apparent effect on mature macrophages. M1 mycer cell lines provide a powerful tool to increase our understanding of the role of c-myc in normal myelopoiesis and in leukemogenesis, also providing a strategy to clone c-myc target genes.

Apoptosis-like programmed cell death induces antisense ribosomal RNA (rRNA) fragmentation and rRNA degradation in Leishmania

PubMed Central

Padmanabhan, P K; Samant, M; Cloutier, S; Simard, M J; Papadopoulou, B

2012-01-01

Few natural antisense (as) RNAs have been reported as yet in the unicellular protozoan Leishmania. Here, we describe that Leishmania produces natural asRNAs complementary to all ribosomal RNA (rRNA) species. Interestingly, we show that drug-induced apoptosis-like programmed cell death triggers fragmentation of asRNA complementary to the large subunit gamma (LSU-γ) rRNA, one of the six 28S rRNA processed fragments in Leishmania. Heat and oxidative stress also induce fragmentation of asrRNA, but to a lesser extent. Extensive asrRNA cleavage correlates with rRNA breakdown and translation inhibition. Indeed, overexpression of asLSU-γ rRNA accelerates rRNA degradation upon induction of apoptosis. In addition, we provide mechanistic insight into the regulation of apoptosis-induced asrRNA fragmentation by a 67 kDa ATP-dependent RNA helicase of the DEAD-box subfamily. This helicase binds both sense (s)LSU-γ and asLSU-γ rRNAs, and appears to have a key role in protecting rRNA from degradation by preventing asrRNA cleavage and thus cell death. Remarkably, the asrRNA fragmentation process operates not only in trypanosomatid protozoa but also in mammals. Our findings uncover a novel mechanism of regulation involving asrRNA fragmentation and rRNA breakdown, that is triggered by apoptosis and conditions of reduced translation under stress, and seems to be evolutionary conserved. PMID:22767185

Caffeine-Induced Premature Chromosome Condensation Results in the Apoptosis-Like Programmed Cell Death in Root Meristems of Vicia faba

PubMed Central

Rybaczek, Dorota; Musiałek, Marcelina Weronika; Balcerczyk, Aneta

2015-01-01

We have demonstrated that the activation of apoptosis-like programmed cell death (AL-PCD) was a secondary result of caffeine (CF) induced premature chromosome condensation (PCC) in hydroxyurea-synchronized Vicia faba root meristem cells. Initiation of the apoptotic-like cell degradation pathway seemed to be the result of DNA damage generated by treatment with hydroxyurea (HU) [double-stranded breaks (DSBs) mostly] and co-treatment with HU/CF [single-stranded breaks (SSBs) mainly]. A single chromosome comet assay was successfully used to study different types of DNA damage (neutral variant–DSBs versus alkaline–DSBs or SSBs). The immunocytochemical detection of H2AXS139Ph and PARP-2 were used as markers for DSBs and SSBs, respectively. Acridine orange and ethidium bromide (AO/EB) were applied for quantitative immunofluorescence measurements of dead, dying and living cells. Apoptotic-type DNA fragmentation and positive TUNEL reaction finally proved that CF triggers AL-PCD in stressed V. faba root meristem cells. In addition, the results obtained under transmission electron microscopy (TEM) further revealed apoptotic-like features at the ultrastructural level of PCC-type cells: (i) extensive vacuolization; (ii) abnormal chromatin condensation, its marginalization and concomitant degradation; (iii) formation of autophagy-like vesicles (iv) protoplast shrinkage (v) fragmentation of cell nuclei and (vi) extensive degeneration of the cells. The results obtained have been discussed with respect to the vacuolar/autolytic type of plant-specific AL-PCD. PMID:26545248

Ras promotes cell survival by antagonizing both JNK and Hid signals in the Drosophila eye.

PubMed

Wu, Yue; Zhuang, Yuan; Han, Min; Xu, Tian; Deng, Kejing

2009-10-20

Programmed cell death, or apoptosis, is a fundamental physiological process during normal development or in pathological conditions. The activation of apoptosis can be elicited by numerous signalling pathways. Ras is known to mediate anti-apoptotic signals by inhibiting Hid activity in the Drosophila eye. Here we report the isolation of a new loss-of-function ras allele, rasKP, which causes excessive apoptosis in the Drosophila eye. This new function is likely to be mediated through the JNK pathway since the inhibition of JNK signalling can significantly suppress rasKP-induced apoptosis, whereas the removal of hid only weakly suppresses the phenotype. Furthermore, the reduction of JNK signalling together with the expression of the baculovirus caspase inhibitor p35, which blocks Hid activity, strongly suppresses the rasKP cell death. In addition, we find a strong correlation between rasKP-induced apoptosis in the eye disc and the activation of JNK signalling. In the Drosophila eye, Ras may protect cells from apoptosis by inhibiting both JNK and Hid activities. Surprisingly, reducing Ras activity in the wing, however, does not cause apoptosis but rather affects cell and organ size. Thus, in addition to its requirement for cell viability, Ras appears to mediate different biological roles depending on the developmental context and on the level of its expression.

Reactive oxygen species are required for zoledronic acid-induced apoptosis in osteoclast precursors and mature osteoclast-like cells

PubMed Central

Tai, Ta-Wei; Chen, Ching-Yu; Su, Fong-Chin; Tu, Yuan-Kun; Tsai, Tsung-Ting; Lin, Chiou-Feng; Jou, I.-Ming

2017-01-01

Inhibiting osteoclasts and osteoclast precursors to reduce bone resorption is an important strategy to treat osteoclast-related diseases, such as osteoporosis, inflammatory bone loss, and malignant bone metastasis. However, the mechanism by which apoptosis is induced in the osteoclasts and their precursors are not completely understood. Here, we used nitrogen-containing bisphosphonate zoledronic acid (ZA) to induce cell apoptosis in human and murine osteoclast precursors and mature osteoclast-like cells. Caspase-3-mediated cell apoptosis occurred following the ZA (100 μM) treatment. Reactive oxygen species (ROS) were also generated in a time-dependent manner. Following knock-down of the p47phox expression, which is required for ROS activation, or co-treatment with the ROS inhibitor, N-acetyl-L-cysteine, ZA-induced apoptosis was significantly suppressed in both osteoclast precursors and mature osteoclast-like cells. The ROS-activated mitogen-activated protein kinases pathways did not trigger cell apoptosis. However, a ROS-regulated Mcl-1 decrease simultaneously with glycogen synthase kinase (GSK)-3β promoted cell apoptosis. These findings show that ZA induces apoptosis in osteoclast precursors and mature osteoclast-like cells by triggering ROS- and GSK-3β-mediated Mcl-1 down-regulation. PMID:28281643

Diabetes and renal tubular cell apoptosis

PubMed Central

Habib, Samy L

2013-01-01

Apoptosis contributes to the development of diabetic nephropathy, but the mechanism by which high glucose induces apoptosis is not fully understood. Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Hyperglycemia and high glucose in vitro also lead to apoptosis, a form of programmed cell death. High glucose similar to those seen with hyperglycemia in people with diabetes mellitus, lead to accelerated apoptosis, a form of programmed cell death characterized by cell shrinkage, chromatin condensation and DNA fragmentation, in variety of cell types, including renal proximal tubular epithelial cells. PMID:23593533

Diabetes and renal tubular cell apoptosis.

PubMed

Habib, Samy L

2013-04-15

Apoptosis contributes to the development of diabetic nephropathy, but the mechanism by which high glucose induces apoptosis is not fully understood. Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Hyperglycemia and high glucose in vitro also lead to apoptosis, a form of programmed cell death. High glucose similar to those seen with hyperglycemia in people with diabetes mellitus, lead to accelerated apoptosis, a form of programmed cell death characterized by cell shrinkage, chromatin condensation and DNA fragmentation, in variety of cell types, including renal proximal tubular epithelial cells.

Plasma membrane changes during programmed cell deaths

PubMed Central

Zhang, Yingying; Chen, Xin; Gueydan, Cyril; Han, Jiahuai

2018-01-01

Ruptured and intact plasma membranes are classically considered as hallmarks of necrotic and apoptotic cell death, respectively. As such, apoptosis is usually considered a non-inflammatory process while necrosis triggers inflammation. Recent studies on necroptosis and pyroptosis, two types of programmed necrosis, revealed that plasma membrane rupture is mediated by MLKL channels during necroptosis but depends on non-selective gasdermin D (GSDMD) pores during pyroptosis. Importantly, the morphology of dying cells executed by MLKL channels can be distinguished from that executed by GSDMD pores. Interestingly, it was found recently that secondary necrosis of apoptotic cells, a previously believed non-regulated form of cell lysis that occurs after apoptosis, can be programmed and executed by plasma membrane pore formation like that of pyroptosis. In addition, pyroptosis is associated with pyroptotic bodies, which have some similarities to apoptotic bodies. Therefore, different cell death programs induce distinctive reshuffling processes of the plasma membrane. Given the fact that the nature of released intracellular contents plays a crucial role in dying/dead cell-induced immunogenicity, not only membrane rupture or integrity but also the nature of plasma membrane breakdown would determine the fate of a cell as well as its ability to elicit an immune response. In this review, we will discuss recent advances in the field of apoptosis, necroptosis and pyroptosis, with an emphasis on the mechanisms underlying plasma membrane changes observed on dying cells and their implication in cell death-elicited immunogenicity. PMID:29076500

Pneumolysin Activates Macrophage Lysosomal Membrane Permeabilization and Executes Apoptosis by Distinct Mechanisms without Membrane Pore Formation

PubMed Central

Bewley, Martin A.; Naughton, Michael; Preston, Julie; Mitchell, Andrea; Holmes, Ashleigh; Marriott, Helen M.; Read, Robert C.; Mitchell, Timothy J.; Whyte, Moira K. B.

2014-01-01

ABSTRACT Intracellular killing of Streptococcus pneumoniae is complemented by induction of macrophage apoptosis. Here, we show that the toxin pneumolysin (PLY) contributes both to lysosomal/phagolysosomal membrane permeabilization (LMP), an upstream event programing susceptibility to apoptosis, and to apoptosis execution via a mitochondrial pathway, through distinct mechanisms. PLY is necessary but not sufficient for the maximal induction of LMP and apoptosis. PLY’s ability to induce both LMP and apoptosis is independent of its ability to form cytolytic pores and requires only the first three domains of PLY. LMP involves TLR (Toll-like receptor) but not NLRP3/ASC (nucleotide-binding oligomerization domain [Nod]-like receptor family, pyrin domain-containing protein 3/apoptosis-associated speck-like protein containing a caspase recruitment domain) signaling and is part of a PLY-dependent but phagocytosis-independent host response that includes the production of cytokines, including interleukin-1 beta (IL-1β). LMP involves progressive and selective permeability to 40-kDa but not to 250-kDa fluorescein isothiocyanate (FITC)-labeled dextran, as PLY accumulates in the cytoplasm. In contrast, the PLY-dependent execution of apoptosis requires phagocytosis and is part of a host response to intracellular bacteria that also includes NO generation. In cells challenged with PLY-deficient bacteria, reconstitution of LMP using the lysomotrophic detergent LeuLeuOMe favored cell necrosis whereas PLY reconstituted apoptosis. The results suggest that PLY contributes to macrophage activation and cytokine production but also engages LMP. Following bacterial phagocytosis, PLY triggers apoptosis and prevents macrophage necrosis as a component of a broad-based antimicrobial strategy. This illustrates how a key virulence factor can become the focus of a multilayered and coordinated innate response by macrophages, optimizing pathogen clearance and limiting inflammation. PMID:25293758

Apoptosis-Like Death, an Extreme SOS Response in Escherichia coli

PubMed Central

Erental, Ariel; Kalderon, Ziva; Saada, Ann; Smith, Yoav

2014-01-01

ABSTRACT In bacteria, SOS is a global response to DNA damage, mediated by the recA-lexA genes, resulting in cell cycle arrest, DNA repair, and mutagenesis. Previously, we reported that Escherichia coli responds to DNA damage via another recA-lexA-mediated pathway resulting in programmed cell death (PCD). We called it apoptosis-like death (ALD) because it is characterized by membrane depolarization and DNA fragmentation, which are hallmarks of eukaryotic mitochondrial apoptosis. Here, we show that ALD is an extreme SOS response that occurs only under conditions of severe DNA damage. Furthermore, we found that ALD is characterized by additional hallmarks of eukaryotic mitochondrial apoptosis, including (i) rRNA degradation by the endoribonuclease YbeY, (ii) upregulation of a unique set of genes that we called extensive-damage-induced (Edin) genes, (iii) a decrease in the activities of complexes I and II of the electron transport chain, and (iv) the formation of high levels of OH˙ through the Fenton reaction, eventually resulting in cell death. Our genetic and molecular studies on ALD provide additional insight for the evolution of mitochondria and the apoptotic pathway in eukaryotes. PMID:25028428

Amphotericin B induces apoptosis-like programmed cell death in Naegleria fowleri and Naegleria gruberi.

PubMed

Cárdenas-Zúñiga, Roberto; Silva-Olivares, Angélica; Villalba-Magdaleno, José D' Artagnan; Sánchez-Monroy, Virginia; Serrano-Luna, Jesús; Shibayama, Mineko

2017-07-01

Naegleria fowleri and Naegleria gruberi belong to the free-living amoebae group. It is widely known that the non-pathogenic species N. gruberi is usually employed as a model to describe molecular pathways in this genus, mainly because its genome has been recently described. However, N. fowleri is an aetiological agent of primary amoebic meningoencephalitis, an acute and fatal disease. Currently, the most widely used drug for its treatment is amphotericin B (AmB). It was previously reported that AmB has an amoebicidal effect in both N. fowleri and N. gruberi trophozoites by inducing morphological changes that resemble programmed cell death (PCD). PCD is a mechanism that activates morphological, biochemical and genetic changes. However, PCD has not yet been characterized in the genus Naegleria. The aim of the present work was to evaluate the typical markers to describe PCD in both amoebae. These results showed that treated trophozoites displayed several parameters of apoptosis-like PCD in both species. We observed ultrastructural changes, an increase in reactive oxygen species, phosphatidylserine externalization and a decrease in intracellular potassium, while DNA degradation was evaluated using the TUNEL assay and agarose gels, and all of these parameters are related to PCD. Finally, we analysed the expression of apoptosis-related genes, such as sir2 and atg8, in N. gruberi. Taken together, our results showed that AmB induces the morphological, biochemical and genetic changes of apoptosis-like PCD in the genus Naegleria.

The effects of selected drugs and dietary compounds on proliferation and apoptosis in colorectal carcinoma.

PubMed

Kiedrowski, Miroslaw; Mroz, Andrzej

2014-01-01

Like many malignancies, the development of colorectal carcinoma (CRC) can be considered as an imbalance between the compromised process of programmed cell death (apoptosis) and excessive, uncontrolled proliferation. Several mutations and epigenetic alterations are acquired during colorectal carcinogenesis. These are responsible for the cell cycle regulation, cellular sensitivity to pro- and antiapoptotic factors, cell proliferation, angiogenesis, invasiveness, as well as metastatic potential. The molecular alterations, along with their morphological expressions, have been recognised in detail, and most of the CRC cases can be attributed to either adenoma-carcinoma or serrated neoplasia pathways: in the first, the antiapoptotic features prevail; while in the second, the proliferative activity is of the utmost importance. The aim of the work is to discuss the influence of selected drugs and dietary compounds on the proliferation and apoptosis in CRC.

High-density lipoproteins protect endothelial cells from tumor necrosis factor-alpha-induced apoptosis.

PubMed

Sugano, M; Tsuchida, K; Makino, N

2000-06-16

High-density lipoproteins (HDL) levels have been shown to be inversely correlated with coronary heart disease, but the mechanisms of the direct protective effect of HDL on endothelial cells are not fully understood. The apoptosis of endothelial cells induced by cytokines and/or oxidized low-density lipoproteins, etc. may provide a mechanistic clue to the "response-to-injury" hypothesis of atherogenesis. Here we report that HDL prevent the apoptosis of human umbilical venous endothelial cells (HUVECs) induced by tumor necrosis factor-alpha (TNF-alpha) via an inhibition of CPP32-like protease activity. The incubation of HUVECs with TNF-alpha significantly increased the CPP32-like protease activity, and induced apoptosis. Preincubation of HUVECs with HDL before incubation with TNF-alpha significantly suppressed the increase in the CPP32-like protease activity, preventing apoptosis in a concentration-dependent manner. These results suggest that HDL prevent the suicide pathway leading to apoptosis of endothelial cells by decreasing the CPP32-like protease activity and that HDL thus play a protective role against the "response-to-injury" hypothesis of atherogenesis. Copyright 2000 Academic Press.

RIG-I-like helicases induce immunogenic cell death of pancreatic cancer cells and sensitize tumors toward killing by CD8+ T cells

PubMed Central

Duewell, P; Steger, A; Lohr, H; Bourhis, H; Hoelz, H; Kirchleitner, S V; Stieg, M R; Grassmann, S; Kobold, S; Siveke, J T; Endres, S; Schnurr, M

2014-01-01

Pancreatic cancer is characterized by a microenvironment suppressing immune responses. RIG-I-like helicases (RLH) are immunoreceptors for viral RNA that induce an antiviral response program via the production of type I interferons (IFN) and apoptosis in susceptible cells. We recently identified RLH as therapeutic targets of pancreatic cancer for counteracting immunosuppressive mechanisms and apoptosis induction. Here, we investigated immunogenic consequences of RLH-induced tumor cell death. Treatment of murine pancreatic cancer cell lines with RLH ligands induced production of type I IFN and proinflammatory cytokines. In addition, tumor cells died via intrinsic apoptosis and displayed features of immunogenic cell death, such as release of HMGB1 and translocation of calreticulin to the outer cell membrane. RLH-activated tumor cells led to activation of dendritic cells (DCs), which was mediated by tumor-derived type I IFN, whereas TLR, RAGE or inflammasome signaling was dispensable. Importantly, CD8α+ DCs effectively engulfed apoptotic tumor material and cross-presented tumor-associated antigen to naive CD8+ T cells. In comparison, tumor cell death mediated by oxaliplatin, staurosporine or mechanical disruption failed to induce DC activation and antigen presentation. Tumor cells treated with sublethal doses of RLH ligands upregulated Fas and MHC-I expression and were effectively sensitized towards Fas-mediated apoptosis and cytotoxic T lymphocyte (CTL)-mediated lysis. Vaccination of mice with RLH-activated tumor cells induced protective antitumor immunity in vivo. In addition, MDA5-based immunotherapy led to effective tumor control of established pancreatic tumors. In summary, RLH ligands induce a highly immunogenic form of tumor cell death linking innate and adaptive immunity. PMID:25012502

The role of necroptosis in pulmonary diseases.

PubMed

Mizumura, Kenji; Maruoka, Shuichiro; Gon, Yasuhiro; Choi, Augustine M K; Hashimoto, Shu

2016-11-01

By regulating the cell number and eliminating harmful cells, programmed cell death plays a critical role in development, homeostasis, and disease. While apoptosis is a recognized form of programmed cell death, necrosis was considered a type of uncontrolled cell death induced by extreme physical or chemical stress. However, recent studies have revealed the existence of a genetically programmed and regulated form of necrosis, termed necroptosis. Necroptosis is defined as necrotic cell death that is dependent on receptor-interacting protein kinase 3 (RIPK3). RIPK3, receptor-interacting protein kinase 1 (RIPK1), and a mixed-lineage kinase domain-like protein (MLKL) form a multiprotein complex called a necrosome. Although necroptosis generally provides a cell-autonomous host defense, on the other hand, cell rupture caused by necroptosis induces inflammation through the release of damage-associated molecular patterns, such as mitochondrial DNA, HMGB1, and IL-1. Previously, necroptosis was considered an alternative to apoptosis, but it is becoming increasingly clear that necroptosis itself is relevant to clinical disease, independent of apoptosis. According to some recent studies, autophagy, a cellular process for organelle and protein turnover, regulates necroptosis. This review outlines the principal components of necroptosis and provides an overview of the emerging importance of necroptosis in the pathogenesis of pulmonary disease, including chronic obstructive pulmonary disease, lung cancer, infection, and sepsis. We also discuss the molecular relationship between necroptosis and autophagy. Strategies targeting necroptosis may yield novel therapies for pulmonary diseases. Copyright © 2016 The Japanese Respiratory Society. Published by Elsevier B.V. All rights reserved.

Harnessing the apoptotic programs in cancer stem-like cells

PubMed Central

Wang, Ying-Hua; Scadden, David T

2015-01-01

Elimination of malignant cells is an unmet challenge for most human cancer types even with therapies targeting specific driver mutations. Therefore, a multi-pronged strategy to alter cancer cell biology on multiple levels is increasingly recognized as essential for cancer cure. One such aspect of cancer cell biology is the relative apoptosis resistance of tumor-initiating cells. Here, we provide an overview of the mechanisms affecting the apoptotic process in tumor cells emphasizing the differences in the tumor-initiating or stem-like cells of cancer. Further, we summarize efforts to exploit these differences to design therapies targeting that important cancer cell population. PMID:26253117

Apoptosis-like death, an extreme SOS response in Escherichia coli.

PubMed

Erental, Ariel; Kalderon, Ziva; Saada, Ann; Smith, Yoav; Engelberg-Kulka, Hanna

2014-07-15

In bacteria, SOS is a global response to DNA damage, mediated by the recA-lexA genes, resulting in cell cycle arrest, DNA repair, and mutagenesis. Previously, we reported that Escherichia coli responds to DNA damage via another recA-lexA-mediated pathway resulting in programmed cell death (PCD). We called it apoptosis-like death (ALD) because it is characterized by membrane depolarization and DNA fragmentation, which are hallmarks of eukaryotic mitochondrial apoptosis. Here, we show that ALD is an extreme SOS response that occurs only under conditions of severe DNA damage. Furthermore, we found that ALD is characterized by additional hallmarks of eukaryotic mitochondrial apoptosis, including (i) rRNA degradation by the endoribonuclease YbeY, (ii) upregulation of a unique set of genes that we called extensive-damage-induced (Edin) genes, (iii) a decrease in the activities of complexes I and II of the electron transport chain, and (iv) the formation of high levels of OH˙ through the Fenton reaction, eventually resulting in cell death. Our genetic and molecular studies on ALD provide additional insight for the evolution of mitochondria and the apoptotic pathway in eukaryotes. Importance: The SOS response is the first described and the most studied bacterial response to DNA damage. It is mediated by a set of two genes, recA-lexA, and it results in DNA repair and thereby in the survival of the bacterial culture. We have shown that Escherichia coli responds to DNA damage by an additional recA-lexA-mediated pathway resulting in an apoptosis-like death (ALD). Apoptosis is a mode of cell death that has previously been reported only in eukaryotes. We found that E. coli ALD is characterized by several hallmarks of eukaryotic mitochondrial apoptosis. Altogether, our results revealed that recA-lexA is a DNA damage response coordinator that permits two opposite responses: life, mediated by the SOS, and death, mediated by the ALD. The choice seems to be a function of the degree of DNA damage in the cell. Copyright © 2014 Erental et al.

Bcl-2 Blocks a Caspase-Dependent Pathway of Apoptosis Activated by Herpes Simplex Virus 1 Infection in HEp-2 Cells

PubMed Central

Galvan, Veronica; Brandimarti, Renato; Munger, Joshua; Roizman, Bernard

2000-01-01

Earlier reports have shown that herpes simplex virus 1 (HSV-1) mutants induce programmed cell death and that wild-type virus blocks the execution of the cell death program triggered by expression of viral genes, by the Fas and tumor necrosis factor pathways, or by nonspecific stress agents. In particular, an earlier report from this laboratory showed that the mutant virus d120 lacking the genes encoding infected cell protein 4 (ICP4), the major regulatory protein of the virus, induces a caspase-3-independent pathway of apoptosis in human SK-N-SH cells. Here we report that the pathway of apoptosis induced by the d120 mutant in human HEp-2 cells is caspase dependent. Specifically, in HEp-2 cells infected with d120, (i) a broad-range inhibitor of caspase activity, z-vad-FMK, efficiently blocked DNA fragmentation, (ii) cytochrome c was released into the cytoplasm, (iii) caspase-3 was activated inasmuch as poly(ADP-ribose) polymerase was cleaved, and (iv) chromatin condensation and fragmentation of cellular DNA were observed. In parallel studies, HEp-2 cells were transfected with a plasmid encoding human Bcl-2 and a clone (VAX-3) expressing high levels of Bcl-2 was selected. This report shows that Bcl-2 blocked all of the manifestations associated with programmed cell death caused by infection with the d120 mutant. Consistent with their resistance to programmed cell death, VAX-3 cells overproduced infected cell protein 0 (ICP0). An unexpected observation was that ICP0 encoded by the d120 mutant accumulated late in infection in small, quasi-uniform vesicle-like structures in all cell lines tested. Immunofluorescence-based colocalization studies indicated that these structures were not mitochondria or components of the endoplasmic reticulum or the late endosomal compartment. These studies affirm the conclusion that HSV can induce programmed cell death at multiple steps in the course of its replication, that the d120 mutant can induce both caspase-dependent and -independent pathways of programmed cell death, and that virus-induced stimuli of programmed cell death may differ with respect to the pathway that they activate. PMID:10644366

Somatostatin protects photoreceptor cells against high glucose-induced apoptosis.

PubMed

Arroba, Ana I; Mazzeo, Aurora; Cazzoni, Daniele; Beltramo, Elena; Hernández, Cristina; Porta, Massimo; Simó, Rafael; Valverde, Ángela M

2016-01-01

Many cellular and molecular studies in experimental animals and early retinal function tests in patients with diabetic retinopathy (DR) have shown that retinal neurodegeneration is an early event in the pathogenesis of the disease. Somatostatin (SST) is one of the most important neuroprotective factors synthesized by the retina: SST levels are decreased in parallel to retinal neurodegeneration in early stages of DR. In this study, we characterized the induction of apoptosis (programmed cell death) in a 661W photoreceptor-like cell line cultured under high glucose (HG) conditions and the effect of SST. A 661W photoreceptor-like cell line and retinal explants from 10-week-old male C57BL/6 mice were cultured under HG conditions and treated with SST. Hyperglycemia significantly reduced the cellular viability by increasing the percentage of apoptotic cells, and this effect was ameliorated by SST (p˂0.05). Activation of caspase-8 by hyperglycemia was found in the 661W cells and retinal explants and decreased in the presence of SST (p˂0.05). Moreover, we detected activation of calpain-2 associated with hyperglycemia-induced cell death, as well as increased protein tyrosine phosphatase 1B (PTP1B) protein levels; both had a pattern of cleavage that was absent in the presence of SST (p˂0.05). Treatment of the 661W cells and retinal explants with SST for 24 h increased the phosphorylation of type 1 insulin-like growth factor receptor (IGF-IR; tyrosine 1165/1166) and protein kinase B (Akt; serine 473), suggesting this survival signaling is activated in the neuroretina by SST (p˂0.05). This study has provided new mechanistic insights first into the involvement of calpain-2 and PTP1B in the loss of cell survival and increased caspase-8-dependent apoptosis induced by hyperglycemia in photoreceptor cells and second, on the protective effect of SST against apoptosis by the enhancement of IGF-IR-mediated Akt phosphorylation.

Somatostatin protects photoreceptor cells against high glucose–induced apoptosis

PubMed Central

Mazzeo, Aurora; Cazzoni, Daniele; Beltramo, Elena; Hernández, Cristina; Porta, Massimo; Simó, Rafael; Valverde, Ángela M.

2016-01-01

Purpose Many cellular and molecular studies in experimental animals and early retinal function tests in patients with diabetic retinopathy (DR) have shown that retinal neurodegeneration is an early event in the pathogenesis of the disease. Somatostatin (SST) is one of the most important neuroprotective factors synthesized by the retina: SST levels are decreased in parallel to retinal neurodegeneration in early stages of DR. In this study, we characterized the induction of apoptosis (programmed cell death) in a 661W photoreceptor-like cell line cultured under high glucose (HG) conditions and the effect of SST. Methods A 661W photoreceptor-like cell line and retinal explants from 10-week-old male C57BL/6 mice were cultured under HG conditions and treated with SST. Results Hyperglycemia significantly reduced the cellular viability by increasing the percentage of apoptotic cells, and this effect was ameliorated by SST (p˂0.05). Activation of caspase-8 by hyperglycemia was found in the 661W cells and retinal explants and decreased in the presence of SST (p˂0.05). Moreover, we detected activation of calpain-2 associated with hyperglycemia-induced cell death, as well as increased protein tyrosine phosphatase 1B (PTP1B) protein levels; both had a pattern of cleavage that was absent in the presence of SST (p˂0.05). Treatment of the 661W cells and retinal explants with SST for 24 h increased the phosphorylation of type 1 insulin-like growth factor receptor (IGF-IR; tyrosine 1165/1166) and protein kinase B (Akt; serine 473), suggesting this survival signaling is activated in the neuroretina by SST (p˂0.05). Conclusions This study has provided new mechanistic insights first into the involvement of calpain-2 and PTP1B in the loss of cell survival and increased caspase-8-dependent apoptosis induced by hyperglycemia in photoreceptor cells and second, on the protective effect of SST against apoptosis by the enhancement of IGF-IR-mediated Akt phosphorylation. PMID:28050125

Toll-like receptor 4 promotes proliferation and apoptosis resistance in human papillomavirus-related cervical cancer cells through the Toll-like receptor 4/nuclear factor-κB pathway.

PubMed

Jiang, Ninghong; Xie, Feng; Guo, Qisang; Li, Ming-Qing; Xiao, Jingjing; Sui, Long

2017-06-01

Toll-like receptor 4 is overexpressed in various tumors, including cervical carcinoma. However, the role of Toll-like receptor 4 in cervical cancer remains controversial, and the underlying mechanisms are largely elusive. Therefore, Toll-like receptor 4 in cervical cancer and related mechanisms were investigated in this study. Quantitative reverse transcription polymerase chain reaction and western blot analyses were used to detect messenger RNA and protein levels in HeLa, Caski, and C33A cells with different treatments. Proliferation was quantified using Cell Counting Kit-8. Cell cycle distribution and apoptosis were assessed by flow cytometry. Higher levels of Toll-like receptor 4 expression were found in human papillomavirus-positive cells compared to human papillomavirus-negative cells. Proliferation of HeLa and Caski cells was promoted in lipopolysaccharide-stimulated groups but suppressed in short hairpin RNA-transfected groups. Apoptosis rates were lower in lipopolysaccharide-stimulated groups relative to short hairpin RNA-transfected groups. In addition, G2-phase distribution was enhanced when Toll-like receptor 4 was downregulated. Moreover, the pNF-κBp65 level was positively correlated with the Toll-like receptor 4 level in HeLa and Caski cells, though when an nuclear factor-κB inhibitor was applied to lipopolysaccharide-stimulated groups, the patterns of proliferation and apoptosis were opposite to those of the lipopolysaccharide-stimulated groups without inhibitor treatment. In conclusion, these data suggest that Toll-like receptor 4 promotes proliferation and apoptosis resistance in human papillomavirus-related cervical cancer cells at least in part through the Toll-like receptor 4/nuclear factor-κB pathway, which may be correlated with the occurrence and development of cervical carcinoma.

Cell proliferation and apoptosis during histogenesis of the guinea pig and rabbit cerebellar cortex.

PubMed

Lossi, Laura; Coli, Alessandra; Giannessi, Elisabetta; Stornelli, Maria Rita; Marroni, Paolo

2002-01-01

Cell proliferation and apoptosis are essential for development of the nervous system. In this study we have investigated the histogenesis of the cerebellar cortex in guinea pig (a precocial species) and rabbit (an altricial species) at different stages of pregnancy and postnatal life. Proliferating cells were identified after labeling with antibodies against the proliferating cell nuclear antigen (PCNA) and/or the Ki-67 antigen. Apoptotic cells were visualized in situ by the TUNEL method and by immunodetection of cleaved caspase 3 and 9. In guinea pigs, both proliferating and apoptotic cells were detected during pre-natal life (E0-E40). Conversely, cell proliferation and apoptosis in rabbits were temporally restricted to early postnatal weeks (P0-P20). In both species cell proliferation was mainly linked to differentiation and migration of the granule cells. In both species, the majority of cells undergoing programmed cell death likely corresponded to granule cells. They were mainly detected in the external granular layer, and were by far more common than previously reported in other locations of the postnatal brain. This study shows that apoptosis is a shared process of cell death during cerebellar development in both altricial and precocial animals, and that there is a direct spatial and temporal correlation between cell proliferation and death in two mammals with different time tables in cerebellar maturation.

The Caenorhabditis elegans mucolipin-like gene cup-5 is essential for viability and regulates lysosomes in multiple cell types.

PubMed

Hersh, Bradley M; Hartwieg, Erika; Horvitz, H Robert

2002-04-02

The misregulation of programmed cell death, or apoptosis, contributes to the pathogenesis of many diseases. We used Nomarski microscopy to screen for mutants containing refractile cell corpses in a C. elegans strain in which all programmed cell death is blocked and such corpses are absent. We isolated a mutant strain that accumulates refractile bodies resembling irregular cell corpses. We rescued this mutant phenotype with the C. elegans mucolipidosis type IV (ML-IV) homolog, the recently identified cup-5 (coelomocyte-uptake defective) gene. ML-IV is a human autosomal recessive lysosomal storage disease characterized by psychomotor retardation and ophthalmological abnormalities. Our null mutations in cup-5 cause maternal-effect lethality. In addition, cup-5 mutants contain excess lysosomes in many and possibly all cell types and contain lamellar structures similar to those observed in ML-IV cell lines. The human ML-IV gene is capable of rescuing both the maternal-effect lethality and the lysosome-accumulation abnormality of cup-5 mutants. cup-5 mutants seem to contain excess apoptotic cells as detected by staining with terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling. We suggest that the increased apoptosis seen in cup-5 mutants is a secondary consequence of the lysosomal defect, and that abnormalities in apoptosis may be associated with human lysosomal storage disorders.

THE PROS AND CONS OF APOPTOSIS ASSAYS FOR USE IN THE STUDY OF CELLS, TISSUES AND ORGANS

EPA Science Inventory

Abstract
Programmed cell death or apoptosis occurs in many tissues during normal development and in the normal homeostasis of adult tissues. Apoptosis also plays a significant role in abnormal development and disease. Increased interest in apoptosis and cell death in general...

Adenosine decreases oxidative stress and protects H2O2-treated neural stem cells against apoptosis through decreasing Mst1 expression.

PubMed

Gholinejad, Masoumeh; Jafari Anarkooli, Iraj; Taromchi, Amirhossein; Abdanipour, Alireza

2018-05-01

Overproduction of free radicals during oxidative stress induces damage to key biomolecules and activates programed cell death pathways. Neuronal cell death in the nervous system leads to a number of neurodegenerative diseases. The aim of the present study was to evaluate the neuroprotective effect of adenosine on inhibition of apoptosis induced by hydrogen peroxide (H 2 O 2 ) in bone marrow-derived neural stem cells (B-dNSCs), with focus on its regulatory effect on the expression of mammalian sterile 20-like kinase 1 ( Mst1 ), as a novel proapoptotic kinase. B-dNSCs were exposed to adenosine at different doses (2, 4, 6, 8 and 10 µM) for 48 h followed by 125 µM H 2 O 2 for 30 min. Using MTT, terminal deoxynucleotidyl transferase dUTP nick-end labeling and real-time reverse transcription polymerase chain reaction assays, the effects of adenosine on cell survival, apoptosis and Mst1 , nuclear factor (erythroid-derived 2)-like 2 and B-cell lymphoma 2 and adenosine A1 receptor expression were evaluated in pretreated B-dNSCs compared with controls (cells treated with H 2 O 2 only). Firstly, results of the MTT assay indicated 6 µM adenosine to be the most protective dose in terms of promotion of cell viability. Subsequent assays using this dosage indicated that apoptosis rate and Mst1 expression in B-dNSCs pretreated with 6 µM adenosine were significantly decreased compared with the control group. These findings suggest that adenosine protects B-dNSCs against oxidative stress-induced cell death, and therefore, that it may be used to promote the survival rate of B-dNSCs and as a candidate for the treatment of oxidative stress-mediated neurological diseases.

Apoptosis and Vocal Fold Disease: Clinically Relevant Implications of Epithelial Cell Death

ERIC Educational Resources Information Center

Novaleski, Carolyn K.; Carter, Bruce D.; Sivasankar, M. Preeti; Ridner, Sheila H.; Dietrich, Mary S.; Rousseau, Bernard

2017-01-01

Purpose: Vocal fold diseases affecting the epithelium have a detrimental impact on vocal function. This review article provides an overview of apoptosis, the most commonly studied type of programmed cell death. Because apoptosis can damage epithelial cells, this article examines the implications of apoptosis on diseases affecting the vocal fold…

Apoptotic signals induce specific degradation of ribosomal RNA in yeast

PubMed Central

Mroczek, Seweryn; Kufel, Joanna

2008-01-01

Organisms exposed to reactive oxygen species, generated endogenously during respiration or by environmental conditions, undergo oxidative stress. Stress response can either repair the damage or activate one of the programmed cell death (PCD) mechanisms, for example apoptosis, and finally end in cell death. One striking characteristic, which accompanies apoptosis in both vertebrates and yeast, is a fragmentation of cellular DNA and mammalian apoptosis is often associated with degradation of different RNAs. We show that in yeast exposed to stimuli known to induce apoptosis, such as hydrogen peroxide, acetic acid, hyperosmotic stress and ageing, two large subunit ribosomal RNAs, 25S and 5.8S, became extensively degraded with accumulation of specific intermediates that differ slightly depending on cell death conditions. This process is most likely endonucleolytic, is correlated with stress response, and depends on the mitochondrial respiratory status: rRNA is less susceptible to degradation in respiring cells with functional defence against oxidative stress. In addition, RNA fragmentation is independent of two yeast apoptotic factors, metacaspase Yca1 and apoptosis-inducing factor Aif1, but it relies on the apoptotic chromatin condensation induced by histone H2B modifications. These data describe a novel phenotype for certain stress- and ageing-related PCD pathways in yeast. PMID:18385160

Apoptosis in lung injury and remodeling.

PubMed

Li, Xiaopeng; Shu, Ruijie; Filippatos, Gerasimos; Uhal, Bruce D

2004-10-01

The mode of cell death termed apoptosis, sometimes referred to as programmed cell death, is as critical a determinant of cell population size as is cell proliferation. Although best characterized in cells of the immune system, apoptosis is now known to be a key factor in the maintenance of normal cell turnover within structural cells in the parenchyma of virtually every organ. Recent interest in apoptosis in the lung has sparked a surge of investigations designed to determine the roles of apoptosis in lung development, injury, and remodeling. Of particular recent interest are the roles of apoptosis in disease pathogenesis and resolution, in which the concept of apoptosis as a "programmed" cell death, i.e., genetically determined, is often more accurately viewed as "inappropriate cell suicide" with regard to its extent and/or timing. Data accumulating over the past decade have made clear the complexity of the control of lung cell apoptosis; concepts of the regulation of apoptosis originally determined in classical cell culture models are often, but not always, applicable to structural cells. For this reason, each of the many cell types of the lung must be studied as a potentially new subject with its own idiosyncrasies yet to be discovered. In light of the large volume of literature now available, this article focuses on the roles of apoptosis in three pathophysiological contexts: acute respiratory distress syndrome, chronic obstructive pulmonary disease, and pulmonary fibrosis. Each section presents key data describing the evidence for apoptosis in the lung, its possible relevance to disease pathogenesis, and proposed mechanisms that might suggest potential avenues for therapeutic intervention.

Smac mimetics and type II interferon synergistically induce necroptosis in various cancer cell lines.

PubMed

Cekay, Michael John; Roesler, Stefanie; Frank, Tanja; Knuth, Anne-Kathrin; Eckhardt, Ines; Fulda, Simone

2017-12-01

Since cancer cells often evade apoptosis, induction of necroptosis as another mode of programmed cell death is considered a promising therapeutic alternative. Here, we identify a novel synergistic interaction of Smac mimetics that antagonize x-linked Inhibitor of Apoptosis (XIAP), cellular Inhibitor of Apoptosis (cIAP) 1 and 2 with interferon (IFN)γ to induce necroptosis in apoptosis-resistant cancer cells in which caspase activation is blocked. This synergism is confirmed by calculation of combination indices (CIs) and found in both solid and hematological cancer cell lines as well as for different Smac mimetics (i.e. BV6, Birinapant), pointing to a broader relevance. Importantly, individual genetic knockdown of key components of necroptosis signaling, i.e. receptor-interacting protein (RIP) 1, RIP3 or mixed lineage kinase domain-like pseudokinase (MLKL), significantly protects from BV6/IFNγ-induced cell death. Similarly, pharmacological inhibitors of RIP1 (necrostatin-1(Nec-1)), RIP3 (GSK'872) or MLKL (necrosulfonamide (NSA)) significantly reduce BV6/IFNγ-stimulated cell death. Of note, IFN-regulatory factor (IRF)1 is required for BV6/IFNγ-mediated necroptosis, as IRF1 silencing provides protection from cell death. By comparison, antibodies blocking tumor necrosis factor (TNF)α, TNF-related apoptosis-inducing ligand (TRAIL) or CD95 ligand fail to inhibit BV6/IFNγ-induced cell death, pointing to a mechanism independently of death receptor ligands. This is the first report showing that Smac mimetics synergize with IFNγ to trigger necroptosis in apoptosis-resistant cancer cells with important implications for Smac mimetic-based strategies for the treatment of cancer. Copyright © 2017 Elsevier B.V. All rights reserved.

Multiple Modes of Cell Death Discovered in a Prokaryotic (Cyanobacterial) Endosymbiont

PubMed Central

Zheng, Weiwen; Rasmussen, Ulla; Zheng, Siping; Bao, Xiaodong; Chen, Bin; Gao, Yuan; Guan, Xiong; Larsson, John; Bergman, Birgitta

2013-01-01

Programmed cell death (PCD) is a genetically-based cell death mechanism with vital roles in eukaryotes. Although there is limited consensus on similar death mode programs in prokaryotes, emerging evidence suggest that PCD events are operative. Here we present cell death events in a cyanobacterium living endophytically in the fern Azolla microphylla, suggestive of PCD. This symbiosis is characterized by some unique traits such as a synchronized development, a vertical transfer of the cyanobacterium between plant generations, and a highly eroding cyanobacterial genome. A combination of methods was used to identify cell death modes in the cyanobacterium. Light- and electron microscopy analyses showed that the proportion of cells undergoing cell death peaked at 53.6% (average 20%) of the total cell population, depending on the cell type and host developmental stage. Biochemical markers used for early and late programmed cell death events related to apoptosis (Annexin V-EGFP and TUNEL staining assays), together with visualization of cytoskeleton alterations (FITC-phalloidin staining), showed that all cyanobacterial cell categories were affected by cell death. Transmission electron microscopy revealed four modes of cell death: apoptotic-like, autophagic-like, necrotic-like and autolytic-like. Abiotic stresses further enhanced cell death in a dose and time dependent manner. The data also suggest that dynamic changes in the peptidoglycan cell wall layer and in the cytoskeleton distribution patterns may act as markers for the various cell death modes. The presence of a metacaspase homolog (domain p20) further suggests that the death modes are genetically programmed. It is therefore concluded that multiple, likely genetically programmed, cell death modes exist in cyanobacteria, a finding that may be connected with the evolution of cell death in the plant kingdom. PMID:23822984

Apoptosis: a mechanism contributing to remodeling of skeletal muscle in response to hindlimb unweighting

NASA Technical Reports Server (NTRS)

Allen, D. L.; Linderman, J. K.; Roy, R. R.; Bigbee, A. J.; Grindeland, R. E.; Mukku, V.; Edgerton, V. R.

1997-01-01

The role of apoptosis in the elimination of myonuclei during hindlimb unloading-induced atrophy and the inhibition of apoptosis in the prevention of muscle atrophy were examined. The number of nuclei demonstrating double-stranded DNA fragmentation seen by terminal deoxynucleotidyl transferase (TDT) histochemical staining, an indicator of apoptosis, was significantly increased after 14 days of suspension. Double staining with TDT and antilaminin immunohistochemistry revealed that some TDT-positive nuclei were within the fiber lamina and were most likely myonuclei. The number of fibers containing morphologically abnormal nuclei was also significantly greater in suspended compared with control rats. Combined treatment with growth hormone and insulin-like growth factor I (GH/ IGF-I) and resistance exercise attenuated the increase in TDT-positive nuclei (approximately 26%, P > 0.05) and significantly decreased the number of fibers with morphologically abnormal nuclei. The data suggest that 1) "programmed nuclear death" contributes to the elimination of myonuclei and/or satellite cells from atrophying fibers, and 2) GH/IGF-I administration plus muscle loading ameliorates the apoptosis associated with hindlimb unloading.

Correlation of 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced apoptotic cell death in the embryonic vasculature with embryotoxicity

USGS Publications Warehouse

Cantrell, Susannah M.; Joy-Schlezinger, Jennifer; Stegeman, John J.; Tillitt, Donald E.; Hannington, Mark D.

1998-01-01

Vertebrate embryos are particularly sensitive to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Identification of tissues that are susceptible to the adverse effects of TCDD is requisite for understanding the embryo toxic effects of TCDD. The objective of the present study was to quantitate the temporal appearance of and dose dependence of apoptosis in TCDD-exposed medaka embryos (Oryzias latipes). A fluorescent-based DNA end-labeling assay provided a sensitive method for detection of TCDD-induced apoptosis in tissue sections of medaka embryos. Apoptotic cells were readily apparent in the medial yolk vein at all observed embryonic stages in TCDD-exposed embryos. Slope-comparison analysis indicated that TCDD-induced programmed cell death in the embryonic medial yolk vein was mechanistically linked to embryo mortality. These data are consistent with the hypothesis that vascular damage contributes to the acute embryo toxic effects of TCDD. However, as sublethal concentrations of dioxin-like compounds are more typical of environmental exposures, tissue damage was also assessed in medaka fry that were exposed to low doses of TCDD during embryonic development. Cell death was detected in gill and digestive tissues in visibly healthy medaka fry that had been exposed to low doses of TCDD during embryonic development. Increased expression of cytochrome P450 1A is a major biochemical consequence of TCDD exposure and is often used as a biomarker for exposure to dioxin-like compounds. Therefore, we compared the tissue distribution of TCDD-induced P450 1A expression and TCDD-induced programmed cell death. TCDD-induced programmed cell death co-localized with TCDD-induced P450 1A expression in both embryos and in visibly healthy post-hatch fry. Our results suggest that aberrant programmed cell death may be a suitable marker for exposure of feral organisms to dioxin-like compounds.

A new chemotherapy agent-free theranostic system composed of graphene oxide nano-complex and aptamers for treatment of cancer cells.

PubMed

Bahreyni, Amirhossein; Yazdian-Robati, Rezvan; Hashemitabar, Shirin; Ramezani, Mohammad; Ramezani, Pouria; Abnous, Khalil; Taghdisi, Seyed Mohammad

2017-06-30

The common cancer treatment strategies like chemotherapy and radiotherapy are nonspecific and can trigger severe side effects by damaging normal cells. So, targeted cancer therapies, such as apoptosis induction, have attracted great attention in recent years. In this project, two nano-complexes, MUC1 aptamer-NAS-24 aptamer-Graphene oxide (GO) and MUC1 aptamer-Cytochrome C aptamer-GO, were designed to induce cell programmed death in MDA-MB-231 and MCF-7 cells (breast cancer cell lines) and to verify the level of apoptosis in both cell lines. MUC1 aptamer was a molecular recognition probe that led the internalization of two nano-complexes into MDA-MB-231 and MCF-7 cells (MUC1 positive cells) but not into HepG2 cell (liver cancer cell line, MUC1 negative cells). The apoptosis induction relied on binding of NAS-24 aptamer to its target, vimentin, in MDA-MB-231 and MCF-7 (target cells) with different levels of vimentin content. The function of first nano-complex was confirmed by binding of FAM-labeled cytochrome C aptamer to its target (cytochrome C) which was released from mitochondria, based on the function of the first nano-complex. Fluorometric analysis and gel retardation assay proved the formation of nano-complexes. The results of flow cytometry and fluorescence microscopy indicated efficient apoptosis induction just in target cells (MDA-MB-231 and MCF-7 cells) but not in non-target cells (HepG2 cell). The results of MTT assay also confirmed cell death process. Overall, our results proved excellent targeted apoptosis in breast cancer cells by designed nano-complexes which can be applied as an efficient cancer therapy method. Copyright © 2017 Elsevier B.V. All rights reserved.

Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie

A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CMmore » treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration.« less

Yeast as a model to study apoptosis?

PubMed

Fleury, Christophe; Pampin, Mathieu; Tarze, Agathe; Mignotte, Bernard

2002-02-01

Programmed cell death (PCD) serves as a major mechanism for the precise regulation of cell numbers, and as a defense mechanism to remove unwanted and potentially dangerous cells. Despite the striking heterogeneity of cell death induction pathways, the execution of the death program is often associated with characteristic morphological and biochemical changes termed apoptosis. Although for a long time the absence of mitochondrial changes was considered as a hallmark of apoptosis, mitochondria appear today as the central executioner of programmed cell death. This crucial position of mitochondria in programmed cell death control is not due to a simple loss of function (deficit in energy supplying), but rather to an active process in the regulation of effector mechanisms. The large diversity of regulators of apoptosis in mammals and their numerous interactions complicate the analysis of their individual functions. Yeast, eukaryotic but unicellular organism, lack the main regulators of apoptosis (caspases, Bcl-2 family members, ...) found in mammals. This absence render them a powerful tool for heterologous expression, functional studies, and even cloning of new regulators of apoptosis. Great advances have thus been made in our understanding of the molecular mechanisms of Bcl-2 family members interactions with themselves and other cellular proteins, specially thanks to the two hybrid system and the easy manipulation of yeast (molecular biology and genetics). This review will focus on the use of yeast as a tool to identify new regulators and study function of mammalian apoptosis regulators.

Salinomycin overcomes ABC transporter-mediated multidrug and apoptosis resistance in human leukemia stem cell-like KG-1a cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fuchs, Dominik; Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg; Daniel, Volker

2010-04-16

Leukemia stem cells are known to exhibit multidrug resistance by expression of ATP-binding cassette (ABC) transporters which constitute transmembrane proteins capable of exporting a wide variety of chemotherapeutic drugs from the cytosol. We show here that human promyeloblastic leukemia KG-1a cells exposed to the histone deacetylase inhibitor phenylbutyrate resemble many characteristics of leukemia stem cells, including expression of functional ABC transporters such as P-glycoprotein, BCRP and MRP8. Consequently, KG-1a cells display resistance to the induction of apoptosis by various chemotherapeutic drugs. Resistance to apoptosis induction by chemotherapeutic drugs can be reversed by cyclosporine A, which effectively inhibits the activity ofmore » P-glycoprotein and BCRP, thus demonstrating ABC transporter-mediated drug resistance in KG-1a cells. However, KG-1a are highly sensitive to apoptosis induction by salinomycin, a polyether ionophore antibiotic that has recently been shown to kill human breast cancer stem cell-like cells and to induce apoptosis in human cancer cells displaying multiple mechanisms of drug and apoptosis resistance. Whereas KG-1a cells can be adapted to proliferate in the presence of apoptosis-inducing concentrations of bortezomib and doxorubicin, salinomycin does not permit long-term adaptation of the cells to apoptosis-inducing concentrations. Thus, salinomycin should be regarded as a novel and effective agent for the elimination of leukemia stem cells and other tumor cells exhibiting ABC transporter-mediated multidrug resistance.« less

Simultaneous induction of apoptotic, autophagic, and necrosis-like cell death by monoclonal antibodies recognizing chicken transferrin receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ohno, Yoshiya; Yagi, Hideki; Nakamura, Masanori

Programmed cell death (PCD) is categorized as apoptotic, autophagic, or necrosis-like. Although the possibility that plural (two or three) death signals could be induced by a given stimulus has been reported, the precise mechanisms regulating PCD are not well understood. Recently, we have obtained two anti-chicken transferrin receptor (TfR) monoclonal antibodies (mAbs; D18 and D19) inducing a unique cell death. Although the cell death had several features of apoptosis, autophagic and necrosis-like morphological alterations were simultaneously observed in electron microphotographs. In addition to cells with condensed chromatin and an intact plasma membrane (apoptotic cells), cells having many vacuoles in themore » cytoplasm (autophagic cells), and enlarged cells with ruptured plasma membranes (necrosis-like cells) were observed in DT40 cells treated with the mAbs, however, the latter two types of dead cells were not detected upon treatment with staurosporine, a typical apoptosis inducer. In autophagic cells, numerous membrane-bound vesicles occupying most of the cytoplasmic space, which frequently contained electron-dense materials from cytoplasmic fragments and organelles, were observed. The simultaneous induction of multiple death signals from a stimulus via the TfR is of great interest to those researching cell death. In addition, activation of caspases was observed in DT40 cells treated with D19, however, the cell death was not inhibited with z-VAD-fmk, a pan-caspase inhibitor, suggesting that at least in part, a caspase-independent pathway is involved in the TfR-mediated cell death.« less

Sheeppox virus SPPV14 encodes a Bcl-2-like cell death inhibitor that counters a distinct set of mammalian proapoptotic proteins.

PubMed

Okamoto, Toru; Campbell, Stephanie; Mehta, Ninad; Thibault, John; Colman, Peter M; Barry, Michele; Huang, David C S; Kvansakul, Marc

2012-11-01

Many viruses express inhibitors of programmed cell death (apoptosis), thereby countering host defenses that would otherwise rapidly clear infected cells. To counter this, viruses such as adenoviruses and herpesviruses express recognizable homologs of the mammalian prosurvival protein Bcl-2. In contrast, the majority of poxviruses lack viral Bcl-2 (vBcl-2) homologs that are readily identified by sequence similarities. One such virus, myxoma virus, which is the causative agent of myxomatosis, expresses a virulence factor that is a potent inhibitor of apoptosis. In spite of the scant sequence similarity to Bcl-2, myxoma virus M11L adopts an almost identical 3-dimensional fold. We used M11L as bait in a sequence similarity search for other Bcl-2-like proteins and identified six putative vBcl-2 proteins from poxviruses. Some are potent inhibitors of apoptosis, in particular sheeppox virus SPPV14, which inhibited cell death induced by multiple agents. Importantly, SPPV14 compensated for the loss of antiapoptotic F1L in vaccinia virus and acts to directly counter the cell death mediators Bax and Bak. SPPV14 also engages a unique subset of the death-promoting BH3-only ligands, including Bim, Puma, Bmf, and Hrk. This suggests that SPPV14 may have been selected for specific biological roles as a virulence factor for sheeppox virus.

The Vibrio alginolyticus T3SS effectors, Val1686 and Val1680, induce cell rounding, apoptosis and lysis of fish epithelial cells

PubMed Central

Zhao, Zhe; Liu, Jinxin; Deng, Yiqin; Huang, Wen; Ren, Chunhua; Call, Douglas R.; Hu, Chaoqun

2018-01-01

ABSTRACT Vibrio alginolyticus is a Gram-negative bacterium that is an opportunistic pathogen of both marine animals and people. Its pathogenesis likely involves type III secretion system (T3SS) mediated induction of rapid apoptosis, cell rounding and osmotic lysis of infected eukaryotic cells. Herein, we report that effector proteins, Val1686 and Val1680 from V. alginolyticus, were responsible for T3SS-mediated death of fish cells. Val1686 is a Fic-domain containing protein that not only contributed to cell rounding by inhibiting Rho guanosine triphosphatases (GTPases), but was requisite for the induction of apoptosis because the deletion mutant (Δval1686) was severely weakened in its ability to induce cell rounding and apoptosis in fish cells. In addition, Val1686 alone was sufficient to induce cell rounding and apoptosis as evidenced by the transfection of Val1686 into fish cells. Importantly, the Fic-domain essential for cell rounding activity was equally important to activation of apoptosis of fish cells, indicating that apoptosis is a downstream event of Val1686-dependent GTPase inhibition. V. alginolyticus infection likely activates JNK and ERK pathways with sequential activation of caspases (caspase-8/-10, -9 and -3) and subsequent apoptosis. Val1680 contributed to T3SS-dependent lysis of fish cells in V. alginolyticus, but did not induce autophagy as has been reported for its homologue (VopQ) in V. parahaemolyticus. Together, Val1686 and Val1680 work together to induce apoptosis, cell rounding and cell lysis of V. alginolyticus-infected fish cells. These findings provide new insights into the mechanism of cell death caused by T3SS of V. alginolyticus. PMID:29252102

[Apoptosis: cellular and clinical aspects].

PubMed

Løvschall, H; Mosekilde, L

1997-04-01

Removal of damaged cells is essential for the maintenance of life in multicellular organisms. The process of self destruction, apoptosis, eliminates surplus or damaged cells as part of the pathophysiological defence system. Apoptosis is essential in structural and functional organogenesis during embryological development. The physiological regulation of tissue kinetics is a product of both cell proliferation and cell death. Internal and external regulatory stimuli regulate the balance between apoptosis and mitosis by genetic interaction. Apoptosis is characterized by condensation of chromatine as a result of DNA degradation, formation of blebs in the plasma and nuclear membranes, condensation of cytoplasma, formation of vesicular apoptotic bodies, and phagocytosis by neighbouring cells without inflammatory response. A number of observations indicate that programmed cell death plays an important role in the regulation of cytofunctional homeostasis and defense against accumulation of damaged cells, eg with DNA alterations. Dysregulation of the apoptotic gene program, eg by mutations, may not only lead to loss or degeneration of tissue, but also to hyperproliferative and tumorigenic disorders. New evidence indicates that apoptosis regulation is important both in aging processes and diseases such as: neuropathies, immunopathies, viral infections, cancer, etc. Pharmacological intervention designed to modulate apoptosis seems to raise new possibilities in the treatment of disease.

What a Shock: No Apoptosis without Heat Shock Protein 90α | Center for Cancer Research

Cancer.gov

Apoptosis, also known as programmed cell death, consists of a series of reactions designed to systematically chop up a cell and its contents. The process is used to eliminate specific cells during development or to remove old or damaged cells without harming any surrounding cells. Since cancer cells can develop mechanisms to avoid apoptosis, researchers may be able to identify

Project INSPIRE-HBCU Undergraduate Collaborative Summer Training Program to Inspire Students in Prostate Cancer Research

DTIC Science & Technology

2008-02-01

gallate ( EGCG ). It has been shown that tea polyphenols such as EGCG potently and specifically inhibit chymotrypsin-like activity of the proteasome...autochthonous mouse model of prostate cancer; 2004 2. Ahmad, Nihal; Green Tea Constituent Epigallocatechin -3- Gallate and Induction of apoptosis and cell...such as multiple myeloma. - It has been shown that tea polyphenols, such as (-)- EGCG , potently and specifically inhibit chymotrypsin-like activity of

Trichostatin A-induced apoptosis is mediated by Kruppel-like factor 4 in ovarian and lung cancer.

PubMed

Zohre, Sadeghi; Kazem, Nejati-Koshki; Abolfazl, Akbarzadeh; Mohammad, Rahmati-Yamchi; Aliakbar, Movassaghpour; Effat, Alizadeh; Zahra, Davoudi; Hassan, Dariushnejad; Nosratollah, Zarghami

2014-01-01

The istone deacetylase (HDAC) inhibitor trichostatin A (TSA) is known to mediate the regulation of gene expression and anti proliferation activity in cancer cells. Kruppel-like factor 4 (klf4) is a zinc finger- containing transcription factor of the SP/KLF family, that is expressed in a variety of tissues and regulates cell proliferation, differentiation, tumorigenesis, and apoptosis. It may either either function as a tumor suppressor or an oncogene depending on genetic context of tumors. In this study, we tested the possibility that TSA may increase klf4 expression and cancer cell growth inhibition and apoptosis in SKOV-3 and A549 cells. The cytotoxicity of TSA was determined using the MTT assay test, while klf4 gene expression was assessed by real time PCR and to ability of TSA to induce apoptosis using a Vybrant Apoptosis Assay kit. Our results showed that TSA exerted dose and time dependent cytotoxicity effect on SKOV-3 and A549 cells. Moreover TSA up-regulated klf4 expression. Flow cytometric analysis demonstrated that apoptosis was increased after TSA treatment. Taken together, this study showed that TSA increased klf4 expression in SKOV3 and A549 cell lines, consequently, klf4 may played a tumor-suppressor role by increasing both cell growth inhibition and apoptosis. This study sheds light on the details of molecular mechanisms of HDACI-induced cell cycle arrest and apoptosis.

α-blockade, apoptosis, and prostate shrinkage: how are they related?

PubMed

Chłosta, Piotr; Drewa, Tomasz; Kaplan, Steven

2013-01-01

The α1-adrenoreceptor antagonists, such as terazosin and doxazosin, induce prostate programmed cell death (apoptosis) within prostate epithelial and stromal cells in vitro. This treatment should cause prostate volume decrease, However, this has never been observed in clinical conditions. The aim of this paper is to review the disconnect between these two processes. PubMed and DOAJ were searched for papers related to prostate, apoptosis, and stem cell death. The following key words were used: prostate, benign prostate hyperplasia, programmed cell death, apoptosis, cell death, α1-adrenoreceptor antagonist, α-blockade, prostate epithelium, prostate stroma, stem cells, progenitors, and in vitro models. We have shown how discoveries related to stem cells can influence our understanding of α-blockade treatment for BPH patients. Prostate epithelial and mesenchymal compartments have stem (progenitors) and differentiating cells. These compartments are described in relation to experimental in vitro and in vivo settings. Apoptosis is observed within prostate tissue, but this effect has no clinical significance and cannot lead to prostate shrinkage. In part, this is due to stem cells that are responsible for prostate tissue regeneration and are resistant to apoptosis triggered by α1-receptor antagonists.

Anti-apoptotic Bcl-XL but not Mcl-1 contributes to protection against virus-induced apoptosis.

PubMed

Ohmer, Michaela; Weber, Arnim; Sutter, Gerd; Ehrhardt, Katrin; Zimmermann, Albert; Häcker, Georg

2016-08-18

Infection of mammalian cells with viruses often induces apoptosis. How the recognition of viruses leads to apoptosis of the infected cell and which host cell factors regulate this cell death is incompletely understood. In this study, we focussed on two major anti-apoptotic proteins of the host cell, whose abundance and activity are important for cell survival, the Bcl-2-like proteins Mcl-1 and Bcl-XL. During infection of epithelial cells and fibroblasts with modified vaccinia virus Ankara (MVA), Mcl-1 protein levels dropped but the MVA Bcl-2-like protein F1L could replace Mcl-1 functionally; a similar activity was found in vaccinia virus (VACV)-infected cells. During infection with murine cytomegalovirus (MCMV), Mcl-1-levels were not reduced but a viral Mcl-1-like activity was also generated. Infection of mouse macrophages with any of these viruses, on the other hand, induced apoptosis. Virus-induced macrophage apoptosis was unaltered in the absence of Mcl-1. However, apoptosis was substantially increased in infected Bcl-XL-deficient macrophages or macrophages treated with the Bcl-2/Bcl-XL-inhibitor ABT-737. Genetic loss of Bcl-XL or treatment of macrophages with ABT-737 reduced the generation of infectious VACV. These data show that Mcl-1 is dispensable for the regulation of apoptosis during infection with different large DNA viruses, either because the viruses replace its function (in fibroblasts and epithelial cells) or because the pro-apoptotic activity generated by the infection appears not to be blocked by it (in macrophages). Bcl-XL, on the other hand, can be important to maintain survival of virus-infected cells, and its activity can determine outcome of the infection.

Anti-apoptotic Bcl-XL but not Mcl-1 contributes to protection against virus-induced apoptosis

PubMed Central

Ohmer, Michaela; Weber, Arnim; Sutter, Gerd; Ehrhardt, Katrin; Zimmermann, Albert; Häcker, Georg

2016-01-01

Infection of mammalian cells with viruses often induces apoptosis. How the recognition of viruses leads to apoptosis of the infected cell and which host cell factors regulate this cell death is incompletely understood. In this study, we focussed on two major anti-apoptotic proteins of the host cell, whose abundance and activity are important for cell survival, the Bcl-2-like proteins Mcl-1 and Bcl-XL. During infection of epithelial cells and fibroblasts with modified vaccinia virus Ankara (MVA), Mcl-1 protein levels dropped but the MVA Bcl-2-like protein F1L could replace Mcl-1 functionally; a similar activity was found in vaccinia virus (VACV)-infected cells. During infection with murine cytomegalovirus (MCMV), Mcl-1-levels were not reduced but a viral Mcl-1-like activity was also generated. Infection of mouse macrophages with any of these viruses, on the other hand, induced apoptosis. Virus-induced macrophage apoptosis was unaltered in the absence of Mcl-1. However, apoptosis was substantially increased in infected Bcl-XL-deficient macrophages or macrophages treated with the Bcl-2/Bcl-XL-inhibitor ABT-737. Genetic loss of Bcl-XL or treatment of macrophages with ABT-737 reduced the generation of infectious VACV. These data show that Mcl-1 is dispensable for the regulation of apoptosis during infection with different large DNA viruses, either because the viruses replace its function (in fibroblasts and epithelial cells) or because the pro-apoptotic activity generated by the infection appears not to be blocked by it (in macrophages). Bcl-XL, on the other hand, can be important to maintain survival of virus-infected cells, and its activity can determine outcome of the infection. PMID:27537523

Endogenous bax translocation in SH-SY5Y human neuroblastoma cells and cerebellar granule neurons undergoing apoptosis.

PubMed

McGinnis, K M; Gnegy, M E; Wang, K K

1999-05-01

Changes at the mitochondria are an early, required step in apoptosis in various cell types. We used western blot analysis to demonstrate that the proapoptotic protein Bax translocated from the cytosolic to the mitochondrial fraction in SH-SY5Y human neuroblastoma cells undergoing staurosporine- or EGTA-mediated apoptosis. Levels of mitochondrial Bax increased 15 min after staurosporine treatment. In EGTA-treated cells, increased levels of mitochondrial Bax were seen at 4 h, consistent with a slower onset of apoptosis in EGTA versus staurosporine treatments. We also demonstrate the concomitant translocation of cytochrome c from the mitochondrial to the cytosolic fractions. We correlated these translocations with changes in caspase-3-like activity. An increase in caspase-3-like activity was evident 2 h after staurosporine treatment. Inhibition of the mitochondrial permeability transition had no effect on Bax translocation or caspase-3-like activity in staurosporine-treated SH-SY5Y cells. In primary cultures of cerebellar granule neurons undergoing low K(+)-mediated apoptosis, Bax translocation to the mitochondrial fraction was evident at 3 h. Cytochrome c release into the cytosol was not significant until 8 h after treatment. These data support a model of apoptosis in which Bax acts directly at the mitochondria to allow the release of cytochrome c.

Death penalty for keratinocytes: apoptosis versus cornification.

PubMed

Lippens, S; Denecker, G; Ovaere, P; Vandenabeele, P; Declercq, W

2005-11-01

Homeostasis implies a balance between cell growth and cell death. This balance is essential for the development and maintenance of multicellular organisms. Homeostasis is controlled by several mechanisms including apoptosis, a process by which cells condemned to death are completely eliminated. However, in some cases, total destruction and removal of dead cells is not desirable, as when they fulfil a specific function such as formation of the skin barrier provided by corneocytes, also known as terminally differentiated keratinocytes. In this case, programmed cell death results in accumulation of functional cell corpses. Previously, this process has been associated with apoptotic cell death. In this overview, we discuss differences and similarities in the molecular regulation of epidermal programmed cell death and apoptosis. We conclude that despite earlier confusion, apoptosis and cornification occur through distinct molecular pathways, and that possibly antiapoptotic mechanisms are implicated in the terminal differentiation of keratinocytes.

Grouper iridovirus GIV66 is a Bcl-2 protein that inhibits apoptosis by exclusively sequestering Bim.

PubMed

Banjara, Suresh; Mao, Jiahao; Ryan, Timothy M; Caria, Sofia; Kvansakul, Marc

2018-04-13

Programmed cell death or apoptosis is a critical mechanism for the controlled removal of damaged or infected cells, and proteins of the Bcl-2 family are important arbiters of this process. Viruses have been shown to encode functional and structural homologs of Bcl-2 to counter premature host-cell apoptosis and ensure viral proliferation or survival. Grouper iridovirus (GIV) is a large DNA virus belonging to the Iridoviridae family and harbors GIV66, a putative Bcl-2-like protein and mitochondrially localized apoptosis inhibitor. However, the molecular and structural basis of GIV66-mediated apoptosis inhibition is currently not understood. To gain insight into GIV66's mechanism of action, we systematically evaluated its ability to bind peptides spanning the BH3 domain of pro-apoptotic Bcl-2 family members. Our results revealed that GIV66 harbors an unusually high level of specificity for pro-apoptotic Bcl-2 and displays affinity only for Bcl-2-like 11 (Bcl2L11 or Bim). Using crystal structures of both apo-GIV66 and GIV66 bound to the BH3 domain from Bim, we unexpectedly found that GIV66 forms dimers via an interface that results in occluded access to the canonical Bcl-2 ligand-binding groove, which breaks apart upon Bim binding. This observation suggests that GIV66 dimerization may affect GIV66's ability to bind host pro-death Bcl-2 proteins and enables highly targeted virus-directed suppression of host apoptosis signaling. Our findings provide a mechanistic understanding for the potent anti-apoptotic activity of GIV66 by identifying it as the first single-specificity, pro-survival Bcl-2 protein and identifying a pivotal role of Bim in GIV-mediated inhibition of apoptosis. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Regulation of programmed cell death or apoptosis in atherosclerosis.

PubMed

Geng, Y J

1997-01-01

Intimal thickening caused by accumulation of cells, lipids, and connective tissue characterizes atherosclerosis, an arterial disease that leads to cardiac and cerebral infarction. Apoptosis, or genetically programmed cell death, is important for the development and morphogenesis of organs and tissues. As in other tissues, cells of cardiovascular tissues can undergo apoptosis. Increased apoptosis has been found in both human and animal atherosclerotic lesions, mediating tissue turnover and lesion development. In addition to vascular cells, many activated immune cells, mainly macrophages and T cells, are present in atherosclerotic lesions, where these cells produce biologically active substances such as the proinflammatory cytokines tumor necrosis factor, interleukin-1 (IL-1), and interferon-gamma. Simultaneous exposure to these cytokines may trigger apoptosis of vascular smooth muscle cells. The products of death-regulating genes including Fas/Fas ligand, members of IL-1 beta cysteinyl protease (caspase) family, the tumor suppressive gene p53, and the protooncogene c-myc have been found in vascular cells and may participate in the regulation of vascular apoptosis during the development of atherosclerosis. Abnormal occurrence of apoptosis may take place in atherosclerotic lesions, including attenuation or acceleration of the apoptotic death process. The former may cause an increase in the cellularity of the lesions, and the latter can reduce cellular components important for maintaining the integrity and stability of the plaques. Clarification of the molecular mechanism that regulates apoptosis may help design a new strategy for treatment of patients with atherosclerosis and its major complications, heart attack and stroke.

Disparate effects of non-steroidal anti-inflammatory drugs on apoptosis in guinea-pig gastric mucous cells: inhibition of basal apoptosis by diclofenac

PubMed Central

Ashton, Miranda; Hanson, Peter J

2002-01-01

Non-steroidal anti-inflammatory drugs (NSAIDs) induce apoptosis in gastrointestinal cancer cell lines. Similar actions on normal gastric epithelial cells could contribute to NSAID gastropathy. The present work therefore compared the actions of diclofenac, ibuprofen, indomethacin, and the cyclo-oxygenase-2 selective inhibitor, NS-398, on a primary culture of guinea-pig gastric mucous epithelial cells. Cell number was assessed by staining with crystal violet. Apoptotic activity was determined by condensation and fragmentation of nuclei and by assay of caspase-3-like activity. Necrosis was evaluated from release of cellular enzymes. Ibuprofen (250 μM for 24 h) promoted cell loss, and apoptosis, under both basal conditions and when apoptosis was increased by 25 μM N-Hexanoyl-D-sphingosine (C6-ceramide). Diclofenac (250 μM for 24 h) reduced the proportion of apoptotic nuclei from 5.2 to 2.1%, and caused inhibition of caspase-3-like activity, without causing necrosis under basal conditions. No such reduction in apoptotic activity was evident in the presence of 25 μM C6-ceramide. The inhibitory effect of diclofenac on basal caspase-3-like activity was also exhibited by the structurally similar mefenamic and flufenamic acids (1–250 μM), but not by niflumic acid. Inhibition of superoxide production by the cells increased caspase-3-like activity, but the inhibitory action of diclofenac on caspase activity remained. Diclofenac did not affect superoxide production. Diclofenac inhibited caspase-3-like activity in cell homogenates and also inhibited human recombinant caspase-3. In conclusion, NSAIDs vary in their effect on apoptotic activity in a primary culture of guinea-pig gastric mucous epithelial cells, and the inhibitory effect of diclofenac on basal apoptosis could involve an action on caspase activity. PMID:11815376

Found in Translation: How Preclinical Research Is Guiding the Clinical Development of the BCL2-Selective Inhibitor Venetoclax.

PubMed

Leverson, Joel D; Sampath, Deepak; Souers, Andrew J; Rosenberg, Saul H; Fairbrother, Wayne J; Amiot, Martine; Konopleva, Marina; Letai, Anthony

2017-12-01

Since the discovery of apoptosis as a form of programmed cell death, targeting the apoptosis pathway to induce cancer cell death has been a high-priority goal for cancer therapy. After decades of effort, drug-discovery scientists have succeeded in generating small-molecule inhibitors of antiapoptotic BCL2 family proteins. Innovative medicinal chemistry and structure-based drug design, coupled with a strong fundamental understanding of BCL2 biology, were essential to the development of BH3 mimetics such as the BCL2-selective inhibitor venetoclax. We review a number of preclinical studies that have deepened our understanding of BCL2 biology and facilitated the clinical development of venetoclax. Significance: Basic research into the pathways governing programmed cell death have paved the way for the discovery of apoptosis-inducing agents such as venetoclax, a BCL2-selective inhibitor that was recently approved by the FDA and the European Medicines Agency. Preclinical studies aimed at identifying BCL2-dependent tumor types have translated well into the clinic thus far and will likely continue to inform the clinical development of venetoclax and other BCL2 family inhibitors. Cancer Discov; 7(12); 1376-93. ©2017 AACR. ©2017 American Association for Cancer Research.

Apoptosis induced by cold shock in vitro is dependent on cell growth phase.

PubMed

Soloff, B L; Nagle, W A; Moss, A J; Henle, K J; Crawford, J T

1987-06-15

Chinese hamster V79 fibroblast cells were exposed to brief periods of cold but non-freezing temperatures at different points on the population growth curve. Upon rewarming, cells at the transition from logarithmic to stationary growth exhibited apoptosis (programmed cell death). Cells in other stages of growth, or after reentry into logarithmic growth by refeeding, did not exhibit apoptosis. Apoptosis was expressed by marked cytoplasmic blebbing, by a characteristic non-random fragmentation of DNA into nucleosomal-sized pieces, and by loss of colony-forming ability. The data suggest that cold shock served as a stimulus for susceptible cells to undergo apoptosis. Thus, the experiments describe a new in vitro system for studying the mechanisms of apoptosis.

Apoptosis in fish: environmental factors and programmed cell death.

PubMed

AnvariFar, Hossein; Amirkolaie, Abdolsamad Keramat; Miandare, Hamed Kolangi; Ouraji, Hossein; Jalali, M Ali; Üçüncü, Sema İşisağ

2017-06-01

Apoptosis, a form of programmed cell death, is a critical component in maintaining homeostasis and growth in all tissues and plays a significant role in immunity and cytotoxicity. In contrast to necrosis or traumatic cell death, apoptosis is a well-controlled and vital process characterized mainly by cytoplasmic shrinkage, chromatin condensation, DNA fragmentation, membrane blebbing and apoptotic bodies. Our understanding of apoptosis is partly based on observations in invertebrates but mainly in mammals. Despite the great advantages of fish models in studying vertebrate development and diseases and the tremendous interest observed in recent years, reports on apoptosis in fish are still limited. Although apoptotic machinery is well conserved between aquatic and terrestrial organisms throughout the history of evolution, some differences exist in key components of apoptotic pathways. Core parts of apoptotic machinery in fish are virtually expressed as equivalent to the mammalian models. Some differences are, however, evident, such as the extrinsic and intrinsic pathways of apoptosis including lack of a C-terminal region in the Fas-associated protein with a death domain in fish. Aquatic species inhabit a complex and highly fluctuating environment, making these species good examples to reveal features of apoptosis that may not be easily investigated in mammals. Therefore, in order to gain a wider view on programmed cell death in fish, interactions between the main environmental factors, chemicals and apoptosis are discussed in this review. It is indicated that apoptosis can be induced in fish by exposure to environmental stressors during different stages of the fish life cycle.

Two types of death of poliovirus-infected cells: caspase involvement in the apoptosis but not cytopathic effect.

PubMed

Agol, V I; Belov, G A; Bienz, K; Egger, D; Kolesnikova, M S; Raikhlin, N T; Romanova, L I; Smirnova, E A; Tolskaya, E A

1998-12-20

The death of poliovirus-infected cells may occur in two forms: canonical cytopathic effect (CPE) (on productive infections) or apoptosis (when the viral reproduction is hindered by certain drugs or some other restrictive conditions). Morphological manifestations of the CPE and apoptosis, being distinct, share some traits (e.g., chromatin condensation and nuclear deformation). It was shown here that a permeable caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-(OMe) fluoromethyl ketone (zVAD.fmk), prevented the development of the poliovirus-induced apoptosis on abortive infection. The apoptotic pathway could be dissected by an inhibitor of chymotrypsin-like serine proteases, N-tosyl-l-phenylalanine chloromethyl ketone (TPCK), which prevented the cleavage of DNA to oligonucleosome-sized pieces and nuclear fragmentation but did not suppress cellular shrinkage, cytoplasmic blebbing, and partial chromatin condensation. These results demonstrate that caspase activation is involved in the execution phase of the viral apoptosis and suggest that a nuclear subset of the apoptotic program is under a separate control, involving a TPCK-sensitive event. Neither zVAD.fmk nor TPCK, at the concentrations affecting the apoptotic response, exerted appreciable influence on the virus growth or cellular pathological changes on productive infection, indicating that the pathways leading to the poliovirus-evoked CPE and apoptosis are different. Copyright 1998 Academic Press.

Autonomous rexinoid death signaling is suppressed by converging signaling pathways in immature leukemia cells.

PubMed

Benoit, G R; Flexor, M; Besançon, F; Altucci, L; Rossin, A; Hillion, J; Balajthy, Z; Legres, L; Ségal-Bendirdjian, E; Gronemeyer, H; Lanotte, M

2001-07-01

On their own, retinoid X receptor (RXR)-selective ligands (rexinoids) are silent in retinoic acid receptor (RAR)-RXR heterodimers, and no selective rexinoid program has been described as yet in cellular systems. We report here on the rexinoid signaling capacity that triggers apoptosis of immature promyelocytic NB4 cells as a default pathway in the absence of survival factors. Rexinoid-induced apoptosis displays all features of bona fide programmed cell death and is inhibited by RXR, but not RAR antagonists. Several types of survival signals block rexinoid-induced apoptosis. RARalpha agonists switch the cellular response toward differentiation and induce the expression of antiapoptosis factors. Activation of the protein kinase A pathway in the presence of rexinoid agonists induces maturation and blocks immature cell apoptosis. Addition of nonretinoid serum factors also blocks cell death but does not induce cell differentiation. Rexinoid-induced apoptosis is linked to neither the presence nor stability of the promyelocytic leukemia-RARalpha fusion protein and operates also in non-acute promyelocytic leukemia cells. Together our results support a model according to which rexinoids activate in certain leukemia cells a default death pathway onto which several other signaling paradigms converge. This pathway is entirely distinct from that triggered by RAR agonists, which control cell maturation and postmaturation apoptosis.

Prediction of apoptosis protein locations with genetic algorithms and support vector machines through a new mode of pseudo amino acid composition.

PubMed

Kandaswamy, Krishna Kumar; Pugalenthi, Ganesan; Möller, Steffen; Hartmann, Enno; Kalies, Kai-Uwe; Suganthan, P N; Martinetz, Thomas

2010-12-01

Apoptosis is an essential process for controlling tissue homeostasis by regulating a physiological balance between cell proliferation and cell death. The subcellular locations of proteins performing the cell death are determined by mostly independent cellular mechanisms. The regular bioinformatics tools to predict the subcellular locations of such apoptotic proteins do often fail. This work proposes a model for the sorting of proteins that are involved in apoptosis, allowing us to both the prediction of their subcellular locations as well as the molecular properties that contributed to it. We report a novel hybrid Genetic Algorithm (GA)/Support Vector Machine (SVM) approach to predict apoptotic protein sequences using 119 sequence derived properties like frequency of amino acid groups, secondary structure, and physicochemical properties. GA is used for selecting a near-optimal subset of informative features that is most relevant for the classification. Jackknife cross-validation is applied to test the predictive capability of the proposed method on 317 apoptosis proteins. Our method achieved 85.80% accuracy using all 119 features and 89.91% accuracy for 25 features selected by GA. Our models were examined by a test dataset of 98 apoptosis proteins and obtained an overall accuracy of 90.34%. The results show that the proposed approach is promising; it is able to select small subsets of features and still improves the classification accuracy. Our model can contribute to the understanding of programmed cell death and drug discovery. The software and dataset are available at http://www.inb.uni-luebeck.de/tools-demos/apoptosis/GASVM.

Apoptosis: Focus on sea urchin development.

PubMed

Agnello, Maria; Roccheri, Maria Carmela

2010-03-01

It has been proposed that the apoptosis is an essential requirement for the evolution of all animals, in fact the apoptotic program is highly conserved from nematodes to mammals. Throughout development, apoptosis is employed by multicellular organisms to eliminate damaged or unnecessary cells. Here, we will discuss both developmental programmed cell death (PCD) under normal conditions and stress induced apoptosis, in sea urchin embryos. Sea urchin represent an excellent model system for studying embryogenesis and cellular processes involved in metamorphosis. PCD plays an essential role in sculpting and remodelling the embryos and larvae undergoing metamorphosis. Moreover, this marine organism directly interacts with its environment, and is susceptible to effects of several aquatic contaminants. Apoptosis can be adopted as a defence mechanism against any environmental chemical, physical and mechanical stress, for removing irreversibly damaged cells. This review, while not comprehensive in its reporting, aims to provide an overview of current knowledge on mechanisms to regulate physiological and the induced apoptotic program in sea urchin embryos.

Anesthetic propofol overdose causes endothelial cytotoxicity in vitro and endothelial barrier dysfunction in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Ming-Chung; Department of Anesthesiology, Chi Mei Medical Center, Liouying, Tainan, Taiwan; Chen, Chia-Ling

An overdose and a prolonged treatment of propofol may cause cellular cytotoxicity in multiple organs and tissues such as brain, heart, kidney, skeletal muscle, and immune cells; however, the underlying mechanism remains undocumented, particularly in vascular endothelial cells. Our previous studies showed that the activation of glycogen synthase kinase (GSK)-3 is pro-apoptotic in phagocytes during overdose of propofol treatment. Regarding the intravascular administration of propofol, we therefore hypothesized that propofol overdose also induces endothelial cytotoxicity via GSK-3. Propofol overdose (100 μg/ml) inhibited growth in human arterial and microvascular endothelial cells. After treatment, most of the endothelial cells experienced caspase-independent necrosis-likemore » cell death. The activation of cathepsin D following lysosomal membrane permeabilization (LMP) determined necrosis-like cell death. Furthermore, propofol overdose also induced caspase-dependent apoptosis, at least in part. Caspase-3 was activated and acted downstream of mitochondrial transmembrane potential (MTP) loss; however, lysosomal cathepsins were not required for endothelial cell apoptosis. Notably, activation of GSK-3 was essential for propofol overdose-induced mitochondrial damage and apoptosis, but not necrosis-like cell death. Intraperitoneal administration of a propofol overdose in BALB/c mice caused an increase in peritoneal vascular permeability. These results demonstrate the cytotoxic effects of propofol overdose, including cathepsin D-regulated necrosis-like cell death and GSK-3-regulated mitochondrial apoptosis, on endothelial cells in vitro and the endothelial barrier dysfunction by propofol in vivo. Highlights: ► Propofol overdose causes apoptosis and necrosis in endothelial cells. ► Propofol overdose triggers lysosomal dysfunction independent of autophagy. ► Glycogen synthase kinase-3 facilitates propofol overdose-induced apoptosis. ► Propofol overdose causes an increase in peritoneal vascular permeability.« less

Patterns of cell death in the embryonic antenna of the grasshopper Schistocerca gregaria.

PubMed

Boyan, George; Graf, Philip; Ehrhardt, Erica

2018-03-01

We have investigated the pattern of apoptosis in the antennal epithelium during embryonic development of the grasshopper Schistocerca gregaria. The molecular labels lachesin and annulin reveal that the antennal epithelium becomes subdivided into segment-like meristal annuli within which sensory cell clusters later differentiate. To determine whether apoptosis is involved in the development of such sensory cell clusters, we examined the expression pattern of the cell death labels acridine orange and TUNEL in the epithelium. We found stereotypic, age-dependent, wave-like patterns of cell death in the antenna. Early in embryogenesis, apoptosis is restricted to the most basal meristal annuli but subsequently spreads to encompass almost the entire antenna. Cell death then declines in more basal annuli and is only found in the tip region later in embryogenesis. Apoptosis is restricted throughout to the midregion of a given annulus and away from its border with neighboring annuli, arguing against a causal role in annular formation. Double-labeling for cell death and sensory cell differentiation reveals apoptosis occurring within bands of differentiating sensory cell clusters, matching the meristal organization of the apical antenna. Examination of the individual epithelial lineages which generate sensory cells reveals that apoptosis begins peripherally within a lineage and with age expands to encompass the differentiated sensory cell at the base. We conclude that complete lineages can undergo apoptosis and that the youngest cells in these lineages appear to die first, with the sensory neuron dying last. Lineage-based death in combination with cell death patterns in different regions of the antenna may contribute to odor-mediated behaviors in the grasshopper.

Overcoming chemotherapy drug resistance by targeting inhibitors of apoptosis proteins (IAPs).

PubMed

Rathore, Rama; McCallum, Jennifer E; Varghese, Elizabeth; Florea, Ana-Maria; Büsselberg, Dietrich

2017-07-01

Inhibitors of apoptosis (IAPs) are a family of proteins that play a significant role in the control of programmed cell death (PCD). PCD is essential to maintain healthy cell turnover within tissue but also to fight disease or infection. Uninhibited, IAPs can suppress apoptosis and promote cell cycle progression. Therefore, it is unsurprising that cancer cells demonstrate significantly elevated expression levels of IAPs, resulting in improved cell survival, enhanced tumor growth and subsequent metastasis. Therapies to target IAPs in cancer has garnered substantial scientific interest and as resistance to anti-cancer agents becomes more prevalent, targeting IAPs has become an increasingly attractive strategy to re-sensitize cancer cells to chemotherapies, antibody based-therapies and TRAIL therapy. Antagonism strategies to modulate the actions of XIAP, cIAP1/2 and survivin are the central focus of current research and this review highlights advances within this field with particular emphasis upon the development and specificity of second mitochondria-derived activator of caspase (SMAC) mimetics (synthetic analogs of endogenously expressed inhibitors of IAPs SMAC/DIABLO). While we highlight the potential of SMAC mimetics as effective single agent or combinatory therapies to treat cancer we also discuss the likely clinical implications of resistance to SMAC mimetic therapy, occasionally observed in cancer cell lines.

Induction of Programmed Cell Death by Parvovirus H-1 in U937 Cells: Connection with the Tumor Necrosis Factor Alpha Signalling Pathway

PubMed Central

Rayet, Béatrice; Lopez-Guerrero, José-Antonio; Rommelaere, Jean; Dinsart, Christiane

1998-01-01

The human promonocytic cell line U937 undergoes apoptosis upon treatment with tumor necrosis factor alpha (TNF-α). This cell line has previously been shown to be very sensitive to the lytic effect of the autonomous parvovirus H-1. Parvovirus infection leads to the activation of the CPP32 ICE-like cysteine protease which cleaves the enzyme poly(ADP-ribose)polymerase and induces morphologic changes that are characteristic of apoptosis in a way that is similar to TNF-α treatment. This effect is also observed when the U937 cells are infected with a recombinant H-1 virus which expresses the nonstructural (NS) proteins but in which the capsid genes are replaced by a reporter gene, indicating that the induction of apoptosis can be assigned to the cytotoxic nonstructural proteins in this cell system. The c-Myc protein, which is overexpressed in U937 cells, is rapidly downregulated during infection, in keeping with a possible role of this product in mediating the apoptotic cell death induced by H-1 virus infection. Interestingly, four clones (designated RU) derived from the U937 cell line and selected for their resistance to H-1 virus (J. A. Lopez-Guerrero et al., Blood 89:1642–1653, 1997) failed to decrease c-Myc expression upon treatment with differentiation agents and also resisted the induction of cell death after TNF-α treatment. Our data suggest that the RU clones have developed defense strategies against apoptosis, either by their failure to downregulate c-Myc and/or by activating antiapoptotic factors. PMID:9765434

Resistance of Actin to Cleavage during Apoptosis

NASA Astrophysics Data System (ADS)

Song, Qizhong; Wei, Tie; Lees-Miller, Susan; Alnemri, Emad; Watters, Dianne; Lavin, Martin F.

1997-01-01

A small number of cellular proteins present in the nucleus, cytosol, and membrane fraction are specifically cleaved by the interleukin-1β -converting enzyme (ICE)-like family of proteases during apoptosis. Previous results have demonstrated that one of these, the cytoskeletal protein actin, is degraded in rat PC12 pheochromocytoma cells upon serum withdrawal. Extracts from etoposide-treated U937 cells are also capable of cleaving actin. It was assumed that cleavage of actin represented a general phenomenon, and a mechanism coordinating proteolytic, endonucleolytic, and morphological aspects of apoptosis was proposed. We demonstrate here that actin is resistant to degradation in several different human cells induced to undergo apoptosis in response to a variety of stimuli, including Fas ligation, serum withdrawal, cytotoxic T-cell killing, and DNA damage. On the other hand, cell-free extracts from these cells and the ICE-like protease CPP32 were capable of cleaving actin in vitro. We conclude that while actin contains cleavage sites for ICE-like proteases, it is not degraded in vivo in human cells either because of lack of access of these proteases to actin or due to the presence of other factors that prevent degradation.

24(S)-hydroxycholesterol induces neuronal cell death through necroptosis, a form of programmed necrosis.

PubMed

Yamanaka, Kazunori; Saito, Yoshiro; Yamamori, Tohru; Urano, Yasuomi; Noguchi, Noriko

2011-07-15

24(S)-Hydroxycholesterol (24S-OHC) produced by cholesterol 24-hydroxylase expressed mainly in neurons plays an important physiological role in the brain. Conversely, it has been reported that 24S-OHC possesses potent cytotoxicity. The molecular mechanisms of 24S-OHC-induced cell death have not yet been fully elucidated. In this study, using human neuroblastoma SH-SY5Y cells and primary cortical neuronal cells derived from rat embryo, we characterized the form of cell death induced by 24S-OHC. SH-SY5Y cells treated with 24S-OHC exhibited neither fragmentation of the nucleus nor caspase activation, which are the typical characteristics of apoptosis. 24S-OHC-treated cells showed necrosis-like morphological changes but did not induce ATP depletion, one of the features of necrosis. When cells were treated with necrostatin-1, an inhibitor of receptor-interacting serine/threonine kinase 1 (RIPK1) required for necroptosis, 24S-OHC-induced cell death was significantly suppressed. The knockdown of RIPK1 by transfection of small interfering RNA of RIPK1 effectively attenuated 24S-OHC-induced cell death. It was found that neither SH-SY5Y cells nor primary cortical neuronal cells expressed caspase-8, which was regulated for RIPK1-dependent apoptosis. Collectively, these results suggest that 24S-OHC induces neuronal cell death by necroptosis, a form of programmed necrosis.

24(S)-Hydroxycholesterol Induces Neuronal Cell Death through Necroptosis, a Form of Programmed Necrosis*

PubMed Central

Yamanaka, Kazunori; Saito, Yoshiro; Yamamori, Tohru; Urano, Yasuomi; Noguchi, Noriko

2011-01-01

24(S)-Hydroxycholesterol (24S-OHC) produced by cholesterol 24-hydroxylase expressed mainly in neurons plays an important physiological role in the brain. Conversely, it has been reported that 24S-OHC possesses potent cytotoxicity. The molecular mechanisms of 24S-OHC-induced cell death have not yet been fully elucidated. In this study, using human neuroblastoma SH-SY5Y cells and primary cortical neuronal cells derived from rat embryo, we characterized the form of cell death induced by 24S-OHC. SH-SY5Y cells treated with 24S-OHC exhibited neither fragmentation of the nucleus nor caspase activation, which are the typical characteristics of apoptosis. 24S-OHC-treated cells showed necrosis-like morphological changes but did not induce ATP depletion, one of the features of necrosis. When cells were treated with necrostatin-1, an inhibitor of receptor-interacting serine/threonine kinase 1 (RIPK1) required for necroptosis, 24S-OHC-induced cell death was significantly suppressed. The knockdown of RIPK1 by transfection of small interfering RNA of RIPK1 effectively attenuated 24S-OHC-induced cell death. It was found that neither SH-SY5Y cells nor primary cortical neuronal cells expressed caspase-8, which was regulated for RIPK1-dependent apoptosis. Collectively, these results suggest that 24S-OHC induces neuronal cell death by necroptosis, a form of programmed necrosis. PMID:21613228

Human Immunodeficiency Virus Envelope Protein Gp120 Induces Proliferation but Not Apoptosis in Osteoblasts at Physiologic Concentrations

PubMed Central

Cummins, Nathan W.; Klicpera, Anna; Sainski, Amy M.; Bren, Gary D.; Khosla, Sundeep; Westendorf, Jennifer J.; Badley, Andrew D.

2011-01-01

Patients with HIV infection have decreased numbers of osteoblasts, decreased bone mineral density and increased risk of fracture compared to uninfected patients; however, the molecular mechanisms behind these associations remain unclear. We questioned whether Gp120, a component of the envelope protein of HIV capable of inducing apoptosis in many cell types, is able to induce cell death in bone-forming osteoblasts. We show that treatment of immortalized osteoblast-like cells and primary human osteoblasts with exogenous Gp120 in vitro at physiologic concentrations does not result in apoptosis. Instead, in the osteoblast-like U2OS cell line, cells expressing CXCR4, a receptor for Gp120, had increased proliferation when treated with Gp120 compared to control (P<0.05), which was inhibited by pretreatment with a CXCR4 inhibitor and a G-protein inhibitor. This suggests that Gp120 is not an inducer of apoptosis in human osteoblasts and likely does not directly contribute to osteoporosis in infected patients by this mechanism. PMID:21931863

[Influence of human gastrointestinal tract bacterial pathogens on host cell apoptosis].

PubMed

Wronowska, Weronika; Godlewska, Renata; Jagusztyn-Krynicka, Elzbieta Katarzyna

2005-01-01

Several pathogenic bacteria are able to trigger apoptosis in the host cell, but the mechanisms by which it occurs differ, and the resulting pathology can take different courses. Induction and/or blockage of programmed cell death upon infection is a result of complex interaction of bacterial proteins with cellular proteins involved in signal transduction and apoptosis. In this review we focus on pro/anti-apoptotic activities exhibited by two enteric pathogens Salmonella enterica, Yersinia spp. and gastric pathogen Helicobacter pylori. We present current knowledge on how interaction between mammalian and bacterial cell relates to the molecular pathways of apoptosis, and what is the role of apoptosis in pathogenesis.

The Microanatomic Segregation of Selection by Apoptosis in the Germinal Center

PubMed Central

Mayer, Christian T.; Gazumyan, Anna; Kara, Ervin E.; Gitlin, Alexander D.; Golijanin, Jovana; Viant, Charlotte; Pai, Joy; Oliveira, Thiago Y.; Wang, Qiao; Escolano, Amelia; Medina-Ramirez, Max; Sanders, Rogier W.; Nussenzweig, Michel C.

2018-01-01

B cells undergo rapid cell division and affinity maturation in anatomically distinct sites in lymphoid organs called germinal centers (GCs). Homeostasis is maintained in part by B-cell apoptosis. However, the precise contribution of apoptosis to GC biology and selection is not well defined. We developed apoptosis-indicator mice and used them to visualize, purify, and characterize dying GC B cells. Apoptosis is prevalent in the GC with up to half of all GC B cells dying every 6h. Moreover, programmed cell death is differentially regulated in the light zone (LZ) and the dark zone (DZ): LZ B cells die by default if they are not positively selected, whereas DZ cells die when their antigen receptors are damaged by activation-induced cytidine deaminase (AID). PMID:28935768

What a Shock: No Apoptosis without Heat Shock Protein 90α | Center for Cancer Research

Cancer.gov

Apoptosis, also known as programmed cell death, consists of a series of reactions designed to systematically chop up a cell and its contents. The process is used to eliminate specific cells during development or to remove old or damaged cells without harming any surrounding cells. Since cancer cells can develop mechanisms to avoid apoptosis, researchers may be able to identify new targets to combat cancer by better understanding the details of the apoptotic process.

THE ROLE OF APOPTOSIS IN NEUROTOXICOLOGY

EPA Science Inventory

Apoptosis, a form of programmed cell death, occurs in the nervous system throughout development, but with a preponderance of cell death occurring during the prenatal and perinatal periods. Aberrant periods of increased or decreased cell death, induced by toxicants in air, water,...

Differential Expression of Programmed Cell Death on the Follicular Development in Normal and Miniature Pig Ovary

PubMed Central

Kim, Sang Hwan; Min, Kwan Sik; Kim, Nam Hyung; Yoon, Jong Taek

2012-01-01

Follicles are important in oocyte maturation. Successful estrous cycle requires remodeling of follicular cells, and proper execution of programmed cell death is crucial for normal follicular development. The objectives of the present study were to understand programmed cell death during follicle development, to analyze the differential follicle development patterns, and to assess the patterns of apoptosis and autophagy expression during follicle development in normal and miniature pigs. Through the analysis of differential patterns of programmed cell death during follicular development in porcine, MAP1LC3A, B and other autophagy-associated genes (ATG5, mTOR, Beclin-1) were found to increase in normal pigs, while it decreased in miniature pigs. However, for the apoptosis-associated genes, progression of genes during follicular development increased in miniature pigs, while it decreased in normal pigs. Thus, results show that normal and miniature pigs showed distinct patterns of follicular remodeling manifesting that programmed cell death largely depends on the types of pathway during follicular development (Type II or autophagy for normal pigs and Type I or apoptosis for miniature pigs). PMID:23056260

Propolis Augments Apoptosis Induced by Butyrate via Targeting Cell Survival Pathways

PubMed Central

Drago, Eric; Bordonaro, Michael; Lee, Seon; Atamna, Wafa; Lazarova, Darina L.

2013-01-01

Diet is one of the major lifestyle factors affecting incidence of colorectal cancer (CC), and despite accumulating evidence that numerous diet-derived compounds modulate CC incidence, definitive dietary recommendations are not available. We propose a strategy that could facilitate the design of dietary supplements with CC-preventive properties. Thus, nutrient combinations that are a source of apoptosis-inducers and inhibitors of compensatory cell proliferation pathways (e.g., AKT signaling) may produce high levels of programmed death in CC cells. Here we report the combined effect of butyrate, an apoptosis inducer that is produced through fermentation of fiber in the colon, and propolis, a honeybee product, on CC cells. We established that propolis increases the apoptosis of CC cells exposed to butyrate through suppression of cell survival pathways such as the AKT signaling. The programmed death of CC cells by combined exposure to butyrate and propolis is further augmented by inhibition of the JNK signaling pathway. Analyses on the contribution of the downstream targets of JNK signaling, c-JUN and JAK/STAT, to the apoptosis of butyrate/propolis-treated CC cells ascertained that JAK/STAT signaling has an anti-apoptotic role; whereas, the role of cJUN might be dependent upon regulatory cell factors. Thus, our studies ascertained that propolis augments apoptosis of butyrate-sensitive CC cells and re-sensitizes butyrate-resistant CC cells to apoptosis by suppressing AKT signaling and downregulating the JAK/STAT pathway. Future in vivo studies should evaluate the CC-preventive potential of a dietary supplement that produces high levels of colonic butyrate, propolis, and diet-derived JAK/STAT inhibitors. PMID:24023824

Rocaglamide overcomes tumor necrosis factor-related apoptosis-inducing ligand resistance in hepatocellular carcinoma cells by attenuating the inhibition of caspase-8 through cellular FLICE-like-inhibitory protein downregulation.

PubMed

Luan, Zhou; He, Ying; He, Fan; Chen, Zhishui

2015-01-01

The enhancement of apoptosis is a therapeutic strategy used in the treatment of cancer. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising antitumor agent. However, hepatocellular carcinoma (HCC) cells exhibit marked resistance to the induction of cell death by TRAIL. The present study investigated whether rocaglamide, a naturally occurring product isolated from the genus Aglaia, is able to sensitize resistant HCC cells to TRAIL-mediated apoptosis. Two HCC cell lines, HepG2 and Huh-7, were treated with rocaglamide and/or TRAIL and the induction of apoptosis and effects on the TRAIL signaling pathway were investigated. The in vivo efficacy of rocaglamide was determined in TRAIL-resistant Huh-7-derived tumor xenografts. Rocaglamide significantly sensitized the TRAIL-resistant HCC cells to apoptosis by TRAIL, which resulted from the rocaglamide-mediated downregulation of cellular FLICE-like inhibitory protein and subsequent caspase-8 activation. Furthermore, rocaglamide markedly inhibited tumor growth from Huh-7 cells propagated in severe combined immunodeficient mice, suggesting that chemosentization also occurred in vivo. These data suggest that rocaglamide acted synergistically with TRAIL against the TRAIL-resistant HCC cells. Thus, it is concluded that rocaglamide as an adjuvant to TRAIL-based therapy may present a promising therapeutic approach for the treatment of HCC.

Inhibition of Ced-3/ICE-related Proteases Does Not Prevent Cell Death Induced by Oncogenes, DNA Damage, or the Bcl-2 Homologue Bak

PubMed Central

McCarthy, Nicola J.; Whyte, Moira K.B.; Gilbert, Christopher S.; Evan, Gerard I.

1997-01-01

There is increasing evidence for a central role in mammalian apoptosis of the interleukin-1β– converting enzyme (ICE) family of cysteine proteases, homologues of the product of the nematode “death” gene, ced-3. Ced-3 is thought to act as an executor rather than a regulator of programmed cell death in the nematode. However, it is not known whether mammalian ICE-related proteases (IRPs) are involved in the execution or the regulation of mammalian apoptosis. Moreover, an absolute requirement for one or more IRPs for mammalian apoptosis has yet to be established. We have used two cell-permeable inhibitors of IRPs, Z-Val-Ala-Asp.fluoromethylketone (ZVAD.fmk) and t-butoxy carbonyl-Asp.fluoromethylketone (BD.fmk), to demonstrate a critical role for IRPs in mammalian apoptosis induced by several disparate mechanisms (deregulated oncogene expression, ectopic expression of the Bcl-2 relative Bak, and DNA damage–induced cell death). In all instances, ZVAD.fmk and BD.fmk treatment inhibits characteristic biochemical and morphological events associated with apoptosis, including cleavage of nuclear lamins and poly-(ADP-ribose) polymerase, chromatin condensation and nucleosome laddering, and external display of phosphatidylserine. However, neither ZVAD.fmk nor BD.fmk inhibits the onset of apoptosis, as characterized by the onset of surface blebbing; rather, both act to delay completion of the program once initiated. In complete contrast, IGF-I and Bcl-2 delay the onset of apoptosis but have no effect on the kinetics of the program once initiated. Our data indicate that IRPs constitute part of the execution machinery of mammalian apoptosis induced by deregulated oncogenes, DNA damage, or Bak but that they act after the point at which cells become committed to apoptosis or can be rescued by survival factors. Moreover, all such blocked cells have lost proliferative potential and all eventually die by a process involving cytoplasmic blebbing. PMID:9008715

Cell death in response to antimetabolites directed at thymidylate synthase.

PubMed

Barbour, Karen W; Berger, Franklin G

2008-02-01

Thymidylate synthase (TS) is an indispensable enzyme in the de novo biosynthesis of TMP during DNA replication and cell growth, and has, therefore, been an important target for several classes of antimetabolites used in cancer chemotherapy. While most investigations of the action of TS-directed agents have focused on apoptosis as the primary means of cell death, little is known regarding the role, if any, of non-apoptotic mechanisms. In the present study, we have examined the mode of cell death induced by several TS inhibitors. Apoptosis and necrosis in response to TS inhibitors was assessed. The roles of caspases and the transcriptional regulator nuclear factor kappa B (NFkappaB) in drug-induced cell death were analyzed. Finally, drug-mediated changes in expression of several proteins involved in regulation of apoptosis were analyzed. Though human colon tumor cells exposed to TS inhibitors undergo classical apoptosis, it is not the predominant mechanism of response; rather, a necrosis-like mechanism prevails. The apoptotic response to TS inhibitors is caspase-dependent, and is promoted by NFkappaB. In contrast, the necrosis-like response is independent of both caspases and NFkappaB. Exposure to TS inhibitors induces PARP cleavage, but does not alter expression of the pro or activated forms of caspases-3 or caspases-8, Fas, or FasL. Treatment with the death-inducing cytokine TNFalpha, like TS inhibitors, results in a limited extent of apoptosis that is both caspase- and NFkappaB-dependent; however, unlike TS inhibitors, the cytokine does not induce necrosis. Classical apoptosis occurs to a limited extent in human colon tumor cells exposed to TS inhibitors, with caspase-independent necrosis being the prinicipal mechanism of cell death. We suggest that the role of necrosis and necrosis-like mechanisms should be considered in future studies of the action of TS-directed antimetabolites, as well as other chemotherapeutic agents.

Opposite effects of flurbiprofen and the nitroxybutyl ester of flurbiprofen on apoptosis in cultured guinea-pig gastric mucous cells

PubMed Central

Johal, Kamaljit; Hanson, Peter J

2000-01-01

The nitric oxide (NO)-donating nitroxybutyl ester of flurbiprofen (NO-flurbiprofen), shows reduced gastro-intestinal toxicity relative to flurbiprofen. NO may exert either pro- or anti-apoptotic effects, while non-steroidal anti-inflammatory drugs may induce apoptosis. The aim of the present work was therefore to compare the effects of flurbiprofen and NO-flurbiprofen on apoptosis in guinea-pig gastric mucous cells. Apoptotic activity was assessed by assay of caspase activity and from the fragmentation and condensation of nuclei. Incubation with flurbiprofen for 24 h produced a concentration-dependent induction of apoptosis in cells attached to the culture plate (caspase 3-like activity increased by 257% at 500 μM), while NO-flurbiprofen inhibited basal apoptosis (caspase 3-like activity decreased by 71% at 500 μM). Caspase activity and nuclear fragmentation were substantially increased in cells that had spontaneously detached from the culture plate. NO-flurbiprofen inhibited caspase activity (55% at 500 μM) but not nuclear fragmentation in these detached cells. NO flurbiprofen inhibited the activation of apoptosis by 25 μM C6-ceramide in cells attached to the culture plate. Inhibition of caspase activity by NO-flurbiprofen was detectable after 6 h of incubation with intact cells, but by contrast with the NO-donor S-nitrosyl-N-acetyl-penicillamine, was not demonstrable with cell homogenates. Activation of caspase 3-like activity by flurbiprofen was slow (>6 h incubation needed) and was inhibited by cycloheximide. The presence of a nitroxybutyl ester moiety on flurbiprofen prevents the pro-apoptotic activity of the parent compound and may contribute to the reduced gastro-intestinal toxicity of NO-flurbiprofen. PMID:10864887

Regulation of Apoptosis during Flavivirus Infection.

PubMed

Okamoto, Toru; Suzuki, Tatsuya; Kusakabe, Shinji; Tokunaga, Makoto; Hirano, Junki; Miyata, Yuka; Matsuura, Yoshiharu

2017-08-28

Apoptosis is a type of programmed cell death that regulates cellular homeostasis by removing damaged or unnecessary cells. Its importance in host defenses is highlighted by the observation that many viruses evade, obstruct, or subvert apoptosis, thereby blunting the host immune response. Infection with Flaviviruses such as Japanese encephalitis virus (JEV), Dengue virus (DENV) and West Nile virus (WNV) has been shown to activate several signaling pathways such as endoplasmic reticulum (ER)-stress and AKT/PI3K pathway, resulting in activation or suppression of apoptosis in virus-infected cells. On the other hands, expression of some viral proteins induces or protects apoptosis. There is a discrepancy between induction and suppression of apoptosis during flavivirus infection because the experimental situation may be different, and strong links between apoptosis and other types of cell death such as necrosis may make it more difficult. In this paper, we review the effects of apoptosis on viral propagation and pathogenesis during infection with flaviviruses.

IL-1β-induced matrix metalloproteinase-13 is activated by a disintegrin and metalloprotease-28-regulated proliferation of human osteoblast-like cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozeki, Nobuaki; Kawai, Rie; Yamaguchi, Hideyuki

2014-04-15

We reported previously that matrix metalloproteinase (MMP)-13 accelerates bone remodeling in oral periradicular lesions, and indicated a potentially unique role for MMP-13 in wound healing and regeneration of alveolar bone. The ADAM (a disintegrin and metalloprotease) family is a set of multifunctional cell surface and secreted glycoproteins, of which ADAM-28 has been localized in bone and bone-like tissues. In this study, we show that interleukin (IL)-1β induces the expression of MMP-13 and ADAM-28 in homogeneous α7 integrin-positive human skeletal muscle stem cell (α7{sup +}hSMSC)-derived osteoblast-like (α7{sup +}hSMSC-OB) cells, and promotes proliferation while inhibiting apoptosis in these cells. At higher concentrations,more » however, IL-1β failed to induce the expression of these genes and caused an increase in apoptosis. We further employed ADAM-28 small interfering RNA (siRNA) to investigate whether IL-1β-induced MMP-13 expression is linked to this IL-1β-mediated changes in cell proliferation and apoptosis. Silencing ADAM-28 expression potently suppressed IL-1β-induced MMP-13 expression and activity, decreased cell proliferation and increased apoptosis in α7{sup +}hSMSC-OB cells. In contrast, MMP-13 siRNA had no effect on ADAM-28 expression, suggesting ADAM-28 regulates MMP-13. Exogenous MMP-13 induced α7{sup +}hSMSC-OB cell proliferation and could rescue ADAM-28 siRNA-induced apoptosis, and we found that proMMP-13 is partially cleaved into its active form by ADAM-28 in vitro. Overall, our results suggest that IL-1β-induced MMP-13 expression and changes in cell proliferation and apoptosis in α7{sup +}hSMSC-OB cells are regulated by ADAM-28. - Highlights: • IL-1β induces the MMP-13 and ADAM-28 expression in human osteoblast-like cells. • IL-1β-induced MMP-13 expression increases proliferation and decreased apoptosis. • MMP-13 expression induced by IL-1β is regulated by ADAM-28. • proMMP-13 appears to be cleaved into its active form via ADAM-28.« less

Potential role of centrioles in determining the morphogenetic status of animal somatic cells.

PubMed

Tkemaladze, J; Chichinadze, K

2005-05-01

Irreversible differentiation (change of morphogenetic status) and programmed death (apoptosis) are observed only in somatic cells. Cell division is the only way by which the morphogenetic status of the offspring cells may be modified. It is known that there is a fixed limit to the number of possible cell divisions, the so-called 'Hayflick limit'. Existing links between cell division, differentiation and apoptosis make it possible to conclude that all these processes could be controlled by a single self-reproducing structure. Potential candidates for this replicable structure in a somatic cell are chromosomes, mitochondria (both contain DNA), and centrioles. Centrioles (diplosome) are the most likely unit that can fully regulate the processes of irreversible differentiation, determination and modification of the morphogenetic status. It may contain differently encoded RNA molecules stacked in a definite order. During mitosis, these RNA molecules are released one by one into the cytoplasm. In the presence of reverse transcriptase and endonuclease, RNA can be embedded in nuclear DNA. This process presumably changes the status of repressed and potentially active genes and, subsequently, the morphogenetic status of a cell.

Identification of RIP1 as a critical mediator of Smac mimetic-mediated sensitization of glioblastoma cells for Drozitumab-induced apoptosis.

PubMed

Cristofanon, S; Abhari, B A; Krueger, M; Tchoghandjian, A; Momma, S; Calaminus, C; Vucic, D; Pichler, B J; Fulda, S

2015-04-16

This study aims at evaluating the combination of the tumor-necrosis-factor-related apoptosis-inducing ligand (TRAIL)-receptor 2 (TRAIL-R2)-specific antibody Drozitumab and the Smac mimetic BV6 in preclinical glioblastoma models. To this end, the effect of BV6 and/or Drozitumab on apoptosis induction and signaling pathways was analyzed in glioblastoma cell lines, primary glioblastoma cultures and glioblastoma stem-like cells. Here, we report that BV6 and Drozitumab synergistically induce apoptosis and reduce colony formation in several glioblastoma cell lines (combination index<0.1). Also, BV6 profoundly enhances Drozitumab-induced apoptosis in primary glioblastoma cultures and glioblastoma stem-like cells. Importantly, BV6 cooperates with Drozitumab to suppress tumor growth in two glioblastoma in vivo models including an orthotopic, intracranial mouse model, underlining the clinical relevance of these findings. Mechanistic studies reveal that BV6 and Drozitumab act in concert to trigger the formation of a cytosolic receptor-interacting protein (RIP) 1/Fas-associated via death domain (FADD)/caspase-8-containing complex and subsequent activation of caspase-8 and -3. BV6- and Drozitumab-induced apoptosis is blocked by the caspase inhibitor zVAD.fmk, pointing to caspase-dependent apoptosis. RNA interference-mediated silencing of RIP1 almost completely abolishes the BV6-conferred sensitization to Drozitumab-induced apoptosis, indicating that the synergism critically depends on RIP1 expression. In contrast, both necrostatin-1, a RIP1 kinase inhibitor, and Enbrel, a TNFα-blocking antibody, do not interfere with BV6/Drozitumab-induced apoptosis, demonstrating that apoptosis occurs independently of RIP1 kinase activity or an autocrine TNFα loop. In conclusion, the rational combination of BV6 and Drozitumab presents a promising approach to trigger apoptosis in glioblastoma, which warrants further investigation.

An Early and Robust Activation of Caspases Heads Cells for a Regulated Form of Necrotic-like Cell Death.

PubMed

Garcia-Belinchón, Mercè; Sánchez-Osuna, María; Martínez-Escardó, Laura; Granados-Colomina, Carla; Pascual-Guiral, Sònia; Iglesias-Guimarais, Victoria; Casanelles, Elisenda; Ribas, Judit; Yuste, Victor J

2015-08-21

Apoptosis is triggered by the activation of caspases and characterized by chromatin condensation and nuclear fragmentation (type II nuclear morphology). Necrosis is depicted by a gain in cell volume (oncosis), swelling of organelles, plasma membrane leakage, and subsequent loss of intracellular contents. Although considered as different cell death entities, there is an overlap between apoptosis and necrosis. In this sense, mounting evidence suggests that both processes can be morphological expressions of a common biochemical network known as "apoptosis-necrosis continuum." To gain insight into the events driving the apoptosis-necrosis continuum, apoptotically proficient cells were screened facing several apoptotic inducers for the absence of type II apoptotic nuclear morphologies. Chelerythrine was selected for further studies based on its cytotoxicity and the lack of apoptotic nuclear alterations. Chelerythrine triggered an early plasma membrane leakage without condensed chromatin aggregates. Ultrastructural analysis revealed that chelerythrine-mediated cytotoxicity was compatible with a necrotic-like type of cell death. Biochemically, chelerythrine induced the activation of caspases. Moreover, the inhibition of caspases prevented chelerythrine-triggered necrotic-like cell death. Compared with staurosporine, chelerythrine induced stronger caspase activation detectable at earlier times. After using a battery of chemicals, we found that high concentrations of thiolic antioxidants fully prevented chelerythrine-driven caspase activation and necrotic-like cell death. Lower amounts of thiolic antioxidants partially prevented chelerythrine-mediated cytotoxicity and allowed cells to display type II apoptotic nuclear morphology correlating with a delay in caspase-3 activation. Altogether, these data support that an early and pronounced activation of caspases can drive cells to undergo a form of necrotic-like regulated cell death. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

An Early and Robust Activation of Caspases Heads Cells for a Regulated Form of Necrotic-like Cell Death*

PubMed Central

Garcia-Belinchón, Mercè; Sánchez-Osuna, María; Martínez-Escardó, Laura; Granados-Colomina, Carla; Pascual-Guiral, Sònia; Iglesias-Guimarais, Victoria; Casanelles, Elisenda; Ribas, Judit; Yuste, Victor J.

2015-01-01

Apoptosis is triggered by the activation of caspases and characterized by chromatin condensation and nuclear fragmentation (type II nuclear morphology). Necrosis is depicted by a gain in cell volume (oncosis), swelling of organelles, plasma membrane leakage, and subsequent loss of intracellular contents. Although considered as different cell death entities, there is an overlap between apoptosis and necrosis. In this sense, mounting evidence suggests that both processes can be morphological expressions of a common biochemical network known as “apoptosis-necrosis continuum.” To gain insight into the events driving the apoptosis-necrosis continuum, apoptotically proficient cells were screened facing several apoptotic inducers for the absence of type II apoptotic nuclear morphologies. Chelerythrine was selected for further studies based on its cytotoxicity and the lack of apoptotic nuclear alterations. Chelerythrine triggered an early plasma membrane leakage without condensed chromatin aggregates. Ultrastructural analysis revealed that chelerythrine-mediated cytotoxicity was compatible with a necrotic-like type of cell death. Biochemically, chelerythrine induced the activation of caspases. Moreover, the inhibition of caspases prevented chelerythrine-triggered necrotic-like cell death. Compared with staurosporine, chelerythrine induced stronger caspase activation detectable at earlier times. After using a battery of chemicals, we found that high concentrations of thiolic antioxidants fully prevented chelerythrine-driven caspase activation and necrotic-like cell death. Lower amounts of thiolic antioxidants partially prevented chelerythrine-mediated cytotoxicity and allowed cells to display type II apoptotic nuclear morphology correlating with a delay in caspase-3 activation. Altogether, these data support that an early and pronounced activation of caspases can drive cells to undergo a form of necrotic-like regulated cell death. PMID:26124276

Escherichia coli STb toxin induces apoptosis in intestinal epithelial cell lines.

PubMed

Syed, H Claudia; Dubreuil, J Daniel

2012-09-01

A previous study conducted in our laboratory demonstrated that cells having internalized Escherichia coli STb toxin display apoptotic-like morphology. We therefore investigated if STb could induce programmed cell death in both a human and an animal intestinal epithelial cell lines. HRT-18 (Human Colon Tumor) and IEC-18 (Rat Ileum Epithelial Cells) cell lines were used. As STb is frequently tested in a rat model, the IEC-18 cell line was most relevant to our work. The cell lines were treated with various amounts of purified STb (nanomole range) for a period of 24 h after which cells were harvested and examined for apoptotic characteristics. Caspase-9, the initiator of mitochondrion-mediated apoptosis, and caspase-3, an effector of caspase-9, were both activated following STb intoxication of HRT-18 and IEC-18 cells whereas caspase-8, the initiator caspase of the extrinsic pathway, was not activated. For both cell lines, agarose gel electrophoresis of the cell DNA content reveals laddering of DNA, resulting from DNA fragmentation, a characteristic of apoptosis. Hoechst 33342-stained DNA of STb-treated cell lines, observed using fluorescence microscopy, revealed condensation and fragmentation of the nuclei. Apoptotic indexes calculated from fragmented nuclei of Hoechst 33342-stained DNA for HRT-18 and IEC-18 cells showed an STb dose-dependent response. Overall, these data indicate that STb toxin induces a mitochondrion-mediated caspase-dependent apoptotic pathway. Copyright © 2012 Elsevier Ltd. All rights reserved.

APOPTOSIS DURING DEVELOPMENT AND AGING AND IN RESPONSE TO MERCURY EXPOSURE.

EPA Science Inventory

In the central nervous system from embryogenesis through senescence, cell number is regulated, in part, by apoptosis. Each region of the nervous system has a characteristic temporal pattern of programmed cell death, which includes far greater numbers of cells undergoing apop...

Calcium-mediated apoptosis in a plant hypersensitive disease resistance response.

PubMed

Levine, A; Pennell, R I; Alvarez, M E; Palmer, R; Lamb, C

1996-04-01

Avirulent pathogens elicit a battery of plant defenses, often accompanied by collapse of the challenged cells. In soybean cells, sustained accumulation of H2O2 from an oxidative burst cues localized host cell death. Such hypersensitive cell death appears to be an active process, but little is known about the mechanisms underlying cellular collapse. We show that H2O2 stimulates a rapid influx of Ca2+ into soybean cells, which activates a physiological cell death program resulting in the generation of large (approximately 50 kb) DNA fragments and cell corpse morphology--including cell shrinkage, plasma membrane blebbing and nuclear condensation--characteristic of apoptosis. In contrast, H2O2 induction of the cellular protectant gene glutathione S-transferase is Ca(2+)-independent. Apoptosis in soybean cells and leaf tissue was induced by avirulent Pseudomonas syringae pv. glycinea but was not observed at comparable stages of the compatible interaction with the isogenic virulent strain, which fails to elicit a hypersensitive response. Apoptosis was also observed at the onset of the hypersensitive response in Arabidopsis leaves inoculated with avirulent P. syringae pv. tomato and in tobacco cells treated with the fungal peptide cryptogein, which is involved in the induction of non-host resistance to Phytophthora cryptogea. These observations establish a signal function for Ca2+ downstream of the oxidative burst in the activation of a physiological cell death program in soybean cells that is similar to apoptosis in animals. That the characteristic cell corpse morphology is also induced in Arabidopsis and tobacco by different avirulence signals suggests that apoptosis may prove to be a common, but not necessarily ubiquitous, feature of incompatible plant-pathogen interactions. Emerging similarities between facets of hypersensitive disease resistance and the mammalian native immune system indicate that apoptosis is a widespread defence mechanism in eukaryotes.

Emerging roles of apoptotic microtubules during the execution phase of apoptosis.

PubMed

Oropesa Ávila, Manuel; Fernández Vega, Alejandro; Garrido Maraver, Juan; Villanueva Paz, Marina; De Lavera, Isabel; De La Mata, Mario; Cordero, Mario D; Alcocer Gómez, Elizabet; Delgado Pavón, Ana; Álvarez Córdoba, Mónica; Cotán, David; Sánchez-Alcázar, José Antonio

2015-09-01

Apoptosis is a genetically programmed energy-dependent process of cell demise, characterized by specific morphological and biochemical events in which the activation of caspases has an essential role. During apoptosis the cytoskeleton participates actively in characteristic morphological rearrangements of the dying cell. This reorganisation has been assigned mainly to actinomyosin ring contraction, while microtubule and intermediate filaments are depolymerized at early stages of apoptosis. However, recent reports have showed that microtubules are reformed during the execution phase of apoptosis organizing an apoptotic microtubule network (AMN). AMN is organized behind plasma membrane, forming a cortical structure. Apoptotic microtubules repolymerization takes place in many cell types and under different apoptotic inducers. It has been hypothesized that AMN is critical for maintaining plasma membrane integrity and cell morphology during the execution phase of apoptosis. AMN disorganization leads apoptotic cells to secondary necrosis and the release of potential toxic molecules which can damage neighbor cells and promotes inflammation. Therefore, AMN formation during physiological apoptosis or in pathological apoptosis induced by anti-cancer treatments is essential for tissue homeostasis and the prevention of additional cell damage and inflammation. © 2015 Wiley Periodicals, Inc.

Antiapoptotic Activity of the Herpesvirus Saimiri-Encoded Bcl-2 Homolog: Stabilization of Mitochondria and Inhibition of Caspase-3-Like Activity

PubMed Central

Derfuss, Tobias; Fickenscher, Helmut; Kraft, Michael S.; Henning, Golo; Lengenfelder, Doris; Fleckenstein, Bernhard; Meinl, Edgar

1998-01-01

Viruses have evolved different strategies to interfere with host cell apoptosis. Herpesvirus saimiri (HVS) and other lymphotropic herpesviruses code for proteins that are homologous to the cellular antiapoptotic Bcl-2. In this study HVS-Bcl-2 was stably expressed in the human leukemia cell line Jurkat and in the murine T-cell hybridoma DO to assess its antiapoptotic spectrum and to gain further insight into its mode of action. HVS- Bcl-2 prevented apoptosis that occurs as a result of a disturbance of intracellular homeostasis by, for example, DNA damage or menadione, which gives rise to oxygen radicals. In Jurkat cells, HVS-Bcl-2 also inhibited apoptosis mediated by the death receptor CD95. In DO cells, HVS-Bcl-2 did not interfere with CD95-mediated apoptosis but blocked dexamethasone-induced cell death. Mitochondrial damage is a central coordinating event in apoptosis induced by different stimuli. To assess the integrity of mitochondria, we used rhodamine 123, which is released upon disturbance of the mitochondrial membrane potential, and determined the release of cytochrome c into the cytosol. Both signs of mitochondrial damage were prevented by HVS-Bcl-2. This viral protein also inhibited the generation of caspase-3-like DEVDase activity and blocked the cleavage of poly(ADP-ribose) polymerase, a natural substrate of caspase-3-like proteases. In conclusion, HVS-Bcl-2 protects against a great variety of apoptotic stimuli, stabilizes mitochondria, and acts upstream of the generation of caspase-3-like activity. PMID:9621051

Modulation of eukaryotic cell apoptosis by members of the bacterial order Actinomycetales.

PubMed

Barry, Daniel P; Beaman, Blaine L

2006-10-01

Apoptosis, or programmed cell death, is normally responsible for the orderly elimination of aged or damaged cells, and is a necessary part of the homeostasis and development of multicellular organisms. Some pathogenic bacteria can disrupt this process by triggering excess apoptosis or by preventing it when appropriate. Either event can lead to disease. There has been extensive research into the modulation of host cell death by microorganisms, and several reviews have been published on the phenomenon. Rather than covering the entire field, this review focuses on the dysregulation of host cell apoptosis by members of the order Actinomycetales, containing the genera Corynebacterium, Mycobacterium, Rhodococcus, and Nocardia.

The Morphogenetic Role of Apoptosis.

PubMed

Monier, Bruno; Suzanne, Magali

2015-01-01

Beyond safeguarding the organism from cell misbehavior and controlling cell number, apoptosis (or programmed cell death) plays key roles during animal development. In particular, it has long been acknowledged that apoptosis participates in tissue remodeling. Yet, until recently, this contribution to morphogenesis was considered as "passive," consisting simply in the local removal of unnecessary cells leading to a new shape. In recent years, applying live imaging methods to study the dynamics of apoptosis in various contexts has considerably modified our vision, revealing that in fact, dying cells remodel their neighborhood actively. Here, we first focus on the intrinsic cellular properties of apoptotic cells during their dismantling, in particular the role of the cytoskeleton during their characteristic morphological changes. Second, we review the various roles of apoptosis during developmental morphogenetic processes and pinpoint the crucial role of live imaging in revealing new concepts, in particular apoptosis as a generator of mechanical forces to control tissue dynamics. © 2015 Elsevier Inc. All rights reserved.

Wavelength-dependent backscattering measurements for quantitative monitoring of apoptosis, Part 1: early and late spectral changes are indicative of the presence of apoptosis in cell cultures

NASA Astrophysics Data System (ADS)

Mulvey, Christine S.; Zhang, Kexiong; Liu, Wei-Han Bobby; Waxman, David J.; Bigio, Irving J.

2011-11-01

Apoptosis, a form of programmed cell death with unique morphological and biochemical features, is dysregulated in cancer and is activated by many cancer chemotherapeutic drugs. Noninvasive assays for apoptosis in cell cultures can aid in screening of new anticancer agents. We have previously demonstrated that elastic scattering spectroscopy can monitor apoptosis in cell cultures. In this report we present data on monitoring the detailed time-course of scattering changes in a Chinese hamster ovary (CHO) and PC-3 prostate cancer cells treated with staurosporine to induce apoptosis. Changes in the backscattering spectrum are detectable within 10 min, and continue to progress up to 48 h after staurosporine treatment, with the magnitude and kinetics of scattering changes dependent on inducer concentration. Similar responses were observed in CHO cells treated with several other apoptosis-inducing protocols. Early and late scattering changes were observed under conditions shown to induce apoptosis via caspase activity assay and were absent under conditions where apoptosis was not induced. Finally, blocking caspase activity and downstream apoptotic morphology changes prevented late scattering changes. These observations demonstrate that early and late changes in wavelength-dependent backscattering correlate with the presence of apoptosis in cell cultures and that the late changes are specific to apoptosis.

Modern Radiobiology: Contention Of Concepts: Advanced Technology And Development Of Effective Prophylaxis, Prevention And Treatment Of Biological Consequences After Irradiation.

NASA Astrophysics Data System (ADS)

Popov, Dmitri; Maliev, Vecheslav; Jones, Jeffrey

"Alle Ding' sind Gift, und nichts ohn' Gift; allein die Dosis macht, daß ein Ding kein Gift ist." Paracelsus Philippus Aureolus Theophrastus Bombastus von Hohenheim. Key worlds: Apoptosis, Necrosis, Domains associated with Cell Death, Caspase (catalytic) Domains, Death Domains (DDs), Death Effector Domains (DEDs), Caspase-Associated Recruitment Domains (CARDs, BIR Domains (IAPs), Bcl-2 Homology (BH) Domains, death ligands - TRAIL (TNF-Related Apoptosis-Inducing Ligand), FasL (Fas Ligand), TNFalpha (Tumor Necrosis Factor alpha), Toll-like receptors (TLR), Systemic inflammatory response syndrome (SIRS), Toxic Multiple Organ Injury (TMOI), Toxic Multiple Organ Dysfunction Syndromes (TMODS), Toxic Multiple Organ Failure (TMOF), Anaphylatoxins, or complement peptides; membrane attack complex (MAC), ROS - Reactive Oxygen Species; ASMase, acid sphingomyelinase; Neurotoxins, Cytotoxins, Haemotoxins. Introduction: Radiation affects many cell structures, organelles and metabolic pathways. Different doses and types of radiation ( gamma-radiation, neutron, heavy ion radiation) progress to reversible and irreversible forms of cell injury. Consideration: Apoptosis and Necrosis, major forms of post-radiation cell death, can be initiated and modulated by programmed control and proceed by similar or different pathways.[Akadi et al.,1993, Dunlacht J., et al. 1999] Radiation induced cell death by triggering apoptosis pathways was described in many articles and supported by many scientists. [Rio et al. 2002, Rakesh et al. 1997.] However some authors present results that two distinct pathways can initiate or apoptotic or necrotic responses: the death receptors and mitochondrial pathways.

Centriole, differentiation, and senescence.

PubMed

Tkemaladze, J; Chichinadze, K

2010-01-01

Irreversible differentiation (change of morphogenetic status) and programmed death (apoptosis) are observed only in somatic cells, and cell division is the only way by which the morphogenetic status of the offspring cells may be modified. It is known that there is a fixed limit to the number of possible cell divisions, the so-called Hayflick limit. Existing links between cell division, differentiation, and apoptosis make it possible to conclude that all of these processes could be controlled by a single self-reproducing structure. Potential candidates for this replicable structure in a somatic cell are the chromosomes, mitochondria (both contain DNA), and centrioles. Centrioles (a diplosome, or pair of centrioles) are the most likely unit that can fully regulate the processes of irreversible differentiation, determination, and modification of the morphogenetic status. Centrioles may contain differently encoded RNA molecules stacked in a definite order, and during mitosis, these RNA molecules are released one by one into the cytoplasm. In the presence of reverse transcriptase and endonuclease, processing of this RNA presumably changes the status of repressed and potentially active genes and, subsequently, the morphogenetic status of a cell.

Necroptosis in cancer: An angel or a demon?

PubMed

Wang, Tianzhen; Jin, Yinji; Yang, Weiwei; Zhang, Lei; Jin, Xiaoming; Liu, Xi; He, Yan; Li, Xiaobo

2017-06-01

In the past few decades, apoptosis has been regarded as the only form of programmed cell death. However, the traditional view has been challenged by the identification of several forms of regulated necrosis, including necroptosis. Necroptosis is typified by a necrotic cell death morphology and is controlled by RIP1, RIP3, and mixed lineage kinase domain-like protein. The physiological role of necroptosis is to serve as a "fail-safe" form of cell death for cells that fail to undergo apoptosis during embryonic development and disease defense. Currently, established studies have indicated that necroptosis is involved in cancer initiation and progression. Although elevated necroptosis contributes to cancer cell death, extensive cell death also increases the risk of proliferation and metastasis of the surviving cells by inducing the generation reactive oxygen species, activation of inflammation, and suppression of the immune response. Thus, questions regarding the overall impact of necroptosis on cancer remain open. In this review, we introduce the basic knowledge regarding necroptosis, summarize its dual effects on cancer progression, and analyze its advantages and disadvantages in clinical applications.

Variability in apoptotic response to poliovirus infection.

PubMed

Romanova, Lyudmila I; Belov, George A; Lidsky, Peter V; Tolskaya, Elena A; Kolesnikova, Marina S; Evstafieva, Alexandra G; Vartapetian, Andrey B; Egger, Denise; Bienz, Kurt; Agol, Vadim I

2005-01-20

In several cell types, poliovirus activates the apoptotic program, implementation of which is suppressed by viral antiapoptotic functions. In such cells, productive infection leads to a necrotic cytopathic effect (CPE), while abortive reproduction, associated with inadequate viral antiapoptotic functions, results in apoptosis. Here, we describe two other types of cell response to poliovirus infection. Murine L20B cells expressing human poliovirus receptor responded to the infection by both CPE and apoptosis concurrently. Interruption of productive infection decreased rather than increased the proportion of apoptotic cells. Productive infection was accompanied by the early efflux of cytochrome c from the mitochondria in a proportion of cells and by activation of DEVD-specific caspases. Inactivation of caspase-9 resulted in a marked, but incomplete, prevention of the apoptotic response of these cells to viral infection. Thus, the poliovirus-triggered apoptotic program in L20B cells was not completely suppressed by the viral antiapoptotic functions. In contrast, human rhabdomyosarcoma RD cells did not develop appreciable apoptosis during productive or abortive infection, exhibiting inefficient efflux of cytochrome c from mitochondria and no marked activation of DEVD-specific caspases. The cells were also refractory to several nonviral apoptosis inducers. Nevertheless, typical caspase-dependent signs of apoptosis in a proportion of RD cells were observed after cessation of viral reproduction. Such "late" apoptosis was also observed in productively infected HeLa cells. In addition, a tiny proportion of all studied cells were TUNEL positive even in the presence of a caspase inhibitor. Degradation of DNA in such cells appeared to be a postmortem phenomenon. Biological relevance of variable host responses to viral infection is discussed.

Activation of BAD by therapeutic inhibition of epidermal growth factor receptor and transactivation by insulin-like growth factor receptor.

PubMed

Gilmore, Andrew P; Valentijn, Anthony J; Wang, Pengbo; Ranger, Ann M; Bundred, Nigel; O'Hare, Michael J; Wakeling, Alan; Korsmeyer, Stanley J; Streuli, Charles H

2002-08-02

Novel cancer chemotherapeutics are required to induce apoptosis by activating pro-apoptotic proteins. Both epidermal growth factor (EGF) and insulin-like growth factor (IGF) provide potent survival stimuli in many epithelia, and activation of their receptors is commonly observed in solid human tumors. Here we demonstrate that blockade of the EGF receptor by a new drug in phase III clinical trails for cancer, ZD1839, potently induces apoptosis in mammary epithelial cell lines and primary cultures, as well as in a primary pleural effusion from a breast cancer patient. We identified the mechanism of apoptosis induction by ZD1839. We showed that it prevents cell survival by activating the pro-apoptotic protein BAD. Moreover, we demonstrate that IGF transactivates the EGF receptor and that ZD1839 blocks IGF-mediated phosphorylation of MAPK and BAD. Many cancer therapies kill tumor cells by inducing apoptosis as a consequence of targeting DNA; however, the threshold at which apoptosis can be triggered through DNA damage is often different from that in normal cells. Our results indicate that by targeting a growth factor-mediated survival signaling pathway, BAD phosphorylation can be manipulated therapeutically to induce apoptosis.

Radiation and stress-induced apoptosis: A role for Fas/Fas ligand interactions

PubMed Central

Reap, Elizabeth A.; Roof, Kevin; Maynor, Kenrick; Borrero, Michelle; Booker, Jessica; Cohen, Philip L.

1997-01-01

The lpr gene encodes a defective form of Fas, a cell surface protein that mediates apoptosis. This defect blocks apoptotic deletion of autoreactive T and B cells, leading to lymphoproliferation and lupus-like autoantibody production. The effects of the lpr Fas mutation on other kinds of physiologically relevant apoptosis are largely undocumented. To assess whether some of the apoptosis known to occur after ionizing radiation might be mediated by Fas/Fas ligand (FasL) interactions, we quantitated in vitro apoptosis by flow cytometry measurement of DNA content in splenic T and B cells from irradiated 5- to 8-month-old B6/lpr mice. Total apoptosis of both lpr and control cells was substantial after treatment; however there was a significant difference between B6 (73%) and lpr (25%) lymphocyte apoptosis. Thy1, CD4, CD8, and IgM cells from lpr showed much lower levels of apoptosis than control cells after irradiation. Apoptosis induced by heat shock was also impaired in lpr. The finding that γ-irradiation increased Fas expression on B6 cells and that irradiation-induced apoptosis could be blocked with a Fas–Fc fusion protein further supported the possible involvement of Fas in this form of apoptosis. Fas/FasL interactions may thus play an important role in identifying and eliminating damaged cells after γ-irradiation and other forms of injury. PMID:9159145

METHYLMERCURY BUT NOT MERCURIC CHLORIDE INDUCES APOPTOTIC CELL DEATH IN PC12 CELLS.

EPA Science Inventory

Normal development of the nervous system requires the process of apoptosis, a form of programmed cell death, to remove superfluous neurons. Abnormal patterns of apoptosis may be a consequence of exposure to environmental neurotoxicants leading to a disruption in the tightly regul...

Characterization of Apoptosis Signaling Cascades During the Differentiation Process of Human Neural ReNcell VM Progenitor Cells In Vitro.

PubMed

Jaeger, Alexandra; Fröhlich, Michael; Klum, Susanne; Lantow, Margareta; Viergutz, Torsten; Weiss, Dieter G; Kriehuber, Ralf

2015-11-01

Apoptosis is an essential physiological process accompanying the development of the central nervous system and human neurogenesis. However, the time scale and the underlying molecular mechanisms are yet poorly understood. Due to this fact, we investigated the functionality and general inducibility of apoptosis in the human neural ReNcell VM progenitor cell line during differentiation and also after exposure to staurosporine (STS) and ultraviolet B (UVB) irradiation. Transmission light microscopy, flow cytometry, and Western-/Immunoblot analysis were performed to compare proliferating and differentiating, in addition to STS- and UVB-treated cells. In particular, from 24 to 72 h post-initiation of differentiation, G0/G1 cell cycle arrest, increased loss of apoptotic cells, activation of pro-apoptotic BAX, Caspase-3, and cleavage of its substrate PARP were observed during cell differentiation and, to a higher extent, after treatment with STS and UVB. We conclude that redundant or defective cells are eliminated by apoptosis, while otherwise fully differentiated cells were less responsive to apoptosis induction by STS than proliferating cells, likely as a result of reduced APAF-1 expression, and increased levels of BCL-2. These data provide the evidence that apoptotic mechanisms in the neural ReNcell VM progenitor cell line are not only functional, but also inducible by external stimuli like growth factor withdrawal or treatment with STS and UVB, which marks this cell line as a suitable model to investigate apoptosis signaling pathways in respect to the differentiation processes of human neural progenitor cells in vitro.

Tumor cell-released TLR4 ligands stimulate Gr-1+CD11b+F4/80+ cells to induce apoptosis of activated T cells.

PubMed

Liu, Yan-Yan; Sun, Ling-Cong; Wei, Jing-Jing; Li, Dong; Yuan, Ye; Yan, Bin; Liang, Zhi-Hui; Zhu, Hui-Fen; Xu, Yong; Li, Bo; Song, Chuan-Wang; Liao, Sheng-Jun; Lei, Zhang; Zhang, Gui-Mei; Feng, Zuo-Hua

2010-09-01

Gr-1(+)CD11b(+)F4/80(+) cells play important roles in tumor development and have a negative effect on tumor immunotherapy. So far, the mechanisms underlying the regulation of their immunosuppressive phenotype by classical and alternative macrophage activation stimuli are not well elucidated. In this study, we found that molecules from necrotic tumor cells (NTC-Ms) stimulated Gr-1(+)CD11b(+)F4/80(+) cells to induce apoptosis of activated T cells but not nonstimulated T cells. The apoptosis-inducing capacity was determined by higher expression levels of arginase I and IL-10 relative to those of NO synthase 2 and IL-12 in Gr-1(+)CD11b(+)F4/80(+) cells, which were induced by NTC-Ms through TLR4 signaling. The apoptosis-inducing capacity of NTC-Ms-stimulated Gr-1(+)CD11b(+)F4/80(+) cells could be enhanced by IL-10. IFN-gamma may reduce the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells only if their response to IFN-gamma was not attenuated. However, the potential of Gr-1(+)CD11b(+)F4/80(+) cells to express IL-12 in response to IFN-gamma could be attenuated by tumor, partially due to the existence of active STAT3 in Gr-1(+)CD11b(+)F4/80(+) cells and NTC-Ms from tumor. In this situation, IFN-gamma could not effectively reduce the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells. Tumor immunotherapy with 4-1BBL/soluble programmed death-1 may significantly reduce, but not abolish the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells in local microenvironment. Blockade of TLR4 signaling could further reduce the apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells and enhance the suppressive effect of 4-1BBL/soluble form of programmed death-1 on tumor growth. These findings indicate the relationship of distinct signaling pathways with apoptosis-inducing capacity of Gr-1(+)CD11b(+)F4/80(+) cells and emphasize the importance of blocking TLR4 signaling to prevent the induction of T cell apoptosis by Gr-1(+)CD11b(+)F4/80(+) cells.

Induction of apoptosis in rhabdomyosarcoma cells through down-regulation of PAX proteins

PubMed Central

Bernasconi, Michele; Remppis, Andrew; Fredericks, William J.; Rauscher, Frank J.; Schäfer, Beat W.

1996-01-01

The expression of a number of human paired box-containing (PAX) genes has been correlated with various types of tumors. Novel fusion genes encoding chimeric fusion proteins have been found in the pediatric malignant tumor alveolar rhabdomyosarcoma (RMS). They are generated by two chromosomal translocations t(2;13) and t(1;13) juxtaposing PAX3 or PAX7, respectively, with a forkhead domain gene FKHR. Here we describe that specific down-regulation of the t(2;13) translocation product in alveolar RMS cells by antisense oligonucleotides results in reduced cellular viability. Cells of embryonal RMS, the other major histiotype of this tumor, were found to express either wild type PAX3 or PAX7 at elevated levels when compared with primary human myoblasts. Treatment of corresponding embryonal RMS cells with antisense olignucleotides directed against the mRNA translational start site of either one of these two transcription factors similarly triggers cell death, which is most likely due to induction of apoptosis. Retroviral mediated ectopic expression of mouse Pax3 in a PAX7 expressing embryonal RMS cell line could partially rescue antisense induced apoptosis. These data suggest that the PAX3/FKHR fusion gene and wild-type PAX genes play a causative role in the formation of RMS and presumably other tumor types, possibly by suppressing the apoptotic program that would normally eliminate these cells. PMID:8917562

Neuroprotective Effect of Resveratrol on Acute Brain Ischemia Reperfusion Injury by Measuring Annexin V, p53, Bcl-2 Levels in Rats.

PubMed

Kizmazoglu, Ceren; Aydin, Hasan Emre; Sevin, Ismail Ertan; Kalemci, Orhan; Yüceer, Nurullah; Atasoy, Metin Ant

2015-12-01

Cerebral ischemia is as a result of insufficient cerebral blood flow for cerebral metabolic functions. Resveratrol is a natural phytoalexin that can be extracted from grape's skin and had potent role in treating the cerebral ischemia. Apoptosis, a genetically programmed cellular event which occurs after ischemia and leads to biochemical and morphological changes in cells. There are some useful markers for apoptosis like Bcl-2, bax, and p53. The last reports, researchers verify the apoptosis with early markers like Annexin V. We preferred in this experimental study a model of global cerebral infarction which was induced by bilateral common carotid artery occlusion method. Rats were randomly divided into 4 groups : sham, ischemia-reperfusion (I/R), I/R plus 20 mg/kg resveratrol and I/R plus 40 mg/kg resveratrol. Statistical analysis was performed using Sigmastat 3.5 ve IBM SPSS Statistics 20. We considered a result significant when p<0.001. After administration of resveratrol, Bcl-2 and Annexin levels were significantly increased (p<0.001). Depending on the dose of resveratrol, Bcl2 levels increased, p53 levels decreased but Annexin V did not effected. P53 levels were significantly increased in ishemia group, so apoptosis is higher compared to other groups. In the acute period, Annexin V levels misleading us because the apoptotic cell counts could not reach a certain level. Therefore we should support our results with bcl-2 and p53.

Deficiency in methionine, tryptophan, isoleucine, or choline induces apoptosis in cultured cells.

PubMed

Yen, Chi-Liang E; Mar, Mei-Heng; Craciunescu, Corneliu N; Edwards, Lloyd J; Zeisel, Steven H

2002-07-01

Cells in culture die by apoptosis when deprived of the essential nutrient choline. We now report that cells (both proliferating PC12 cells and postmitotic neurons isolated from fetal rat brains) undergo apoptosis when deprived of other individual essential nutrients (methionine, tryptophan or isoleucine). In PC12 cells, deficiencies of each nutrient independently led to ceramide accumulation and to caspase activation, both recognized signals of several apoptotic pathways. A similar profile of caspases was activated in PC12 cells deprived of choline, methionine, tryptophan or isoleucine. More than one caspase was involved and these caspases appeared to transmit parallel signals for apoptosis induction because only broad-spectrum caspase inhibitors, but not inhibitors for specific individual caspases inhibited apoptosis in choline- or methionine-deprived cells. The induction of these caspase-dependent apoptosis pathways likely did not involve the same upstream signals. Choline deficiency perturbed choline metabolism but did not affect protein synthesis, whereas amino acid deficiencies inhibited protein synthesis but did not perturb choline metabolism. In addition, a subclone of PC12 cells that was resistant to choline deficiency-induced apoptosis was not resistant to tryptophan deficiency-induced apoptosis. These observations suggest that deficiency of each studied nutrient activates different pathways for signaling apoptosis that ultimately converge on a common execution pathway.

Ligation of Siglec-8: a selective mechanism for induction of human eosinophil apoptosis.

PubMed

Nutku, Esra; Aizawa, Hideyuki; Hudson, Sherry A; Bochner, Bruce S

2003-06-15

Sialic acid binding immunoglobulin-like lectin 8 (Siglec-8), which exists in 2 isoforms including one possessing cytoplasmic tyrosine motifs, is expressed only on human eosinophils, basophils, and mast cells. Until now, its function was unknown. Here we define a novel function of Siglec-8 on eosinophils. Siglec-8 cross-linking with antibodies rapidly generated caspase-3-like activity and reduced eosinophil viability through induction of apoptosis. The pancaspase inhibitor benzyloxycarbonyl (Cbz)-Val-Ala-Asp-(Ome)-fluoromethyl ketone (zVAD-FMK) completely blocked this response, implicating caspases in Siglec-8 cross-linking-induced apoptosis. Eosinophil survival-promoting cytokines such as interleukin 5 (IL-5) and granulocyte-macrophage colony-stimulating factor (GM-CSF) failed to block apoptosis and instead enhanced the sensitivity of eosinophils to undergo apoptosis in response to Siglec-8 antibody. Siglec-8 activation may provide a useful therapeutic approach to reduce numbers of eosinophils (and perhaps basophils and mast cells) in disease states where these cells are important.

An apoptosis targeted stimulus with nanosecond pulsed electric fields (nsPEFs) in E4 squamous cell carcinoma.

PubMed

Ren, Wei; Beebe, Stephen J

2011-04-01

Stimuli directed towards activation of apoptosis mechanisms are an attractive approach to eliminate evasion of apoptosis, a ubiquitous cancer hallmark. In these in vitro studies, kinetics and electric field thresholds for several apoptosis characteristics are defined in E4 squamous carcinoma cells (SCC) exposed to ten 300 ns pulses with increasing electric fields. Cell death was >95% at the highest electric field and coincident with phosphatidylserine externalization, caspase and calpain activation in the presence and absence of cytochrome c release, decreases in Bid and mitochondria membrane potential (Δψm) without apparent changes reactive oxygen species levels or in Bcl2 and Bclxl levels. Bid cleavage was caspase-dependent (55-60%) and calcium-dependent (40-45%). Intracellular calcium as an intrinsic mechanism and extracellular calcium as an extrinsic mechanism were responsible for about 30 and 70% of calcium dependence for Bid cleavage, respectively. The results reveal electric field-mediated cell death induction and progression, activating pro-apoptotic-like mechanisms and affecting plasma membrane and intracellular functions, primarily through extrinsic-like pathways with smaller contributions from intrinsic-like pathways. Nanosecond second pulsed electric fields trigger heterogeneous cell death mechanisms in E4 SCC populations to delete them, with caspase-associated cell death as a predominant, but not an unaccompanied event.

How Trypanosoma cruzi handles cell cycle arrest promoted by camptothecin, a topoisomerase I inhibitor.

PubMed

Zuma, Aline Araujo; Mendes, Isabela Cecília; Reignault, Lissa Catherine; Elias, Maria Carolina; de Souza, Wanderley; Machado, Carlos Renato; Motta, Maria Cristina M

2014-02-01

The protozoan Trypanosoma cruzi is the etiological agent of Chagas disease, which affects approximately 8 million people in Latin America. This parasite contains a single nucleus and a kinetoplast, which harbors the mitochondrial DNA (kDNA). DNA topoisomerases act during replication, transcription and repair and modulate DNA topology by reverting supercoiling in the DNA double-strand. In this work, we evaluated the effects promoted by camptothecin, a topoisomerase I inhibitor that promotes protozoan proliferation impairment, cell cycle arrest, ultrastructure alterations and DNA lesions in epimastigotes of T. cruzi. The results showed that inhibition of cell proliferation was reversible only at the lowest drug concentration (1μM) used. The unpacking of nuclear heterochromatin and mitochondrion swelling were the main ultrastructural modifications observed. Inhibition of parasite proliferation also led to cell cycle arrest, which was most likely caused by nuclear DNA lesions. Following camptothecin treatment, some of the cells restored their DNA, whereas others entered early apoptosis but did not progress to late apoptosis, indicating that the protozoa stay alive in a "senescence-like" state. This programmed cell death may be associated with a decrease in mitochondrial membrane potential and an increase in the production of reactive oxygen species. Taken together, these results indicate that the inhibition of T. cruzi proliferation is related to events capable of affecting cell cycle, DNA organization and mitochondrial activity. Copyright © 2014. Published by Elsevier B.V.

Activin A prevents neuron-like PC12 cell apoptosis after oxygen-glucose deprivation☆

PubMed Central

Xu, Guihua; He, Jinting; Guo, Hongliang; Mei, Chunli; Wang, Jiaoqi; Li, Zhongshu; Chen, Han; Mang, Jing; Yang, Hong; Xu, Zhongxin

2013-01-01

In this study, PC12 cells were induced to differentiate into neuron-like cells using nerve growth factor, and were subjected to oxygen-glucose deprivation. Cells were treated with 0, 10, 20, 30, 50, 100 ng/mL exogenous Activin A. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay and Hoechst 33324 staining showed that the survival percentage of PC12 cells significantly decreased and the rate of apoptosis significantly increased after oxygen-glucose deprivation. Exogenous Activin A significantly increased the survival percentage of PC12 cells in a dose-dependent manner. Reverse transcription-PCR results revealed a significant increase in Activin receptor IIA, Smad3 and Smad4 mRNA levels, which are key sites in the Activin A/Smads signaling pathway, in neuron-like cells subjected to oxygen-glucose deprivation, while mRNA expression of the apoptosis-regulation gene caspase-3 decreased. Our experimental findings indicate that exogenous Activin A plays an anti-apoptotic role and protects neurons by means of activating the Activin A/Smads signaling pathway. PMID:25206395

The selective progesterone receptor modulator CDB4124 inhibits proliferation and induces apoptosis in uterine leiomyoma cells.

PubMed

Luo, Xia; Yin, Ping; Coon V, John S; Cheng, You-Hong; Wiehle, Ronald D; Bulun, Serdar E

2010-05-15

To evaluate the effects of selective P receptor (PR) modulator CDB4124 on cell proliferation and apoptosis in cultured human uterine leiomyoma smooth muscle (LSM) cells and control myometrial smooth muscle (MSM) cells in matched uteri. Laboratory research. Academic medical center. Premenopausal women (n = 12) undergoing hysterectomy for leiomyoma-related symptoms. Treatment of primary LSM and MSM cells with CDB4124 (10(-8)-10(-6) M) or vehicle for 24, 48, or 72 hours. Western blot for protein expression of proliferating cell nuclear antigen, cleaved polyadenosine 5'-diphosphate-ribose polymerase, Bcl-2, and Krüppel-like transcription factor 11; 93-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide (MTT) assay to evaluate viable cell numbers; and real-time polymerase chain reaction (PCR) to quantify messenger RNA (mRNA) levels. Treatment with CDB4124 significantly decreased levels of the proliferation marker proliferating cell nuclear antigen, the number of viable LSM cells, and the antiapoptotic protein Bcl-2. On the other hand, treatment with CDB4124 increased levels of the apoptosis marker cleaved polyadenosine 5'-diphosphate-ribose polymerase and the tumor suppressor Krüppel-like transcription factor 11 in a dose- and time-dependent manner in LSM cells. In matched MSM cells, however, CDB4124 did not affect cell proliferation or apoptosis. CDB4124 selectively inhibits proliferation and induces apoptosis in LSM but not in MSM cells. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Apoptosis in capillary endothelial cells in ageing skeletal muscle

PubMed Central

Wang, Huijuan; Listrat, Anne; Meunier, Bruno; Gueugneau, Marine; Coudy-Gandilhon, Cécile; Combaret, Lydie; Taillandier, Daniel; Polge, Cécile; Attaix, Didier; Lethias, Claire; Lee, Kijoon; Goh, Kheng Lim; Béchet, Daniel

2014-01-01

The age-related loss of skeletal muscle mass and function (sarcopenia) is a consistent hallmark of ageing. Apoptosis plays an important role in muscle atrophy, and the intent of this study was to specify whether apoptosis is restricted to myofibre nuclei (myonuclei) or occurs in satellite cells or stromal cells of extracellular matrix (ECM). Sarcopenia in mouse gastrocnemius muscle was characterized by myofibre atrophy, oxidative type grouping, delocalization of myonuclei and ECM fibrosis. Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) indicated a sharp rise in apoptosis during ageing. TUNEL coupled with immunostaining for dystrophin, paired box protein-7 (Pax7) or laminin-2α, respectively, was used to identify apoptosis in myonuclei, satellite cells and stromal cells. In adult muscle, apoptosis was not detected in myofibres, but was restricted to stromal cells. Moreover, the age-related rise in apoptotic nuclei was essentially due to stromal cells. Myofibre-associated apoptosis nevertheless occurred in old muscle, but represented < 20% of the total muscle apoptosis. Specifically, apoptosis in old muscle affected a small proportion (0.8%) of the myonuclei, but a large part (46%) of the Pax7+ satellite cells. TUNEL coupled with CD31 immunostaining further attributed stromal apoptosis to capillary endothelial cells. Age-dependent rise in apoptotic capillary endothelial cells was concomitant with altered levels of key angiogenic regulators, perlecan and a perlecan domain V (endorepellin) proteolytic product. Collectively, our results indicate that sarcopenia is associated with apoptosis of satellite cells and impairment of capillary functions, which is likely to contribute to the decline in muscle mass and functionality during ageing. PMID:24245531

FLIP the Switch: Regulation of Apoptosis and Necroptosis by cFLIP

PubMed Central

Tsuchiya, Yuichi; Nakabayashi, Osamu; Nakano, Hiroyasu

2015-01-01

cFLIP (cellular FLICE-like inhibitory protein) is structurally related to caspase-8 but lacks proteolytic activity due to multiple amino acid substitutions of catalytically important residues. cFLIP protein is evolutionarily conserved and expressed as three functionally different isoforms in humans (cFLIPL, cFLIPS, and cFLIPR). cFLIP controls not only the classical death receptor-mediated extrinsic apoptosis pathway, but also the non-conventional pattern recognition receptor-dependent apoptotic pathway. In addition, cFLIP regulates the formation of the death receptor-independent apoptotic platform named the ripoptosome. Moreover, recent studies have revealed that cFLIP is also involved in a non-apoptotic cell death pathway known as programmed necrosis or necroptosis. These functions of cFLIP are strictly controlled in an isoform-, concentration- and tissue-specific manner, and the ubiquitin-proteasome system plays an important role in regulating the stability of cFLIP. In this review, we summarize the current scientific findings from biochemical analyses, cell biological studies, mathematical modeling, and gene-manipulated mice models to illustrate the critical role of cFLIP as a switch to determine the destiny of cells among survival, apoptosis, and necroptosis. PMID:26694384

Effects of Nano-CeO₂ with Different Nanocrystal Morphologies on Cytotoxicity in HepG2 Cells.

PubMed

Wang, Lili; Ai, Wenchao; Zhai, Yanwu; Li, Haishan; Zhou, Kebin; Chen, Huiming

2015-09-02

Cerium oxide nanoparticles (nano-CeO₂) have been reported to cause damage and apoptosis in human primary hepatocytes. Here, we compared the toxicity of three types of nano-CeO₂ with different nanocrystal morphologies (cube-, octahedron-, and rod-like crystals) in human hepatocellular carcinoma cells (HepG2). The cells were treated with the nano-CeO₂ at various concentrations (6.25, 12.5, 25, 50, 100 μg/mL). The crystal structure, size and morphology of nano-CeO₂ were investigated by X-ray diffractometry and transmission electron microscopy. The specific surface area was detected using the Brunauer, Emmet and Teller method. The cellular morphological and internal structure were observed by microscopy; apoptotic alterations were measured using flow cytometry; nuclear DNA, mitochondrial membrane potential (MMP), reactive oxygen species (ROS) and glutathione (GSH) in HepG2 cells were measured using high content screening technology. The scavenging ability of hydroxyl free radicals and the redox properties of the nano-CeO₂ were measured by square-wave voltammetry and temperature-programmed-reduction methods. All three types of nano-CeO₂ entered the HepG2 cells, localized in the lysosome and cytoplasm, altered cellular shape, and caused cytotoxicity. The nano-CeO₂ with smaller specific surface areas induced more apoptosis, caused an increase in MMP, ROS and GSH, and lowered the cell's ability to scavenge hydroxyl free radicals and antioxidants. In this work, our data demonstrated that compared with cube-like and octahedron-like nano-CeO₂, the rod-like nano-CeO₂ has lowest toxicity to HepG2 cells owing to its larger specific surface areas.

Die another way – non-apoptotic mechanisms of cell death

PubMed Central

Tait, Stephen W. G.; Ichim, Gabriel; Green, Douglas R.

2014-01-01

ABSTRACT Regulated, programmed cell death is crucial for all multicellular organisms. Cell death is essential in many processes, including tissue sculpting during embryogenesis, development of the immune system and destruction of damaged cells. The best-studied form of programmed cell death is apoptosis, a process that requires activation of caspase proteases. Recently it has been appreciated that various non-apoptotic forms of cell death also exist, such as necroptosis and pyroptosis. These non-apoptotic cell death modalities can be either triggered independently of apoptosis or are engaged should apoptosis fail to execute. In this Commentary, we discuss several regulated non-apoptotic forms of cell death including necroptosis, autophagic cell death, pyroptosis and caspase-independent cell death. We outline what we know about their mechanism, potential roles in vivo and define outstanding questions. Finally, we review data arguing that the means by which a cell dies actually matters, focusing our discussion on inflammatory aspects of cell death. PMID:24833670

HAMLET triggers apoptosis but tumor cell death is independent of caspases, Bcl-2 and p53.

PubMed

Hallgren, O; Gustafsson, L; Irjala, H; Selivanova, G; Orrenius, S; Svanborg, C

2006-02-01

HAMLET (Human alpha-lactalbumin Made Lethal to Tumor cells) triggers selective tumor cell death in vitro and limits tumor progression in vivo. Dying cells show features of apoptosis but it is not clear if the apoptotic response explains tumor cell death. This study examined the contribution of apoptosis to cell death in response to HAMLET. Apoptotic changes like caspase activation, phosphatidyl serine externalization, chromatin condensation were detected in HAMLET-treated tumor cells, but caspase inhibition or Bcl-2 over-expression did not prolong cell survival and the caspase response was Bcl-2 independent. HAMLET translocates to the nuclei and binds directly to chromatin, but the death response was unrelated to the p53 status of the tumor cells. p53 deletions or gain of function mutations did not influence the HAMLET sensitivity of tumor cells. Chromatin condensation was partly caspase dependent, but apoptosis-like marginalization of chromatin was also observed. The results show that tumor cell death in response to HAMLET is independent of caspases, p53 and Bcl-2 even though HAMLET activates an apoptotic response. The use of other cell death pathways allows HAMLET to successfully circumvent fundamental anti-apoptotic strategies that are present in many tumor cells.

Regulation of Apoptosis during Flavivirus Infection

PubMed Central

Okamoto, Toru; Suzuki, Tatsuya; Kusakabe, Shinji; Tokunaga, Makoto; Hirano, Junki; Miyata, Yuka; Matsuura, Yoshiharu

2017-01-01

Apoptosis is a type of programmed cell death that regulates cellular homeostasis by removing damaged or unnecessary cells. Its importance in host defenses is highlighted by the observation that many viruses evade, obstruct, or subvert apoptosis, thereby blunting the host immune response. Infection with Flaviviruses such as Japanese encephalitis virus (JEV), Dengue virus (DENV) and West Nile virus (WNV) has been shown to activate several signaling pathways such as endoplasmic reticulum (ER)-stress and AKT/PI3K pathway, resulting in activation or suppression of apoptosis in virus-infected cells. On the other hands, expression of some viral proteins induces or protects apoptosis. There is a discrepancy between induction and suppression of apoptosis during flavivirus infection because the experimental situation may be different, and strong links between apoptosis and other types of cell death such as necrosis may make it more difficult. In this paper, we review the effects of apoptosis on viral propagation and pathogenesis during infection with flaviviruses. PMID:28846635

Cancer’s Achilles’ Heel: Apoptosis and Necroptosis to the Rescue

PubMed Central

Dasgupta, Atreyi; Nomura, Motonari; Shuck, Ryan; Yustein, Jason

2016-01-01

Apoptosis, and the more recently discovered necroptosis, are two avenues of programmed cell death. Cancer cells survive by evading these two programs, driven by oncogenes and tumor suppressor genes. While traditional therapy using small molecular inhibitors and chemotherapy are continuously being utilized, a new and exciting approach is actively underway by identifying and using synergistic relationship between driver and rescue genes in a cancer cell. Through these synthetic lethal relationships, we are gaining tremendous insights into tumor vulnerabilities and specific molecular avenues for induction of programmed cell death. In this review, we briefly discuss the two cell death processes and cite examples of such synergistic manipulations for therapeutic purposes. PMID:28025559

Magnolol protects Ctenopharyngodon idella kidney cells from apoptosis induced by grass carp reovirus.

PubMed

Chen, Xiaohui; Hao, Kai; Yu, Xiaobo; Huang, Aiguo; Zhu, Bin; Wang, Gao-Xue; Ling, Fei

2018-03-01

Many natural products from medicinal plants are small molecular weight compounds with enormous structural diversity and show various biological activities. Magnolol is a biphenol compound rich in the stem bark of Magnolia officinalis Rehd et Wils., and is able to suppress viral replication in GCRV-infected grass carp (Ctenopharyngodon idella) kidney (CIK) cells in the previous study. In this study, in vivo studies demonstrated that magnolol was efficient to restrain the replication of GCRV and repair the low level of superoxide dismutase and total antioxidant capacity in serum at the non-toxic concentration in vivo. Furthermore, magnolol inhibited CIK cell apoptosis induced by GCRV and kept the normal cellular morphological structure, reflecting in the protection of CIK cells from cell swelling, the formation of apoptotic bodies, the disappearance of cellular morphology and nuclear fragmentation. Reverse transcript quantitative polymerase chain reaction (RT-qPCR) showed that magnolol facilitated the expression of apoptosis-inhibiting gene bcl-2, while suppressed the expression of apoptosis-promoting gene bax in GCRV-infected cells. Besides, RT-qPCR and enzyme activity assays proved that magnolol suppressed the expression of caspase 3, caspase 8 and caspase 9. Moreover, interactions between magnolol and proteins were predicted by using the STITCH program, which revealed that ten proteins including caspase 3, were involved in the apoptosis pathway, p53 signaling pathway, mitogen-activated protein kinase (MAPK) signaling pathway and toll-like receptor signaling pathway. Further assays were performed to test the effect of magnolol on apoptosis pathway, which showed that magnolol dramatically inhibited the activity of caspase 3 rather than those of caspase 8 and caspase 9. Collectively, the present study revealed that magnolol heightened the resistance of grass carp against GCRV infection and refrained GCRV-induced apoptosis, which may be attributed to the direct interaction of magnolol with caspase 3. The present results make a contribution to understanding the mechanisms by which small-molecule drugs possess antiviral activities, and lay a foundation for the development of broad-spectrum antiviral compounds in aquaculture industry. Copyright © 2017 Elsevier Ltd. All rights reserved.

Pan-Cancer Analysis Links PARK2 to BCL-XL-Dependent Control of Apoptosis.

PubMed

Gong, Yongxing; Schumacher, Steven E; Wu, Wei H; Tang, Fanying; Beroukhim, Rameen; Chan, Timothy A

2017-02-01

Mutation of the PARK2 gene can promote both Parkinson's Disease and cancer, yet the underlying mechanisms of how PARK2 controls cellular physiology is incompletely understood. Here, we show that the PARK2 tumor suppressor controls the apoptotic regulator BCL-XL and modulates programmed cell death. Analysis of approximately 10,000 tumor genomes uncovers a striking pattern of mutual exclusivity between PARK2 genetic loss and amplification of BCL2L1, implicating these genes in a common pathway. PARK2 directly binds to and ubiquitinates BCL-XL. Inactivation of PARK2 leads to aberrant accumulation of BCL-XL both in vitro and in vivo, and cancer-specific mutations in PARK2 abrogate the ability of the ubiquitin E3 ligase to target BCL-XL for degradation. Furthermore, PARK2 modulates mitochondrial depolarization and apoptosis in a BCL-XL-dependent manner. Thus, like genes at the nodal points of growth arrest pathways such as p53, the PARK2 tumor suppressor is able to exert its antiproliferative effects by regulating both cell cycle progression and programmed cell death. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Correlation of lung surface area to apoptosis and proliferation in human emphysema.

PubMed

Imai, K; Mercer, B A; Schulman, L L; Sonett, J R; D'Armiento, J M

2005-02-01

Pulmonary emphysema is associated with alterations in matrix proteins and protease activity. These alterations may be linked to programmed cell death by apoptosis, potentially influencing lung architecture and lung function. To evaluate apoptosis in emphysema, lung tissue was analysed from 10 emphysema patients and six individuals without emphysema (normal). Morphological analysis revealed alveolar cells in emphysematous lungs with convoluted nuclei characteristic of apoptosis. DNA fragmentation was detected using terminal deoxynucleotide transferase-mediated dUTP nick-end labelling (TUNEL) and gel electrophoresis. TUNEL revealed higher apoptosis in emphysematous than normal lungs. Markers of apoptosis, including active caspase-3, proteolytic fragment of poly (ADP-ribose) polymerase, Bax and Bad, were detected in emphysematous lungs. Linear regression showed that apoptosis was inversely correlated with surface area. Emphysematous lungs demonstrated lower surface areas and increased cell proliferation. There was no correlation between apoptosis and proliferation, suggesting that, although both events increase during emphysema, they are not in equilibrium, potentially contributing to reduced lung surface area. In summary, cell-based mechanisms associated with emphysematous parenchymal damage include increased apoptosis and cell proliferation. Apoptosis correlated with airspace enlargement, supporting epidemiological evidence of the progressive nature of emphysema. These data extend the understanding of cell dynamics and structural changes within the lung during emphysema pathogenesis.

Mitomycin C induces apoptosis in rheumatoid arthritis fibroblast-like synoviocytes via a mitochondrial-mediated pathway.

PubMed

Yan, Chuqi; Kong, Dechao; Ge, Dong; Zhang, Yanming; Zhang, Xishan; Su, Changhui; Cao, Xiaojian

2015-01-01

Rheumatoid arthritis (RA) is a systemic chronic inflammatory disease characterised by prominent synoviocyte hyperplasia and a potential imbalance between the growth and death of fibroblast-like synoviocytes (FLS). Mitomycin C (MMC) has previously been demonstrated to inhibit fibroblast proliferation and to induce fibroblast apoptosis. However, the effects of MMC on the proliferation and apoptosis of human RA FLS and the potential mechanisms underlying its effects remain unknown. Cell viability was determined using the Cell Counting Kit-8 assay. Apoptotic cell death was analysed via Annexin V-FITC/PI double staining and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labelling. The production of intracellular reactive oxygen species (ROS) was assessed via flow cytometry, and the changes in mitochondrial membrane potential (ΔΨm) were visualized based on JC-1 staining via fluorescence microscopy. The expression of apoptosis-related proteins was determined via Western blot. Treatment with MMC significantly reduced cell viability and induced apoptosis in RA FLS. Furthermore, MMC exposure was found to stimulate the production of ROS and to disrupt the ΔΨm compared to the control treatment. Moreover, MMC increased the release of mitochondrial cytochrome c, the ratio of Bax/Bcl-2, the activation of caspase-9 and caspase-3, and the subsequent cleavage of poly(ADP-ribose) polymerase. Our findings suggest that MMC inhibits cell proliferation and induces apoptosis in RA FLS, and the mechanism underlying this MMC-induced apoptosis may involve a mitochondrial signalling pathway. © 2015 S. Karger AG, Basel.

Changes in Structure, Interstitial Cajal-like Cells and Apoptosis of Smooth Muscle Cells in Congenital Ureteropelvic Junction Obstruction

PubMed Central

Mehrazma, Mitra; Tanzifi, Parin; Rakhshani, Naser

2014-01-01

Objective: The goal of this study is to evaluate some structural changes in muscular, collagenous and neural components as well as expression of Cajal-like cells and apoptosis of smooth muscle cells in congenital ureteropelvic junction obstruction (UPJO). Methods: Tissue specimens were obtained from 25 patients with UPJO and compared with normal ureteropelvic junction regions of 19 autopsies. In paraffin embedded sections the amount of Cajal-like cells, density of nerve fibers and smooth muscle cell apoptosis (using immunohistochemical staining) were determined. Collagen deposition and muscular components were stained by Trichrome-Masson staining and evaluated by image analysis techniques. Arrangement of muscular bundles was also evaluated qualitatively. Findings : The number of Cajal-like cells was significantly lower in patients than in controls. The apoptotic score and mean number of nerve fibers were not statistically different for the two groups. Arrangement of muscular fibers was more irregular in patients than in controls (P<0.001). Collagen deposition was significantly higher in patients than in controls (P<0.001). The mean amount of muscular component was lower in patients than in normal ones. (P= 0.09) Conclusion: We found significant pathologic changes in congenital ureteropelvic junction obstruction such as decrease in Cajal-like cells, increase in collagen deposition and irregular arrangement of muscle fibers. PMID:25793054

The effects of humanin and its analogues on male germ cell apoptosis induced by chemotherapeutic drugs.

PubMed

Jia, Yue; Ohanyan, Aikoui; Lue, Yan-He; Swerdloff, Ronald S; Liu, Peter Y; Cohen, Pinchas; Wang, Christina

2015-04-01

Human (HN) prevents stress-induced apoptosis in many cells/tissues. In this study we showed that HN ameliorated chemotherapy [cyclophosphamide (CP) and Doxorubicin (DOX)]-induced male germ cell apoptosis both ex vivo in seminiferous tubule cultures and in vivo in the testis. HN acts by several putative mechanisms via binding to: an IL-12 like trimeric membrane receptor; BAX; or insulin-like growth factor binding protein-3 (IGFBP-3, a proapoptotic factor). To understand the mechanisms of HN on male germ cell apoptosis, we studied five HN analogues including: HNG (HN-S14G, a potent agonist), HNG-F6A (no binding to IGFBP-3), HN-S7A (no self-dimerization), HN-C8P (no binding to BAX), and HN-L12A (a HN antagonist) on CP-induced male germ cell apoptosis in mice. CP-induced germ cell apoptosis was inhibited by HN, HNG, HNG-F6A, HN-S7A, and HN-C8P (less effective); but not by HN-L12A. HN-L12A, but not HN-S7A or HN-C8P, blocked the protective effect of HN against CP-induced male germ cell apoptosis. HN, HN-S7A, and HN-C8P restored CP-suppressed STAT3 phosphorylation. These results suggest that HN: (1) decreases DOX (ex vivo) and CP (in vivo) induced male germ cell apoptosis; (2) action is mediated by the membrane receptor/STAT3 with minor contribution by BAX-binding pathway; (3) self-dimerization or binding to IGFBP-3 may not be involved in HN's effect in testis. HN is an important molecule in the regulation of germ cell homeostasis after injury and agonistic analogues may be developed for treating male infertility or protection against chemotherapy side effects.

The Effects of Humanin and Its Analogues on Male Germ Cell Apoptosis Induced by Chemotherapeutic Drugs

PubMed Central

Jia, Yue; Ohanyan, Aikoui; Lue, Yan-He; Swerdloff, Ronald S.; Liu, Peter Y.; Cohen, Pinchas; Wang, Christina

2015-01-01

Human (HN) prevents stress-induced apoptosis in many cells/tissues. In this study we showed that HN ameliorated chemotherapy (Cyclophosphamide, CP and Doxorubicin, DOX)-induced male germ cell apoptosis both ex vivo in seminiferous tubule cultures and in vivo in the testis. HN acts by several putative mechanisms via binding to: an IL-12 like trimeric membrane receptor; BAX; or Insulin-Like Growth Factor Binding Protein-3 (IGFBP-3, a proapoptotic factor). To understand the mechanisms of HN on male germ cell apoptosis, we studied five HN analogues including: HNG (HN-S14G, a potent agonist), HNG-F6A (no binding to IGFBP-3), HN-S7A (no self-dimerization), HN-C8P (no binding to BAX), and HN-L12A (a HN antagonist) on CP-induced male germ cell apoptosis in mice. CP-induced germ cell apoptosis was inhibited by HN, HNG, HNG-F6A, HN-S7A, and HN-C8P (less effective); but not by HN-L12A. HN-L12A, but not HN-S7A or HN-C8P, blocked the protective effect of HN against CP-induced male germ cell apoptosis. HN, HN-S7A, and HN-C8P restored CP-suppressed STAT3 phosphorylation. These results suggest that HN: 1) decreases DOX (ex vivo) and CP (in vivo) induced male germ cell apoptosis; 2) action is mediated by the membrane receptor/STAT3 with minor contribution by BAX-binding pathway; 3) self-dimerization or binding to IGFBP-3 may not be involved in HN’s effect in testis. HN is an important molecule in the regulation of germ cell homeostasis after injury and agonistic analogues may be developed for treating male infertility or protection against chemotherapy side effects. PMID:25666707

Silencing of Hsp27 and Hsp72 in glioma cells as a tool for programmed cell death induction upon temozolomide and quercetin treatment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jakubowicz-Gil, Joanna, E-mail: jjgil@poczta.umcs.lublin.pl; Langner, Ewa; Bądziul, Dorota

The aim of the present study was to investigate whether silencing of Hsp27 or Hsp72 expression in glioblastoma multiforme T98G and anaplastic astrocytoma MOGGCCM cells increases their sensitivity to programmed cell death induction upon temozolomide and/or quercetin treatment. Transfection with specific siRNA was performed for the Hsp gene silencing. As revealed by microscopic observation and flow cytometry, the inhibition of Hsp expression was correlated with severe apoptosis induction upon the drug treatment studied. No signs of autophagy were detected. This was correlated with a decreased mitochondrial membrane potential, increased level of cytochrome c in the cytoplasm, and activation of caspasemore » 3 and caspase 9. All these results suggest that the apoptotic signal was mediated by an internal pathway. Additionally, in a large percentage of cells treated with temozolomide, with or without quercetin, granules within the ER system were found, which was accompanied by an increased level of caspase 12 expression. This might be correlated with ER stress. Quercetin and temozolomide also changed the shape of nuclei from circular to “croissant like” in both transfected cell lines. Our results indicate that blocking of Hsp27 and Hsp72 expression makes T98G cells and MOGGCCM cells extremely vulnerable to apoptosis induction upon temozolomide and quercetin treatment and that programmed cell death is initiated by an internal signal. - Highlights: • Hsps gene silencing induced severe apoptosis upon temozolomide–quercetin treatment • Apoptosis in transfected glioma cells was initiated by internal signal • Programmed cell death was preceded by ER stress • Temozolomide–quercetin treatment changed nuclei shape in transfected glioma cells.« less

Mycobacterium avium MAV2052 protein induces apoptosis in murine macrophage cells through Toll-like receptor 4.

PubMed

Lee, Kang-In; Choi, Han-Gyu; Son, Yeo-Jin; Whang, Jake; Kim, Kwangwook; Jeon, Heat Sal; Park, Hye-Soo; Back, Yong Woo; Choi, Seunga; Kim, Seong-Woo; Choi, Chul Hee; Kim, Hwa-Jung

2016-04-01

Mycobacterium avium and its sonic extracts induce apoptosis in macrophages. However, little is known about the M. avium components regulating macrophage apoptosis. In this study, using multidimensional fractionation, we identified MAV2052 protein, which induced macrophage apoptosis in M. avium culture filtrates. The recombinant MAV2052 induced macrophage apoptosis in a caspase-dependent manner. The loss of mitochondrial transmembrane potential (ΔΨm), mitochondrial translocation of Bax, and release of cytochrome c from mitochondria were observed in macrophages treated with MAV2052. Further, reactive oxygen species (ROS) production was required for the apoptosis induced by MAV2052. In addition, ROS and mitogen-activated protein kinases were involved in MAV2052-mediated TNF-α and IL-6 production. ROS-mediated activation of apoptosis signal-regulating kinase 1 (ASK1)-JNK pathway was a major signaling pathway for MAV2052-induced apoptosis. Moreover, MAV2052 bound to Toll-like receptor (TLR) 4 molecule and MAV2052-induced ROS production, ΔΨm loss, and apoptosis were all significantly reduced in TLR4(-/-) macrophages. Altogether, our results suggest that MAV2052 induces apoptotic cell death through TLR4 dependent ROS production and JNK pathway in murine macrophages.

c-Jun induces apoptosis of starved BM2 monoblasts by activating cyclin A-CDK2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vanhara, Petr; Bryja, Vitezslav; Horvath, Viktor

2007-02-02

c-Jun is one of the major components of the activating protein-1 (AP-1), the transcription factor that participates in regulation of proliferation, differentiation, and apoptosis. In this study, we explored functional interactions of the c-Jun protein with several regulators of the G1/S transition in serum-deprived v-myb-transformed chicken monoblasts BM2. We show that the c-Jun protein induces expression of cyclin A, thus up-regulating activity of cyclin A-associated cyclin-dependent kinase 2 (CDK2), and causing massive programmed cell death of starved BM2cJUN cells. Specific inhibition of CDK2 suppresses frequency of apoptosis of BM2cJUN cells. We conclude that up-regulation of cyclin A expression and CDK2more » activity can represent important link between the c-Jun protein, cell cycle machinery, and programmed cell death pathway in leukemic cells.« less

Periostin inhibits mechanical stretch-induced apoptosis in osteoblast-like MG-63 cells.

PubMed

Yu, Kai-Wen; Yao, Chung-Chen; Jeng, Jiiang-Huei; Shieh, Hao-Ying; Chen, Yi-Jane

2018-04-01

Appropriate mechanical stress plays an important role in regulating the proliferation and differentiation of osteoblasts, whereas high-level mechanical stress may be harmful and compromise cell survival. Periostin, a matricellular protein, is essential in maintaining functional integrity of bone and collagen-rich connective tissue in response to mechanical stress. This study investigated whether or not high-level mechanical stretch induces cell apoptosis and the regulatory role of periostin in mechanical stretch-induced apoptosis in osteoblastic cells. Osteoblast-like MG-63 cells were seeded onto Bio-Flex I culture plates and subjected to cyclic mechanical stretching (15% elongation, 0.1 Hz) in a Flexercell tension plus system-5000. The same process was applied to cells pre-treated with exogenous human recombinant periostin before mechanical stretching. We used a chromatin condensation and membrane permeability dead cell apoptosis kit to evaluate the stretch-induced cell responses. Expression of caspase-3 and cPARP was examined by immunofluorescent stain and flow cytometry. The expression of periostin in MG-63 cells is involved in the TGF-β signaling pathway. High-level cyclic mechanical stretch induced apoptotic responses in MG-63 osteoblastic cells. The percentages of apoptotic cells and cells expressing cPARP protein increased in the groups of cells subjected to mechanical stretch, but these responses were absent in the presence of exogenous periostin. Our study revealed that high-level mechanical stretch induces apoptotic cell death, and that periostin plays a protective role against mechanical stretch-induced apoptosis in osteoblastic cells. Copyright © 2017. Published by Elsevier B.V.

Depletion of mitochondrial fission factor DRP1 causes increased apoptosis in human colon cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Inoue-Yamauchi, Akane, E-mail: ainoyama@research.twmu.ac.jp; Oda, Hideaki

2012-04-27

Highlights: Black-Right-Pointing-Pointer DRP1 is required for mitochondrial fission in colon cancer cells. Black-Right-Pointing-Pointer DRP1 participates in inhibition of colon cancer cell apoptosis. Black-Right-Pointing-Pointer DRP1 can inhibit apoptosis through the regulation of cytochrome c release. -- Abstract: Mitochondria play a critical role in regulation of apoptosis, a form of programmed cell death, by releasing apoptogenic factors including cytochrome c. Growing evidence suggests that dynamic changes in mitochondrial morphology are involved in cellular apoptotic response. However, whether DRP1-mediated mitochondrial fission is required for induction of apoptosis remains speculative. Here, we show that siRNA-mediated DRP1 knockdown promoted accumulation of elongated mitochondria in HCT116more » and SW480 human colon cancer cells. Surprisingly, DRP1 down-regulation led to decreased proliferation and increased apoptosis of these cells. A higher rate of cytochrome c release and reductions in mitochondrial membrane potential were also revealed in DRP1-depleted cells. Taken together, our present findings suggest that mitochondrial fission factor DRP1 inhibits colon cancer cell apoptosis through the regulation of cytochrome c release and mitochondrial membrane integrity.« less

TLK1 and Rad9 Cooperate in XRT-Refractory CaP

DTIC Science & Technology

2011-04-01

groups that are confirmatory of our work). Silencing of Tousled-like kinase 1 sensitizes cholangiocarcinoma cells to cisplatin-induced apoptosis... cholangiocarcinoma cells to cisplatin-induced apoptosis. Cancer Letters 2010;296:27–34. 10. De Benedetti A. Tousled kinase TLK1B mediates chromatin

Apoptosis: its role in pituitary development and neoplastic pituitary tissue.

PubMed

Guzzo, M F; Carvalho, L R S; Bronstein, M D

2014-04-01

Apoptosis, also known as programmed cell death, is a phenomenon in which different stimuli trigger cellular mechanisms that culminate in death, in the absence of inflammatory cell response. Two different activation pathways are known, the intrinsic pathway (or mitochondrial) and extrinsic (or death-receptor pathway), both pathways trigger enzymatic reactions that lead cells to break up and be phagocytized by neighboring cells. This process is a common occurrence in physiological and pathological states, participating in the control of cell proliferation, differentiation and remodeling of organs. In the early steps of pituitary gland formation, numerous apoptotic cells are detected in the separation of Rathke's pouch from the roof of oral ectoderm. In the distal part of the gland, which will form the adenohypophysis, the ratio of apoptosis was significantly lower. However, there is evidence that neoplastic pituitary cells undergo unbalance in genes that control apoptosis leading to uncontrolled cell growth. No direct evidence of apoptosis was found in the drugs used for tumors producing prolactin and growth hormone. In conclusion, an unbalancing in the apoptosis process is the boundary between development and tumor growth.

Molecular basis for the interplay of apoptosis and proliferation mediated by Bcl-xL:Bim interactions in pancreatic cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Abrol, Ravinder, E-mail: abrol@wag.caltech.edu; Edderkaoui, Mouad; Goddard, William A.

2012-06-15

Highlights: Black-Right-Pointing-Pointer Direct role of Bcl-2 protein interactions in cell proliferation is not clear. Black-Right-Pointing-Pointer Designed Bcl-xL mutants show opposite effects on apoptosis and proliferation. Black-Right-Pointing-Pointer Disrupting Bcl-xL:Bim interaction increased apoptosis in pancreatic cancer. Black-Right-Pointing-Pointer Disrupting Bcl-xL:Bim interaction decreased proliferation in pancreatic cancer. Black-Right-Pointing-Pointer Bcl-xL:Bim interaction can control both apoptosis and proliferation. -- Abstract: A major mechanism through which cancer cells avoid apoptosis is by promoting the association of anti-apoptotic members of the pro-survival Bcl-2 protein family (like Bcl-2 and Bcl-xL) with BH{sub 3} domain-only proteins (like Bim and Bid). Apoptosis and cell proliferation have been shown to be linkedmore » for many cancers but the molecular basis for this link is far from understood. We have identified the Bcl-xL:Bim protein-protein interface as a direct regulator of proliferation and apoptosis in pancreatic cancer cells. We were able to predict and subsequently verify experimentally the effect of various Bcl-xL single-point mutants (at the position A142) on binding to Bim by structural analysis and computational modeling of the inter-residue interactions at the Bcl-xL:Bim protein-protein interface. The mutants A142N, A142Q, and A142Y decreased binding of Bim to Bcl-xL and A142S increased this binding. The Bcl-xL mutants, with decreased affinity for Bim, caused an increase in apoptosis and a corresponding decrease in cell proliferation. However, we could prevent these effects by introducing a small interfering RNA (siRNA) targeted at Bim. These results show a novel role played by the Bcl-xL:Bim interaction in regulating proliferation of pancreatic cancer cells at the expense of apoptosis. This study presents a physiologically relevant model of the Bcl-xL:Bim interface that can be used for rational therapeutic design for the inhibition of proliferation and cancer cell resistance to apoptosis.« less

Broad targeting of resistance to apoptosis in cancer

PubMed Central

Mohammad, Ramzi M.; Muqbil, Irfana; Lowe, Leroy; Yedjou, Clement; Hsu, Hsue-Yin; Lin, Liang-Tzung; Siegelin, Markus David; Fimognari, Carmela; Kumar, Nagi B.; Dou, Q. Ping; Yang, Huanjie; Samadi, Abbas K.; Russo, Gian Luigi; Spagnuolo, Carmela; Ray, Swapan K.; Chakrabarti, Mrinmay; Morre, James D.; Coley, Helen M.; Honoki, Kanya; Fujii, Hiromasa; Georgakilas, Alexandros G.; Amedei, Amedeo; Niccolai, Elena; Amin, Amr; Ashraf, S. Salman; Helferich, William G.; Yang, Xujuan; Boosani, Chandra S.; Guha, Gunjan; Bhakta, Dipita; Ciriolo, Maria Rosa; Aquilano, Katia; Chen, Sophie; Mohammed, Sulma I.; Keith, W. Nicol; Bilsland, Alan; Halicka, Dorota; Nowsheen, Somaira; Azmi, Asfar S.

2015-01-01

Apoptosis or programmed cell death is natural way of removing aged cells from the body. Most of the anti-cancer therapies trigger apoptosis induction and related cell death networks to eliminate malignant cells. However, in cancer, de-regulated apoptotic signaling, particularly the activation of an anti-apoptotic systems, allows cancer cells to escape this program leading to uncontrolled proliferation resulting in tumor survival, therapeutic resistance and recurrence of cancer. This resistance is a complicated phenomenon that emanates from the interactions of various molecules and signaling pathways. In this comprehensive review we discuss the various factors contributing to apoptosis resistance in cancers. The key resistance targets that are discussed include (1) Bcl-2 and Mcl-1 proteins; (2) autophagy processes; (3) necrosis and necroptosis; (4) heat shock protein signaling; (5) the proteasome pathway; (6) epigenetic mechanisms; and (7) aberrant nuclear export signaling. The shortcomings of current therapeutic modalities are highlighted and a broad spectrum strategy using approaches including (a) gossypol; (b) epigallocatechin-3-gallate; (c) UMI-77 (d) triptolide and (e) selinexor that can be used to overcome cell death resistance is presented. This review provides a roadmap for the design of successful anti-cancer strategies that overcome resistance to apoptosis for better therapeutic outcome in patients with cancer. PMID:25936818

Alpha cyano-4-hydroxy-3-methoxycinnamic acid inhibits proliferation and induces apoptosis in human breast cancer cells.

PubMed

Hamdan, Lamia; Arrar, Zoheir; Al Muataz, Yacoub; Suleiman, Lutfi; Négrier, Claude; Mulengi, Joseph Kajima; Boukerche, Habib

2013-01-01

This study investigated the underlying mechanism of 4-hydroxy-3-methoxycinnamic acid (ACCA), on the growth of breast cancer cells and normal immortal epithelial cells, and compared their cytotoxic effects responses. Treatment of breast cancer cells (MCF-7, T47D, and MDA-231) with ACCA resulted in dose- and time-dependent decrease of cell proliferation, viability in colony formation assay, and programmed cell death (apoptosis) with minimal effects on non-tumoral cells. The ability of ACCA to suppress growth in cancer cells not expressing or containing defects in p53 gene indicates a lack of involvement of this critical tumor suppressor element in mediating ACCA-induced growth inhibition. Induction of apoptosis correlated with an increase in Bax protein, an established inducer of programmed cell death, and the ratio of Bax to Bcl-2, an established inhibitor of apoptosis. We also documented the ability of ACCA to inhibit the migration and invasion of MDA-231 cells with ACCA in vitro. Additionally, tumor growth of MDA-231 breast cancer cells in vivo was dramatically affected with ACCA. On the basis of its selective anticancer inhibitory activity on tumor cells, ACCA may represent a promising therapeutic drug that should be further evaluated as a chemotherapeutic agent for human breast cancer.

Alpha Cyano-4-Hydroxy-3-Methoxycinnamic Acid Inhibits Proliferation and Induces Apoptosis in Human Breast Cancer Cells

PubMed Central

Hamdan, Lamia; Arrar, Zoheir; Al Muataz, Yacoub; Suleiman, Lutfi; Négrier, Claude; Mulengi, Joseph Kajima; Boukerche, Habib

2013-01-01

This study investigated the underlying mechanism of 4-hydroxy-3-methoxycinnamic acid (ACCA), on the growth of breast cancer cells and normal immortal epithelial cells, and compared their cytotoxic effects responses. Treatment of breast cancer cells (MCF-7, T47D, and MDA-231) with ACCA resulted in dose- and time-dependent decrease of cell proliferation, viability in colony formation assay, and programmed cell death (apoptosis) with minimal effects on non-tumoral cells. The ability of ACCA to suppress growth in cancer cells not expressing or containing defects in p53 gene indicates a lack of involvement of this critical tumor suppressor element in mediating ACCA-induced growth inhibition. Induction of apoptosis correlated with an increase in Bax protein, an established inducer of programmed cell death, and the ratio of Bax to Bcl-2, an established inhibitor of apoptosis. We also documented the ability of ACCA to inhibit the migration and invasion of MDA-231 cells with ACCA in vitro. Additionally, tumor growth of MDA-231 breast cancer cells in vivo was dramatically affected with ACCA. On the basis of its selective anticancer inhibitory activity on tumor cells, ACCA may represent a promising therapeutic drug that should be further evaluated as a chemotherapeutic agent for human breast cancer. PMID:24039831

DOE Office of Scientific and Technical Information (OSTI.GOV)

Galán-Malo, Patricia; Vela, Laura; Gonzalo, Oscar

Microtubule poisons and other anti-mitotic drugs induce tumor death but the molecular events linking mitotic arrest to cell death are still not fully understood. We have analyzed cell fate after mitotic arrest produced by the microtubule-destabilizing drug vincristine in a panel of human tumor cell lines showing different response to vincristine. In Jurkat, RPMI 8226 and HeLa cells, apoptosis was triggered shortly after vincristine-induced mitotic arrest. However, A549 cells, which express a great amount of Bcl-x{sub L} and undetectable amounts of Bak, underwent mitotic slippage prior to cell death. However, when Bcl-x{sub L} gene was silenced in A549 cells, vincristinemore » induced apoptosis during mitotic arrest. Another different behavior was found in MiaPaca2 cells, where vincristine caused death by mitotic catastrophe that switched to apoptosis when cyclin B1 degradation was prevented by proteasome inhibition. Overexpression of Bcl-x{sub L} or silencing Bax and Bak expression delayed the onset of apoptosis in Jurkat and RPMI 8226 cells, enabling mitotic slippage and endoreduplication. In HeLa cells, overexpression of Bcl-x{sub L} switched cell death from apoptosis to mitotic catastrophe. Mcl-1 offered limited protection to vincristine-induced cell death and Mcl-1 degradation was not essential for vincristine-induced death. All these results, taken together, indicate that the Bcl-x{sub L}/Bak ratio and the ability to degrade cyclin B1 determine cell fate after mitotic arrest in the different tumor cell types. Highlights: ► Vincristine induces cell death by apoptosis or mitotic catastrophe. ► Apoptosis-proficient cells die by apoptosis during mitosis upon vincristine treatment. ► p53wt apoptosis-deficient cells undergo apoptosis from a G1-like tetraploid state. ► p53mt apoptosis-deficient cells can survive and divide giving rise to 8N cells.« less

Induction of cell death by the lysosomotropic detergent MSDH.

PubMed

Li, W; Yuan, X; Nordgren, G; Dalen, H; Dubowchik, G M; Firestone, R A; Brunk, U T

2000-03-17

Controlled lysosomal rupture was initiated in lysosome-rich, macrophage-like cells by the synthetic lysosomotropic detergent, O-methyl-serine dodecylamide hydrochloride (MSDH). When MSDH was applied at low concentrations, resulting in partial lysosomal rupture, activation of pro-caspase-3-like proteases and apoptosis followed after some hours. Early during apoptosis, but clearly secondary to lysosomal destabilization, the mitochondrial transmembrane potential declined. At high concentrations, MSDH caused extensive lysosomal rupture and necrosis. It is suggested that lysosomal proteases, if released to the cytosol, may cause apoptosis directly by pro-caspase activation and/or indirectly by mitochondrial attack with ensuing discharge of pro-apoptotic factors.

Necroptosis: Fifty shades of RIPKs.

PubMed

Ichim, Gabriel; Tait, Stephen Wg

2015-01-01

Apoptosis and necroptosis are 2 major, yet distinct, forms of regulated cell death. Whereas apoptosis requires caspase protease function, necroptosis requires activation of the receptor interacting protein kinases 1 (RIPK1) and RIPK3. Following activation, RIPK3 phosphorylates mixed-lineage kinase domain-like (MLKL), leading to cell death. Apoptosis and necroptosis are deeply intertwined such that a given death stimulus can often engage either form of cell death. Recent studies published in Cell Death and Differentiation by the Han, Oberst, and Vaux laboratories provide exciting new insights into necroptosis and how it interconnects with apoptosis. As we will discuss, their findings address key questions including: How does a cell choose between apoptosis or necroptosis? How can RIPK3 also induce apoptosis? What is the nature of the RIPK1-3 signaling cascade leading to necroptosis? Finally, data from the Oberst and Han groups strongly argue that RIPK1 is not only involved in executing necroptosis, but also protects against this process in some settings.

Sall2 knockdown exacerbates palmitic acid induced dysfunction and apoptosis of pancreatic NIT-1 beta cells.

PubMed

Wang, Ye; Liu, Jie; Liu, Zheng; Chen, Jing; Hu, Xuemei; Hu, Yimeng; Yuan, Yin; Wu, Guijun; Dai, Zhe; Xu, Yancheng

2018-05-18

Spalt-like (Sall) proteins are a class of transcription factors. The role of Sall2 in beta cells remain poorly understood. Here, we aimed to explore whether Sall2 involved in lipotoxicity-mediated dysfunction and apoptosis in pancreatic NIT-1 beta cells. Our results showed that high concentrations of palmitic acid (PA) led to impaired cell viability and decreased Sall2 expression in NIT-1 cells. Knocking down of Sall2 in NIT-1 cells resulted in increased sensitivity to lipotoxicity and caused higher rates of cell apoptosis following PA treatment. Additionally, Sall2 Knockdown impaired insulin synthesis and secretion in response to glucose. Further research indicated Sall2 knockdown attenuate antioxidant capacity and decreased expression level of Peroxiredoxin 2 in NIT-1 cells. These finding implicate that Sall2 may play a significant role in NIT-1 cell function and cell apoptosis under lipotoxic conditions. Therefore, the study of Sall2 in NIT-1 cells provided a new perspective for molecular mechanism of lipotoxicity mediating dysfunction and apoptosis of beta cells. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Resveratrol suppresses growth of cancer stem-like cells by inhibiting fatty acid synthase.

PubMed

Pandey, Puspa R; Okuda, Hiroshi; Watabe, Misako; Pai, Sudha K; Liu, Wen; Kobayashi, Aya; Xing, Fei; Fukuda, Koji; Hirota, Shigeru; Sugai, Tamotsu; Wakabayashi, Go; Koeda, Keisuke; Kashiwaba, Masahiro; Suzuki, Kazuyuki; Chiba, Toshimi; Endo, Masaki; Fujioka, Tomoaki; Tanji, Susumu; Mo, Yin-Yuan; Cao, Deliang; Wilber, Andrew C; Watabe, Kounosuke

2011-11-01

Resveratrol is a natural polyphenolic compound and has been shown to exhibit cardio-protective as well as anti-neoplastic effects on various types of cancers. However, the exact mechanism of its anti-tumor effect is not clearly defined. Resveratrol has been shown to have strong hypolipidemic effect on normal adipocytes and as hyper-lipogenesis is a hallmark of cancer cell physiology, the effect of resveratrol on lipid synthesis in cancer stem-like cells (CD24(-)/CD44(+)/ESA(+)) that were isolated from both ER+ and ER- breast cancer cell lines was examined. The authors found that resveratrol significantly reduced the cell viability and mammosphere formation followed by inducing apoptosis in cancer stem-like cells. This inhibitory effect of resveratrol is accompanied by a significant reduction in lipid synthesis which is caused by the down-regulation of the fatty acid synthase (FAS) gene followed by up-regulation of pro-apoptotic genes, DAPK2 and BNIP3. The activation of apoptotic pathway in the cancer stem-like cells was suppressed by TOFA and by Fumonisin B1, suggesting that resveratrol-induced apoptosis is indeed through the modulation of FAS-mediated cell survival signaling. Importantly, resveratrol was able to significantly suppress the growth of cancer stem-like cells in an animal model of xenograft without showing apparental toxicity. Taken together, the results of this study indicate that resveratrol is capable of inducing apoptosis in the cancer stem-like cells through suppression of lipogenesis by modulating FAS expression, which highlights a novel mechanism of anti-tumor effect of resveratrol.

Resveratrol suppresses growth of cancer stem-like cells by inhibiting fatty acid synthase

PubMed Central

Pandey, Puspa R.; Okuda, Hiroshi; Watabe, Misako; Pai, Sudha K.; Liu, Wen; Kobayashi, Aya; Xing, Fei; Fukuda, Koji; Hirota, Shigeru; Sugai, Tamotsu; Wakabayashi, Go; Koeda, Keisuke; Kashiwaba, Masahiro; Suzuki, Kazuyuki; Chiba, Toshimi; Endo, Masaki; Fujioka, Tomoaki; Tanji, Susumu; Mo, Yin-Yuan; Cao, Deliang; Wilber, Andrew C.; Watabe, Kounosuke

2012-01-01

Resveratrol is a natural polyphenolic compound and has been shown to exhibit cardio-protective as well as anti-neoplastic effects on various types of cancers. However, the exact mechanism of its anti-tumor effect is not clearly defined. Resveratrol has been shown to have strong hypolipidemic effect on normal adipocytes and as hyper-lipogenesis is a hallmark of cancer cell physiology, we examined the effect of resveratrol on lipid synthesis in cancer stem-like cells (CD24−/CD44+/ESA+) that were isolated from both ER+ and ER− breast cancer cell lines. We found that resveratrol significantly reduced the cell viability and mammosphere formation followed by inducing apoptosis in cancer stem-like cells. This inhibitory effect of resveratrol is accompanied by a significant reduction in lipid synthesis which is caused by the down-regulation of the fatty acid synthase (FAS) gene followed by up-regulation of pro-apoptotic genes, DAPK2 and BNIP3. The activation of apoptotic pathway in the cancer stem-like cells was suppressed by TOFA and by Fumonisin B1, suggesting that resveratrol-induced apoptosis is indeed through the modulation of FAS-mediated cell survival signaling. Importantly, resveratrol was able to significantly suppress the growth of cancer stem-like cells in an animal model of xenograft without showing apparental toxicity. Taken together, our results indicate that resveratrol is capable of inducing apoptosis in the cancer stem-like cells through suppression of lipogenesis by modulating FAS expression, which highlights a novel mechanism of anti-tumor effect of resveratrol. PMID:21188630

Persian shallot, Allium hirtifolium Boiss, induced apoptosis in human hepatocellular carcinoma cells.

PubMed

Hosseini, Farzaneh Sadat; Falahati-Pour, Soudeh Khanamani; Hajizadeh, Mohammad Reza; Khoshdel, Alireza; Mirzaei, Mohammad Reza; Ahmadirad, Hadis; Behroozi, Reza; Jafari, Nesa; Mahmoodi, Mehdi

2017-08-01

This study investigated the potential of Persian shallot extract as an anticancer agent in HepG2 tumor cell line, an in vitro human hepatoma cancer model system. The inhibitory effect of Persian shallot on the growth of HepG2 cells was measured by MTT assay. To explore the underlying mechanism of cell growth inhibition of Persian shallot, the activity of Persian shallot in inducing apoptosis was investigated through the detection of annexin V signal by flow cytometry and expression of some apoptosis related genes such p21, p53, puma, caspase-8 family-Bcl-2 proteins like bid, bim, bcl-2 and bax were measured by real-time PCR in HepG2 cells. Persian shallot extract inhibited the growth of HepG2 cells in a dose-dependent manner. The IC 50 value (inhibiting cell growth by 50%) was 149 μg/ml. The results of real-time PCR revealed a significant up-regulation of bid, bim, caspase-8, puma, p53, p21 and bax genes and a significant downregulation of bcl-2 gene in HepG2 cells treated with Persian shallot extract significantly. Therefore, this is the first report on an increased expression of bid, bim, caspase-8, puma, p53, p21 and bax genes and down regulation of bcl-2 gene indicating that the Persian shallot extract possibly induced the process of cell death through the intrinsic and extrinsic apoptosis pathways and triggers the programmed cell death in HepG2 tumor cell lines by modulating the expression of pro-/anti-apoptotic genes. Furthermore, we showed that Persian shallot extract increased annexin V signal and expression, resulting in apoptotic cell death of HepG2 cells after 24 h treatment. Therefore, according to the results of this study, the Persian shallot extract could be considered as a potential candidate for production of drug for the prevention or treatment of human hepatoma.

Endoplasmic reticulum stress is involved in the lidocaine-induced apoptosis in SH-SY5Y neuroblastoma cells.

PubMed

Li, Kehan; Han, Xuechang

2015-05-01

Lidocaine has been indicated to promote apoptosis and to promote endoplasmic reticulum (ER) stress. However, the mechanism underlining ER stress-mediated apoptosis is unclear. In the present study, we investigated the promotion to ER stress in the lidocaine-induced apoptosis in human neuroblastoma SH-SY5Y cells. Firstly, we confirmed that lidocaine treatment induced apoptosis in SH-SY5Y cells, time-dependently and dose-dependently, via MTT cell viability assay and annexin V/FITC apoptosis detection with a FACScan flow cytometer. And the anti-apoptosis Bcl-2 and Bcl-xL were downregulated, whereas the apoptosis-executive caspase 3 was promoted through Western blot assay and caspase 3 activity assay. Moreover, the ER stress-associated binding immunoglobulin protein (BiP), PKR-like ER kinase (PERK), activating transcription factor 4 (ATF4) and CCAAT/enhancer-binding protein homologous protein (CHOP) were also upregulated at both mRNA and protein levels by lidocaine treatment. On the other hand, downregulation of the ER stress-associated BiP by RNAi method not only blocked the lidocaine-promoted ER stress but also attenuated the lidocaine-induced SH-SY5Y cell apoptosis. In conclusion, the present study confirmed the involvement of ER stress in the lidocaine-induced apoptosis in human neuroblastoma SH-SY5Y cells. Our study provides a better understanding on the mechanism of lidocaine's neurovirulence.

Tumors acquire inhibitor of apoptosis protein (IAP)-mediated apoptosis resistance through altered specificity of cytosolic proteolysis.

PubMed

Hong, Xu; Lei, Lu; Glas, Rickard

2003-06-16

Many tumors overexpress members of the inhibitor of apoptosis protein (IAP) family. IAPs contribute to tumor cell apoptosis resistance by the inhibition of caspases, and are degraded by the proteasome to allow further progression of apoptosis. Here we show that tumor cells can alter the specificity of cytosolic proteolysis in order to acquire apoptosis resistance, which promotes formation of rapidly growing tumors. Survival of tumor cells with low proteasomal activity can occur in the presence of high expression of Tri-peptidyl-peptidase II (TPP II), a large subtilisin-like peptidase that complements proteasomal activity. We find that this state leaves tumor cells unable of effectively degrading IAPs, and that cells in this state form rapidly growing tumors in vivo. We also find, in studies of apoptosis resistant cells derived from large in vivo tumors, that these have acquired an altered peptidase activity, with up-regulation of TPP II activity and decreased proteasomal activity. Importantly, we find that growth of subcutaneous tumors is limited by maintenance of the apoptosis resistant phenotype. The apoptosis resistant phenotype was reversed by increased expression of Smac/DIABLO, an antagonist of IAP molecules. Our data suggest a reversible mechanism in regulation of apoptosis resistance that drives tumor progression in vivo. These data are relevant in relation to the multitude of therapy-resistant clinical tumors that have increased levels of IAP molecules.

Dimethoxycurcumin-induced cell death in human breast carcinoma MCF7 cells: evidence for pro-oxidant activity, mitochondrial dysfunction, and apoptosis.

PubMed

Kunwar, A; Jayakumar, S; Srivastava, A K; Priyadarsini, K I

2012-04-01

The factors responsible for the induction of cell death by dimethoxycurcumin (Dimc), a synthetic analog of curcumin, were assessed in human breast carcinoma MCF7 cells. Initial cytotoxic studies with both curcumin and Dimc using MTT assay indicated their comparable effects. Further, the mechanism of action was explored in terms of oxidative stress, mitochondrial dysfunction, and modulation in the expression of proteins involved in cell cycle regulation and apoptosis. Dimc (5-50 μM) caused generation of reactive oxygen species, reduction in glutathione level, and induction of DNA damage. The mitochondrial dysfunction induced by Dimc was evidenced by the reduction in mitochondrial membrane potential and decrease in cellular energy status (ATP/ADP) monitored by HPLC analysis. The observed decrease in ATP was also supported by the significant suppression of different (α, β, γ, and ε) subunits of ATP synthase. The cytotoxic effect of Dimc was further characterized in terms of induction of S-phase cell cycle arrest and apoptosis, and their relative contribution was found to vary with the treatment concentration of Dimc. The S-phase arrest and apoptosis could also be correlated with the changes in the expressions of cell cycle proteins like p53, p21, CDK4, and cyclin-D1 and apoptotic markers like Bax and Bcl-2. Overall, the results demonstrated that Dimc induced cell death in MCF7 cells through S-phase arrest and apoptosis.

Activation of cellular death programs associated with immunosenescence-like phenotype in TPPII knockout mice

PubMed Central

Huai, Jisen; Firat, Elke; Nil, Ahmed; Million, Daniele; Gaedicke, Simone; Kanzler, Benoit; Freudenberg, Marina; van Endert, Peter; Kohler, Gabriele; Pahl, Heike L.; Aichele, Peter; Eichmann, Klaus; Niedermann, Gabriele

2008-01-01

The giant cytosolic protease tripeptidyl peptidase II (TPPII) has been implicated in the regulation of proliferation and survival of malignant cells, particularly lymphoma cells. To address its functions in normal cellular and systemic physiology we have generated TPPII-deficient mice. TPPII deficiency activates cell type-specific death programs, including proliferative apoptosis in several T lineage subsets and premature cellular senescence in fibroblasts and CD8+ T cells. This coincides with up-regulation of p53 and dysregulation of NF-κB. Prominent degenerative alterations at the organismic level were a decreased lifespan and symptoms characteristic of immunohematopoietic senescence. These symptoms include accelerated thymic involution, lymphopenia, impaired proliferative T cell responses, extramedullary hematopoiesis, and inflammation. Thus, TPPII is important for maintaining normal cellular and systemic physiology, which may be relevant for potential therapeutic applications of TPPII inhibitors. PMID:18362329

Topological control of life and death in non-proliferative epithelia.

PubMed

Martinand-Mari, Camille; Maury, Benoit; Rousset, François; Sahuquet, Alain; Mennessier, Gérard; Rochal, Sergei; Lorman, Vladimir; Mangeat, Paul; Baghdiguian, Stephen

2009-01-01

Programmed cell death is one of the most fascinating demonstrations of the plasticity of biological systems. It is classically described to act upstream of and govern major developmental patterning processes (e.g. inter-digitations in vertebrates, ommatidia in Drosophila). We show here the first evidence that massive apoptosis can also be controlled and coordinated by a pre-established pattern of a specific 'master cell' population. This new concept is supported by the development and validation of an original model of cell patterning. Ciona intestinalis eggs are surrounded by a three-layered follicular organization composed of 60 elongated floating extensions made of as many outer and inner cells, and indirectly spread through an extracellular matrix over 1200 test cells. Experimental and selective ablation of outer and inner cells results in the abrogation of apoptosis in respective remaining neighbouring test cells. In addition incubation of outer/inner follicular cell-depleted eggs with a soluble extract of apoptotic outer/inner cells partially restores apoptosis to apoptotic-defective test cells. The 60 inner follicular cells were thus identified as 'apoptotic master' cells which collectively are induction sites for programmed cell death of the underlying test cells. The position of apoptotic master cells is controlled by topological constraints exhibiting a tetrahedral symmetry, and each cell spreads over and can control the destiny of 20 smaller test cells, which leads to optimized apoptosis signalling.

Virus-like particles in venom of Meteorus pulchricornis induce host hemocyte apoptosis.

PubMed

Suzuki, M; Tanaka, T

2006-06-01

Ultrastructural studies on the reproductive tract and venom apparatus of a female braconid, Meteorus pulchricornis, revealed that the parasitoid lacks the calyx region in its oviduct, but possesses a venom gland with two venom gland filaments and a venom reservoir filled with white and cloudy fluid. Its venom gland cell is concaved and has a lumen filled with numerous granules. Transmisson electron microscopic (TEM) observation revealed that virus-like particles (VLPs) were produced in venom gland cells. The virus-like particle observed in M. pulchricornis (MpVLP) is composed of membranous envelopes with two different parts: a high-density core and a whitish low-density part. The VLPs of M. pulchricornis is also found assembling ultimately in the lumen of venom gland cell. Microvilli were found thrusting into the lumen of the venom gland cell and seem to aid in driving the matured MpVLPs to the common duct of the venom gland filament. Injection of MpVLPs into non-parasitized Pseudaletia separata hosts induced apoptosis in hemocytes, particularly granulocytes (GRs). Rate of apoptosis induced in GRs peaked 48h after VLP injection. While a large part of the GR population collapsed due to apoptosis caused by MpVLPs, the plasmatocyte population was minimally affected. The capacity of MpVLPs to cause apoptosis in host's hemocytes was further demonstrated by a decrease ( approximately 10-fold) in ability of host hemocytes to encapsulate fluorescent latex beads when MpVLPs were present. Apparently, the reduced encapsulation ability was due to a decrease in the GR population resulting from MpVLP-induced apoptosis.

6-shogaol induces autophagic cell death then triggered apoptosis in colorectal adenocarcinoma HT-29 cells.

PubMed

Li, Ting-Yi; Chiang, Been-Huang

2017-09-01

6-shogaol is a phytochemical of dietary ginger, we found that 6-shogaol could induced both autophagic and apoptotic death in human colon adenocarcinoma (HT-29) cells. Results of this study showed that 6-shogal induced cell cycle arrest, autophagy, and apoptosis in HT-29 cells in a time sequence. After 6h, 6-shogal induced apparent G2/M arrest, then the HT-29 cells formed numerous autophagosomes in each phase of the cell cycle. After 18h, increases in acidic vesicles and LAMP-1 (Lysosome-associated membrane proteins 1) showed that 6-shogaol had caused autophagic cell death. After 24h, cell shrinkage and Caspase-3/7 activities rising, suggesting that apoptotic cell death had increased. And after 48h, the result of TUNEL assay indicated the highest occurrence of apoptosis upon 6-shogaol treatment. It appeared that apoptosis is triggered by autophagy in 6-shogaol treated HT-29 cells, the damage of autophagic cell death initiated apoptosis program. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Distant homologs of anti-apoptotic factor HAX1 encode parvalbumin-like calcium binding proteins.

PubMed

Kokoszyńska, Katarzyna; Rychlewski, Leszek; Wyrwicz, Lucjan S

2010-07-15

Apoptosis is a highly ordered and orchestrated multiphase process controlled by the numerous cellular and extra-cellular signals, which executes the programmed cell death via release of cytochrome c alterations in calcium signaling, caspase-dependent limited proteolysis and DNA fragmentation. Besides the general modifiers of apoptosis, several tissue-specific regulators of this process were identified including HAX1 (HS-1 associated protein X-1) - an anti-apoptotic factor active in myeloid cells. Although HAX1 was the subject of various experimental studies, the mechanisms of its action and a functional link connected with the regulation of apoptosis still remains highly speculative. Here we provide the data which suggests that HAX1 may act as a regulator or as a sensor of calcium. On the basis of iterative similarity searches, we identified a set of distant homologs of HAX1 in insects. The applied fold recognition protocol gives us strong evidence that the distant insects' homologs of HAX1 are novel parvalbumin-like calcium binding proteins. Although the whole three EF-hands fold is not preserved in vertebrate our analysis suggests that there is an existence of a potential single EF-hand calcium binding site in HAX1. The molecular mechanism of its action remains to be identified, but the risen hypothesis easily translates into previously reported lines of various data on the HAX1 biology as well as, provides us a direct link to the regulation of apoptosis. Moreover, we also report that other family of myeloid specific apoptosis regulators - myeloid leukemia factors (MLF1, MLF2) share the homologous C-terminal domain and taxonomic distribution with HAX1. Performed structural and active sites analyses gave new insights into mechanisms of HAX1 and MLF families in apoptosis process and suggested possible role of HAX1 in calcium-binding, still the analyses require further experimental verification.

Distant homologs of anti-apoptotic factor HAX1 encode parvalbumin-like calcium binding proteins

PubMed Central

2010-01-01

Background Apoptosis is a highly ordered and orchestrated multiphase process controlled by the numerous cellular and extra-cellular signals, which executes the programmed cell death via release of cytochrome c alterations in calcium signaling, caspase-dependent limited proteolysis and DNA fragmentation. Besides the general modifiers of apoptosis, several tissue-specific regulators of this process were identified including HAX1 (HS-1 associated protein X-1) - an anti-apoptotic factor active in myeloid cells. Although HAX1 was the subject of various experimental studies, the mechanisms of its action and a functional link connected with the regulation of apoptosis still remains highly speculative. Findings Here we provide the data which suggests that HAX1 may act as a regulator or as a sensor of calcium. On the basis of iterative similarity searches, we identified a set of distant homologs of HAX1 in insects. The applied fold recognition protocol gives us strong evidence that the distant insects' homologs of HAX1 are novel parvalbumin-like calcium binding proteins. Although the whole three EF-hands fold is not preserved in vertebrate our analysis suggests that there is an existence of a potential single EF-hand calcium binding site in HAX1. The molecular mechanism of its action remains to be identified, but the risen hypothesis easily translates into previously reported lines of various data on the HAX1 biology as well as, provides us a direct link to the regulation of apoptosis. Moreover, we also report that other family of myeloid specific apoptosis regulators - myeloid leukemia factors (MLF1, MLF2) share the homologous C-terminal domain and taxonomic distribution with HAX1. Conclusions Performed structural and active sites analyses gave new insights into mechanisms of HAX1 and MLF families in apoptosis process and suggested possible role of HAX1 in calcium-binding, still the analyses require further experimental verification. PMID:20633251

Bim directly antagonizes Bcl-xl in doxorubicin-induced prostate cancer cell apoptosis independently of p53.

PubMed

Yang, Min-Chi; Lin, Ru-Wei; Huang, Shih-Bo; Huang, Shin-Yuan; Chen, Wen-Jie; Wang, Shiaw; Hong, Yi-Ren; Wang, Chihuei

2016-01-01

Doxorubicin and other anthracycline compounds exert their anti-cancer effects by causing DNA damage and initiating cell cycle arrest in cancer cells, followed by apoptosis. DNA damage generally activates a p53-mediated pathway to initiate apoptosis by increasing the level of the BH3-only protein, Puma. However, p53-mediated apoptosis in response to DNA damage has not yet been validated in prostate cancers. In the current study, we used LNCaP and PC3 prostate cancer cells, representing wild type p53 and a p53-null model, to determine if DNA damage activates p53-mediated apoptosis in prostate cancers. Our results revealed that PC3 cells were 4 to 8-fold less sensitive than LNCaP cells to doxorubicin-inuced apoptosis. We proved that the differential response of LNCaP and PC3 to doxorubicin was p53-independent by introducing wild-type or dominant negative p53 into PC3 or LNCaP cells, respectively. By comparing several apoptosis-related proteins in both cell lines, we found that Bcl-xl proteins were much more abundant in PC3 cells than in LNCaP cells. We further demonstrated that Bcl-xl protects LNCaP and PC3 cells from doxorubicin-induced apoptosis by using ABT-263, an inhibitor of Bcl-xl, as a single agent or in combination with doxorubicin to treat LNCaP or PC3 cells. Bcl-xl rather than p53, likely contributes to the differential response of LNCaP and PC3 to doxorubicin in apoptosis. Finally, co-immunoprecipitation and siRNA analysis revealed that a BH3-only protein, Bim, is involved in doxorubicin-induced apoptosis by directly counteracting Bcl-xl.

Apoptosis and Molecular Targeting Therapy in Cancer

PubMed Central

Hassan, Mohamed; Watari, Hidemichi; AbuAlmaaty, Ali; Ohba, Yusuke; Sakuragi, Noriaki

2014-01-01

Apoptosis is the programmed cell death which maintains the healthy survival/death balance in metazoan cells. Defect in apoptosis can cause cancer or autoimmunity, while enhanced apoptosis may cause degenerative diseases. The apoptotic signals contribute into safeguarding the genomic integrity while defective apoptosis may promote carcinogenesis. The apoptotic signals are complicated and they are regulated at several levels. The signals of carcinogenesis modulate the central control points of the apoptotic pathways, including inhibitor of apoptosis (IAP) proteins and FLICE-inhibitory protein (c-FLIP). The tumor cells may use some of several molecular mechanisms to suppress apoptosis and acquire resistance to apoptotic agents, for example, by the expression of antiapoptotic proteins such as Bcl-2 or by the downregulation or mutation of proapoptotic proteins such as BAX. In this review, we provide the main regulatory molecules that govern the main basic mechanisms, extrinsic and intrinsic, of apoptosis in normal cells. We discuss how carcinogenesis could be developed via defective apoptotic pathways or their convergence. We listed some molecules which could be targeted to stimulate apoptosis in different cancers. Together, we briefly discuss the development of some promising cancer treatment strategies which target apoptotic inhibitors including Bcl-2 family proteins, IAPs, and c-FLIP for apoptosis induction. PMID:25013758

Overcoming chemotherapy resistance of ovarian cancer cells by liposomal cisplatin: molecular mechanisms unveiled by gene expression profiling.

PubMed

Koch, Martin; Krieger, Michaela L; Stölting, Daniel; Brenner, Norbert; Beier, Manfred; Jaehde, Ulrich; Wiese, Michael; Royer, Hans-Dieter; Bendas, Gerd

2013-04-15

Previously we reported that liposomal cisplatin (CDDP) overcomes CDDP resistance of ovarian A2780cis cancer cells (Krieger et al., Int. J. Pharm. 389, 2010, 10-17). Here we find that the cytotoxic activity of liposomal CDDP is not associated with detectable DNA platination in resistant ovarian cancer cells. This suggests that the mode of action of liposomal CDDP is different from the free drug. To gain insight into mechanisms of liposomal CDDP activity, we performed a transcriptome analysis of untreated A2780cis cells, and A2780cis cells in response to exposure with IC50 values of free or liposomal CDDP. A process network analysis of upregulated genes showed that liposomal CDDP induced a highly different gene expression profile in comparison to the free drug. p53 was identified as a key player directing transcriptional responses to free or liposomal CDDP. The free drug induced expression of essential genes of the intrinsic (mitochondrial) apoptosis pathway (BAX, BID, CASP9) most likely through p38MAPK activation. In contrast, liposomal CDDP induced expression of genes from DNA damage pathways and several genes of the extrinsic pathway of apoptosis (TNFRSF10B-DR5, CD70-TNFSF7). It thus appears that liposomal CDDP overcomes CDDP resistance by inducing DNA damage and in consequence programmed cell death by the extrinsic pathway. Predictions from gene expression data with respect to apoptosis activation were confirmed at the protein level by an apoptosis antibody array. This sheds new light on liposomal drug carrier approaches in cancer and suggests liposomal CDDP as promising strategy for the treatment of CDDP resistant ovarian carcinomas. Copyright © 2013 Elsevier Inc. All rights reserved.

ACLY and ACC1 Regulate Hypoxia-Induced Apoptosis by Modulating ETV4 via α-ketoglutarate.

PubMed

Keenan, Melissa M; Liu, Beiyu; Tang, Xiaohu; Wu, Jianli; Cyr, Derek; Stevens, Robert D; Ilkayeva, Olga; Huang, Zhiqing; Tollini, Laura A; Murphy, Susan K; Lucas, Joseph; Muoio, Deborah M; Kim, So Young; Chi, Jen-Tsan

2015-10-01

In order to propagate a solid tumor, cancer cells must adapt to and survive under various tumor microenvironment (TME) stresses, such as hypoxia or lactic acidosis. To systematically identify genes that modulate cancer cell survival under stresses, we performed genome-wide shRNA screens under hypoxia or lactic acidosis. We discovered that genetic depletion of acetyl-CoA carboxylase (ACACA or ACC1) or ATP citrate lyase (ACLY) protected cancer cells from hypoxia-induced apoptosis. Additionally, the loss of ACLY or ACC1 reduced levels and activities of the oncogenic transcription factor ETV4. Silencing ETV4 also protected cells from hypoxia-induced apoptosis and led to remarkably similar transcriptional responses as with silenced ACLY or ACC1, including an anti-apoptotic program. Metabolomic analysis found that while α-ketoglutarate levels decrease under hypoxia in control cells, α-ketoglutarate is paradoxically increased under hypoxia when ACC1 or ACLY are depleted. Supplementation with α-ketoglutarate rescued the hypoxia-induced apoptosis and recapitulated the decreased expression and activity of ETV4, likely via an epigenetic mechanism. Therefore, ACC1 and ACLY regulate the levels of ETV4 under hypoxia via increased α-ketoglutarate. These results reveal that the ACC1/ACLY-α-ketoglutarate-ETV4 axis is a novel means by which metabolic states regulate transcriptional output for life vs. death decisions under hypoxia. Since many lipogenic inhibitors are under investigation as cancer therapeutics, our findings suggest that the use of these inhibitors will need to be carefully considered with respect to oncogenic drivers, tumor hypoxia, progression and dormancy. More broadly, our screen provides a framework for studying additional tumor cell stress-adaption mechanisms in the future.

ACLY and ACC1 Regulate Hypoxia-Induced Apoptosis by Modulating ETV4 via α-ketoglutarate

PubMed Central

Keenan, Melissa M.; Liu, Beiyu; Tang, Xiaohu; Wu, Jianli; Cyr, Derek; Stevens, Robert D.; Ilkayeva, Olga; Huang, Zhiqing; Tollini, Laura A.; Murphy, Susan K.; Lucas, Joseph; Muoio, Deborah M.; Kim, So Young; Chi, Jen-Tsan

2015-01-01

In order to propagate a solid tumor, cancer cells must adapt to and survive under various tumor microenvironment (TME) stresses, such as hypoxia or lactic acidosis. To systematically identify genes that modulate cancer cell survival under stresses, we performed genome-wide shRNA screens under hypoxia or lactic acidosis. We discovered that genetic depletion of acetyl-CoA carboxylase (ACACA or ACC1) or ATP citrate lyase (ACLY) protected cancer cells from hypoxia-induced apoptosis. Additionally, the loss of ACLY or ACC1 reduced levels and activities of the oncogenic transcription factor ETV4. Silencing ETV4 also protected cells from hypoxia-induced apoptosis and led to remarkably similar transcriptional responses as with silenced ACLY or ACC1, including an anti-apoptotic program. Metabolomic analysis found that while α-ketoglutarate levels decrease under hypoxia in control cells, α-ketoglutarate is paradoxically increased under hypoxia when ACC1 or ACLY are depleted. Supplementation with α-ketoglutarate rescued the hypoxia-induced apoptosis and recapitulated the decreased expression and activity of ETV4, likely via an epigenetic mechanism. Therefore, ACC1 and ACLY regulate the levels of ETV4 under hypoxia via increased α-ketoglutarate. These results reveal that the ACC1/ACLY-α-ketoglutarate-ETV4 axis is a novel means by which metabolic states regulate transcriptional output for life vs. death decisions under hypoxia. Since many lipogenic inhibitors are under investigation as cancer therapeutics, our findings suggest that the use of these inhibitors will need to be carefully considered with respect to oncogenic drivers, tumor hypoxia, progression and dormancy. More broadly, our screen provides a framework for studying additional tumor cell stress-adaption mechanisms in the future. PMID:26452058

Tissue Inhibitor of Metalloproteinase-3 (TIMP-3) induces FAS dependent apoptosis in human vascular smooth muscle cells.

PubMed

English, William R; Ireland-Zecchini, Heather; Baker, Andrew H; Littlewood, Trevor D; Bennett, Martin R; Murphy, Gillian

2018-01-01

Over expression of Tissue Inhibitor of Metalloproteinases-3 (TIMP-3) in vascular smooth muscle cells (VSMCs) induces apoptosis and reduces neointima formation occurring after saphenous vein interposition grafting or coronary stenting. In studies to address the mechanism of TIMP-3-driven apoptosis in human VSMCs we find that TIMP-3 increased activation of caspase-8 and apoptosis was inhibited by expression of Cytokine response modifier A (CrmA) and dominant negative FAS-Associated protein with Death Domain (FADD). TIMP-3 induced apoptosis did not cause mitochondrial depolarisation, increase activation of caspase-9 and was not inhibited by over-expression of B-cell Lymphoma 2 (Bcl2), indicating a mitochondrial independent/type-I death receptor pathway. TIMP-3 increased levels of the First Apoptosis Signal receptor (FAS) and depletion of FAS with shRNA showed TIMP-3-induced apoptosis was FAS dependent. TIMP-3 induced formation of the Death-Inducing Signalling Complex (DISC), as detected by immunoprecipitation and by immunofluorescence. Cellular-FADD-like IL-1 converting enzyme-Like Inhibitory Protein (c-FLIP) localised with FAS at the cell periphery in the absence of TIMP-3 and this localisation was lost on TIMP-3 expression with c-FLIP adopting a perinuclear localisation. Although TIMP-3 inhibited FAS shedding, this did not increase total surface levels of FAS but instead increased FAS levels within localised regions at the cell surface. A Disintegrin And Metalloproteinase 17 (ADAM17) is inhibited by TIMP-3 and depletion of ADAM17 with shRNA significantly decreased FAS shedding. However ADAM17 depletion did not induce apoptosis or replicate the effects of TIMP-3 by increasing localised clustering of cell surface FAS. ADAM17-depleted cells could activate caspase-3 when expressing levels of TIMP-3 that were otherwise sub-apoptotic, suggesting a partial role for ADAM17 mediated ectodomain shedding in TIMP-3 mediated apoptosis. We conclude that TIMP-3 induced apoptosis in VSMCs is highly dependent on FAS and is associated with changes in FAS and c-FLIP localisation, but is not solely dependent on shedding of the FAS ectodomain.

Tissue Inhibitor of Metalloproteinase–3 (TIMP-3) induces FAS dependent apoptosis in human vascular smooth muscle cells

PubMed Central

Ireland-Zecchini, Heather; Baker, Andrew H.; Littlewood, Trevor D.; Bennett, Martin R.; Murphy, Gillian

2018-01-01

Over expression of Tissue Inhibitor of Metalloproteinases-3 (TIMP-3) in vascular smooth muscle cells (VSMCs) induces apoptosis and reduces neointima formation occurring after saphenous vein interposition grafting or coronary stenting. In studies to address the mechanism of TIMP-3-driven apoptosis in human VSMCs we find that TIMP-3 increased activation of caspase-8 and apoptosis was inhibited by expression of Cytokine response modifier A (CrmA) and dominant negative FAS-Associated protein with Death Domain (FADD). TIMP-3 induced apoptosis did not cause mitochondrial depolarisation, increase activation of caspase-9 and was not inhibited by over-expression of B-cell Lymphoma 2 (Bcl2), indicating a mitochondrial independent/type-I death receptor pathway. TIMP-3 increased levels of the First Apoptosis Signal receptor (FAS) and depletion of FAS with shRNA showed TIMP-3-induced apoptosis was FAS dependent. TIMP-3 induced formation of the Death-Inducing Signalling Complex (DISC), as detected by immunoprecipitation and by immunofluorescence. Cellular-FADD-like IL-1 converting enzyme-Like Inhibitory Protein (c-FLIP) localised with FAS at the cell periphery in the absence of TIMP-3 and this localisation was lost on TIMP-3 expression with c-FLIP adopting a perinuclear localisation. Although TIMP-3 inhibited FAS shedding, this did not increase total surface levels of FAS but instead increased FAS levels within localised regions at the cell surface. A Disintegrin And Metalloproteinase 17 (ADAM17) is inhibited by TIMP-3 and depletion of ADAM17 with shRNA significantly decreased FAS shedding. However ADAM17 depletion did not induce apoptosis or replicate the effects of TIMP-3 by increasing localised clustering of cell surface FAS. ADAM17-depleted cells could activate caspase-3 when expressing levels of TIMP-3 that were otherwise sub-apoptotic, suggesting a partial role for ADAM17 mediated ectodomain shedding in TIMP-3 mediated apoptosis. We conclude that TIMP-3 induced apoptosis in VSMCs is highly dependent on FAS and is associated with changes in FAS and c-FLIP localisation, but is not solely dependent on shedding of the FAS ectodomain. PMID:29617412

Porphyra-334, a mycosporine-like amino acid, attenuates UV-induced apoptosis in HaCaT cells.

PubMed

Suh, Sung-Suk; Oh, Se Kyung; Lee, Sung Gu; Kim, Il-Chan; Kim, Sanghee

2017-06-27

The main aim of the current research was to study the effect of porphyra-334, one of mycosporine-like amino acids (MAAs), well known as UV-absorbing compounds, on UVinduced apoptosis in human immortalized keratinocyte (HaCaT) cells. Due to their UV-screening capacity and ability to prevent UV-induced DNA damage, MAAs have recently attracted considerable attention in both industry and research in pharmacology. Herein, human HaCaT cells were used to determine the biological activities of porphyra- 334 by various in vitro assays, including proliferation, apoptosis and Western blot assays. The proliferation rate of UV-irradiated HaCaT cells was significantly decreased compared to the control group. Pretreatment with porphyra- 334 markedly attenuated the inhibitory effect of UV and induced a dramatic decrease in the apoptotic rate. Expression of active caspase-3 protein was increased in response to UV irradiation, while caspase-3 levels were similar between cells treated with porphyra-334 and the non-irradiated control group. Taken together, our data suggest that porphyra-334 inhibits UV-induced apoptosis in HaCaT cells through attenuation of the caspase pathway.

Brazilian Red Propolis Induces Apoptosis-Like Cell Death and Decreases Migration Potential in Bladder Cancer Cells

PubMed Central

Begnini, Karine Rech; Moura de Leon, Priscila Marques; Thurow, Helena; Schultze, Eduarda; Campos, Vinicius Farias; Borsuk, Sibele; Dellagostin, Odir Antônio; Savegnago, Lucielli; Moura, Sidnei; Padilha, Francine F.; Pêgas Henriques, João Antonio; Seixas, Fabiana Kömmling

2014-01-01

Natural products continue to be an invaluable resource of anticancer drug discovery in recent years. Propolis is known for its biological activities such as antimicrobial and antitumor effects. This study assessed the effects of Brazilian red propolis (BRP) on apoptosis and migration potential in human bladder cancer cells. The effect of BRP ethanolic extract (25, 50, and 100 μg/mL) on 5637 cells was determined by MTT, LIVE/DEAD, and migration (scratch assay) assays. Apoptosis induction was investigated through flow cytometry and gene expression profile was investigated by qRT-PCR. Results showed cytotoxicity on MTT and LIVE/DEAD assays, with IC50 values of 95 μg/mL in 24 h of treatment. Cellular migration of 5637 cells was significantly inhibited through lower doses of BRP ethanolic extract (25 and 50 μg/mL). Flow cytometry analyses showed that BRP induced cytotoxicity through apoptosis-like mechanisms in 5637 cells and qRT-PCR revealed increased levels of Bax/Bcl-2 ratio, p53, AIF, and antioxidant enzymes genes. Data suggest that BRP may be a potential source of drugs to bladder cancer treatment. PMID:25530785

Brazilian red propolis induces apoptosis-like cell death and decreases migration potential in bladder cancer cells.

PubMed

Begnini, Karine Rech; Moura de Leon, Priscila Marques; Thurow, Helena; Schultze, Eduarda; Campos, Vinicius Farias; Martins Rodrigues, Fernanda; Borsuk, Sibele; Dellagostin, Odir Antônio; Savegnago, Lucielli; Roesch-Ely, Mariana; Moura, Sidnei; Padilha, Francine F; Collares, Tiago; Pêgas Henriques, João Antonio; Seixas, Fabiana Kömmling

2014-01-01

Natural products continue to be an invaluable resource of anticancer drug discovery in recent years. Propolis is known for its biological activities such as antimicrobial and antitumor effects. This study assessed the effects of Brazilian red propolis (BRP) on apoptosis and migration potential in human bladder cancer cells. The effect of BRP ethanolic extract (25, 50, and 100 μg/mL) on 5637 cells was determined by MTT, LIVE/DEAD, and migration (scratch assay) assays. Apoptosis induction was investigated through flow cytometry and gene expression profile was investigated by qRT-PCR. Results showed cytotoxicity on MTT and LIVE/DEAD assays, with IC50 values of 95 μg/mL in 24 h of treatment. Cellular migration of 5637 cells was significantly inhibited through lower doses of BRP ethanolic extract (25 and 50 μg/mL). Flow cytometry analyses showed that BRP induced cytotoxicity through apoptosis-like mechanisms in 5637 cells and qRT-PCR revealed increased levels of Bax/Bcl-2 ratio, p53, AIF, and antioxidant enzymes genes. Data suggest that BRP may be a potential source of drugs to bladder cancer treatment.

MicroRNA-195 targets ADP-ribosylation factor-like protein 2 to induce apoptosis in human embryonic stem cell-derived neural progenitor cells

PubMed Central

Zhou, Y; Jiang, H; Gu, J; Tang, Y; Shen, N; Jin, Y

2013-01-01

Neural progenitor cells (NPCs) derived from human embryonic stem cells (hESCs) have great potential in cell therapy, drug screening and toxicity testing of neural degenerative diseases. However, the molecular regulation of their proliferation and apoptosis, which needs to be revealed before clinical application, is largely unknown. MicroRNA miR-195 is known to be expressed in the brain and is involved in a variety of proapoptosis or antiapoptosis processes in cancer cells. Here, we defined the proapoptotic role of miR-195 in NPCs derived from two independent hESC lines (human embryonic stem cell-derived neural progenitor cells, hESC-NPCs). Overexpression of miR-195 in hESC-NPCs induced extensive apoptotic cell death. Consistently, global transcriptional microarray analyses indicated that miR-195 primarily regulated genes associated with apoptosis in hESC-NPCs. Mechanistically, a small GTP-binding protein ADP-ribosylation factor-like protein 2 (ARL2) was identified as a direct target of miR-195. Silencing ARL2 in hESC-NPCs provoked an apoptotic phenotype resembling that of miR-195 overexpression, revealing for the first time an essential role of ARL2 for the survival of human NPCs. Moreover, forced expression of ALR2 could abolish the cell number reduction caused by miR-195 overexpression. Interestingly, we found that paraquat, a neurotoxin, not only induced apoptosis but also increased miR-195 and reduced ARL2 expression in hESC-NPCs, indicating the possible involvement of miR-195 and ARL2 in neurotoxin-induced NPC apoptosis. Notably, inhibition of miR-195 family members could block neurotoxin-induced NPC apoptosis. Collectively, miR-195 regulates cell apoptosis in a context-dependent manner through directly targeting ARL2. The finding of the critical role of ARL2 for the survival of human NPCs and association of miR-195 and ARL2 with neurotoxin-induced apoptosis have important implications for understanding molecular mechanisms that control NPC survival and would facilitate our manipulation of the neurological pathogenesis. PMID:23807224

A mesenchymal-like phenotype and expression of CD44 predict lack of apoptotic response to sorafenib in liver tumor cells.

PubMed

Fernando, Joan; Malfettone, Andrea; Cepeda, Edgar B; Vilarrasa-Blasi, Roser; Bertran, Esther; Raimondi, Giulia; Fabra, Àngels; Alvarez-Barrientos, Alberto; Fernández-Salguero, Pedro; Fernández-Rodríguez, Conrado M; Giannelli, Gianluigi; Sancho, Patricia; Fabregat, Isabel

2015-02-15

The multikinase inhibitor sorafenib is the only effective drug in advanced cases of hepatocellular carcinoma (HCC). However, response differs among patients and effectiveness only implies a delay. We have recently described that sorafenib sensitizes HCC cells to apoptosis. In this work, we have explored the response to this drug of six different liver tumor cell lines to define a phenotypic signature that may predict lack of response in HCC patients. Results have indicated that liver tumor cells that show a mesenchymal-like phenotype, resistance to the suppressor effects of transforming growth factor beta (TGF-β) and high expression of the stem cell marker CD44 were refractory to sorafenib-induced cell death in in vitro studies, which correlated with lack of response to sorafenib in nude mice xenograft models of human HCC. In contrast, epithelial-like cells expressing the stem-related proteins EpCAM or CD133 were sensitive to sorafenib-induced apoptosis both in vitro and in vivo. A cross-talk between the TGF-β pathway and the acquisition of a mesenchymal-like phenotype with up-regulation of CD44 expression was found in the HCC cell lines. Targeted CD44 knock-down in the mesenchymal-like cells indicated that CD44 plays an active role in protecting HCC cells from sorafenib-induced apoptosis. However, CD44 effect requires a TGF-β-induced mesenchymal background, since the only overexpression of CD44 in epithelial-like HCC cells is not sufficient to impair sorafenib-induced cell death. In conclusion, a mesenchymal profile and expression of CD44, linked to activation of the TGF-β pathway, may predict lack of response to sorafenib in HCC patients. © 2014 UICC.

Apoptosis: a guide for the perplexed.

PubMed

Sloviter, Robert S

2002-01-01

The term 'apoptosis' describes an active process of cellular deconstruction originally contrasted morphologically with necrosis. The mistaken equivalence of the terms apoptosis and 'programmed cell death' has caused confusion and implied that apoptosis is an identifiable therapeutic target rather than a name of a type of cell death. The roots of confusion are suggested to lie not in superficial disagreements about the morphology and biochemistry of cell death, but in the lamentable disconnection of modern science from its philosophical foundations (i.e. Socratic definition, nominalism versus realism, and William of Ockham's advocacy of Aristotelian metaphysics over Plato's Theory of Forms). Renewed awareness of these issues might be the key to understanding that apoptosis is a created concept, not a real entity, and that the use of terms that defy definition has become an obstacle to clear thinking about preventable cell death.

Resistance to etoposide-induced apoptosis in a Burkitt's lymphoma cell line.

PubMed

Zhao, E G; Song, Q; Cross, S; Misko, I; Lees-Miller, S P; Lavin, M F

1998-08-31

Burkitt's lymphoma cells that vary in their phenotypic characteristics show significantly different degrees of susceptibility to radiation-induced apoptosis. Propensity to undergo apoptosis is reflected in the degradation of substrates such as DNA-dependent protein kinase but the status of bcl-2, c-myc and p53 has been uninformative. In this study, we have focused on 2 Epstein-Barr virus (EBV)-associated Burkitt's cell lines, one (WW2) susceptible and the other (BL29) resistant to etoposide-induced apoptosis. Differences in expression of BHRF1, an EBV gene that is homologous to the Bcl-2 proto-oncogene and known to inhibit apoptosis, or changes in apoptosis inhibitory proteins (IAPs), did not appear to account for the difference in susceptibility in the 2 cell lines. Cytoplasmic extracts from etoposide-treated WW2 cells caused apoptotic changes in nuclei isolated from either BL29 or WW2 cells, whereas extracts from BL29 cells failed to do so. In addition, extracts from etoposide-treated WW2 cells degraded the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), an important indicator of apoptosis, but this protein was resistant to degradation by BL29 extracts. It appears likely that caspase 3 (CPP32) is involved in this degradation since it was activated only in the apoptosis susceptible cells and the pattern of cleavage of DNA-PKcs was similar to that reported previously with recombinant caspase 3. As observed previously, addition of caspase 3 to nuclei failed to induce morphological changes indicative of apoptosis, but addition of caspase 3 to nuclei in the presence of extract from the resistant cells led to apoptotic changes. We conclude that resistance to apoptosis in BL29 cells is due to a failure of etoposide to activate upstream effectors of caspase activity.

Light scattering application for quantitative estimation of apoptosis

NASA Astrophysics Data System (ADS)

Bilyy, Rostyslav O.; Stoika, Rostyslav S.; Getman, Vasyl B.; Bilyi, Olexander I.

2004-05-01

Estimation of cell proliferation and apoptosis are in focus of instrumental methods used in modern biomedical sciences. Present study concerns monitoring of functional state of cells, specifically the development of their programmed death or apoptosis. The available methods for such purpose are either very expensive, or require time-consuming operations. Their specificity and sensitivity are frequently not sufficient for making conclusions which could be used in diagnostics or treatment monitoring. We propose a novel method for apoptosis measurement based on quantitative determination of cellular functional state taking into account their physical characteristics. This method uses the patented device -- laser microparticle analyser PRM-6 -- for analyzing light scattering by the microparticles, including cells. The method gives an opportunity for quick, quantitative, simple (without complicated preliminary cell processing) and relatively cheap measurement of apoptosis in cellular population. The elaborated method was used for studying apoptosis expression in murine leukemia cells of L1210 line and human lymphoblastic leukemia cells of K562 line. The results obtained by the proposed method permitted measuring cell number in tested sample, detecting and quantitative characterization of functional state of cells, particularly measuring the ratio of the apoptotic cells in suspension.

Long noncoding RNA Saf and splicing factor 45 increase soluble Fas and resistance to apoptosis

PubMed Central

Riberdy, Janice M.; Persons, Derek A.; Wilber, Andrew

2016-01-01

In multicellular organisms, cell growth and differentiation is controlled in part by programmed cell death or apoptosis. One major apoptotic pathway is triggered by Fas receptor (Fas)-Fas ligand (FasL) interaction. Neoplastic cells are frequently resistant to Fas-mediated apoptosis, evade Fas signals through down regulation of Fas and produce soluble Fas proteins that bind FasL thereby blocking apoptosis. Soluble Fas (sFas) is an alternative splice product of Fas pre-mRNA, commonly created by exclusion of transmembrane spanning sequences encoded within exon 6 (FasΔEx6). Long non-coding RNAs (lncRNAs) interact with other RNAs, DNA, and proteins to regulate gene expression. One lncRNA, Fas-antisense or Saf, was shown to participate in alternative splicing of Fas pre-mRNA through unknown mechanisms. We show that Saf is localized in the nucleus where it interacts with Fas receptor pre-mRNA and human splicing factor 45 (SPF45) to facilitate alternative splicing and exclusion of exon 6. The product is a soluble Fas protein that protects cells against FasL-induced apoptosis. Collectively, these studies reveal a novel mechanism to modulate this critical cell death program by an lncRNA and its protein partner. PMID:26885613

Technical note: a noninvasive method for measuring mammary apoptosis and epithelial cell activation in dairy animals using microparticles extracted from milk.

PubMed

Pollott, G E; Wilson, K; Jerram, L; Fowkes, R C; Lawson, C

2014-01-01

Milk production from dairy animals has been described in terms of 3 processes: the increase in secretory cell numbers in late pregnancy and early lactation, secretion rate of milk per cell, and the decline in cell numbers as lactation progresses. This latter process is thought to be determined by the level of programmed cell death (apoptosis) found in the animal. Until now, apoptosis has been measured by taking udder biopsies, using magnetic resonance imaging scans, or using animals postmortem. This paper describes an alternative, noninvasive method for estimating apoptosis by measuring microparticles in milk samples. Microparticles are the product of several processes in dairy animals, including apoptosis. Milk samples from 12 Holstein cows, at or past peak lactation, were collected at 5 monthly samplings. The samples (n=57) were used to measure the number of microparticles and calculate microparticle density for 4 metrics: annexin V positive and merocyanine 540 dye positive, for both and total particles, in both whole milk (WM) and spun milk. Various measures of milk production were also recorded for the 12 cows, including daily milk yield, fat and protein percentage in the milk, somatic cell count, and the days in milk when the samples were taken. A high correlation was found between the 4 WM microparticle densities and days in milk (0.46 to 0.64), and a moderate correlation between WM microparticle densities and daily milk yield (-0.33 to -0.44). No significant relationships were found involving spun milk samples, somatic cell count, or fat and protein percentage. General linear model analyses revealed differences between cows for both level of microparticle density and its rate of change in late lactation. Persistency of lactation was also found to be correlated with the WM microparticle traits (-0.65 to -0.32). As apoptosis is likely to be the major contributor to microparticle numbers in late lactation, this work found a noninvasive method for estimating apoptosis that gave promising results. Further investigation is required to find out the factors affecting microparticle production and how it changes throughout lactation. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Magnolol inhibits angiogenesis by regulating ROS-mediated apoptosis and the PI3K/AKT/mTOR signaling pathway in mES/EB-derived endothelial-like cells.

PubMed

Kim, Gi Dae; Oh, Jedo; Park, Hyen-Joo; Bae, Kihwan; Lee, Sang Kook

2013-08-01

Magnolol, a neolignan from the traditional medicinal plant Magnolia obovata, has been shown to possess neuroprotective, anti-inflammatory, anticancer and anti-angiogenic activities. However, the precise mechanism of the anti-angiogenic activity of magnolol remains to be elucidated. In the present study, the anti-angiogenic effect of magnolol was evaluated in mouse embryonic stem (mES)/embryoid body (EB)-derived endothelial-like cells. The endothelial-like cells were obtained by differentiation from mES/EB cells. Magnolol (20 µM) significantly suppressed the transcriptional and translational expression of platelet endothelial cell adhesion molecule (PECAM), an endothelial biomarker, in mES/EB-derived endothelial-like cells. To further understand the molecular mechanism of the suppression of PECAM expression, signaling pathways were analyzed in the mES/EB-derived endothelial-like cells. Magnolol induced the generation of reactive oxygen species (ROS) by mitochondria, a process that was associated with the induction of apoptosis as determined by positive Annexin V staining and the activation of cleaved caspase-3. The involvement of ROS generation by magnolol was confirmed by treatment with an antioxidant, N-acetyl-cysteine (NAC). NAC inhibited the magnolol-mediated induction of ROS generation and suppression of PECAM expression. In addition, magnolol suppressed the activation of MAPKs (ERK, JNK and p38) and the PI3K/AKT/mTOR signaling pathway in mES/EB-derived endothelial-like cells. Taken together, these findings demonstrate for the first time that the anti-angiogenic activity of magnolol may be associated with ROS-mediated apoptosis and the suppression of the PI3K/AKT/mTOR signaling pathway in mES/EB-derived endothelial-like cells.

The Activity of Differentiation Factors Induces Apoptosis in Polyomavirus Large T-Expressing Myoblasts

PubMed Central

Fimia, Gian Maria; Gottifredi, Vanesa; Bellei, Barbara; Ricciardi, Maria Rosaria; Tafuri, Agostino; Amati, Paolo; Maione, Rossella

1998-01-01

It is commonly accepted that pathways that regulate proliferation/differentiation processes, if altered in their normal interplay, can lead to the induction of programmed cell death. In a previous work we reported that Polyoma virus Large Tumor antigen (PyLT) interferes with in vitro terminal differentiation of skeletal myoblasts by binding and inactivating the retinoblastoma antioncogene product. This inhibition occurs after the activation of some early steps of the myogenic program. In the present work we report that myoblasts expressing wild-type PyLT, when subjected to differentiation stimuli, undergo cell death and that this cell death can be defined as apoptosis. Apoptosis in PyLT-expressing myoblasts starts after growth factors removal, is promoted by cell confluence, and is temporally correlated with the expression of early markers of myogenic differentiation. The block of the initial events of myogenesis by transforming growth factor β or basic fibroblast growth factor prevents PyLT-induced apoptosis, while the acceleration of this process by the overexpression of the muscle-regulatory factor MyoD further increases cell death in this system. MyoD can induce PyLT-expressing myoblasts to accumulate RB, p21, and muscle- specific genes but is unable to induce G00 arrest. Several markers of different phases of the cell cycle, such as cyclin A, cdk-2, and cdc-2, fail to be down-regulated, indicating the occurrence of cell cycle progression. It has been frequently suggested that apoptosis can result from an unbalanced cell cycle progression in the presence of a contrasting signal, such as growth factor deprivation. Our data involve differentiation pathways, as a further contrasting signal, in the generation of this conflict during myoblast cell apoptosis. PMID:9614186

Cellular commitment in the developing cerebellum

PubMed Central

Marzban, Hassan; Del Bigio, Marc R.; Alizadeh, Javad; Ghavami, Saeid; Zachariah, Robby M.; Rastegar, Mojgan

2014-01-01

The mammalian cerebellum is located in the posterior cranial fossa and is critical for motor coordination and non-motor functions including cognitive and emotional processes. The anatomical structure of cerebellum is distinct with a three-layered cortex. During development, neurogenesis and fate decisions of cerebellar primordium cells are orchestrated through tightly controlled molecular events involving multiple genetic pathways. In this review, we will highlight the anatomical structure of human and mouse cerebellum, the cellular composition of developing cerebellum, and the underlying gene expression programs involved in cell fate commitments in the cerebellum. A critical evaluation of the cell death literature suggests that apoptosis occurs in ~5% of cerebellar cells, most shortly after mitosis. Apoptosis and cellular autophagy likely play significant roles in cerebellar development, we provide a comprehensive discussion of their role in cerebellar development and organization. We also address the possible function of unfolded protein response in regulation of cerebellar neurogenesis. We discuss recent advancements in understanding the epigenetic signature of cerebellar compartments and possible connections between DNA methylation, microRNAs and cerebellar neurodegeneration. Finally, we discuss genetic diseases associated with cerebellar dysfunction and their role in the aging cerebellum. PMID:25628535

Measuring Apoptosis by Microscopy and Flow Cytometry.

PubMed

Hollville, Emilie; Martin, Seamus J

2016-02-02

Apoptosis is a mode of programmed cell death that plays an important role during development and in the maintenance of tissue homeostasis. Numerous physiological as well as pathological stimuli trigger apoptosis such as engagement of Fas, TRAIL, or TNF receptors, growth factor deprivation, hypoxia, or exposure to cytotoxic drugs. Apoptosis is coordinated from within by members of the caspase family of cysteine proteases that, upon activation, trigger a series of morphological changes including cell shrinkage, extensive plasma membrane blebbing, chromatin condensation, DNA hydrolysis, and nuclear fragmentation. These dramatic structural and biochemical alterations result not only in the controlled dismantling of the cell, but also in the efficient recognition and removal of apoptotic cells by phagocytes. Necrosis, which is typically nonprogrammed or imposed upon the cell by overwhelming membrane or organelle damage, is characterized by rapid plasma membrane rupture followed by organelle and cell swelling. Necrosis is often provoked by infectious agents or a severe departure from physiological conditions. This unit describes protocols for the measurement of apoptosis and for distinguishing apoptosis from necrosis. Copyright © 2016 John Wiley & Sons, Inc.

LncRNA-LET inhibits cell viability, migration and EMT while induces apoptosis by up-regulation of TIMP2 in human granulosa-like tumor cell line KGN.

PubMed

Han, Qingfang; Zhang, Wenke; Meng, Jinlai; Ma, Li; Li, Aihua

2018-04-01

Polycystic ovary syndrome (PCOS) is a common endocrine disease characterized by hyperandrogenism, irregular menses, and polycystic ovaries. Several long non-coding RNAs (lncRNAs) are aberrantly expressed in PCOS patients; however, little is known about the effects of the lncRNA-low expression in tumor (lncRNA-LET) on PCOS. We aimed to explore the effects of lncRNA-LET on human granulosa-like tumor cell line, KGN. Expression of lncRNA-LET in normal IOSE80 cells and granulosa cells was determined by qRT-PCR. KGN cell viability, apoptosis and migration were measured by trypan blue exclusion method, flow cytometry assay and wound healing assay, respectively. TGF-β1 was used to induce epithelial-mesenchymal transition (EMT) process. LncRNA-LET expression and mRNA expressions of TIMP2 and EMT-related proteins were measured by qRT-PCR. Western blot analysis was used to measure the protein expression of apoptosis-related proteins, EMT-related proteins, TIMP2, and the proteins in the Wnt/β-catenin and Notch signaling pathways. lncRNA-LET was down-regulated in KGN cells, and its overexpression inhibited cell viability and migration, and promoted apoptosis in KGN cells. Overexpression of lncRNA-LET increased the expression of E-cadherin and decreased the expressions of N-cadherin and vimentin in KGN cells. These effects of lncRNA-LET on KGN cells were reversed by TIMP2 suppression. Overexpression of TIMP2 inhibited cell viability, migration and EMT process, and increased apoptosis by activating the Wnt/β-catenin and Notch pathways. Overexpression of lncRNA-LET inhibits cell viability, migration and EMT process, and increases apoptosis in KGN cells by up-regulating the expression of TIMP2 and activating the Wnt/β-catenin and notch signaling pathways. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Kinetics of apoptotic markers in exogeneously induced apoptosis of EL4 cells.

PubMed

Jessel, Robert; Haertel, Steffen; Socaciu, Carmen; Tykhonova, Svetlana; Diehl, Horst A

2002-01-01

We investigated the time-dependence of apoptotic events in EL4 cells by monitoring plasma membrane changes in correlation to DNA fragmentation and cell shrinkage. We applied three apoptosis inducers (staurosporine, tubericidine and X-rays) and we looked at various markers to follow the early-to-late apoptotic events: phospholipid translocation (identified through annexin V-fluorescein assay and propidium iodide), lipid package (via merocyanine assay), membrane fluidity and anisotropy (via fluorescent measurements), DNA fragmentation by the fluorescence-labeling test and cell size measurements. The different apoptotic inducers caused different reactions of the cells: staurosporine induced apoptosis most rapidly in a high number of cells, tubercidine triggered apoptosis only in the S phase cells, while X-rays caused a G2/M arrest and subsequently apoptosis. Loss of lipid asymmetry is promptly detectable after one hour of incubation time. The phosphatidylserine translocation, decrease of lipid package and anisotropy, and the increase of membrane fluidity appeared to be based on the same process of lipid asymmetry loss. Therefore, the DNA fragmentation and the cell shrinkage appear to be parallel and independent processes running on different time scales but which are kinetically inter-related. The results indicate different signal steps to apoptosis dependent on inducer characteristics but the kinetics of "early-to-late" apoptosis appears to be a fixed program.

2-aryl benzimidazole conjugate induced apoptosis in human breast cancer MCF-7 cells through caspase independent pathway.

PubMed

Nayak, V Lakshma; Nagesh, Narayana; Ravikumar, A; Bagul, Chandrakant; Vishnuvardhan, M V P S; Srinivasulu, Vunnam; Kamal, Ahmed

2017-01-01

Apoptosis is a representative form of programmed cell death, which has been assumed to be critical for cancer prevention. Thus, any agent that can induce apoptosis may be useful for cancer treatment and apoptosis induction is arguably the most potent defense against cancer promotion. In our previous studies, 2-aryl benzimidazole conjugates were synthesized and evaluated for their antiproliferative activity and one of the new molecule (2f) was considered as a potential lead. This lead molecule showed significant antiproliferative activity against human breast cancer cell line, MCF-7. The results of the present study revealed that this compound arrested the cell cycle at G2/M phase. Topoisomerase II inhibition assay and Western blot analysis suggested that this compound effectively inhibits topoisomerase II activity which leads to apoptotic cell death. Apoptosis induction in MCF-7 cells was further confirmed by loss of mitochondrial membrane potential (∆Ψm), release of cytochrome c from mitochondria, an increase in the level of apoptosis inducing factor (AIF), generation of reactive oxygen species (ROS), up regulation of proapoptotic protein Bax and down regulation of anti apoptotic protein Bcl-2. Apoptosis assay using Annexin V-FITC assay also suggested that this compound induced cell death by apoptosis. However, compound 2f induced apoptosis could not be reversed by Z-VAD-FMK (a pan-caspase inhibitor) demonstrated that the 2f induced apoptosis was caspase independent. Further, 2f treatment did not activate caspase-7 and caspase-9 activity, suggesting that this compound induced apoptosis in breast cancer cells via a caspase independent pathway. Most importantly, this compound was less toxic towards non-tumorigenic breast epithelial cells, MCF-10A. Furthermore, docking studies also support the potentiality of this molecule to bind to the DNA topoisomerase II.

Molecular aspects of ultraviolet radiation-induced apoptosis in the skin.

PubMed

Chow, Jeffrey; Tron, Victor A

2005-12-01

Apoptosis, or programmed cell death, is an essential physiological process that controls cell numbers during physiological processes, and eliminates abnormal cells that can potentially harm an organism. This review summarizes our current state of knowledge of apoptosis induction in skin by UV radiation. A review of the literature was undertaken focusing on cell death in the skin secondary to UV radiation. It is evident that a number of apoptotic pathways, both intrinsic and extrinsic, are induced following exposure to damaging UV radiation. Although our understanding of the apoptotic processes is gradually increasing, many important aspects remain obscure. These include interconnections between pathways, wavelength-specific differences and cell type differences.

Transglutaminase induction by various cell death and apoptosis pathways.

PubMed

Fesus, L; Madi, A; Balajthy, Z; Nemes, Z; Szondy, Z

1996-10-31

Clarification of the molecular details of forms of natural cell death, including apoptosis, has become one of the most challenging issues of contemporary biomedical sciences. One of the effector elements of various cell death pathways is the covalent cross-linking of cellular proteins by transglutaminases. This review will discuss the accumulating data related to the induction and regulation of these enzymes, particularly of tissue type transglutaminase, in the molecular program of cell death. A wide range of signalling pathways can lead to the parallel induction of apoptosis and transglutaminase, providing a handle for better understanding the exact molecular interactions responsible for the mechanism of regulated cell death.

Peptide Logic Circuits Based on Chemoenzymatic Ligation for Programmable Cell Apoptosis.

PubMed

Li, Yong; Sun, Sujuan; Fan, Lin; Hu, Shanfang; Huang, Yan; Zhang, Ke; Nie, Zhou; Yao, Shouzhou

2017-11-20

A novel and versatile peptide-based bio-logic system capable of regulating cell function is developed using sortase A (SrtA), a peptide ligation enzyme, as a generic processor. By modular peptide design, we demonstrate that mammalian cells apoptosis can be programmed by peptide-based logic operations, including binary and combination gates (AND, INHIBIT, OR, and AND-INHIBIT), and a complex sequential logic circuit (multi-input keypad lock). Moreover, a proof-of-concept peptide regulatory circuit was developed to analyze the expression profile of cell-secreted protein biomarkers and trigger cancer-cell-specific apoptosis. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

N-Desmethyldauricine Induces Autophagic Cell Death in Apoptosis-Defective Cells via Ca2+ Mobilization.

PubMed

Law, Betty Y K; Mok, Simon W F; Chen, Juan; Michelangeli, Francesco; Jiang, Zhi-Hong; Han, Yu; Qu, Yuan Q; Qiu, Alena C L; Xu, Su-Wei; Xue, Wei-Wei; Yao, Xiao-Jun; Gao, Jia Y; Javed, Masood-Ul-Hassan; Coghi, Paolo; Liu, Liang; Wong, Vincent K W

2017-01-01

Resistance of cancer cells to chemotherapy remains a significant problem in oncology. Mechanisms regulating programmed cell death, including apoptosis, autophagy or necrosis, in the treatment of cancers have been extensively investigated over the last few decades. Autophagy is now emerging as an important pathway in regulating cell death or survival in cancer therapy. Recent studies demonstrated variety of natural small-molecules could induce autophagic cell death in apoptosis-resistant cancer cells, therefore, discovery of novel autophagic enhancers from natural products could be a promising strategy for treatment of chemotherapy-resistant cancer. By computational virtual docking analysis, biochemical assays, and advanced live-cell imaging techniques, we have identified N -desmethyldauricine (LP-4), isolated from rhizoma of Menispermum dauricum DC as a novel inducer of autophagy. LP-4 was shown to induce autophagy via the Ulk-1-PERK and Ca 2+ /Calmodulin-dependent protein kinase kinase β (CaMKKβ)-AMPK-mTOR signaling cascades, via mobilizing calcium release through inhibition of SERCA, and importantly, lead to autophagic cell death in a panel of cancer cells, apoptosis-defective and apoptosis-resistant cells. Taken together, this study provides detailed insights into the cytotoxic mechanism of a novel autophagic compound that targeting the apoptosis resistant cancer cells, and new implication on drug discovery from natural products for drug resistant cancer therapy.

The Bcl-2-Beclin 1 interaction in (-)-gossypol-induced autophagy versus apoptosis in prostate cancer cells.

PubMed

Lian, Jiqin; Karnak, David; Xu, Liang

2010-11-01

Bcl-2 is a key dual regulator of autophagy and apoptosis, but how the level of Bcl-2 influences the cellular decision between autophagy and apoptosis is unclear. The natural BH3-mimetic (-)-gossypol preferentially induces autophagy in androgen-independent (AI) prostate cancer cells that have high levels of Bcl-2 and are resistant to apoptosis, whereas apoptosis is preferentially induced in androgen-dependent or -independent cells with low Bcl-2. (-)-Gossypol induces autophagy via blocking Bcl-2-Beclin 1 interaction at the endoplasmic reticulum (ER), together with downregulating Bcl-2, upregulating Beclin 1 and activating the autophagic pathway. Furthermore, (-)-gossypol-induced autophagy is Beclin 1- and Atg5-dependent. These results provide new insights into the mode of cell death induced by Bcl-2 inhibitors, which could facilitate the rational design of clinical trials by selecting patients who are most likely to benefit from the Bcl-2-targeted molecular therapy.

Fluoride Induces Apoptosis in Mammalian Cells: In Vitro and In Vivo Studies.

PubMed

Ribeiro, Daniel Araki; Cardoso, Caroline Margonato; Yujra, Veronica Quispe; DE Barros Viana, Milena; Aguiar, Odair; Pisani, Luciana Pellegrini; Oshima, Celina Tizuko Fujiyama

2017-09-01

Apoptosis is genetically programmed cell death, an irreversible process of cell senescence with characteristic features different from other cellular mechanisms of death such as necrosis. In the last years, apoptosis has been extensively studied in the scientific literature, because it has been established that apoptosis plays a crucial role following the time course of chronic degenerative diseases, such as cancer. Thus, several researchers have strugged to detect what chemical agents are able to inter fere with the apoptotic process. Thus, the purpose of this literature review is to assess if fluoride induces apoptosis in mammalian cells using in vivo and in vitro test systems. Certain mammalian cell types such as oral cells, blood and brain were exetensively investigated; the results showed that fluoride is able to induce apoptosis in both intrinsinc and extrinsic pathways. Moreover, other cells types have been poorly investigated such as bone, kidney and reproductive cells with conflicting results so far. Therefore, this area needs further investigation for the safety of human populations exposed to fluoride in a chronic way, as for example in developing countries. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Regulatory RNA Key Player in p53-Mediated Apoptosis in Embryonic Stem Cells | Center for Cancer Research

Cancer.gov

Embryonic stem cells (ESCs) must maintain the integrity of their genomes or risk passing potentially deleterious mutations on to numerous tissues. Thus, ESCs have a unique genome surveillance system and easily undergo apoptosis or differentiation when DNA damage is detected. The protein p53 is known to promote differentiation in mouse ESCs (mESCs), but its role in DNA damage-induced apoptosis (DIA) is unclear. p53 may have a pro-apoptotic function since it can regulate apoptotic genes in embryonal cells. Given that ESCs have a distinct transcriptional program, Jing Huang, Ph.D., of CCR’s Laboratory of Cancer Biology and Genetics, and his colleagues wondered whether p53 might regulate DIA in ESCs by utilizing the ESC-specific expression program.

Involvement of Apoptosis in Host-Parasite Interactions in the Zebra Mussel

PubMed Central

Minguez, Laëtitia; Brulé, Nelly; Sohm, Bénédicte; Devin, Simon; Giambérini, Laure

2013-01-01

The question of whether cell death by apoptosis plays a biological function during infection is key to understanding host-parasite interactions. We investigated the involvement of apoptosis in several host-parasite systems, using zebra mussels Dreissena polymorpha as test organisms and their micro- and macroparasites. As a stress response associated with parasitism, heat shock proteins (Hsp) can be induced. In this protein family, Hsp70 are known to be apoptosis inhibitors. Mussels were diagnosed for their respective infections by standard histological methods; apoptosis was detected using the TUNEL methods on paraffin sections and Hsp70 by immunohistochemistry on cryosections. Circulating hemocytes were the main cells observed in apoptosis whereas infected tissues displayed no or few apoptotic cells. Parasitism by intracellular bacteria Rickettsiales-like and the trematode Bucephalus polymorphus were associated with the inhibition of apoptosis whereas ciliates Ophryoglena spp. or the trematode Phyllodistomum folium did not involve significant differences in apoptosis. Even if some parasites were able to modulate apoptosis in zebra mussels, we did not see evidence of any involvement of Hsp70 on this mechanism. PMID:23785455

Pulsed electromagnetic field affects intrinsic and endoplasmatic reticulum apoptosis induction pathways in MonoMac6 cell line culture.

PubMed

Kaszuba-Zwoinska, J; Chorobik, P; Juszczak, K; Zaraska, W; Thor, P J

2012-10-01

Current studies were aimed to elucidate influence of pulsed electromagnetic field stimulation on cell viability and apoptosis induction pathways. For the experimental model we have chosen monocytic cell line MonoMac6 and several apoptosis inducers with different mechanism of death induction like puromycin, colchicine, cyclophosphamide, minocycline and hydrogen peroxide. MonoMac6 cell line was grown at density 1x10(5) cells/well in 96-well culture plates. To induce cell death cell cultures were treated with different apoptosis inducers like puromycin, colchicine, cyclophosphamide, minocycline, hydrogen peroxide and at the same time with pulsed electromagnetic field 50 Hz, 45±5 mT (PEMF) for 4 hour per each stimulation, three times, in 24 hours intervals. Afterwards, cells were harvested for flow cytometry analysis of cell viability measured by annexin V-APC labeled and propidium iodide staining. Expression of apoptosis related genes was evaluated by semi quantitative reverse transcription (RT)-PCR assay. NuPAGE Novex Western blot analysis was carried out for apoptosis inducing factor (AIF) abundance in cytosolic and nuclear extracts of MonoMac6 cells. Puromycin, colchicine and minocycline activated cells and simultaneously treated with PEMF have shown out diminished percentage of annexinV positive (AnV+) cells comparing to controls without PEMF stimulation. MonaMac6 cells puromycin/colchicyne and PEMF treated were to a higher extent double stained (AnV+,PI+), which means increased late apoptotic as well as necrotic (PI+) cells, than non-stimulated controls. On the other hand, minocycline activated cells prior to PEMF treatment showed diminished amount of apoptotic and necrotic (annexin V, annexin V and propidium iodide, propidium iodide positive staining) cells. The opposite effect of PEMF on the percentage of annexin V positively stained cells has been achieved after treatment of MonoMac6 culture with cyclophoshamide and hydrogen peroxide. PEMF enhanced early phase of apoptosis induced by both apoptosis inducing agents. The analysis of expression of the apoptosis related genes in MonoMac6 cultures treated with puromycin and exposed to PEMF performed in reverse transcription of polymerase chain reaction (PCR) assay has shown changes in mRNA of genes engaged in intrinsic apoptotic pathway and pathway with AIF abundance. The most influenced was expression of gene belonging to pro-apoptotic family of Bcl-2 and AIF agent. Examination of immunoblots developed with anti-AIF antibody showed that cytosol content of AIF protein was diminished after puromycin and PEMF treatment of MonoMac6 cells. The obtained results indicate that PEMF affects induction of apoptosis in MonoMac6 cells stimulated to death with inducing agents to a different extent. Main finding of the current results is that, PEMF stimulation of MonoMac6 cells simultaneously treated with puromycin caused changes in the Bcl-family genes expression as well as in caspase independent pathway of apoptosis inducing factor (AIF).

The sweet taste of death: glucose triggers apoptosis during yeast chronological aging.

PubMed

Ruckenstuhl, Christoph; Carmona-Gutierrez, Didac; Madeo, Frank

2010-10-01

As time goes by, a postmitotic cell ages following a degeneration process ultimately ending in cell death. This phenomenon is evolutionary conserved and present in unicellular eukaryotes as well, making the yeast chronological aging system an appreciated model. Here, single cells die in a programmed fashion (both by apoptosis and necrosis) for the benefit of the whole population. Besides its meaning for aging and cell death research, age-induced programmed cell death represents the first experimental proof for the so-called group selection theory: Apoptotic genes became selected during evolution because of the benefits they might render to the whole cell culture and not to the individual cell. Many anti‐aging stimuli have been discovered in the yeast chronological aging system and have afterwards been confirmed in higher cells or organisms. New work from the Burhans group (this issue) now demonstrates that glucose signaling has a progeriatric effect on chronologically aged yeast cells: Glucose administration results in a diminished efficacy of cells to enter quiescence, finally causing superoxide‐mediated replication stress and apoptosis.

P143 proteins from heterologous nucleopolyhedroviruses induce apoptosis in BM-N cells derived from the silkworm Bombyx mori.

PubMed

Hamajima, Rina; Kobayashi, Michihiro; Ikeda, Motoko

2017-04-02

We previously demonstrated that ribosomal RNA (rRNA) of Bombyx mori BM-N cells is rapidly degraded upon infection with heterologous nucleopolyhedroviruses (NPVs), including Autographa californica multiple NPV (AcMNPV), Hyphantria cunea MNPV, Spodoptera exigua MNPV and S. litura MNPV, and that this response is triggered by viral P143 proteins. The transient expression of P143 proteins from heterologous NPVs was also shown to induce apoptosis and caspase-3-like protease activation in BM-N cells. In the present study, we conducted a transient expression assay using BM-N cells expressing mutant AcMNPV P143 (Ac-P143) proteins and demonstrated that five amino acid residues cooperatively participate in Ac-P143 protein-triggered apoptosis of BM-N cells. Notably, these five residues were previously shown to be required for triggering rRNA degradation in BM-N cells. As rRNA degradation in BM-N cells does not result from apoptosis, the present results suggest that Ac-P143-triggered rRNA degradation is the upstream signal for apoptosis induction in BM-N cells. We further showed that P143 protein-triggered apoptosis does not occur in S. frugiperda Sf9 or Lymantria dispar Ld652Y cells, indicating that apoptosis induction by heterologous P143 proteins is a BM-N cell-specific response. In addition, the observed induction of apoptosis in BM-N cells was found to be mediated by activation of the initiator caspase Bm-Dronc. Taken together, these results suggest that BM-N cells evolved a unique antiviral system that recognizes heterologous NPV P143 proteins to induce rRNA degradation and caspase-dependent apoptosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Possible association between stem-like hallmark and radioresistance in human cervical carcinoma cells.

PubMed

Kumazawa, Shoko; Kajiyama, Hiroaki; Umezu, Tomokazu; Mizuno, Mika; Suzuki, Shiro; Yamamoto, Eiko; Mitsui, Hiroko; Sekiya, Ryuichiro; Shibata, Kiyosumi; Kikkawa, Fumitaka

2014-05-01

We aimed to investigate the possibility of an association between a stem-like hallmark and radiotherapeutic sensitivity in human cervical carcinoma cells. Side-population (SP) cells and non-SP (NSP) cells in HeLa cells were isolated using flow cytometry and Hoechst 33342 efflux. We performed Western blot analysis to evaluate the expression of stem cell markers (CXCR4, Oct3/4, CD133, and SOX2) and apoptosis markers after irradiation. In addition, SP and NSP cells were injected into nude mice and we assessed subcutaneous tumor formation. To examine tolerance of irradiation, colony formation and apoptosis change were confirmed in the SP and NSP cells. SP cells showed a higher expression of CXCR4, Oct3/4, CD133, and SOX2 than NSP cells. The colony size of SP cells cultured on non-coated dishes was larger than that of NSP cells, and NSP cells were easily induced to undergo apoptosis. SP cells tended to form spheroids and showed a higher level of tumorigenicity compared with NSP cells. In addition, nude mice inoculated with SP cells showed greater tumor growth compared with NSP cells. SP cells showed a higher tumorigenicity and lower apoptotic potential, leading to enhanced radiotolerance. Tumor SP cells showed higher-level stem-cell-like characters and radioresistance than NSP cells. SP cells may be useful for new therapeutic approaches for radiation-resistant cervical cancer. © 2014 The Authors. Journal of Obstetrics and Gynaecology Research © 2014 Japan Society of Obstetrics and Gynecology.

Osthole enhances TRAIL-mediated apoptosis through downregulation of c-FLIP expression in renal carcinoma Caki cells.

PubMed

Min, Kyoung-Jin; Han, Min Ae; Kim, Shin; Park, Jong-Wook; Kwon, Taeg Kyu

2017-04-01

Osthole, an active constituent isolated from the fruit of Cnidium monnieri (L.) Cusson, has been shown to induce various beneficial biochemical effects such as anti-inflammatory and antitumor. In the present study, we examined whether osthole could sensitize TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in human renal carcinoma Caki cells. We found that osthole and TRAIL alone, had no effect on apoptosis, but combined treatment with osthole and TRAIL markedly induced apoptosis in Caki (renal carcinoma), U251MG (glioma) and MDA-MB-231 (breast carcinoma) cells. In contrast, combined treatment with osthole and TRAIL did not induce apoptosis in normal human skin fibroblast cells. Osthole induced downregulation of cellular FLICE-like inhibitory protein (c-FLIP) expression, and overexpression of c-FLIP markedly blocked apoptosis induced by the combined treatment with osthole and TRAIL. In addition, osthole markedly reduced mitochondrial membrane potential levels, and increased cytosolic cytochrome c release in combined treatment with osthole and TRAIL. Therefore, these data suggest that osthole may be an efficient TRAIL sensitizer.

c-Myc-induced apoptosis in fibroblasts is inhibited by specific cytokines.

PubMed Central

Harrington, E A; Bennett, M R; Fanidi, A; Evan, G I

1994-01-01

We have investigated the mechanism by which deregulated expression of c-Myc induces death by apoptosis in serum-deprived fibroblasts. We demonstrate that Myc-induced apoptosis in low serum is inhibited by a restricted group of cytokines, principally the insulin-like growth factors and PDGF. Cytokine-mediated protection from apoptosis is not linked to the cytokines' abilities to promote growth. Protection from apoptosis is evident in the post-commitment (mitogen-independent) S/G2/M phases of the cell cycle and also in cells that are profoundly blocked in cell cycle progression by drugs. Moreover, IGF-I inhibition of apoptosis occurs in the absence of protein synthesis, and so does not require immediate early gene expression. We conclude that c-Myc-induced apoptosis does not result from a conflict of growth signals but appears to be a normal physiological aspect of c-Myc function whose execution is regulated by the availability of survival factors. We discuss the possible implications of these findings for models of mammalian cell growth in vivo. Images PMID:8045259

Phloretin induces apoptosis of human esophageal cancer via a mitochondria-dependent pathway.

PubMed

Duan, Hongtao; Wang, Ruixuan; Yan, Xiaolong; Liu, Honggang; Zhang, Yong; Mu, Deguang; Han, Jing; Li, Xiaofei

2017-12-01

2,4,6-trihydroxy-3-(4-hydroxyphenyl)-propiophenone (phloretin) is found in apple tree leaves and the Manchurian apricot, and is a potent compound that exhibits anti-inflammatory, antioxidant and antitumor activities. However, the effect of phloretin on esophageal cancer cells is not well-defined. The present study aimed to examine whether and how phloretin induced apoptosis in human esophageal cancer cells. EC-109 cells were cultured in Dulbecco's modified Eagle's medium and incubated with 60, 70, 80, 90 and 100 µg/ml phloretin for 6, 12, 24 and 48 h. Cell proliferation was measured by an MTT assay. Cell apoptosis rate was measured using flow cytometric analysis subsequent to propidium iodide (PI) staining. The protein expression levels were determined by western blot analysis. It was found that phloretin significantly decreased viable cell numbers in a dose- and time-dependent manner and induced apoptosis in EC-109 cells. Additionally, phloretin exhibited potent anticancer activity in vitro , as evidenced by the downregulation of the anti-apoptosis-associated molecule B-cell lymphoma 2 (bcl-2) and an increase in the levels of the apoptosis-associated molecules bcl-2-like protein 4 and tumor protein p53. Phloretin treatment also affected the expression of apoptotic protease activating factor-1, the protein product of the direct binding of the inhibitor of apoptosis protein with low PI to the X-linked inhibitor of apoptosis protein. The present results indicated that phloretin may inhibit EC-109 cell growth by inducing apoptosis, which may be mediated through a mitochondria-dependent pathway.

Centchroman induces redox-dependent apoptosis and cell-cycle arrest in human endometrial cancer cells.

PubMed

Shyam, Hari; Singh, Neetu; Kaushik, Shweta; Sharma, Ramesh; Balapure, Anil K

2017-04-01

Centchroman (CC) or Ormeloxifene has been shown to induce apoptosis and cell cycle arrest in various types of cancer cells. This has, however, not been addressed for endometrial cancer cells where its (CC) mechanism of action remains unclear. This study focuses on the basis of antineoplasticity of CC by blocking the targets involved in the cell cycle, survival and apoptosis in endometrial cancer cells. Ishikawa Human Endometrial Cancer Cells were cultured under estrogen deprived medium, exposed to CC and analyzed for proliferation and apoptosis. Additionally, we also analyzed oxidative stress induced by CC. Cell viability studies confirmed the IC 50 of CC in Ishikawa cells to be 20 µM after 48 h treatment. CC arrests the cells in G0/G1 phase through cyclin D1 and cyclin E mediated pathways. Phosphatidylserine externalization, nuclear morphology changes, DNA fragmentation, PARP cleavage, and alteration of Bcl-2 family protein expression clearly suggest ongoing apoptosis in the CC treated cells. Activation of caspase 3 & 9, up-regulation of AIF and inhibition of apoptosis by z-VAD-fmk clearly explains the participation of the intrinsic pathway of programmed cell death. Further, the increase of ROS, loss of MMP, inhibition of antioxidant (MnSOD, Cu/Zn-SOD and GST) and inhibition of apoptosis with L-NAC suggests CC induced oxidative stress leading to apoptosis via mitochondria mediated pathway. Therefore, CC could be a potential therapeutic agent for the treatment of Endometrial Cancer adjunct to its utility as a contraceptive and an anti-breast cancer agent.

Programmed cell death in vegetative development: apoptosis during the colonial life cycle of the ascidian Botryllus schlosseri.

PubMed

Tiozzo, S; Ballarin, L; Burighel, P; Zaniolo, G

2006-06-01

Programmed cell death (PCD) by apoptosis is a physiological mechanism by which cells are eliminated during embryonic and post-embryonic stages of animal life cycle. During asexual reproduction, the zooids of colonial ascidians originate from an assorted cell population instead of a single zygote, so that we assume that regulation of the equilibrium among proliferation, differentiation and cell death may follow different pathways in comparison to the embryonic development. Here we investigate the presence of apoptotic events throughout the blastogenetic life cycle of the colonial ascidian Botryllus schlosseri, by means of terminal deoxynucleotidyl transferase dUTP Nick End Labeling (TUNEL) coupled with histochemical and electron microscopy techniques. The occurrence of low levels of morphogenetic cell death suggests that, in contrast to what happens during sexual development (embryogenesis and metamorphosis), apoptosis does not play a pivotal role during asexual propagation in botryllid ascidian. Nevertheless, PCD emerges as a key force to regulate homeostasis in adult zooids and to shape and modulate the growth of the whole colony.

Synergistic apoptosis in head and neck squamous cell carcinoma cells by co-inhibition of insulin-like growth factor-1 receptor signaling and compensatory signaling pathways.

PubMed

Axelrod, Mark J; Mendez, Rolando E; Khalil, Ashraf; Leimgruber, Stephanie S; Sharlow, Elizabeth R; Capaldo, Brian; Conaway, Mark; Gioeli, Daniel G; Weber, Michael J; Jameson, Mark J

2015-12-01

In head and neck squamous cell carcinoma (HNSCC), resistance to single-agent targeted therapy may be overcome by co-targeting of compensatory signaling pathways. A targeted drug screen with 120 combinations was used on 9 HNSCC cell lines. Multiple novel drug combinations demonstrated synergistic growth inhibition. Combining the insulin-like growth factor-1 receptor (IGF-1R) inhibitor, BMS754807, with either the human epidermal growth factor receptor (HER)-family inhibitor, BMS599626, or the Src-family kinase inhibitor, dasatinib, resulted in substantial synergy and growth inhibition. Depending on the cell line, these combinations induced synergistic or additive apoptosis; when synergistic apoptosis was observed, AKT phosphorylation was inhibited to a greater extent than either drug alone. Conversely, when additive apoptosis occurred, AKT phosphorylation was not reduced by the drug combination. Combined IGF-1R/HER family and IGF-1R/Src family inhibition may have therapeutic potential in HNSCC. AKT may be a node of convergence between IGF-1R signaling and pathways that compensate for IGF-1R inhibition. © 2015 Wiley Periodicals, Inc.

Examining live cell cultures during apoptosis by digital holographic phase imaging and Raman spectroscopy

NASA Astrophysics Data System (ADS)

Khmaladze, Alexander

2017-11-01

Cellular apoptosis is a unique, organized series of events, leading to programmed cell death. In this work, we present a combined digital holography/Raman spectroscopy technique to study live cell cultures during apoptosis. Digital holographic microscopy measurements of live cell cultures yield information about cell shape and volume, changes to which are indicative of alterations in cell cycle and initiation of cell death mechanisms. Raman spectroscopic measurements provide complementary information about cells, such as protein, lipid and nucleic acid content, and the spectral signatures associated with structural changes in molecules. Our work indicates that the chemical changes in proteins, which were detected by Raman measurements, preceded morphological changes, which were seen with digital holographic microscopy.

Escin-induced DNA damage promotes escin-induced apoptosis in human colorectal cancer cells via p62 regulation of the ATM/γH2AX pathway.

PubMed

Wang, Zhong; Chen, Qiang; Li, Bin; Xie, Jia-Ming; Yang, Xiao-Dong; Zhao, Kui; Wu, Yong; Ye, Zhen-Yu; Chen, Zheng-Rong; Qin, Zheng-Hong; Xing, Chun-Gen

2018-05-31

Escin, a triterpene saponin isolated from horse chestnut seed, has been used to treat encephaledema, tissue swelling and chronic venous insufficiency. Recent studies show that escin induces cell cycle arrest, tumor proliferation inhibition and tumor cell apoptosis. But the relationship between escin-induced DNA damage and cell apoptosis in tumor cells remains unclear. In this study, we investigated whether and how escin-induced DNA damage contributed to escin-induced apoptosis in human colorectal cancer cells. Escin (5-80 μg/mL) dose-dependently inhibited the cell viability and colony formation in HCT116 and HCT8 cells. Escin treatment induced DNA damage, leading to p-ATM and γH2AX upregulation. Meanwhile, escin treatment increased the expression of p62, an adaptor protein, which played a crucial role in controlling cell survival and tumorigenesis, and had a protective effect against escin-induced DNA damage: knockdown of p62 apparently enhanced escin-induced DNA damage, whereas overexpression of p62 reduced escin-induced DNA damage. In addition, escin treatment induced concentration- and time-dependent apoptosis. Similarly, knockdown of p62 significantly increased escin-induced apoptosis in vitro and produced en escin-like antitumor effect in vivo. Overexpression of p62 decreased the rate of apoptosis. Further studies revealed that the functions of p62 in escin-induced DNA damage were associated with escin-induced apoptosis, and p62 knockdown combined with the ATM inhibitor KU55933 augmented escin-induced DNA damage and further increased escin-induced apoptosis. In conclusion, our results demonstrate that p62 regulates ATM/γH2AX pathway-mediated escin-induced DNA damage and apoptosis.

Molecular mechanisms of celery seed extract induced apoptosis via s phase cell cycle arrest in the BGC-823 human stomach cancer cell line.

PubMed

Gao, Lin-Lin; Feng, Lei; Yao, Shu-Tong; Jiao, Peng; Qin, Shu-Cun; Zhang, Wei; Zhang, Ya-Bin; Li, Fu-Rong

2011-01-01

Mechanisms of apoptosis in tumor cells is an important field of tumor therapy and cancer molecular biology. Loss of cell cycle control, leading to uncontrolled proliferation, is common in cancer. Therefore, the identification of potent and selective cyclin dependent kinase inhibitors is a priority for anti-cancer drug discovery. There are at least two major apoptotic pathways, initiated by caspase-8 and caspase-9, respectively, which can activate caspase cascades. Apoptosis triggered by activation of the mitochondrial-dependent caspase pathway represents the main programmed cell death mechanism. This is activated by various intracellular stresses that induce permeabilization of the mitochondrial membrane. Anti-tumor effects of celery seed extract (CSE) and related mechanisms regarding apoptosis were here investigated in human gastric cancer BGC-823 cells. CSE was produced by supercritical fluid extraction. Cell viability was analyzed by 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl-tetrazolium bromide (MTT) assay and apoptosis by flow cytometry using Annexin/PI staining and DAPI staining and a laser scanning confocal microscope (LSCM). Cell cycling was evaluated using PI staining with flow cytometry and expression of cell cycle and apoptosis-related proteins cyclin A, CDK2, bcl-2 and bax was assessed by immunohistochemical staining. CSE had an anti-proliferation effect on human gastric cancer BGC-823 cells in a dose- and time-dependent manner. After treatment, the apoptotic rate significantly increased, with morphological changes typical of apoptosis observed with LSCM by DAPI staining. Cell cycle and apoptosis related proteins, such as cyclin A, CDK2 and bcl-2 were all down-regulated, whereas bax was up-regulated. The molecular determinants of inhibition of cell proliferation as well as apoptosis of CSE may be associated with cycle arrest in the S phase.

Apoptosis and proliferation of oligodendrocyte progenitor cells in the irradiated rodent spinal cord

DOE Office of Scientific and Technical Information (OSTI.GOV)

Atkinson, Shelley L.; Li Yuqing; Wong, C. Shun

2005-06-01

Purpose: Oligodendrocytes undergo early apoptosis after irradiation. The aim of this study was to determine the relationship between oligodendroglial apoptosis and proliferation of oligodendrocyte progenitor cells (OPC) in the irradiated central nervous system. Methods and Materials: Adult rats and p53 transgenic mice were given single doses of 2 Gy, 8 Gy, or 22 Gy to the cervical spinal cord. Apoptosis was assessed using TUNEL (Tdt-mediated dUTP terminal nick-end labeling) staining or by examining nuclear morphology. Oligodendrocyte progenitor cells were identified with an NG2 antibody or by in situ hybridization for platelet-derived growth factor receptor {alpha}. Proliferation of OPC was assessedmore » by in vivo bromodeoxyuridine (BrdU) labeling and subsequent immunohistochemistry. Because radiation-induced apoptosis of oligodendroglial cells is p53 dependent, p53 transgenic mice were used to study the relationship between apoptosis and cell proliferation. Results: Oligodendrocyte progenitor cells underwent apoptosis within 24 h of irradiation in the rat. That did not result in a change in OPC density at 24 h. Oligodendrocyte progenitor cell density was significantly reduced by 2-4 weeks, but showed recovery by 6 weeks after irradiation. An increase in BrdU-labeled cells was observed at 2 weeks after 8 Gy or 22 Gy, and proliferating cells in the rat spinal cord were immunoreactive for NG2. The mouse spinal cord showed a similar early cell proliferation after irradiation. No difference was observed in the proliferation response in the spinal cord of p53 -/- mice compared with wild type animals. Conclusions: Oligodendroglial cells undergo early apoptosis and OPC undergo early proliferation after ionizing radiation. However, apoptosis is not likely to be the trigger for early proliferation of OPC in the irradiated central nervous system.« less

Anti-CHMP5 single chain variable fragment antibody retrovirus infection induces programmed cell death of AML leukemic cells in vitro.

PubMed

Wang, Hai-rong; Xiao, Zhen-yu; Chen, Miao; Wang, Fei-long; Liu, Jia; Zhong, Hua; Zhong, Ji-hua; Ou-Yang, Ren-rong; Shen, Yan-lin; Pan, Shu-ming

2012-06-01

Over-expressed CHMP5 was found to act as oncogene that probably participated in leukemogenesis. In this study, we constructed the CHMP5 single chain variable fragment antibody (CHMP5-scFv) retrovirus and studied the changes of programmed cell death (PCD) of AML leukemic cells after infection by the retrovirus. The anti-CHMP5 KC14 hybridoma cell line was constructed to generate monoclonal antibody of CHMP5. The protein expression of CHMP5 was studied using immunofluorescence analysis. pMIG-CHMP5 scFv antibody expressible retroviral vector was constructed to prepare CHMP5-scFv retrovirus. AML leukemic U937 cells were infected with the retrovirus, and programmed cell death was studied using confocal microscope, FCM and Western blot. We obtained a monoclonal antibody of CHMP5, and found the expression of CHMP5 was up-regulated in the leukemic cells. After U937 cells were infected with CHMP5-scFv retrovirus, CHMP5 protein was neutralized. Moreover, the infection resulted in a significant increase in apoptosis and necrosis of U937 cells. In U937 cells infected with CHMP5-scFv retrovirus, apoptosis-inducing factor (AIF)-mediated caspase-independent necrotic PCD was activated, but autophagic programmed cell death was not observed. Neither the intrinsic nor extrinsic apoptotic PCD pathway was activated. The granzyme B/perforin-mediated caspase-dependent apoptotic PCD pathway was not activated. CHMP5-scFv retrovirus can neutralize the abnormally high levels of the CHMP5 protein in the cytosol of AML leukemic U937 cells, thereby inducing the programmed cell death of the leukemic cells via AIF-mediated caspase-independent necrosis and apoptosis.

Mammalian Ste20-like protein kinase 3 mediates trophoblast apoptosis in spontaneous delivery.

PubMed

Wu, Hung-Yi; Lin, Chia-Ying; Lin, Tze-Yi; Chen, Tai-Chang; Yuan, Chiun-Jye

2008-02-01

The placenta is essential in transferring gases and nutrients from the mother to the developing fetus. Trophoblast apoptosis may cause labor or other pregnancy-related disorders. This study demonstrated the essential role of Mst3, a human Ste20-like protein kinase, in the oxidative stress-induced apoptosis of trophoblasts of term placenta in normal spontaneous delivery. Oxidative stress, but not hormones released during labor such as prostaglandin E1, oxytocin or angiotensin II, induces the expression of Mst3 and apoptosis of human term placenta after elective Cesarean section without labor pain. The role of Mst3 in oxidative stress-induced apoptosis was further demonstrated in the 3A-sub-E, a human trophoblast cell line. The H2O2-induced apoptosis of 3A-sub-E cells was largely suppressed by overexpressed Mst3KR, the kinase-dead mutant or by selective knockdown of endogenous Mst3. Further studies showed that Jun N-terminal kinase (JNK) may participate in the signaling pathway of H2O2-induced apoptosis by mediating the level of Mst3. Subsequently, caspase 3 and other downstream apoptotic components may be activated by Mst3 and trigger the apoptotic process in human trophoblasts.

Verocytotoxin-induced apoptosis of human microvascular endothelial cells.

PubMed

Pijpers, A H; van Setten, P A; van den Heuvel, L P; Assmann, K J; Dijkman, H B; Pennings, A H; Monnens, L A; van Hinsbergh, V W

2001-04-01

The pathogenesis of the epidemic form of hemolytic uremic syndrome is characterized by endothelial cell damage. In this study, the role of apoptosis in verocytotoxin (VT)-mediated endothelial cell death in human glomerular microvascular endothelial cells (GMVEC), human umbilical vein endothelial cells, and foreskin microvascular endothelial cells (FMVEC) was investigated. VT induced apoptosis in GMVEC and human umbilical vein endothelial cells when the cells were prestimulated with the inflammatory mediator tumor necrosis factor-alpha (TNF-alpha). FMVEC displayed strong binding of VT and high susceptibility to VT under basal conditions, which made them suitable for the study of VT-induced apoptosis without TNF-alpha interference. On the basis of functional (flow cytometry and immunofluorescence microscopy using FITC-conjugated annexin V and propidium iodide), morphologic (transmission electron microscopy), and molecular (agarose gel electrophoresis of cellular DNA fragments) criteria, it was documented that VT induced programmed cell death in microvascular endothelial cells in a dose- and time-dependent manner. Furthermore, whereas partial inhibition of protein synthesis by VT was associated with a considerable number of apoptotic cells, comparable inhibition of protein synthesis by cycloheximide was not. This suggests that additional pathways, independent of protein synthesis inhibition, may be involved in VT-mediated apoptosis in microvascular endothelial cells. Specific inhibition of caspases by Ac-Asp-Glu-Val-Asp-CHO, but not by Ac-Tyr-Val-Ala-Asp-CHO, was accompanied by inhibition of VT-induced apoptosis in FMVEC and TNF-alpha-treated GMVEC. These data indicate that VT can induce apoptosis in human microvascular endothelial cells.

Inhibition of tumor cellular proteasome activity by triptolide extracted from the Chinese medicinal plant 'thunder god vine'.

PubMed

Lu, Li; Kanwar, Jyoti; Schmitt, Sara; Cui, Qiuzhi Cindy; Zhang, Chuanyin; Zhao, Cong; Dou, Q Ping

2011-01-01

The molecular mechanisms of triptolide responsible for its antitumor properties are not yet fully understood. The ubiquitin/proteasome system is an important pathway of protein degradation in cells. This study investigated whether triptolide may inhibit proteasomal activity and induce apoptosis in human cancer cells. In vitro proteasome inhibition was measured by incubation of a purified 20S proteasome with triptolide. Human breast and prostate cancer cell lines were also treated with different doses of triptolide for different times, followed by measurement of proteasome inhibition (levels of the chymotrypsin-like activity, ubiquitinated proteins and three well-known proteasome target proteins, p27, IκB-α and Bax) and apoptosis induction (caspase-3 activity and PARP cleavage). Triptolide did not inhibit the chymotrypsin-like activity of purified 20S proteasome. However, treatment of triptolide was able to cause decreased levels of cellular proteasomal chymotrypsin-like activity and accumulation of ubiquitinated proteins and three well-known proteasome target proteins in human breast and prostate cancer cells, associated with apoptosis induction. It is possible that at least one of metabolites of triptolide has proteasome-inhibitory activity.

Apoptosis of circulating lymphocytes during pediatric cardiac surgery

NASA Astrophysics Data System (ADS)

Bocsi, J.; Pipek, M.; Hambsch, J.; Schneider, P.; Tárnok, A.

2006-02-01

There is a constant need for clinical diagnostic systems that enable to predict disease course for preventative medicine. Apoptosis, programmed cell death, is the end point of the cell's response to different induction and leads to changes in the cell morphology that can be rapidly detected by optical systems. We tested whether apoptosis of T-cells in the peripheral blood is useful as predictor and compared different preparation and analytical techniques. Surgical trauma is associated with elevated apoptosis of circulating leukocytes. Increased apoptosis leads to partial removal of immune competent cells and could therefore in part be responsible for reduced immune defence. Cardiovascular surgery with but not without cardiopulmonary bypass (CPB) induces transient immunosuppression. Its effect on T-cell apoptosis has not been shown yet. Flow-cytometric data of blood samples from 107 children (age 3-16 yr.) who underwent cardiac surgery with (78) or without (29) CPB were analysed. Apoptotic T-lymphocytes were detected based on light scatter and surface antigen (CD45/CD3) expression (ClinExpImmunol2000;120:454). Results were compared to staining with CD3 antibodies alone and in the absence of antibodies. T-cell apoptosis rate was comparable when detected with CD45/CD3 or CD3 alone, however not in the absence of CD3. Patients with but not without CPB surgery had elevated lymphocyte apoptosis. T-cell apoptosis increased from 0.47% (baseline) to 0.97% (1 day postoperatively). In CPB patients with complication 1.10% significantly higher (ANOVA p=0.01) comparing to CPB patients without complications. Quantitation of circulating apoptotic cells based on light scatter seems an interesting new parameter for diagnosis. Increased apoptosis of circulating lymphocytes and neutrophils further contributes to the immune suppressive response to surgery with CPB. (Support: MP, Deutsche Herzstiftung, Frankfurt, Germany)

Lack of HXK2 induces localization of active Ras in mitochondria and triggers apoptosis in the yeast Saccharomyces cerevisiae.

PubMed

Amigoni, Loredana; Martegani, Enzo; Colombo, Sonia

2013-01-01

We recently showed that activated Ras proteins are localized to the plasma membrane and in the nucleus in wild-type cells growing exponentially on glucose, while in the hxk2Δ strain they accumulated mainly in mitochondria. An aberrant accumulation of activated Ras in these organelles was previously reported and correlated to mitochondrial dysfunction, accumulation of ROS, and cell death. Here we show that addition of acetic acid to wild-type cells results in a rapid recruitment of Ras-GTP from the nucleus and the plasma membrane to the mitochondria, providing a further proof that Ras proteins might be involved in programmed cell death. Moreover, we show that Hxk2 protects against apoptosis in S. cerevisiae. In particular, cells lacking HXK2 and showing a constitutive accumulation of activated Ras at the mitochondria are more sensitive to acetic-acid-induced programmed cell death compared to the wild type strain. Indeed, deletion of HXK2 causes an increase of apoptotic cells with several morphological and biochemical changes that are typical of apoptosis, including DNA fragmentation, externalization of phosphatidylserine, and ROS production. Finally, our results suggest that apoptosis induced by lack of Hxk2 may not require the activation of Yca1, the metacaspase homologue identified in yeast.

Drosophila MOF regulates DIAP1 and induces apoptosis in a JNK dependent pathway.

PubMed

Pushpavalli, Sreerangam N C V L; Sarkar, Arpita; Ramaiah, M Janaki; Koteswara Rao, G; Bag, Indira; Bhadra, Utpal; Pal-Bhadra, Manika

2016-03-01

Histone modulations have been implicated in various cellular and developmental processes where in Drosophila Mof is involved in acetylation of H4K16. Reduction in the size of larval imaginal discs is observed in the null mutants of mof with increased apoptosis. Deficiency involving Hid, Reaper and Grim [H99] alleviated mof (RNAi) induced apoptosis in the eye discs. mof (RNAi) induced apoptosis leads to activation of caspases which is suppressed by over expression of caspase inhibitors like P35 and Diap1 clearly depicting the role of caspases in programmed cell death. Also apoptosis induced by knockdown of mof is rescued by JNK mutants of bsk and tak1 indicating the role of JNK in mof (RNAi) induced apoptosis. The adult eye ablation phenotype produced by ectopic expression of Hid, Rpr and Grim, was restored by over expression of Mof. Accumulation of Mof at the Diap1 promoter 800 bp upstream of the transcription start site in wild type larvae is significantly higher (up to twofolds) compared to mof (1) mutants. This enrichment coincides with modification of histone H4K16Ac indicating an induction of direct transcriptional up regulation of Diap1 by Mof. Based on these results we propose that apoptosis triggered by mof (RNAi) proceeds through a caspase-dependent and JNK mediated pathway.

How do viruses control mitochondria-mediated apoptosis?

PubMed

Neumann, Simon; El Maadidi, Souhayla; Faletti, Laura; Haun, Florian; Labib, Shirin; Schejtman, Andrea; Maurer, Ulrich; Borner, Christoph

2015-11-02

There is no doubt that viruses require cells to successfully reproduce and effectively infect the next host. The question is what is the fate of the infected cells? All eukaryotic cells can "sense" viral infections and exhibit defence strategies to oppose viral replication and spread. This often leads to the elimination of the infected cells by programmed cell death or apoptosis. This "sacrifice" of infected cells represents the most primordial response of multicellular organisms to viruses. Subverting host cell apoptosis, at least for some time, is therefore a crucial strategy of viruses to ensure their replication, the production of essential viral proteins, virus assembly and the spreading to new hosts. For that reason many viruses harbor apoptosis inhibitory genes, which once inside infected cells are expressed to circumvent apoptosis induction during the virus reproduction phase. On the other hand, viruses can take advantage of stimulating apoptosis to (i) facilitate shedding and hence dissemination, (ii) to prevent infected cells from presenting viral antigens to the immune system or (iii) to kill non-infected bystander and immune cells which would limit viral propagation. Hence the decision whether an infected host cell undergoes apoptosis or not depends on virus type and pathogenicity, its capacity to oppose antiviral responses of the infected cells and/or to evade any attack from immune cells. Viral genomes have therefore been adapted throughout evolution to satisfy the need of a particular virus to induce or inhibit apoptosis during its life cycle. Here we review the different strategies used by viruses to interfere with the two major apoptosis as well as with the innate immune signaling pathways in mammalian cells. We will focus on the intrinsic mitochondrial pathway and discuss new ideas about how particular viruses could activately engage mitochondria to induce apoptosis of their host. Copyright © 2015 Elsevier B.V. All rights reserved.

SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bonifati, Serena; Daly, Michele B.; St Gelais, Corine

SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G{sub 1}/G{sub 0} phase and reduces apoptosis. These alterationsmore » correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.« less

Exosomes Secreted by Apoptosis-Resistant Acute Myeloid Leukemia (AML) Blasts Harbor Regulatory Network Proteins Potentially Involved in Antagonism of Apoptosis*

PubMed Central

Wojtuszkiewicz, Anna; Schuurhuis, Gerrit J.; Kessler, Floortje L.; Piersma, Sander R.; Knol, Jaco C.; Pham, Thang V.; Jansen, Gerrit; Musters, René J. P.; van Meerloo, Johan; Assaraf, Yehuda G.; Kaspers, Gertjan J. L.; Zweegman, Sonja; Cloos, Jacqueline; Jimenez, Connie R.

2016-01-01

Expression of apoptosis-regulating proteins (B-cell CLL/lymphoma 2 - BCL-2, Myeloid Cell Leukemia 1 - MCL-1, BCL-2 like 1 - BCL-X and BCL-2-associated X protein - BAX) in acute myeloid leukemia (AML) blasts at diagnosis is associated with disease-free survival. We previously found that the initially high apoptosis-resistance of AML cells decreased after therapy, while regaining high levels at relapse. Herein, we further explored this aspect of dynamic apoptosis regulation in AML. First, we showed that the intraindividual ex vivo apoptosis-related profiles of normal lymphocytes and AML blasts within the bone marrow of AML patients were highly correlated. The expression values of apoptosis-regulating proteins were far beyond healthy control lymphocytes, which implicates the influence of microenvironmental factors. Second, we demonstrated that apoptosis-resistant primary AML blasts, as opposed to apoptosis-sensitive cells, were able to up-regulate BCL-2 expression in sensitive AML blasts in contact cultures (p = 0.0067 and p = 1.0, respectively). Using secretome proteomics, we identified novel proteins possibly engaged in apoptosis regulation. Intriguingly, this analysis revealed that major functional protein clusters engaged in global gene regulation, including mRNA splicing, protein translation, and chromatin remodeling, were more abundant (p = 4.01E-06) in secretomes of apoptosis-resistant AML. These findings were confirmed by subsequent extracellular vesicle proteomics. Finally, confocal-microscopy-based colocalization studies show that splicing factors-containing vesicles secreted by high AAI cells are taken up by low AAI cells. The current results constitute the first comprehensive analysis of proteins released by apoptosis-resistant and sensitive primary AML cells. Together, the data point to vesicle-mediated release of global gene regulatory protein clusters as a plausible novel mechanism of induction of apoptosis resistance. Deciphering the modes of communication between apoptosis-resistant blasts may in perspective lead to the discovery of prognostic tools and development of novel therapeutic interventions, aimed at limiting or overcoming therapy resistance. PMID:26801919

[Role of p38MAPK/eNOS signaling pathway in the inhibition of AGEs-induced apoptosis of human umbilical vein endothelial cells by glucagon-like peptide-1].

PubMed

Zeng, Hailong; Huang, Zhiqiu; Zhang, Yineng; Sun, Huilin

2016-01-01

To investigate the role of p38MAPK signaling pathway in the mechanism by which glucagon-like peptide-1 (GLP-1) inhibits endothelial cell damage induced by AGEs. Human umbilical vein endothelial cells were divided into control group, AGEs group, GLP-1 group, AGEs+GLP-1 group, AGEs+inhibitor group, and AGEs+GLP-1+inhibitor group. The expressions of p-p38MAPK/p38MAPK and p-eNOS/eNOS protein were examined by Western blotting, and the cell apoptosis rates were tested by flow cytometry. Compared with the control group, AGEs significantly enhanced the expression of p-p38 MAPK protein (P=0.001) while GLP-1 significantly inhibited its expression (P<0.001). AGEs significantly inhibited the expression of p-eNOS protein (P=0.007), which was enhanced by GLP-1 and p38 MAPK inhibitor (SB203580) (P=0.004). Both SB203580 and GLP-1 treatment decreased the apoptosis rate of AGEs-treated cells (P<0.001). GLP-1 can protect human umbilical vein endothelial cells against AGEs-induced apoptosis partially by inhibiting the phosphorylation of p38MAPK protein and promoting the expression of p-eNOS protein.

Key apoptotic pathways for heat-induced programmed germ cell death in the testis.

PubMed

Hikim, Amiya P Sinha; Lue, Yanhe; Yamamoto, Cindy M; Vera, Yanira; Rodriguez, Susana; Yen, Pauline H; Soeng, Kevin; Wang, Christina; Swerdloff, Ronald S

2003-07-01

Short-term exposure (43 C for 15 min) of the rat testis to mild heat results within 6 h in stage- and cell-specific activation of germ cell apoptosis. Initiation of apoptosis was preceded by a redistribution of Bax from a cytoplasmic to paranuclear localization in heat-susceptible germ cells. Here we show that the relocation of Bax is accompanied by cytosolic translocation of cytochrome c and is associated with activation of the initiator caspase 9 and the executioner caspases 3, 6, and 7 and cleavage of poly(ADP) ribose polymerase. Furthermore, early in apoptosis, a significant amount of Bax also accumulates in endoplasmic reticulum, as assessed by Western blot analyses of fractionated testicular lysates. In additional studies using the FasL-defective gld mice, we have shown that heat-induced germ cell apoptosis is not blocked, thus providing evidence that the Fas signaling system may be dispensable for heat-induced germ cell apoptosis in the testis. Taken together, these results demonstrate that the mitochondria- and possibly also endoplasmic reticulum-dependent pathways are the key apoptotic pathways for heat-induced germ cell death in the testis.

Mille modis morimur: We die in a thousand ways.

PubMed

Banfalvi, Gaspar

2017-02-01

Dying cells subjected to apoptotic programs are engulfed by neighboring cells or by professional phagocytes, without inflammation or immunological reactions in the tissue where apoptosis takes place. Apoptotic cells release danger-associated project signals to their neighbours, through different molecular patterns, stimulate antigen production and immune responses. Microenvironmental effects with several functional consequences indicate that cell death is a complex process and may take place in several ways. This idea is expressed by the title of the Special Issue and by the title of the guest editorial "Mille modis morimur" meaning that not only multicellular organisms, but also single cells may die in a thousand ways. This idea is demonstrated by the papers serving as examples for cell death. Apoptosis was induced by clary sage oil in Candida cells. Heavy metal (Gd) induced cell motility and apoptosis was found in mammalian cells. RNA oxidation enhanced the reversion frequency of apoptosis in yeast mutants. The frequency of apoptotic micronucleus formation increased in a concentration-dependent manner by methotrexate. The antioxidant coenzyme Q10 protected renal proximal tubule cells against nicotine-induced apoptosis. The synergy of 2-deoxy-D-glucose combined with berberine induced lysosome/autophagy. The mitochondrial apoptotic pathway could be regulated by glucocorticoid receptor in collaboration with Bcl-2 family proteins in developing T cells. Cylindrospermopsin induced biochemical changes led to apoptosis in plants. Mechanisms of stress seriously impacted the risk of apoptosis. Transcriptional control of apoptotic cell clearance was achieved by macrophage nuclear receptors. Finally, the clinical aspects of apoptosis-induced lymphopenia were reviewed in sepsis and other severe injuries. These examples not only support the view of many ways of cell death, but predict further potential ways to induce or reduce the risk of cell death.

Epidermal growth factor-like growth factors prevent apoptosis of alcohol-exposed human placental cytotrophoblast cells.

PubMed

Wolff, Garen S; Chiang, Po Jen; Smith, Susan M; Romero, Roberto; Armant, D Randall

2007-07-01

Maternal alcohol abuse during pregnancy can produce an array of birth defects comprising fetal alcohol syndrome. A hallmark of fetal alcohol syndrome is intrauterine growth retardation, which is associated with elevated apoptosis of placental cytotrophoblast cells. Using a human first trimester cytotrophoblast cell line, we examined the relationship between exposure to ethanol and cytotrophoblast survival, as well as the ameliorating effects of epidermal growth factor (EGF)-like growth factors produced by human cytotrophoblast cells. After exposure to 0-100 mM ethanol, cell death was quantified by the TUNEL method, and expression of the nuclear proliferation marker, Ki67, was measured by immunohistochemistry. The mode of cell death was determined by assessing annexin V binding, caspase 3 activation, pyknotic nuclear morphology, reduction of TUNEL by caspase inhibition, and cellular release of lactate dehydrogenase. Ethanol significantly reduced proliferation and increased cell death approximately 2.5-fold through the apoptotic pathway within 1-2 h of exposure to 50 mM alcohol. Exposure to 25-50 mM ethanol significantly increased transforming growth factor alpha (TGFA) and heparin-binding EGF-like growth factor (HBEGF), but not EGF or amphiregulin (AREG). When cytotrophoblasts were exposed concurrently to 100 mM ethanol and 1 nM HBEGF or TGFA, the increase in apoptosis was prevented, while EGF ameliorated at 10 nM and AREG was weakly effective. HBEGF survival-promoting activity required ligation of either of its cognate receptors, HER1 or HER4. These findings reveal the potential for ethanol to rapidly induce cytotrophoblast apoptosis. However, survival factor induction could provide cytotrophoblasts with an endogenous cytoprotective mechanism.

Necroptosis, necrostatins and tissue injury

PubMed Central

Smith, Christopher CT; Yellon, Derek M

2011-01-01

Abstract Cell death is an integral part of the life of an organism being necessary for the maintenance of organs and tissues. If, however, cell death is allowed to proceed unrestricted, tissue damage and degenerative disease may ensue. Until recently, three morphologically distinct types of cell death were recognized, apoptosis (type I), autophagy (type II) and necrosis (type III). Apoptosis is a highly regulated, genetically determined mechanism designed to dismantle cells systematically (e.g. cells that are no longer functionally viable), via protease (caspase) action, and maintain homeostasis. Autophagy is responsible for the degradation of cytoplasmic material, e.g. proteins and organelles, through autophagosome formation and subsequent proteolytic degradation by lysosomes, and is normally considered in the context of survival although it is sometimes associated with cell death. Necrosis was formerly considered to be an accidental, unregulated form of cell death resulting from excessive stress, although it has been suggested that this is an over-simplistic view as necrosis may under certain circumstances involve the mobilization of specific transduction mechanisms. Indeed, recently, an alternative death pathway, termed necroptosis, was delineated and proposed as a form of ‘programmed necrosis’. Identified with the aid of specific inhibitors called necrostatins, necroptosis shares characteristics with both necrosis and apoptosis. Necroptosis involves Fas/tumour necrosis factor-α death domain receptor activation and inhibition of receptor-interacting protein I kinase, and it has been suggested that it may contribute to the development of neurological and myocardial diseases. Significantly, necrostatin-like drugs have been mooted as possible future therapeutic agents for the treatment of degenerative conditions. PMID:21564515

EF24, a novel synthetic curcumin analog, induces apoptosis in cancer cells via a redox-dependent mechanism.

PubMed

Adams, Brian K; Cai, Jiyang; Armstrong, Jeff; Herold, Marike; Lu, Yang J; Sun, Aiming; Snyder, James P; Liotta, Dennis C; Jones, Dean P; Shoji, Mamoru

2005-03-01

In this study, we show that the novel synthetic curcumin analog, EF24, induces cell cycle arrest and apoptosis by means of a redox-dependent mechanism in MDA-MB-231 human breast cancer cells and DU-145 human prostate cancer cells. Cell cycle analysis demonstrated that EF24 causes a G2/M arrest in both cell lines, and that this cell cycle arrest is followed by the induction of apoptosis as evidenced by caspase-3 activation, phosphatidylserine externalization and an increased number of cells with a sub-G1 DNA fraction. In addition, we demonstrate that EF24 induces a depolarization of the mitochondrial membrane potential, suggesting that the compound may also induce apoptosis by altering mitochondrial function. EF24, like curcumin, serves as a Michael acceptor reacting with glutathione (GSH) and thioredoxin 1. Reaction of EF24 with these agents in vivo significantly reduced intracellular GSH as well as oxidized GSH in both the wild-type and Bcl-xL overexpressing HT29 human colon cancer cells. We therefore propose that the anticancer effect of a novel curcumin analog, EF24, is mediated in part by redox-mediated induction of apoptosis.

Live to die another way: modes of programmed cell death and the signals emanating from dying cells

PubMed Central

Fuchs, Yaron; Steller, Hermann

2015-01-01

Preface All life ends in death, but perhaps one of life’s grander ironies is that it also depends on death. Cell-intrinsic suicide pathways, termed programmed cell death (PCD), are crucial for animal development, tissue homeostasis and pathogenesis. Originally, PCD was virtually synonymous with apoptosis, but recently, alternative PCD mechanisms have been reported. Here, we provide an overview of several distinct PCD mechanisms, namely apoptosis, autophagy and necroptosis. In addition, we discuss the complex signals emanating from dying cells, which can either fuel regeneration or instruct additional killing. Further advances in understanding the physiological role of multiple cell death mechanisms and associated signals will be important to selectively manipulate PCD for therapeutic purposes. PMID:25991373

Combinatorial treatment with anacardic acid followed by TRAIL augments induction of apoptosis in TRAIL resistant cancer cells by the regulation of p53, MAPK and NFκβ pathways.

PubMed

Harsha Raj, M; Yashaswini, B; Rössler, Jochen; Salimath, Bharathi P

2016-05-01

TRAIL, an apoptosis inducing cytokine currently in phase II clinical trial, was investigated for its capability to induce apoptosis in six different human tumor cell lines out of which three cell lines showed resistance to TRAIL induced apoptosis. To investigate whether Anacardic acid (A1) an active component of Anacardium occidentale can sensitize the resistant cell lines to TRAIL induced apoptosis, we treated the resistant cells with suboptimal concentration of A1 and showed that it is a potent enhancer of TRAIL induced apoptosis which up-regulates the expression of both DR4 and DR5 receptors, which has been observed in the cellular, protein and mRNA levels. The death receptors upregulation consequent to A1 treatment was corroborated by the activation of p53 as well as phosphorylation of p38 and JNK MAP kinases and concomitant inactivation of NFκβ and ERK signaling cascades. Also, A1 modulated the expression of key apoptotic players like Bax, Bcl-2 and CAD along with the abatement of tumor angiogenesis in vivo in EAT mouse model. Thus, post A1 treatment the TRAIL resistant cells turned into TRAIL sensitive cells. Hence our results demonstrate that A1 can synergize TRAIL induced apoptosis through the upregulation of death receptors and downregulation of anti-apoptotic proteins in cancer context.

Control of apoptosis by Drosophila DCAF12.

PubMed

Hwangbo, Dae-Sung; Biteau, Benoit; Rath, Sneha; Kim, Jihyun; Jasper, Heinrich

2016-05-01

Regulated Apoptosis (Programmed Cell Death, PCD) maintains tissue homeostasis in adults, and ensures proper growth and morphogenesis of tissues during development of metazoans. Accordingly, defects in cellular processes triggering or executing apoptotic programs have been implicated in a variety of degenerative and neoplastic diseases. Here, we report the identification of DCAF12, an evolutionary conserved member of the WD40-motif repeat family of proteins, as a new regulator of apoptosis in Drosophila. We find that DCAF12 is required for Diap1 cleavage in response to pro-apoptotic signals, and is thus necessary and sufficient for RHG (Reaper, Hid, and Grim)-mediated apoptosis. Loss of DCAF12 perturbs the elimination of supernumerary or proliferation-impaired cells during development, and enhances tumor growth induced by loss of neoplastic tumor suppressors, highlighting the wide requirement for DCAF12 in PCD. Copyright © 2016 Elsevier Inc. All rights reserved.

Demyelination as a Target for Cell-Based Therapy of Chronic Blast-Induced Traumatic Brain Injury

DTIC Science & Technology

2014-10-01

like behavior was observed in 17*3psi pressure group at day 3 (Fig. 5). Figure 5: Anxiety-like behavior following BOP exposure (*p < 0.05 at day...3, a marker of apoptosis, in the 17*3 pressure group as depicted in figure 7. 9 Figure 7: An increase in marker of apoptosis, caspase-3 was...capase-3 levels, representative images of control- 0 psi (B) and 17*3 psi pressure group (C). Arrows represents caspase-3 positive cells. 6

Identification of a novel putative gastrointestinal stem cell and adenoma stem cell marker, doublecortin and CaM kinase-like-1, following radiation injury and in adenomatous polyposis coli/multiple intestinal neoplasia mice.

PubMed

May, Randal; Riehl, Terrence E; Hunt, Clayton; Sureban, Sripathi M; Anant, Shrikant; Houchen, Courtney W

2008-03-01

In the gut, tumorigenesis arises from intestinal or colonic crypt stem cells. Currently, no definitive markers exist that reliably identify gut stem cells. Here, we used the putative stem cell marker doublecortin and CaM kinase-like-1 (DCAMKL-1) to examine radiation-induced stem cell apoptosis and adenomatous polyposis coli (APC)/multiple intestinal neoplasia (min) mice to determine the effects of APC mutation on DCAMKL-1 expression. Immunoreactive DCAMKL-1 staining was demonstrated in the intestinal stem cell zone. Furthermore, we observed apoptosis of the cells negative for DCAMKL-1 at 6 hours. We found DNA damage in all the cells in the crypt region, including the DCAMKL-1-positive cells. We also observed stem cell apoptosis and mitotic DCAMKL-1-expressing cells 24 hours after irradiation. Moreover, in APC/min mice, DCAMKL-1-expressing cells were negative for proliferating cell nuclear antigen and nuclear beta-catenin in normal-appearing intestine. However, beta-catenin was nuclear in DCAMKL-1-positive cells in adenomas. Thus, nuclear translocation of beta-catenin distinguishes normal and adenoma stem cells. Targeting DCAMKL-1 may represent a strategy for developing novel chemotherapeutic agents.

Competing Death Programs in Poliovirus-Infected Cells: Commitment Switch in the Middle of the Infectious Cycle

PubMed Central

Agol, Vadim I.; Belov, George A.; Bienz, Kurt; Egger, Denise; Kolesnikova, Marina S.; Romanova, Lyudmila I.; Sladkova, Larissa V.; Tolskaya, Elena A.

2000-01-01

Productive poliovirus infection of HeLa cells leads to the canonical cytopathic effect (CPE), whereas certain types of abortive infection result in apoptosis. To define the time course of commitment to the different types of poliovirus-induced death, inhibitors of viral replication (guanidine HCl) or translation (cycloheximide) were added at different times postinfection (p.i.). Early in the infection (during the first ∼2 h p.i.), predominantly proapoptotic viral function was expressed, rendering the cells committed to apoptosis, which developed several hours after viral expression was arrested. In the middle of infection, concomitantly with the onset of fast generation of viral progeny, the implementation of the viral apoptotic program was abruptly interrupted. In particular, activation of an Asp-Glu-Val-Asp (DEVD)-specific caspase(s) occurring in the apoptosis-committed cells was prevented by the ongoing productive infection. Simultaneously, the cells retaining normal or nearly normal morphology became committed to CPE, which eventually developed regardless of whether or not further viral expression was allowed to proceed. The implementation of the poliovirus-induced apoptotic program was suppressed in HeLa cells overexpressing the Bcl-2 protein, indicating that the fate of poliovirus-infected cells depends on the balance of host and viral pro- and antiapoptotic factors. PMID:10823859

Interference of silibinin with IGF-1R signalling pathways protects human epidermoid carcinoma A431 cells from UVB-induced apoptosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Weiwei; Otkur, Wuxiyar; Li, Lingzhi

Highlights: ► Silibinin protects A431 cells from UVB irradiation-induced apoptosis. ► Up-regulation of the IGF-1R-JNK/ERK pathways by UVB induces cell apoptosis. ► Silibinin inhibits IGF-1R pathways to repress caspase-8-mediated apoptosis. -- Abstract: Ultraviolet B (UVB) from sunlight is a major cause of cutaneous lesion. Silibinin, a traditional hepatic protectant, elicits protective effects against UVB-induced cellular damage. In A431 cells, the insulin-like growth factor-1 receptor (IGF-1R) was markedly up-regulated by UVB irradiation. The activation of the IGF-1R signalling pathways contributed to apoptosis of the cells rather than rescuing the cells from death. Up-regulated IGF-1R stimulated downstream mitogen-activated protein kinases (MAPKs), suchmore » as c-Jun N-terminal kinases (JNK) and extracellular signal-regulated protein kinases 1/2 (ERK1/2). The subsequent activation of caspase-8 and caspase-3 led to apoptosis. The activation of IGF-1R signalling pathways is the cause of A431 cell death. The pharmacological inhibitors and the small interfering RNA (siRNA) targeting IGF-1R suppressed the downstream activation of JNK/ERK-caspases to help the survival of the UVB-irradiated A431 cells. Indeed, silibinin treatment suppressed the IGF-1R-JNK/ERK pathways and thus protected the cells from UVB-induced apoptosis.« less

Absence of autophagy promotes apoptosis by modulating the ROS-dependent RLR signaling pathway in classical swine fever virus-infected cells

PubMed Central

Pei, Jingjing; Deng, Jieru; Ye, Zuodong; Wang, Jiaying; Gou, Hongchao; Liu, Wenjun; Zhao, Mingqiu; Liao, Ming; Yi, Lin; Chen, Jinding

2016-01-01

ABSTRACT A growing number of studies have demonstrated that both macroautophagy/autophagy and apoptosis are important inner mechanisms of cell to maintain homeostasis and participate in the host response to pathogens. We have previously reported that a functional autophagy pathway is trigged by infection of classical swine fever virus (CSFV) and is required for viral replication and release in host cells. However, the interplay of autophagy and apoptosis in CSFV-infected cells has not been clarified. In the present study, we demonstrated that autophagy induction with rapamycin facilitates cellular proliferation after CSFV infection, and that autophagy inhibition by knockdown of essential autophagic proteins BECN1/Beclin 1 or MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) promotes apoptosis via fully activating both intrinsic and extrinsic mechanisms in CSFV-infected cells. We also found that RIG-I-like receptor (RLR) signaling was amplified in autophagy-deficient cells during CSFV infection, which was closely linked to the activation of the intrinsic apoptosis pathway. Moreover, we discovered that virus infection of autophagy-impaired cells results in an increase in copy number of mitochondrial DNA and in the production of reactive oxygen species (ROS), which plays a significant role in enhanced RLR signaling and the activated extrinsic apoptosis pathway in cultured cells. Collectively, these data indicate that CSFV-induced autophagy delays apoptosis by downregulating ROS-dependent RLR signaling and thus contributes to virus persistent infection in host cells. PMID:27463126

Absence of autophagy promotes apoptosis by modulating the ROS-dependent RLR signaling pathway in classical swine fever virus-infected cells.

PubMed

Pei, Jingjing; Deng, Jieru; Ye, Zuodong; Wang, Jiaying; Gou, Hongchao; Liu, Wenjun; Zhao, Mingqiu; Liao, Ming; Yi, Lin; Chen, Jinding

2016-10-02

A growing number of studies have demonstrated that both macroautophagy/autophagy and apoptosis are important inner mechanisms of cell to maintain homeostasis and participate in the host response to pathogens. We have previously reported that a functional autophagy pathway is trigged by infection of classical swine fever virus (CSFV) and is required for viral replication and release in host cells. However, the interplay of autophagy and apoptosis in CSFV-infected cells has not been clarified. In the present study, we demonstrated that autophagy induction with rapamycin facilitates cellular proliferation after CSFV infection, and that autophagy inhibition by knockdown of essential autophagic proteins BECN1/Beclin 1 or MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) promotes apoptosis via fully activating both intrinsic and extrinsic mechanisms in CSFV-infected cells. We also found that RIG-I-like receptor (RLR) signaling was amplified in autophagy-deficient cells during CSFV infection, which was closely linked to the activation of the intrinsic apoptosis pathway. Moreover, we discovered that virus infection of autophagy-impaired cells results in an increase in copy number of mitochondrial DNA and in the production of reactive oxygen species (ROS), which plays a significant role in enhanced RLR signaling and the activated extrinsic apoptosis pathway in cultured cells. Collectively, these data indicate that CSFV-induced autophagy delays apoptosis by downregulating ROS-dependent RLR signaling and thus contributes to virus persistent infection in host cells.

[Apoptosis and pathological process].

PubMed

Rami, Mukhammed Salim Iusef

2007-01-01

Apoptosis (programmed cell death) occurs normally for maitenance of tissue homeostasis and play an important role in morphogenesis, embriogenesis and tissue growth. On the other hand, apoptosis may be involved in different pathological processes such as malignancy, infectious diseases and autoimmune disorders. Apoptosis is regulated by various mediators. Caspases, death receptors, mitochondria, Bcl-2 protoncogenes and tumor supressor genes are considered to be the most important of them. Advance in apoptosis regulation research suggests enormouse facilities for therapy of wide range of human illnesses.

Glucagon-Like Peptide-1 Regulates Cholecystokinin Production in β-Cells to Protect From Apoptosis.

PubMed

Linnemann, Amelia K; Neuman, Joshua C; Battiola, Therese J; Wisinski, Jaclyn A; Kimple, Michelle E; Davis, Dawn Belt

2015-07-01

Cholecystokinin (CCK) is a classic gut hormone that is also expressed in the pancreatic islet, where it is highly up-regulated with obesity. Loss of CCK results in increased β-cell apoptosis in obese mice. Similarly, islet α-cells produce increased amounts of another gut peptide, glucagon-like peptide 1 (GLP-1), in response to cytokine and nutrient stimulation. GLP-1 also protects β-cells from apoptosis via cAMP-mediated mechanisms. Therefore, we hypothesized that the activation of islet-derived CCK and GLP-1 may be linked. We show here that both human and mouse islets secrete active GLP-1 as a function of body mass index/obesity. Furthermore, GLP-1 can rapidly stimulate β-cell CCK production and secretion through direct targeting by the cAMP-modulated transcription factor, cAMP response element binding protein (CREB). We find that cAMP-mediated signaling is required for Cck expression, but CCK regulation by cAMP does not require stimulatory levels of glucose or insulin secretion. We also show that CREB directly targets the Cck promoter in islets from obese (Leptin(ob/ob)) mice. Finally, we demonstrate that the ability of GLP-1 to protect β-cells from cytokine-induced apoptosis is partially dependent on CCK receptor signaling. Taken together, our work suggests that in obesity, active GLP-1 produced in the islet stimulates CCK production and secretion in a paracrine manner via cAMP and CREB. This intraislet incretin loop may be one mechanism whereby GLP-1 protects β-cells from apoptosis.

Glucagon-Like Peptide-1 Regulates Cholecystokinin Production in β-Cells to Protect From Apoptosis

PubMed Central

Linnemann, Amelia K.; Neuman, Joshua C.; Battiola, Therese J.; Wisinski, Jaclyn A.; Kimple, Michelle E.

2015-01-01

Cholecystokinin (CCK) is a classic gut hormone that is also expressed in the pancreatic islet, where it is highly up-regulated with obesity. Loss of CCK results in increased β-cell apoptosis in obese mice. Similarly, islet α-cells produce increased amounts of another gut peptide, glucagon-like peptide 1 (GLP-1), in response to cytokine and nutrient stimulation. GLP-1 also protects β-cells from apoptosis via cAMP-mediated mechanisms. Therefore, we hypothesized that the activation of islet-derived CCK and GLP-1 may be linked. We show here that both human and mouse islets secrete active GLP-1 as a function of body mass index/obesity. Furthermore, GLP-1 can rapidly stimulate β-cell CCK production and secretion through direct targeting by the cAMP-modulated transcription factor, cAMP response element binding protein (CREB). We find that cAMP-mediated signaling is required for Cck expression, but CCK regulation by cAMP does not require stimulatory levels of glucose or insulin secretion. We also show that CREB directly targets the Cck promoter in islets from obese (Leptinob/ob) mice. Finally, we demonstrate that the ability of GLP-1 to protect β-cells from cytokine-induced apoptosis is partially dependent on CCK receptor signaling. Taken together, our work suggests that in obesity, active GLP-1 produced in the islet stimulates CCK production and secretion in a paracrine manner via cAMP and CREB. This intraislet incretin loop may be one mechanism whereby GLP-1 protects β-cells from apoptosis. PMID:25984632

Co-Operation Between FADD and Bin1 in Prostate Cancer Apoptosis

DTIC Science & Technology

2005-04-01

an early step in prostate (and also breast ) cancer development, it may represent an early link between cell growth 9 DAMD17-03-1-0049 PI Andrew M...apoptosis is combined. TRAIL-induced autophagy occurs prostate cancer research programs grants DAMD17-02-1-0612 and DAMD17- during breast epithelial cell...Hahn, W. C., and Weinberg, R. A. (2001). Human breast are involved in mammalian cell immortalization, including cancer cells generated by oncogenic

LPS-induced inflammatory response triggers cell cycle reactivation in murine neuronal cells through retinoblastoma proteins induction.

PubMed

D'Angelo, Barbara; Astarita, Carlo; Boffo, Silvia; Massaro-Giordano, Mina; Antonella Ianuzzi, Carmelina; Caporaso, Antonella; Macaluso, Marcella; Giordano, Antonio

2017-01-01

Cell cycle reactivation in adult neurons is an early hallmark of neurodegeneration. The lipopolysaccharide (LPS) is a well-known pro-inflammatory factor that provokes neuronal cell death via glial cells activation. The retinoblastoma (RB) family includes RB1/p105, retinoblastoma-like 1 (RBL1/p107), and retinoblastoma-like 2 (Rb2/p130). Several studies have indicated that RB proteins exhibit tumor suppressor activities, and play a central role in cell cycle regulation. In this study, we assessed LPS-mediated inflammatory effect on cell cycle reactivation and apoptosis of neuronally differentiated cells. Also, we investigated whether the LPS-mediated inflammatory response can influence the function and expression of RB proteins. Our results showed that LPS challenges triggered cell cycle reactivation of differentiated neuronal cells, indicated by an accumulation of cells in S and G2/M phase. Furthermore, we found that LPS treatment also induced apoptotic death of neurons. Interestingly, we observed that LPS-mediated inflammatory effect on cell cycle re-entry and apoptosis was concomitant with the aberrant expression of RBL1/p107 and RB1/p105. To the best of our knowledge, our study is the first to indicate a role of LPS in inducing cell cycle re-entry and/or apoptosis of differentiated neuronal cells, perhaps through mechanisms altering the expression of specific members of RB family proteins. This study provides novel information on the biology of post-mitotic neurons and could help in identifying novel therapeutic targets to prevent de novo cell cycle reactivation and/or apoptosis of neurons undergoing neurodegenerative processes.

Physician Education: Apoptosis.

PubMed

Kataoka; Tsuruo

1996-01-01

We have come to understand apoptosis as not merely a single form of cell death, but as a fundamental theme in cell biology that has far-reaching implications in the fields of physiology and pathology. At the present time, however, the mechanism of apoptosis is not clearly understood, as research into apoptosis is still at the initial stages. Nevertheless, the links between apoptosis and a variety of pathological conditions are gradually becoming clearer. In this article, we will provide a simple explanation of apoptosis and its mechanism as a novel concept of cell death and discuss the way in which apoptosis has been linked to a variety of pathological conditions. WHAT IS APOPTOSIS?: In normal tissue, cells that are no longer needed are rapidly eliminated without affecting the overall function of the tissue. In this process cells undergo an active and spontaneous suicide called programmed cell death. In fact, the majority of physiological cell deaths take the form of apoptosis. The word apoptosis is used, in contrast to necrosis, to describe the situation in which a cell actively pursues a course toward death upon receiving certain stimuli [1]. The morphological changes of apoptosis found in most cell types first involve contraction in cell volume and condensation of the nucleus. When this happens the intracellular organelles such as the mitochondria retain their normal morphology. As apoptosis proceeds, blebbing of the plasma membrane occurs, and the nucleus becomes fragmented. Finally, the cell itself fragments to form apoptotic bodies that are engulfed by nearby phagocytes. With respect to biochemical changes, it is known that the chromosomes become fragmented into nucleosome units, and DNA forms characteristic ladder patterns when subjected to agarose gel electrophoresis. MECHANISM OF APOPTOSIS: It has been reported that apoptosis is induced in various cells by many kinds of irritations, but the precise mechanism is still unclear. Cell injuries that induce apoptosis include those that cause DNA damage such as radiation and anticancer drugs, those that are mediated by the TNF receptor and Fas receptor (the so-called "death signal receptors"), and the deprivation of cytokines that supply survival signals such as IL-3 and erythropoietin. The tumor suppressor gene p53 plays a very important role in apoptosis induced by damage to DNA. This has been demonstrated by studying resistance to apoptosis of cells derived from p53 knockout mice [2]. Other than the irritations that induce apoptosis, molecules that have been strongly implicated as major players in the drama of apoptosis include the Bcl-2 family proteins and the IL-1 converting enzyme (ICE) and its homolog proteases (caspase family). Both groups of proteins show homology with proteins that affect cell death in nematodes. It is believed that molecules that contribute to cell death have been well conserved in multicellular organisms all the way from the relatively primitive nematodes to mammals including humans. It was discovered that Bcl-2 suppressed apoptosis induced in IL-3 dependent cells by deprivation of IL-3 [3]. It has since become the gene around which apoptosis research revolves. Recently, it has become clear that cell death involving the Bcl-2 protein is under the control of similar proteins from the same family [4]. It is interesting that the phenomenon of cell death may be regulated by the balance of the molecules involved in it. APOPTOSIS ABNORMALITIES AND DISEASE: Physiological cell death plays a major role in the growth and permanent maintenance of the human body [5]. In the process of forming the nervous system, neurons that do not form proper connections die. Physiological cell death also accompanies the removal of virus-infected cells by cytotoxic T cells, the elimination of autoreactive immune cells, the formation of the gut, the reconstitution of cartilage and bone, etc. When physiological cell death that normally should occur is inhibited, inappropriate physiological cell death may occur that is harmful to the body and forms the basis of disease. For example, in patients with neural degenerative disorders such as Alzheimer's disease and Parkinson's disease, we can find premature cell death in a particular subset of neurons. The death of T cells in AIDS patients is also a form of physiological cell death. Inhibition of cell death in the immune system enables the survival of autoreactive B cells and T cells, and is therefore a cause of autoimmune disorders. Apoptosis has been particularly linked to cancer. Normal cells are programmed for death if they are subjected to many types of non-physiological stress such as anticancer drugs or radiation, if they become isolated from surrounding cells and are unable to receive their tissue-specific survival signals [6], or if oncogenes are expressed haphazardly [7]. On the other hand, it is believed that the ability to survive is enhanced in transformed cancer cells because they are more resistant to apoptosis, they exhibit resistance to anticancer drugs, they are no longer dependent on survival signals, and they can metastasize. Therefore, the cancer progresses as the cancer cells maintain the proliferative superiority they acquire from their oncogenes. In other words, when cancer cells become resistant to apoptosis, they become resistant to treatment, metastasize, and proliferate destructively. The concept that the malignancy of cancer is due to its resistance to apoptosis is a relatively new one and is worthy of further study.

Endoplasmic reticulum stress signaling is involved in mitomycin C (MMC)-induced apoptosis in human fibroblasts via PERK pathway.

PubMed

Shi, Kun; Wang, Daode; Cao, Xiaojian; Ge, Yingbin

2013-01-01

Endoplasmic reticulum (ER) stress-mediated cell apoptosis has been implicated in various cell types, including fibroblasts. Previous studies have shown that mitomycin C (MMC)-induced apoptosis occurs in fibroblasts, but the effects of MMC on ER stress-mediated apoptosis in fibroblasts have not been examined. Here, MMC-induced apoptosis in human primary fibroblasts was investigated by exposing cells to a single dose of MMC for 5 minutes. Significant inhibition of cell proliferation and increased apoptosis were observed using a cell viability assay, Annexin V/propidium iodide double staining, cell cycle analysis, and TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labeling) staining. Upregulation of proapoptotic factors, including cleaved caspase-3 and poly ADP-ribose polymerase (PARP), was detected by Western blotting. MMC-induced apoptosis was correlated with elevation of 78-kDa glucose-regulated protein (GRP78) and C/EBP homologous protein (CHOP), which are hallmarks of ER stress. Three unfolded protein response (UPR) sensors (inositol-requiring enzyme 1, IRE1; activating transcription factor 6, ATF6; and PKR-like ER kinase, PERK) and their downstream signaling pathways were also activated. Knockdown of CHOP attenuated MMC-induced apoptosis by increasing the ratio of BCL-2/BAX and decreasing BIM expression, suggesting that ER stress is involved in MMC-induced fibroblast apoptosis. Interestingly, knockdown of PERK significantly decreased ER stress-mediated apoptosis by reducing the expression of CHOP, BIM and cleaved caspase-3. Reactive oxygen species (ROS) scavenging also decreased the expression of GRP78, phospho-PERK, CHOP, and BIM. These results demonstrate that MMC-induced apoptosis is triggered by ROS generation and PERK activation.

Endoplasmic Reticulum Stress Signaling Is Involved in Mitomycin C(MMC)-Induced Apoptosis in Human Fibroblasts via PERK Pathway

PubMed Central

Cao, Xiaojian; Ge, Yingbin

2013-01-01

Endoplasmic reticulum (ER) stress-mediated cell apoptosis has been implicated in various cell types, including fibroblasts. Previous studies have shown that mitomycin C (MMC)-induced apoptosis occurs in fibroblasts, but the effects of MMC on ER stress-mediated apoptosis in fibroblasts have not been examined. Here, MMC-induced apoptosis in human primary fibroblasts was investigated by exposing cells to a single dose of MMC for 5 minutes. Significant inhibition of cell proliferation and increased apoptosis were observed using a cell viability assay, Annexin V/propidium iodide double staining, cell cycle analysis, and TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labeling) staining. Upregulation of proapoptotic factors, including cleaved caspase-3 and poly ADP-ribose polymerase (PARP), was detected by Western blotting. MMC-induced apoptosis was correlated with elevation of 78-kDa glucose-regulated protein (GRP78) and C/EBP homologous protein (CHOP), which are hallmarks of ER stress. Three unfolded protein response (UPR) sensors (inositol-requiring enzyme 1, IRE1; activating transcription factor 6, ATF6; and PKR-like ER kinase, PERK) and their downstream signaling pathways were also activated. Knockdown of CHOP attenuated MMC-induced apoptosis by increasing the ratio of BCL-2/BAX and decreasing BIM expression, suggesting that ER stress is involved in MMC-induced fibroblast apoptosis. Interestingly, knockdown of PERK significantly decreased ER stress-mediated apoptosis by reducing the expression of CHOP, BIM and cleaved caspase-3. Reactive oxygen species (ROS) scavenging also decreased the expression of GRP78, phospho-PERK, CHOP, and BIM. These results demonstrate that MMC-induced apoptosis is triggered by ROS generation and PERK activation. PMID:23533616

TGF-beta-induced apoptosis in human thyrocytes is mediated by p27kip1 reduction and is overridden in neoplastic thyrocytes by NF-kappaB activation.

PubMed

Bravo, Susana B; Pampín, Sandra; Cameselle-Teijeiro, José; Carneiro, Carmen; Domínguez, Fernando; Barreiro, Francisco; Alvarez, Clara V

2003-10-30

Millions of people worldwide suffer goiter, a proliferative disease of the follicular cells of the thyroid that may become neoplastic. Thyroid neoplasms have low proliferative index, low apoptotic index and a high incidence of metastasis. TGF-beta is overexpressed in thyroid follicular tumor cells. To investigate the role of TGF-beta in thyroid tumor progression, we established cultures of human thyrocytes from different proliferative pathologies (Grave's disease, multinodular goiter, follicular adenoma, papillary carcinoma), lymph node metastasis, and a normal thyroid sample. All cultures maintained the thyrocyte phenotype. TGF-beta induced cell-cycle arrest in all cultures, in contrast with results reported for other epithelial tumors. In deprived medium, TGF-beta induced apoptosis in normal thyrocyte cultures and all neoplastic cultures except the metastatic cultures. This apoptosis was mediated by a reduction in p27kip1 levels, inducing cell-cycle initiation. Antisense p27 expression induced apoptosis in the absence of TGF-beta. By contrast, in cells in which p27 was overexpressed, TGF-beta had a survival effect. In growth medium, a net survival effect occurs in neoplastic thyrocytes only, not normal thyrocytes, due to activation of the NF-kappaB survival program. Together, these findings suggest that (a) thyroid neoplasms are due to reduced apoptosis, not increased division, in line with the low proliferative index of these pathologies, and (b) TGF-beta induces apoptosis in normal thyrocytes via p27 reduction, but that in neoplastic thyrocytes this effect is overridden by activation of the NF-kappaB program.

Targeting colorectal cancer cells by a novel sphingosine kinase 1 inhibitor PF-543

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ju, TongFa; Gao, DaQuan; Fang, Zheng-yu, E-mail: fangzhengyu158@sina.com

In this study, we showed that PF-543, a novel sphingosine kinase 1 (SphK1) inhibitor, exerted potent anti-proliferative and cytotoxic effects against a panel of established (HCT-116, HT-29 and DLD-1) and primary human colorectal cancer (CRC) cells. Its sensitivity was negatively associated with SphK1 expression level in the CRC cells. Surprisingly, PF-543 mainly induced programmed necrosis, but not apoptosis, in the CRC cells. CRC cell necrotic death was detected by lactate dehydrogenase (LDH) release, mitochondrial membrane potential (MMP) collapse and mitochondrial P53-cyclophilin-D (Cyp-D) complexation. Correspondingly, the necrosis inhibitor necrostatin-1 largely attenuated PF-543-induced cytotoxicity against CRC cells. Meanwhile, the Cyp-D inhibitors (sanglifehrinmore » A and cyclosporin A), or shRNA-mediated knockdown of Cyp-D, remarkably alleviated PF-543-induced CRC cell necrotic death. Reversely, over-expression of wild-type Cyp-D in HCT-116 cells significantly increased PF-543's sensitivity. In vivo, PF-543 intravenous injection significantly suppressed HCT-116 xenograft growth in severe combined immunodeficient (SCID) mice, whiling remarkably improving the mice survival. The in vivo activity by PF-543 was largely attenuated when combined with the Cyp-D inhibitor cyclosporin A. Collectively, our results demonstrate that PF-543 exerts potent anti-CRC activity in vitro and in vivo. Mitochondrial programmed necrosis pathway is likely the key mechanism responsible for PF-543's actions in CRC cells. - Highlights: • PF-543 is anti-proliferative and cytotoxic to established and primary CRC cells. • PF-543 induces programmed necrosis, but not apoptosis, in CRC cells. • Modulation of mitochondrial protein cyclophilin-D alters PF-543's sensitivity. • PF-543 inhibits HCT-116 xenograft growth in SCID mice, improving mice survival. • Co-administration of cyclophilin-D inhibitor CsA inhibits PF-543's activity in vivo.« less

Wavelength-dependent backscattering measurements for quantitative real-time monitoring of apoptosis in living cells

NASA Astrophysics Data System (ADS)

Mulvey, Christine S.; Sherwood, Carly A.; Bigio, Irving J.

2009-11-01

Apoptosis-programmed cell death-is a cellular process exhibiting distinct biochemical and morphological changes. An understanding of the early morphological changes that a cell undergoes during apoptosis can provide the opportunity to monitor apoptosis in tissue, yielding diagnostic and prognostic information. There is avid interest regarding the involvement of apoptosis in cancer. The initial response of a tumor to successful cancer treatment is often massive apoptosis. Current apoptosis detection methods require cell culture disruption. Our aim is to develop a nondisruptive optical method to monitor apoptosis in living cells and tissues. This would allow for real-time evaluation of apoptotic progression of the same cell culture over time without alteration. Elastic scattering spectroscopy (ESS) is used to monitor changes in light-scattering properties of cells in vitro due to apoptotic morphology changes. We develop a simple instrument capable of wavelength-resolved ESS measurements from cell cultures in the backward direction. Using Mie theory, we also develop an algorithm that extracts the size distribution of scatterers in the sample. The instrument and algorithm are validated with microsphere suspensions. For cell studies, Chinese hamster ovary (CHO) cells are cultured to confluence on plates and are rendered apoptotic with staurosporine. Backscattering measurements are performed on pairs of treated and control samples at a sequence of times up to 6-h post-treatment. Initial results indicate that ESS is capable of discriminating between treated and control samples as early as 10- to 15-min post-treatment, much earlier than is sensed by standard assays for apoptosis. Extracted size distributions from treated and control samples show a decrease in Rayleigh and 150-nm scatterers, relative to control samples, with a corresponding increase in 200-nm particles. Work continues to correlate these size distributions with underlying morphology. To our knowledge, this is the first report of the use of backscattering spectral measurements to quantitatively monitor apoptosis in viable cell cultures in vitro.

A self-defeating anabolic program leads to β-cell apoptosis in endoplasmic reticulum stress-induced diabetes via regulation of amino acid flux.

PubMed

Krokowski, Dawid; Han, Jaeseok; Saikia, Mridusmita; Majumder, Mithu; Yuan, Celvie L; Guan, Bo-Jhih; Bevilacqua, Elena; Bussolati, Ovidio; Bröer, Stefan; Arvan, Peter; Tchórzewski, Marek; Snider, Martin D; Puchowicz, Michelle; Croniger, Colleen M; Kimball, Scot R; Pan, Tao; Koromilas, Antonis E; Kaufman, Randal J; Hatzoglou, Maria

2013-06-14

Endoplasmic reticulum (ER) stress-induced responses are associated with the loss of insulin-producing β-cells in type 2 diabetes mellitus. β-Cell survival during ER stress is believed to depend on decreased protein synthesis rates that are mediated via phosphorylation of the translation initiation factor eIF2α. It is reported here that chronic ER stress correlated with increased islet protein synthesis and apoptosis in β-cells in vivo. Paradoxically, chronic ER stress in β-cells induced an anabolic transcription program to overcome translational repression by eIF2α phosphorylation. This program included expression of amino acid transporter and aminoacyl-tRNA synthetase genes downstream of the stress-induced ATF4-mediated transcription program. The anabolic response was associated with increased amino acid flux and charging of tRNAs for branched chain and aromatic amino acids (e.g. leucine and tryptophan), the levels of which are early serum indicators of diabetes. We conclude that regulation of amino acid transport in β-cells during ER stress involves responses leading to increased protein synthesis, which can be protective during acute stress but can lead to apoptosis during chronic stress. These studies suggest that the increased expression of amino acid transporters in islets can serve as early diagnostic biomarkers for the development of diabetes.

A Self-defeating Anabolic Program Leads to β-Cell Apoptosis in Endoplasmic Reticulum Stress-induced Diabetes via Regulation of Amino Acid Flux*

PubMed Central

Krokowski, Dawid; Han, Jaeseok; Saikia, Mridusmita; Majumder, Mithu; Yuan, Celvie L.; Guan, Bo-Jhih; Bevilacqua, Elena; Bussolati, Ovidio; Bröer, Stefan; Arvan, Peter; Tchórzewski, Marek; Snider, Martin D.; Puchowicz, Michelle; Croniger, Colleen M.; Kimball, Scot R.; Pan, Tao; Koromilas, Antonis E.; Kaufman, Randal J.; Hatzoglou, Maria

2013-01-01

Endoplasmic reticulum (ER) stress-induced responses are associated with the loss of insulin-producing β-cells in type 2 diabetes mellitus. β-Cell survival during ER stress is believed to depend on decreased protein synthesis rates that are mediated via phosphorylation of the translation initiation factor eIF2α. It is reported here that chronic ER stress correlated with increased islet protein synthesis and apoptosis in β-cells in vivo. Paradoxically, chronic ER stress in β-cells induced an anabolic transcription program to overcome translational repression by eIF2α phosphorylation. This program included expression of amino acid transporter and aminoacyl-tRNA synthetase genes downstream of the stress-induced ATF4-mediated transcription program. The anabolic response was associated with increased amino acid flux and charging of tRNAs for branched chain and aromatic amino acids (e.g. leucine and tryptophan), the levels of which are early serum indicators of diabetes. We conclude that regulation of amino acid transport in β-cells during ER stress involves responses leading to increased protein synthesis, which can be protective during acute stress but can lead to apoptosis during chronic stress. These studies suggest that the increased expression of amino acid transporters in islets can serve as early diagnostic biomarkers for the development of diabetes. PMID:23645676

Single-cell-precision microplasma-induced cancer cell apoptosis.

PubMed

Tan, Xiao; Zhao, Shasha; Lei, Qian; Lu, Xinpei; He, Guangyuan; Ostrikov, Kostya

2014-01-01

The issue of single-cell control has recently attracted enormous interest. However, in spite of the presently achievable intracellular-level physiological probing through bio-photonics, nano-probe-based, and some other techniques, the issue of inducing selective, single-cell-precision apoptosis, without affecting neighbouring cells remains essentially open. Here we resolve this issue and report on the effective single-cell-precision cancer cell treatment using the reactive chemistry of the localized corona-type plasma discharge around a needle-like electrode with the spot size ∼1 µm. When the electrode is positioned with the micrometer precision against a selected cell, a focused and highly-localized micro-plasma discharge induces apoptosis in the selected individual HepG2 and HeLa cancer cells only, without affecting any surrounding cells, even in small cell clusters. This is confirmed by the real-time monitoring of the morphological and structural changes at the cellular and cell nucleus levels after the plasma exposure.

The Life and Death of a Plant Cell.

PubMed

Kabbage, Mehdi; Kessens, Ryan; Bartholomay, Lyric C; Williams, Brett

2017-04-28

Like all eukaryotic organisms, plants possess an innate program for controlled cellular demise termed programmed cell death (PCD). Despite the functional conservation of PCD across broad evolutionary distances, an understanding of the molecular machinery underpinning this fundamental program in plants remains largely elusive. As in mammalian PCD, the regulation of plant PCD is critical to development, homeostasis, and proper responses to stress. Evidence is emerging that autophagy is key to the regulation of PCD in plants and that it can dictate the outcomes of PCD execution under various scenarios. Here, we provide a broad and comparative overview of PCD processes in plants, with an emphasis on stress-induced PCD. We also discuss the implications of the paradox that is functional conservation of apoptotic hallmarks in plants in the absence of core mammalian apoptosis regulators, what that means, and whether an equivalent form of death occurs in plants.

Chromatin status of apoptosis genes correlates with sensitivity to chemo-, immune- and radiation therapy in colorectal cancer cell lines.

PubMed

Benard, Anne; Janssen, Connie M; van den Elsen, Peter J; van Eggermond, Marja C J A; Hoon, Dave S B; van de Velde, Cornelis J H; Kuppen, Peter J K

2014-12-01

The apoptosis pathway of programmed cell death is frequently deregulated in cancer. An intact apoptosis pathway is required for proper response to anti-cancer treatment. We investigated the chromatin status of key apoptosis genes in the apoptosis pathway in colorectal cancer cell lines in relation to apoptosis induced by chemo-, immune- or radiation therapy. Using chromatin immunoprecipitation (ChIP), we measured the presence of transcription-activating histone modifications H3Ac and H3K4me3 and silencing modifications H3K9me3 and H3K27me3 at the gene promoter regions of key apoptosis genes Bax, Bcl2, Caspase-9, Fas (CD95) and p53. Cell lines DLD1, SW620, Colo320, Caco2, Lovo and HT29 were treated with cisplatin, anti-Fas or radiation. The apoptotic response was measured by flow cytometry using propidium iodide and annexin V-FITC. The chromatin status of the apoptosis genes reflected the activation status of the intrinsic (Bax, Bcl2, Caspase-9 and p53) and extrinsic (Fas) pathways. An active intrinsic apoptotic pathway corresponded to sensitivity to cisplatin and radiation treatment of cell lines DLD1, SW620 and Colo320. An active Fas promoter corresponded to an active extrinsic apoptotic pathway in cell line DLD1. mRNA expression data correlated with the chromatin status of the apoptosis genes as measured by ChIP. In conclusion, the results presented in this study indicate that the balance between activating and silencing histone modifications, reflecting the chromatin status of apoptosis genes, can be used to predict the response of tumor cells to different anti-cancer therapies and could provide a novel target to sensitize tumors to obtain adequate treatment responses.

Michelob_x is the missing inhibitor of apoptosis protein antagonist in mosquito genomes

PubMed Central

Zhou, Lei; Jiang, Guohua; Chan, Gina; Santos, Carl P; Severson, David W; Xiao, Lei

2005-01-01

Apoptosis is implicated in the life cycle of the malaria parasite in mosquitoes. The genome project for the primary malaria vector Anopheles gambiae showed a significant expansion of the inhibitor of apoptosis protein (IAP) and caspase gene families in comparison with Drosophila. However, because of extensive sequence divergence, no orthologue was identified for the reaper/grim-like IAP antagonist genes that have a pivotal role in cell death regulation in Drosophila. Using a customized searching strategy, we identified michelob_x(mx), a gene not predicted by the genome project, as the missing IAP antagonist in the An. gambiae and other mosquito genomes. Mx has a highly conserved amino-terminal IAP-binding motif. Expression of Mx induces rapid cell death in insect cell lines and is a potent tissue ablator in vivo. Its proapoptotic activity is totally dependent on the IAP-binding motif. Like reaper in Drosophila, mx is transcriptionally induced by ultraviolet irradiation to mediate cell death. PMID:16041319

Visualization of proteolytic activity associated with the apoptotic response in cancer cells

NASA Astrophysics Data System (ADS)

Tice, Brian George

Caspases execute programmed cell death, where low levels of caspase activity are linked to cancer. Chemotherapies utilize induction of apoptosis as a key mechanism for cancer treatment, where caspase-3 is a major player involved in dismantling these aberrant cells. The ability to sensitively measure the initial caspase-3 cleavage events during apoptosis is important for understanding the initiation of this complex cellular process, however, current ensemble methods are not sensitive enough to measure single cleavage events in cells. By utilizing the optical properties of plasmon coupling, peptide-linked gold nanoparticles were developed to enable single molecule imaging of caspase-3 activity in two different cancer systems. Au crown nanoparticles were assembled in a multimeric fashion to overcome the high and heterogeneous background scattering of live cells. In a colon cancer (SW620) cell line challenged with tumor necrosis factor-alpha (TNF-alpha), single molecule trajectories show early stage caspase-3 activation within minutes, which was not detectable by ensemble assays until 23 hours. Variability in caspase-3 activation among the population of cells was identified and likely a result of each cell's specific resistance to death receptor-induced apoptosis. Following these studies, improvements by way of sensitivity and selectivity were tailored into an improved nanosensor construct. Au nanoshell dimers were prepared as a comparably bright construct with 1) reduced heterogeneity compared to the synthesis of the crown nanoparticles and 2) a peptide sequence highly selective for caspase-3. Chronic myeloid leukemia (CML) K562 cells were assessed for their early apoptotic response upon treatment with dasatinib, a clinically approved tyrosine kinase inhibitor that specifically targets BCR-ABL. It has been demonstrated that inhibition of BCR-ABL by dasatinib commits K562 cells to apoptosis. Single molecule experiments with Au nanoshell dimers show caspase-3 activation as early as 8 hours than previously reported. This suggests an early commitment to apoptosis that precedes the competing fate of growth factor mediated survival in CML patient-derived BCR-ABL cells. These nanosensors are sensitive and selective in observing caspase-3 activation compared to ensemble methods; and allow the possibility to detect caspase-3 activity for use as a drug screening or diagnostic tool for personalized care in the treatment of cancer.

Pro-apoptotic signaling induced by Retinoic acid and dsRNA is under the control of Interferon Regulatory Factor-3 in breast cancer cells.

PubMed

Bernardo, Ana R; Cosgaya, José M; Aranda, Ana; Jiménez-Lara, Ana M

2017-07-01

Breast cancer is one of the most lethal malignancies for women. Retinoic acid (RA) and double-stranded RNA (dsRNA) are considered signaling molecules with potential anticancer activity. RA, co-administered with the dsRNA mimic polyinosinic-polycytidylic acid (poly(I:C)), synergizes to induce a TRAIL (Tumor-Necrosis-Factor Related Apoptosis-Inducing Ligand)- dependent apoptotic program in breast cancer cells. Here, we report that RA/poly(I:C) co-treatment, synergically, induce the activation of Interferon Regulatory Factor-3 (IRF3) in breast cancer cells. IRF3 activation is mediated by a member of the pathogen recognition receptors, Toll-like receptor-3 (TLR3), since its depletion abrogates IRF3 activation by RA/poly(I:C) co-treatment. Besides induction of TRAIL, apoptosis induced by RA/poly(I:C) correlates with the increased expression of pro-apoptotic TRAIL receptors, TRAIL-R1/2, and the inhibition of the antagonistic receptors TRAIL-R3/4. IRF3 plays an important role in RA/poly(I:C)-induced apoptosis since IRF3 depletion suppresses caspase-8 and caspase-3 activation, TRAIL expression upregulation and apoptosis. Interestingly, RA/poly(I:C) combination synergizes to induce a bioactive autocrine/paracrine loop of type-I Interferons (IFNs) which is ultimately responsible for TRAIL and TRAIL-R1/2 expression upregulation, while inhibition of TRAIL-R3/4 expression is type-I IFN-independent. Our results highlight the importance of IRF3 and type-I IFNs signaling for the pro-apoptotic effects induced by RA and synthetic dsRNA in breast cancer cells.

Mechanical and hypoxia stress can cause chondrocytes apoptosis through over-activation of endoplasmic reticulum stress.

PubMed

Huang, Ziwei; Zhou, Min; Wang, Qian; Zhu, Mengjiao; Chen, Sheng; Li, Huang

2017-12-01

To examine the role of mechanical force and hypoxia on chondrocytes apoptosis and osteoarthritis (OA)-liked pathological change on mandibular cartilage through over-activation of endoplasmic reticulum stress (ERS). We used two in vitro models to examine the effect of mechanical force and hypoxia on chondrocytes apoptosis separately. The mandibular condylar chondrocytes were obtained from three-week-old male Sprague-Dawley rats. Flexcell 5000T apparatus was used to produce mechanical forces (12%, 0.5Hz, 24h vs 20%, 0.5Hz, 24h) on chondrocytes. For hypoxia experiment, the concentration of O 2 was down regulated to 5% or 1%. Cell apoptosis rates were quantified by annexin V and propidium iodide (PI) double staining and FACS analysis. Quantitative real-time PCR and western blot were performed to evaluate the activation of ERS and cellular hypoxia. Then we used a mechanical stress loading rat model to verify the involvement of ERS in OA-liked mandibular cartilage pathological change. Histological changes in mandibular condylar cartilage were assessed via hematoxylin & eosin (HE) staining. Immunohistochemistry of GRP78, GRP94, HIF-1α, and HIF-2α were performed to evaluate activation of the ERS and existence of hypoxia. Apoptotic cells were detected by the TUNEL method. Tunicamycin, 20% mechanical forces and hypoxia (1% O 2 ) all significantly increased chondrocytes apoptosis rates and expression of ERS markers (GRP78, GRP94 and Caspase 12). However, 12% mechanical forces can only increase the apoptotic sensitivity of chondrocytes. Mechanical stress resulted in OA-liked pathological change on rat mandibular condylar cartilage which included thinning cartilage and bone erosion. The number of apoptotic cells increased. ERS and hypoxia markers expressions were also enhanced. Salubrinal, an ERS inhibitor, can reverse these effects in vitro and in vivo through the down-regulation of ERS markers and hypoxia markers. We confirmed that mechanical stress and local hypoxia both contributed to the chondrocytes apoptosis. Mechanical stress can cause OA-like pathological change in rat mandibular condylar cartilage via ERS activation and hypoxia existed in the meantime. Both mechanical forces and hypoxia can induce ERS and cause chondrocytes apoptosis only if the stimulate was in higher level. Salubrinal can protect chondrocytes from apoptosis, and relieve OA-liked pathological change on mandibular condylar cartilage under mechanical stress stimulation. Copyright © 2017. Published by Elsevier Ltd.

Toll-like receptor 7 promotes the apoptosis of THP-1-derived macrophages through the CHOP-dependent pathway.

PubMed

Yu, Xiaochen; Wang, Yang; Zhao, Wenhui; Zhou, Haizhou; Yang, Wei; Guan, Xiuru

2014-09-01

Macrophage apoptosis is a prominent characteristic of advanced atherosclerotic plaques and leads to plaque destabilization. Certain studies have confirmed that influenza virus A (IVA) infection is related to acute myocardial infarction (AMI). However, it remains unknown as to whether this phenomenon is associated with Toll-like receptor (TLR)7, since single-stranded RNA (ssRNA) of IVA is a natural ligand of TLR7. Thus, in the present study, THP-1‑derived macrophages were infected with IVA or treated with imiquimod (IMQ) in the presence or absence of pre-treatment with oxidized low-density lipoprotein (oxLDL). The macrophages were pre-treated with oxLDL (5 µg/ml) for 24 h to mimic high lipid conditions. Cell viability and apoptosis were detected by 3-(4,5-dimethylthiazol-2-y-1)‑2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and flow cytometry, respectively. Our results revealed that TLR7 played an important role in macrophage apoptosis and cytokine secretion. Both IVA infection and IMQ treatment increased TLR7 expression, as well as the secretion of pro-inflammatory cytokines [interleukin (IL)-6, monocyte chemotactic protein (MCP)-1] and apoptosis. However, this increase in cytokine secretion occurred independently of cell apoptosis. oxLDL had potential synergistic pro-apoptotic effects combined with TLR7 activation. To determine whether endoplasmic reticulum (ER) stress plays a role in cell apoptosis, the mRNA and protein expression of known markers of ER stress [glucose-regulated protein (GRP)78 and C/EBP homologous protein (CHOP)] was detected by reverse transcription PCR (RT-PCR), quantitative reverse transcription PCR (qRT-PCR) and western blot analysis. Our results revealed that apoptosis aggravated ER stress, as shown by the overexpression of the pro-apoptotic sensor, CHOP. In conclusion, our study demonstrates the converging role of oxLDL pre-treatment, IVA infection and IMQ in ER stress-induced cell apoptosis.

beta-Adrenoceptor blockers protect against staurosporine-induced apoptosis in SH-SY5Y neuroblastoma cells.

PubMed

Mikami, Maya; Goubaeva, Farida; Song, Joseph H; Lee, H T; Yang, Jay

2008-07-28

The beta-adrenoceptor blockers exhibit a well-characterized anti-apoptotic property in the heart and kidney while less is known about the effect of this class of drugs on neuronal apoptosis. We studied the effects of three beta-adrenoceptor blockers propranolol (1-(isoproplyamino)-3-(naphthalene-1-yloxy)propan-2-ol), atenolol (2-[4-[2-hydroxy-3-(1-methylethylamino)propoxyl]phenyl]ehanamide), and ICI 118551 (1-[2,3-(dihydro-7-methyl-1H-iden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol), against staurosporine-induced apoptosis in SH-SY5Y human neuroblastoma cells. Staurosporine increased caspase 3-like activity, DNA fragmentation, PARP cleavage, and the number of TUNEL positive cells consistent with the induction of apoptosis. Propranolol and ICI 118551, but not atenolol, demonstrated a concentration-dependent inhibition of caspase 3-like activity. Propranolol and ICI 118551 directly inhibited the enzymatic activity of recombinant caspase 9 while atenolol did not; however, none of the beta-adrenoceptor blockers that were examined directly blocked caspases 2 or 3 activity. In isolated mitochondria, propranolol and ICI 118551 inhibited staurosporine-induced cytochrome c release while atenolol did not. We conclude that propranolol and ICI 118551 protect SH-SY5Y cells against staurosporine-induced apoptosis through a dual action on the mitochondria and on caspase 9 in a cell type and an apoptotic paradigm where the conventional inhibitors of mitochondrial permeability transition such as cyclosporin A and bongkrekic acid demonstrate no protection.

Two distinct phases of apoptosis in mammary gland involution: proteinase-independent and -dependent pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lund, Leif R; Romer, John; Thomasset, Nicole

Postlactational involution of the mammary gland is characterized by two distinct physiological events: apoptosis of the secretory, epithelial cells undergoing programmed cell death, and proteolytic degradation of the mammary gland basement membrane. We examined the spatial and temporal patterns of apoptotic cells in relation to those of proteinases during involution of the BALB/c mouse mammary gland. Apoptosis was almost absent during lactation but became evident at day 2 of involution, when {beta}-casein gene expression was still high. Apoptotic cells were then seen at least up to day 8 of involution, when {beta}-casein gene expression was being extinguished. Expression of sulfatedmore » glycoprotein-2 (SGP-2), interleukin-1{beta} converting enzyme (ICE) and tissue inhibitor of metalloproteinases-1 was upregulated at day 2, when apoptotic cells were seen initially. Expression of the matrix metalloproteinases gelatinase A and stromelysin-1 and the serine proteinase urokinase-type plasminogen activator, which was low during lactation, was strongly upregulated in parallel starting at day 4 after weaning, coinciding with start of the collapse of the lobulo-alveolar structures and the intensive tissue remodeling in involution. The major sites of mRNA synthesis for these proteinases were fibroblast-like cells in the periductal stroma and stromal cells surrounding the collapsed alveoli, suggesting that the degradative phase of involution is due to a specialized mesenchymal-epithelial interaction. To elucidate the functional role of these proteinases during involution, at the onset of weaning we treated mice systemically with the glucocorticoid hydrocortisone, which is known to inhibit mammary gland involution. Although the initial wave of apoptotic cells appeared in the lumina of the gland, the dramatic regression and tissue remodeling usually evident by day 5 was substantially inhibited by systemic treatment with hydrocortisone. mRNA and protein for gelatinase A, stromelysin-1 and uPA were weakly induced, if at all, in hydrocortisonetreated mice. Furthermore, mRNA for membrane-type matrix metalloproteinase decreased after hydrocortisone treatment and paralleled the almost complete inhibition of activation of latent gelatinase A. Concomitantly, the gland filled with an overabundance of milk. Our data support the hypothesis that there are at least two distinct phases of involution: an initial phase, characterized by induction of the apoptosis-associated genes SGP-2 and ICE and apoptosis of fully differentiated mammary epithelial cells without visible degradation of the extracellular matrix, and a second phase, characterized by extracellular matrix remodeling and altered mesenchymal-epithelial interactions, followed by apoptosis of cells that are losing differentiated functions.« less

Cancer dormancy and cell signaling: Induction of p21waf1 initiated by membrane IgM engagement increases survival of B lymphoma cells

PubMed Central

Marches, Radu; Hsueh, Robert; Uhr, Jonathan W.

1999-01-01

The p21WAF1 (p21) cyclin-dependent kinase inhibitor plays a major role in regulating cell cycle arrest. It was recently reported that the p53-independent elevation of p21 protein levels is essential in mediating the G1 arrest resulting from signal transduction events initiated by the crosslinking of membrane IgM on Daudi Burkitt lymphoma cells. Although the role of p21 in cell cycle regulation is well documented, there is little information concerning its role in antibody-mediated apoptosis. In the present study, we examined the involvement of p21 in the regulation of apoptosis by suppressing its induction in anti-IgM-treated Daudi cells through a p21 antisense expression construct approach. Reduction in induced p21 protein levels resulted in diminished G1 arrest and increased apoptosis. The increased susceptibility to anti-IgM-mediated apoptosis was associated with increased caspase-3-like activity and poly-(ADP)ribose polymerase cleavage. These data suggest that p21 may directly interfere with the caspase cascade, thus playing a dual role in regulating both cell cycle progression and apoptosis. PMID:10411940

Effects of FR235222, a novel HDAC inhibitor, in proliferation and apoptosis of human leukaemia cell lines: role of annexin A1.

PubMed

Petrella, Antonello; D'Acunto, Cosimo Walter; Rodriquez, Manuela; Festa, Michela; Tosco, Alessandra; Bruno, Ines; Terracciano, Stefania; Taddei, Maurizio; Paloma, Luigi Gomez; Parente, Luca

2008-03-01

FR235222, a novel histone deacetylase inhibitor (HDACi), at 50nM caused accumulation of acetylated histone H4, inhibition of cell proliferation and G1 cycle arrest accompanied by increase of p21 and down-regulation of cyclin E in human promyelocytic leukaemia U937 cells. The compound was also able to increase the protein and mRNA levels of annexin A1 (ANXA1) without effects on apoptosis. Similar effects were observed in human chronic myelogenous leukaemia K562 cells and human T cell leukaemia Jurkat cells. Cycle arrest and ANXA1 expression, without significant effects on apoptosis, were also induced by different HDACi like suberoylanilide hydroxamic acid (SAHA) and trichostatin-A (TSA). FR235222 at 0.5 microM stimulated apoptosis of all leukaemia cell lines associated to an increased expression of the full-length (37kDa) protein and the appearance of a 33kDa N-terminal cleavage product in both cytosol and membrane. These results suggest that ANXA1 expression may mediate cycle arrest induced by low doses FR235222, whereas apoptosis induced by high doses FR235222 is associated to ANXA1 processing.

5-Fluorouracil-induced apoptosis in cultured oral cancer cells.

PubMed

Tong, D; Poot, M; Hu, D; Oda, D

2000-03-01

Chemotherapy is commonly used to treat advanced oral squamous cell carcinoma (SCC) and is known to kill cancer cells through apoptosis. Our hypothesis states that 5-fluorouracil (5FU) also kills cultured oral epithelial cells through programmed cell death or apoptosis. Cultured oral cancer cells were exposed to an optimum dose of 20 mg/ml of 5FU. Cells were analyzed for changes in cell cycle distribution and induction of cell death including apoptosis. Normal control, human papilloma virus-immortalized (PP), ATCC SCC cell line (CA1) and two primary oral SCC cell lines (CA3 and -4) were studied. Inhibition of apoptosis by a pan-caspase inhibitor was used. SYTO 11 flow cytometry showed increased apoptosis in all 5FU-treated cell cultures compared to untreated controls. The results show biological variation in apoptotic response. CA1 had the lowest apoptotic rate of the cancer cell lines at 1.5%. Next lowest was CA3, followed by CA4 and PP. In addition, alteration in the G1 and S phase fractions were found. Untreated CA1 showed 28% G1, 53% S compared to 43% G1, and 40% S of treated. We investigated the pathway of apoptosis using the pan-caspase inhibitor IDN-1529 by methylthiazolyl diphenyl tetrazolium bromide (MTT) colorimetric analysis. Results showed mild inhibition of cell death when cells were incubated with 50 microM IDN-1529 for 24 h. This suggests a probable caspase-dependent apoptotic pathway. In conclusion, our data suggest that 5FU induces oral cancer cell death through apoptosis and that biological variation exists between normal and cancer cells and between different types of cancer cells themselves. Our data indicate that cultures of a useful in vitro model for chemosensitivity assays are possible. Our results also suggest a caspase-dependent pathway for chemocytotoxicity in oral SCC.

Mechanisms of apoptosis in Crustacea: What conditions induce versus suppress cell death?

PubMed

Menze, Michael A; Fortner, Grady; Nag, Suman; Hand, Steven C

2010-03-01

Arthropoda is the largest of all animal phyla and includes about 90% of extant species. Our knowledge about regulation of apoptosis in this phylum is largely based on findings for the fruit fly Drosophila melanogaster. Recent work with crustaceans shows that apoptotic proteins, and presumably mechanisms of cell death regulation, are more diverse in arthropods than appreciated based solely on the excellent work with fruit flies. Crustacean homologs exist for many major proteins in the apoptotic networks of mammals and D. melanogaster, but integration of these proteins into the physiology and pathophysiology of crustaceans is far from complete. Whether apoptosis in crustaceans is mainly transcriptionally regulated as in D. melanogaster (e.g., RHG 'killer' proteins), or rather is controlled by pro- and anti-apoptotic Bcl-2 family proteins as in vertebrates needs to be clarified. Some phenomena like the calcium-induced opening of the mitochondrial permeability transition pore (MPTP) are apparently lacking in crustaceans and may represent a vertebrate invention. We speculate that differences in regulation of the intrinsic pathway of crustacean apoptosis might represent a prerequisite for some species to survive harsh environmental insults. Pro-apoptotic stimuli described for crustaceans include UV radiation, environmental toxins, and a diatom-produced chemical that promotes apoptosis in offspring of a copepod. Mechanisms that serve to depress apoptosis include the inhibition of caspase activity by high potassium in energetically healthy cells, alterations in nucleotide abundance during energy-limited states like diapause and anoxia, resistance to opening of the calcium-induced MPTP, and viral accommodation during persistent viral infection. Characterization of the players, pathways, and their significance in the core machinery of crustacean apoptosis is revealing new insights for the field of cell death.

Mechanisms of apoptosis in Crustacea: what conditions induce versus suppress cell death?

PubMed Central

Menze, Michael A.; Fortner, Grady; Nag, Suman; Hand, Steven C.

2014-01-01

Arthropoda is the largest of all animal phyla and includes about 90% of extant species. Our knowledge about regulation of apoptosis in this phylum is largely based on findings for the fruit fly Drosophila melanogaster. Recent work with crustaceans shows that apoptotic proteins, and presumably mechanisms of cell death regulation, are more diverse in arthropods than appreciated based solely on the excellent work with fruit flies. Crustacean homologs exist for many major proteins in the apoptotic networks of mammals and D. melanogaster, but integration of these proteins into the physiology and pathophysiology of crustaceans is far from complete. Whether apoptosis in crustaceans is mainly transcriptionally regulated as in D. melanogaster (e.g., RHG ‘killer’ proteins), or rather is controlled by pro- and anti-apoptotic Bcl-2 family proteins as in vertebrates needs to be clarified. Some phenomena like the calcium-induced opening of the mitochondrial permeability transition pore (MPTP) are apparently lacking in crustaceans and may represent a vertebrate invention. We speculate that differences in regulation of the intrinsic pathway of crustacean apoptosis might represent a prerequisite for some species to survive harsh environmental insults. Pro-apoptotic stimuli described for crustaceans include UV radiation, environmental toxins, and a diatom-produced chemical that promotes apoptosis in offspring of a copepod. Mechanisms that serve to depress apoptosis include the inhibition of caspase activity by high potassium in energetically healthy cells, alterations in nucleotide abundance during energy-limited states like diapause and anoxia, resistance to opening of the calcium-induced MPTP, and viral accommodation during persistent viral infection. Characterization of the players, pathways, and their significance in the core machinery of crustacean apoptosis is revealing new insights for the field of cell death. PMID:20043212

ER Stress-Mediated Signaling: Action Potential and Ca(2+) as Key Players.

PubMed

Bahar, Entaz; Kim, Hyongsuk; Yoon, Hyonok

2016-09-15

The proper functioning of the endoplasmic reticulum (ER) is crucial for multiple cellular activities and survival. Disturbances in the normal ER functions lead to the accumulation and aggregation of unfolded proteins, which initiates an adaptive response, the unfolded protein response (UPR), in order to regain normal ER functions. Failure to activate the adaptive response initiates the process of programmed cell death or apoptosis. Apoptosis plays an important role in cell elimination, which is essential for embryogenesis, development, and tissue homeostasis. Impaired apoptosis can lead to the development of various pathological conditions, such as neurodegenerative and autoimmune diseases, cancer, or acquired immune deficiency syndrome (AIDS). Calcium (Ca(2+)) is one of the key regulators of cell survival and it can induce ER stress-mediated apoptosis in response to various conditions. Ca(2+) regulates cell death both at the early and late stages of apoptosis. Severe Ca(2+) dysregulation can promote cell death through apoptosis. Action potential, an electrical signal transmitted along the neurons and muscle fibers, is important for conveying information to, from, and within the brain. Upon the initiation of the action potential, increased levels of cytosolic Ca(2+) (depolarization) lead to the activation of the ER stress response involved in the initiation of apoptosis. In this review, we discuss the involvement of Ca(2+) and action potential in ER stress-mediated apoptosis.

Apoptosis of Alcohol-Exposed Human Placental Cytotrophoblast Cells is Downstream of Intracellular Calcium Signaling

PubMed Central

Bolnick, Jay M.; Karana, Rita; Chiang, Po Jen; Kilburn, Brian A.; Romero, Roberto; Diamond, Michael P.; Smith, Susan M.; Armant, D. Randall

2014-01-01

Background Apoptosis is induced by ethanol in human placental trophoblast cells, possibly disrupting placentation and contributing to intrauterine growth restriction in fetal alcohol spectrum disorder (FASD). Ethanol induces programmed cell death in several embryonic tissues by raising intracellular Ca2+. Therefore, the role of Ca2+ signaling in ethanol-induced apoptosis was examined using human first trimester cytotrophoblast cell lines, examining the hypothesis that apoptosis is dependent on intracellular Ca2+ signaling. Methods Using HTR-8/SVneo and SW.71 cytotrophoblast cell lines, real-time intracellular Ca2+ concentration was monitored by fluo-4 epifluorescence microscopy and apoptosis was assessed by flow cytometry of cells fluorescently labeled for DNA fragmentation (TUNEL) and annexin V binding. Results Intracellular Ca2+ concentrations increased synchronously in all cells within 10 s of exposure to 50 mM ethanol, but not at lower ethanol concentrations (10–25 mM) incapable of inducing apoptosis. Trophoblast cells treated with inhibitors of Ca2+ signaling (BAPTA-AM, U73122, xestospongin D, BAPTA, SKF-96365) produced no intracellular Ca2+ transients after exposure to 50 mM ethanol and were protected from cell death induced by ethanol. Conclusions Ethanol-induced apoptosis in human cytotrophoblast cells, identified by DNA fragmentation and externalized phosphatidylserine, was dependent upon Ca2+ signaling. Both intracellular Ca2+ mobilization and extracellular Ca2+ influx were required, as well as phosphatidylinositol signaling. Inhibition by SKF-96365 suggests that the capacitative Ca2+ entry mechanism that utilizes TRPC channels was activated by ethanol. Apoptosis occurs downsteam of Ca2+ signaling in trophoblasts, and may contribute to placental insufficiency and poor fetal growth associated with FASD. PMID:24889927

Apoptosis induced in Jurkat cells by several agents is preceded by intracellular acidification.

PubMed Central

Gottlieb, R A; Nordberg, J; Skowronski, E; Babior, B M

1996-01-01

We have previously shown that in neutrophils deprived of granulocyte colony-stimulating factor, apoptosis is preceded by acidification and that the protection against apoptosis conferred on neutrophils by granulocyte colony-stimulating factor is dependent upon delay of this acidification. To test the hypothesis that acidification could be a general feature of apoptosis, we examined intracellular pH changes in another cell line. Jurkat cells, a T-lymphoblastoid line, were induced to undergo apoptosis with anti-Fas IgM, cycloheximide, or exposure to short-wavelength UV light. We found that acidification occurred in response to treatment with these agents and that acidification preceded DNA fragmentation. Jurkat cells were also found to possess an acid endonuclease that is active below pH 6.8, compatible with a possible role for this enzyme in chromatin digestion during apoptosis. Incubation of the cells with the bases imidazole or chloroquine during treatment with anti-Fas antibody or cycloheximide or after UV exposure decreased apoptosis as assessed by nuclear morphology and DNA content. The alkalinizing effect of imidazole and chloroquine was shown by the demonstration that the percentage of cells with an intracellular pH below 6.8 after treatment with anti-Fas antibody, cycloheximide, or UV was diminished in the presence of base as compared with similarly treated cells incubated in the absence of base. We conclude that acidification is an early event in programmed cell death and may be essential for genome destruction. Images Fig. 5 PMID:8570610

Gefitinib enhances human colon cancer cells to TRAIL-induced apoptosis of via autophagy- and JNK-mediated death receptors upregulation.

PubMed

Chen, Lei; Meng, Yue; Guo, Xiaoqing; Sheng, Xiaotong; Tai, Guihua; Zhang, Fenglei; Cheng, Hairong; Zhou, Yifa

2016-11-01

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a potent cancer cell-specific apoptosis-inducing cytokine with little toxicity to most normal cells. Here, we report that gefitinib and TRAIL in combination produce a potent synergistic effect on TRAIL-sensitive human colon cancer HCT116 cells and an additive effect on TRAIL-resistant HT-29 cells. Interestingly, gefitinib increases the expression of cell surface receptors DR4 and DR5, possibly explaining the synergistic effect. Knockdown of DR4 and DR5 by siRNA significantly decreases gefitinib- and TRAIL-mediated cell apoptosis, supporting this idea. Because the inhibition of gefitinib-induced autophagy by 3-MA significantly decreases DR4 and DR5 upregulation, as well as reduces gefitinib- and TRAIL-induced apoptosis, we conclude that death receptor upregulation is autophagy mediated. Furthermore, our results indicate that death receptor expression may also be regulated by JNK activation, because pre-treatment of cells with JNK inhibitor SP600125 significantly decreases gefitinib-induced death receptor upregulation. Interestingly, SP600125 also inhibits the expression CHOP, yet CHOP has no impact on death receptor expressions. We also find here that phosphorylation of Akt and ERK might also be required for TRAIL sensitization. In summary, our results indicate that gefitinib effectively enhances TRAIL-induced apoptosis, likely via autophagy and JNK- mediated death receptor expression and phosphorylation of Akt and ERK.

CF3DODA-Me induces apoptosis, degrades Sp1, and blocks the transformation phase of the blebbishield emergency program.

PubMed

Taoka, Rikiya; Jinesh, Goodwin G; Xue, Wenrui; Safe, Stephen; Kamat, Ashish M

2017-05-01

Cancer stem cells are capable of undergoing cellular transformation after commencement of apoptosis through the blebbishield emergency program in a VEGF-VEGFR2-dependent manner. Development of therapeutics targeting the blebbishield emergency program would thus be important in cancer therapy. Specificity protein 1 (Sp1) orchestrates the transcription of both VEGF and VEGFR2; hence, Sp1 could act as a therapeutic target. Here, we demonstrate that CF 3 DODA-Me induced apoptosis, degraded Sp1, inhibited the expression of multiple drivers of the blebbishield emergency program such as VEGFR2, p70S6K, and N-Myc through activation of caspase-3, inhibited reactive oxygen species; and inhibited K-Ras activation to abolish transformation from blebbishields as well as transformation in soft agar. These findings confirm CF 3 DODA-Me as a potential therapeutic candidate that can induce apoptosis and block transformation from blebbishields.

Lysosomal ceramide generated by acid sphingomyelinase triggers cytosolic cathepsin B-mediated degradation of X-linked inhibitor of apoptosis protein in natural killer/T lymphoma cell apoptosis.

PubMed

Taniguchi, M; Ogiso, H; Takeuchi, T; Kitatani, K; Umehara, H; Okazaki, T

2015-04-09

We previously reported that IL-2 deprivation induced acid sphingomyelinase-mediated (ASM-mediated) ceramide elevation and apoptosis in an NK/T lymphoma cell line KHYG-1. However, the molecular mechanism of ASM-ceramide-mediated apoptosis during IL-2 deprivation is poorly understood. Here, we showed that IL-2 deprivation induces caspase-dependent apoptosis characterized by phosphatidylserine externalization, caspase-8, -9, and -3 cleavage, and degradation of X-linked inhibitor of apoptosis protein (XIAP). IL-2 re-supplementation rescued apoptosis via inhibition of XIAP degradation without affecting caspase cleavage. However, IL-2 deprivation induced ceramide elevation via ASM in lysosomes and activated lysosomal cathepsin B (CTSB) but not cathepsin D. A CTSB inhibitor CA-074 Me and knockdown of CTSB inhibited ceramide-mediated XIAP degradation and apoptosis. Inhibition of ceramide accumulation in lysosomes using an ASM inhibitor, desipramine, decreased cytosolic activation of CTSB by inhibiting its transfer into cytosol from the lysosome. Knockdown of ASM also inhibited XIAP degradation and apoptosis. Furthermore, cell permeable N-acetyl sphingosine (C2-ceramide), which increases mainly endogenous d18:1/16:0 and d18:1/24:1 ceramide-like IL-2 deprivation, induced caspase-dependent apoptosis with XIAP degradation through CTSB. These findings suggest that lysosomal ceramide produced by ASM mediates XIAP degradation by activation of cytosolic CTSB and caspase-dependent apoptosis. The ASM-ceramide-CTSB signaling axis is a novel pathway of ceramide-mediated apoptosis in IL-2-deprived NK/T lymphoma cells.

Lysosomal ceramide generated by acid sphingomyelinase triggers cytosolic cathepsin B-mediated degradation of X-linked inhibitor of apoptosis protein in natural killer/T lymphoma cell apoptosis

PubMed Central

Taniguchi, M; Ogiso, H; Takeuchi, T; Kitatani, K; Umehara, H; Okazaki, T

2015-01-01

We previously reported that IL-2 deprivation induced acid sphingomyelinase-mediated (ASM-mediated) ceramide elevation and apoptosis in an NK/T lymphoma cell line KHYG-1. However, the molecular mechanism of ASM–ceramide-mediated apoptosis during IL-2 deprivation is poorly understood. Here, we showed that IL-2 deprivation induces caspase-dependent apoptosis characterized by phosphatidylserine externalization, caspase-8, -9, and -3 cleavage, and degradation of X-linked inhibitor of apoptosis protein (XIAP). IL-2 re-supplementation rescued apoptosis via inhibition of XIAP degradation without affecting caspase cleavage. However, IL-2 deprivation induced ceramide elevation via ASM in lysosomes and activated lysosomal cathepsin B (CTSB) but not cathepsin D. A CTSB inhibitor CA-074 Me and knockdown of CTSB inhibited ceramide-mediated XIAP degradation and apoptosis. Inhibition of ceramide accumulation in lysosomes using an ASM inhibitor, desipramine, decreased cytosolic activation of CTSB by inhibiting its transfer into cytosol from the lysosome. Knockdown of ASM also inhibited XIAP degradation and apoptosis. Furthermore, cell permeable N-acetyl sphingosine (C2-ceramide), which increases mainly endogenous d18:1/16:0 and d18:1/24:1 ceramide-like IL-2 deprivation, induced caspase-dependent apoptosis with XIAP degradation through CTSB. These findings suggest that lysosomal ceramide produced by ASM mediates XIAP degradation by activation of cytosolic CTSB and caspase-dependent apoptosis. The ASM–ceramide–CTSB signaling axis is a novel pathway of ceramide-mediated apoptosis in IL-2-deprived NK/T lymphoma cells. PMID:25855965

Radiosensitivity and Induction of Apoptosis by High LET Carbon Ion Beam and Low LET Gamma Radiation: A Comparative Study

PubMed Central

Ghorai, Atanu; Bhattacharyya, Nitai P.; Sarma, Asitikantha; Ghosh, Utpal

2014-01-01

Cancer treatment with high LET heavy ion beam, especially, carbon ion beam (12C), is becoming very popular over conventional radiotherapy like low LET gamma or X-ray. Combination of Poly(ADP-ribose) polymerase (PARP) inhibitor with xenotoxic drugs or conventional radiation (gamma or X-ray) is the newer approach for cancer therapy. The aim of our study was to compare the radiosensitivity and induction of apoptosis by high LET 12C and low LET gamma radiation in HeLa and PARP-1 knocked down cells. We did comet assay to detect DNA breaks, clonogenic survival assay, and cell cycle analysis to measure recovery after DNA damage. We measured apoptotic parameters like nuclear fragmentation and caspase-3 activation. DNA damage, cell killing, and induction of apoptosis were significantly higher for 12C than gamma radiation in HeLa. Cell killing and apoptosis were further elevated upon knocking down of PARP-1. Both 12C and gamma induced G2/M arrest although the 12C had greater effect. Unlike the gamma, 12C irradiation affects DNA replication as detected by S-phase delay in cell cycle analysis. So, we conclude that high LET 12C has greater potential over low LET gamma radiation in killing cells and radiosensitization upon PARP-1 inhibition was several folds greater for 12C than gamma. PMID:25018892

Hypoxia preconditioning increases survival and decreases expression of Toll-like receptor 4 in pulmonary artery endothelial cells exposed to lipopolysaccharide.

PubMed

Ali, Irshad; Nanchal, Rahul; Husnain, Fouad; Audi, Said; Konduri, G Ganesh; Densmore, John C; Medhora, Meetha; Jacobs, Elizabeth R

2013-09-01

Abstract Pulmonary or systemic infections and hypoxemic respiratory failure are among the leading causes of admission to intensive care units, and these conditions frequently exist in sequence or in tandem. Inflammatory responses to infections are reproduced by lipopolysaccharide (LPS) engaging Toll-like receptor 4 (TLR4). Apoptosis is a hallmark of lung injury in sepsis. This study was conducted to determine whether preexposure to LPS or hypoxia modulated the survival of pulmonary artery endothelial cells (PAECs). We also investigated the role TLR4 receptor expression plays in apoptosis due to these conditions. Bovine PAECs were cultured in hypoxic or normoxic environments and treated with LPS. TLR4 antagonist TAK-242 was used to probe the role played by TLR4 receptors in cell survival. Cell apoptosis and survival were measured by caspase 3 activity and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) incorporation. TLR4 expression and tumor necrosis factor α (TNF-α) production were also determined. LPS increased caspase 3 activity in a TAK-242-sensitive manner and decreased MTT incorporation. Apoptosis was decreased in PAECs preconditioned with hypoxia prior to LPS exposure. LPS increased TNF-α production, and hypoxic preconditioning blunted it. Hypoxic preconditioning reduced LPS-induced TLR4 messenger RNA and TLR4 protein. TAK-242 decreased to baseline the LPS-stimulated expression of TLR4 messenger RNA regardless of environmental conditions. In contrast, LPS followed by hypoxia substantially increased apoptosis and cell death. In conclusion, protection from LPS-stimulated PAEC apoptosis by hypoxic preconditioning is attributable in part to reduction in TLR4 expression. If these signaling pathways apply to septic patients, they may account for differing sensitivities of individuals to acute lung injury depending on oxygen tensions in PAECs in vivo.

Transactivation of inducible nitric oxide synthase gene by Kruppel-like factor 6 regulates apoptosis during influenza A virus infection

PubMed Central

Mgbemena, Victoria; Segovia, Jesus A.; Chang, Te-Hung; Tsai, Su-Yu; Cole, Garry T.; Hung, Chiung-Yu; Bose, Santanu

2012-01-01

Influenza A virus (flu) is a respiratory tract pathogen causing high morbidity and mortality among the human population. Nitric oxide (NO) is a cellular mediator involved in tissue damage due to apoptosis of target cells and resulting enhancement of local inflammation. Inducible nitric oxide (iNOS) is involved in the production of NO following infection. Although NO is a key player in the development of exaggerated lung disease during flu infection, the underlying mechanism including the role of NO in apoptosis during infection has not been reported. Similarly, the mechanism of iNOS gene induction during flu infection is not well defined in terms of host trans-activator(s) required for iNOS gene expression. In the current study we have identified kruppel-like factor 6 (KLF6) as a critical transcription factor essential for iNOS gene expression during flu infection. We have also underscored the requirement of iNOS in inducing apoptosis during infection. KLF6 gene silencing in human lung epithelial cells resulted in drastic loss of NO production, iNOS-promoter specific luciferase activity and expression of iNOS mRNA following flu infection. Chromatin immuno-precipitation assay revealed a direct interaction of KLF6 with iNOS promoter during both in vitro and in vivo flu infection of human lung cells and mouse respiratory tract, respectively. Significant reduction in flu mediated apoptosis was noted in KLF6 silenced cells, cells treated with iNOS inhibitor and in primary murine macrophages derived from iNOS knock-out (KO) mice. A similar reduction in apoptosis was noted in the lungs following intra-tracheal flu infection of iNOS KO mice. PMID:22711891

Interrogating the relevance of mitochondrial apoptosis for vertebrate development and postnatal tissue homeostasis.

PubMed

Tuzlak, Selma; Kaufmann, Thomas; Villunger, Andreas

2016-10-01

"Programmed cell death or 'apoptosis' is critical for organogenesis during embryonic development and tissue homeostasis in the adult. Its deregulation can contribute to a broad range of human pathologies, including neurodegeneration, cancer, or autoimmunity…" These or similar phrases have become generic opening statements in many reviews and textbooks describing the physiological relevance of apoptotic cell death. However, while the role in disease has been documented beyond doubt, facilitating innovative drug discovery, we wonder whether the former is really true. What goes wrong in vertebrate development or in adult tissue when the main route to apoptotic cell death, controlled by the BCL2 family, is impaired? Such scenarios have been mimicked by deletion of one or more prodeath genes within the BCL2 family, and gene targeting studies in mice exploring the consequences have been manifold. Many of these studies were geared toward understanding the role of BCL2 family proteins and mitochondrial apoptosis in disease, whereas fewer focused in detail on their role during normal development or tissue homeostasis, perhaps also due to an irritating lack of phenotype. Looking at these studies, the relevance of classical programmed cell death by apoptosis for development appears rather limited. Together, these many studies suggest either highly selective and context-dependent contributions of mitochondrial apoptosis or significant redundancy with alternative cell death mechanisms, as summarized and discussed here. © 2016 Tuzlak et al.; Published by Cold Spring Harbor Laboratory Press.

Differences in the rate of oestrogen-induced apoptosis in breast cancer by oestradiol and the triphenylethylene bisphenol

PubMed Central

Obiorah, I E; Jordan, V C

2014-01-01

Background and Purpose Triphenylethylene (TPE)-like compounds were the first agents to be used in the treatment of metastatic breast cancer in postmenopausal women. Although structurally related to the anti-oestrogen, 4-hydroxytamoxifen, TPEs possess oestrogenic properties in fully oestrogenized breast cancer cells but do not induce apoptosis with short-term treatment in long-term oestrogen-deprived breast cancer cells. This study determined the differential effects of bisphenol, a TPE, on growth and apoptosis based on the modulation of the shape of the ligand–oestrogen receptor complex. Experimental Approach Apoptotic flow cytometric studies were used to evaluate apoptosis over time. Proliferation of the breast cancer cells was assessed using DNA quantification and cell cycle analysis. Real-time PCR was performed to quantify mRNA levels of apoptotic genes. Regulation of cell cycle and apoptotic genes was determined using PCR-based arrays. Key Results Bisphenol induced an up-regulation of cell cycle genes similar to those induced by 17β oestradiol (E2). Unlike the changes induced by E2 that occur after 24 h, the apoptosis evoked by bisphenol occurred after 4 days, with quantifiable apoptotic changes noted at 6 days. A prolonged up-regulation of endoplasmic reticulum stress and inflammatory stress response genes was observed with subsequent activation of apoptosis-related genes in the second week of treatment with bisphenol. Conclusions and Implications The bisphenol: ERα complex induces delayed biological effects on the growth and apoptosis of breast cancer cells. Both the shape of the complex and the duration of treatment control the initiation of apoptosis. PMID:24819221

Sensitization of cervix cancer cells to Adriamycin by Pentoxifylline induces an increase in apoptosis and decrease senescence.

PubMed

Bravo-Cuellar, Alejandro; Ortiz-Lazareno, Pablo C; Lerma-Diaz, Jose M; Dominguez-Rodriguez, Jorge R; Jave-Suarez, Luis F; Aguilar-Lemarroy, Adriana; del Toro-Arreola, Susana; de Celis-Carrillo, Ruth; Sahagun-Flores, Jose E; de Alba-Garcia, Javier E Garcia; Hernandez-Flores, Georgina

2010-05-19

Chemotherapeutic drugs like Adriamycin (ADR) induces apoptosis or senescence in cancer cells but these cells often develop resistance and generate responses of short duration or complete failure. The methylxantine drug Pentoxifylline (PTX) used routinely in the clinics setting for circulatory diseases has been recently described to have antitumor properties. We evaluated whether pretreatment with PTX modifies apoptosis and senescence induced by ADR in cervix cancer cells. HeLa (HPV 18+), SiHa (HPV 16+) cervix cancer cells and non-tumorigenic immortalized HaCaT cells (control) were treated with PTX, ADR or PTX + ADR. The cellular toxicity of PTX and survival fraction were determinated by WST-1 and clonogenic assay respectively. Apoptosis, caspase activation and ADR efflux rate were measured by flow cytometry, senescence by microscopy. IkappaBalpha and DNA fragmentation were determinated by ELISA. Proapoptotic, antiapoptotic and senescence genes, as well as HPV-E6/E7 mRNA expression, were detected by time real RT-PCR. p53 protein levels were assayed by Western blot. PTX is toxic (WST-1), affects survival (clonogenic assay) and induces apoptosis in cervix cancer cells. Additionally, the combination of this drug with ADR diminished the survival fraction and significantly increased apoptosis of HeLa and SiHa cervix cancer cells. Treatments were less effective in HaCaT cells. We found caspase participation in the induction of apoptosis by PTX, ADR or its combination. Surprisingly, in spite of the antitumor activity displayed by PTX, our results indicate that methylxantine, per se does not induce senescence; however it inhibits senescence induced by ADR and at the same time increases apoptosis. PTX elevates IkappaBalpha levels. Such sensitization is achieved through the up-regulation of proapoptotic factors such as caspase and bcl family gene expression. PTX and PTX + ADR also decrease E6 and E7 expression in SiHa cells, but not in HeLa cells. p53 was detected only in SiHa cells treated with ADR. PTX is a good inducer of apoptosis but does not induce senescence. Furthermore, PTX reduced the ADR-induced senescence and increased apoptosis in cervix cancer cells.

Sensitization of cervix cancer cells to Adriamycin by Pentoxifylline induces an increase in apoptosis and decrease senescence

PubMed Central

2010-01-01

Background Chemotherapeutic drugs like Adriamycin (ADR) induces apoptosis or senescence in cancer cells but these cells often develop resistance and generate responses of short duration or complete failure. The methylxantine drug Pentoxifylline (PTX) used routinely in the clinics setting for circulatory diseases has been recently described to have antitumor properties. We evaluated whether pretreatment with PTX modifies apoptosis and senescence induced by ADR in cervix cancer cells. Methods HeLa (HPV 18+), SiHa (HPV 16+) cervix cancer cells and non-tumorigenic immortalized HaCaT cells (control) were treated with PTX, ADR or PTX + ADR. The cellular toxicity of PTX and survival fraction were determinated by WST-1 and clonogenic assay respectively. Apoptosis, caspase activation and ADR efflux rate were measured by flow cytometry, senescence by microscopy. IκBα and DNA fragmentation were determinated by ELISA. Proapoptotic, antiapoptotic and senescence genes, as well as HPV-E6/E7 mRNA expression, were detected by time real RT-PCR. p53 protein levels were assayed by Western blot. Results PTX is toxic (WST-1), affects survival (clonogenic assay) and induces apoptosis in cervix cancer cells. Additionally, the combination of this drug with ADR diminished the survival fraction and significantly increased apoptosis of HeLa and SiHa cervix cancer cells. Treatments were less effective in HaCaT cells. We found caspase participation in the induction of apoptosis by PTX, ADR or its combination. Surprisingly, in spite of the antitumor activity displayed by PTX, our results indicate that methylxantine, per se does not induce senescence; however it inhibits senescence induced by ADR and at the same time increases apoptosis. PTX elevates IκBα levels. Such sensitization is achieved through the up-regulation of proapoptotic factors such as caspase and bcl family gene expression. PTX and PTX + ADR also decrease E6 and E7 expression in SiHa cells, but not in HeLa cells. p53 was detected only in SiHa cells treated with ADR. Conclusion PTX is a good inducer of apoptosis but does not induce senescence. Furthermore, PTX reduced the ADR-induced senescence and increased apoptosis in cervix cancer cells. PMID:20482878

Ferritin expression in rat hepatocytes and Kupffer cells after lead nitrate treatment.

PubMed

Fan, Yang; Yamada, Toshiyuki; Shimizu, Takeshi; Nanashima, Naoki; Akita, Miki; Suto, Kohji; Tsuchida, Shigeki

2009-02-01

Lead nitrate induces hepatocyte proliferation and subsequent apoptosis in rat livers. Iron is a constituent of heme and is also required for cell proliferation. In this study, the expression of ferritin light-chain (FTL), the major iron storage protein, was investigated in rat livers after a single intravenous injection of lead nitrate. Western blotting and immunohistochemistry revealed that FTL was increased in hepatocytes around the central veins and strongly expressed in nonparenchymal cells. Some FTL-positive nonparenchymal cells were identified as Kupffer cells that were positive for CD68. FTL-positive Kupffer cells occupied about 60% of CD68-positive cells in the periportal and perivenous areas. The relationships between FTL expression and apoptosis induction or the engulfment of apoptotic cells were examined. TUNEL-positive cells were increased in the treatment group, and enhanced expression of milk fat globule EGF-like 8 was demonstrated in some Kupffer cells and hepatocytes, indicating enhanced apoptosis induction and phagocytosis of apoptotic cells. FTL-positive Kupffer cells were not detected without lead nitrate treatment or in rat livers treated with clofibrate, which induces hepatocyte proliferation but not apoptosis. These results suggest that FTL expression in Kupffer cells after lead treatment is dependent on phagocytosis of apoptotic cells.

Effects on growth of human osteoblast-like cells of three nonsteroidal anti-inflammatory drugs: metamizole, dexketoprofen, and ketorolac.

PubMed

De Luna-Bertos, Elvira; Ramos-Torrecillas, Javier; Manzano-Moreno, Francisco Javier; García-Martínez, Olga; Ruiz, Concepción

2015-01-01

Some nonsteroidal anti-inflammatory drugs (NSAIDs) have adverse effects on bone tissue. The objective of this study was to determine the effect of different doses of dexketoprofen, ketorolac, and metamizole on growth of the osteoblast MG63 cell line. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide spectrophotometry results showed that MG63 cell growth was significantly inhibited after 24 hr of culture with doses of 10, 20, 100, or 1,000 µM of each NSAID and with doses of 0.1, 1, or 5 µM of dexketoprofen and ketorolac but not metamizole. Cell-cycle studies revealed that dexketoprofen and ketorolac treatments significantly arrested the cell cycle in phase G0/G1, increasing the percentage of cells in this phase. Apoptosis/necrosis studies showed significant changes versus control cells, with an increased percentage of cells in apoptosis after treatment with 10, 100, or 1,000 µM of metamizole and after treatment with 1, 10, 100, or 1,000 µM of dexketoprofen or ketorolac. In conclusion, treatment of osteoblast-like cells with high doses of the NSAIDs tested increased not only the percentage of cells in apoptosis but also the percentage of necrotic cells. © The Author(s) 2014.

'Hints' in the killer protein gasdermin D: unveiling the secrets of gasdermins driving cell death.

PubMed

Qiu, Shiqiao; Liu, Jing; Xing, Feiyue

2017-04-01

Pyroptosis is a lytic form of cell death distinguished from apoptosis, ferroptosis, necrosis, necroptosis, NETosis, oncosis, pyronecrosis and autophagy. Proinflammatory caspases cleave a gasdermin D (GSDMD) protein to generate a 31 kDa N-terminal domain. The cleavage relieves the intramolecular inhibition on the gasdermin-N domain, which then moves to the plasma membrane to exhibit pore-forming activity. Thus, GSDMD acts as the final and direct executor of pyroptotic cell death. Owing to the selective targeting of the inner leaflet of the plasma membrane with the pore-forming that determines pyroptotic cell death, GSDMD could be a potential target to control cell death or extracellular bacterial infections. Intriguingly, other gasdermin family members also share similar N-terminal domains, but they present different cell death programs. Herein, we summarize features and functions of the novel player proteins in cell death, including GSDMD triggering pyroptosis, Gsdma3/GSDMA initiating autophagy/apoptosis and DFNA5 inducing apoptosis/secondary necrosis. The gasdermin N terminus appears to be a novel pore-forming protein. This provides novel insight into the underlying roles and mechanisms of lytic or nonlytic forms of programmed cell death, as well as their potential applications in inflammation-associated diseases.

BH3-Only Protein BIM Mediates Heat Shock-Induced Apoptosis

PubMed Central

Mahajan, Indra M.; Chen, Miao-Der; Muro, Israel; Robertson, John D.; Wright, Casey W.; Bratton, Shawn B.

2014-01-01

Acute heat shock can induce apoptosis through a canonical pathway involving the upstream activation of caspase-2, followed by BID cleavage and stimulation of the intrinsic pathway. Herein, we report that the BH3-only protein BIM, rather than BID, is essential to heat shock-induced cell death. We observed that BIM-deficient cells were highly resistant to heat shock, exhibiting short and long-term survival equivalent to Bax−/−Bak−/− cells and better than either Bid−/− or dominant-negative caspase-9-expressing cells. Only Bim−/− and Bax−/−Bak−/− cells exhibited resistance to mitochondrial outer membrane permeabilization and loss of mitochondrial inner membrane potential. Moreover, while dimerized caspase-2 failed to induce apoptosis in Bid−/− cells, it readily did so in Bim−/− cells, implying that caspase-2 kills exclusively through BID, not BIM. Finally, BIM reportedly associates with MCL-1 following heat shock, and Mcl-1−/− cells were indeed sensitized to heat shock-induced apoptosis. However, pharmacological inhibition of BCL-2 and BCL-XL with ABT-737 also sensitized cells to heat shock, most likely through liberation of BIM. Thus, BIM mediates heat shock-induced apoptosis through a BAX/BAK-dependent pathway that is antagonized by antiapoptotic BCL-2 family members. PMID:24427286

Human Embryonic and Induced Pluripotent Stem Cells Express TRAIL Receptors and Can Be Sensitized to TRAIL-Induced Apoptosis

PubMed Central

Vinarsky, Vladimir; Krivanek, Jan; Rankel, Liina; Nahacka, Zuzana; Barta, Tomas; Jaros, Josef; Andera, Ladislav

2013-01-01

Death ligands and their tumor necrosis factor receptor (TNFR) family receptors are the best-characterized and most efficient inducers of apoptotic signaling in somatic cells. In this study, we analyzed whether these prototypic activators of apoptosis are also expressed and able to be activated in human pluripotent stem cells. We examined human embryonic stem cells (hESC) and human-induced pluripotent stem cells (hiPSC) and found that both cell types express primarily TNF-related apoptosis-inducing ligand (TRAIL) receptors and TNFR1, but very low levels of Fas/CD95. We also found that although hESC and hiPSC contain all the proteins required for efficient induction and progression of extrinsic apoptotic signaling, they are resistant to TRAIL-induced apoptosis. However, both hESC and hiPSC can be sensitized to TRAIL-induced apoptosis by co-treatment with protein synthesis inhibitors such as the anti-leukemia drug homoharringtonine (HHT). HHT treatment led to suppression of cellular FLICE inhibitory protein (cFLIP) and Mcl-1 expression and, in combination with TRAIL, enhanced processing of caspase-8 and full activation of caspase-3. cFLIP likely represents an important regulatory node, as its shRNA-mediated down-regulation significantly sensitized hESC to TRAIL-induced apoptosis. Thus, we provide the first evidence that, irrespective of their origin, human pluripotent stem cells express canonical components of the extrinsic apoptotic system and on stress can activate death receptor-mediated apoptosis. PMID:23806100

Formononetin induces the mitochondrial apoptosis pathway in prostate cancer cells via downregulation of the IGF-1/IGF-1R signaling pathway.

PubMed

Huang, Wen-Jun; Bi, Ling-Yun; Li, Zhen-Zhao; Zhang, Xing; Ye, Yu

2013-12-20

Abstract Context: Formononetin, an isoflavone, can inhibit the proliferation of cancer cells, including those of the prostate. However, its antitumor mechanism remains unclear. Aim: To investigate whether the insulin-like growth factor 1 (IGF-1)/insulin-like growth factor 1 receptor (IGF-1 R) signaling pathway mediates the formononetin antitumor effect on prostate cancer cells. Materials and methods: The viability of PC-3 cells was measured by MTT assay 48 h after formononetin treatment (25, 50 and 100 μM). Formononetin-induced cell apoptosis was measured by Hoechst 33258 staining and flow cytometry. Expression of Bax mRNA was detected by real-time PCR, and the expression levels of Bax and IGF-1 R proteins were detected by western blots. Results: At concentrations >12.5 μM, formononetin significantly inhibited the proliferation of human prostate cancer cells. Formononetin increased Bax mRNA and protein expression levels and decreased the expression levels of pIGF-1 R protein in a dose-dependent manner. Conclusion: High concentrations of formononetin-induced apoptosis in androgen-independent prostate cancer cells through inhibition of the IGF-1/IGF-1 R pathway.

Fibrocyte migration, differentiation and apoptosis during the corneal wound healing response to injury.

PubMed

Lassance, Luciana; Marino, Gustavo K; Medeiros, Carla S; Thangavadivel, Shanmugapriya; Wilson, Steven E

2018-05-01

The aim of this study was to determine whether bone marrow-derived fibrocytes migrate into the cornea after stromal scar-producing injury and differentiate into alpha-smooth muscle actin (αSMA) + myofibroblasts. Chimeric mice expressing green fluorescent protein (GFP) bone marrow cells had fibrosis (haze)-generating irregular phototherapeutic keratectomy (PTK). Multiplex immunohistochemistry (IHC) for GFP and fibrocyte markers (CD34, CD45, and vimentin) was used to detect fibrocyte infiltration into the corneal stroma and the development of GFP+ αSMA+ myofibroblasts. IHC for activated caspase-3, GFP and CD45 was used to detect fibrocyte and other hematopoietic cells undergoing apoptosis. Moderate haze developed in PTK-treated mouse corneas at 14 days after surgery and worsened, and persisted, at 21 days after surgery. GFP+ CD34+ CD45+ fibrocytes, likely in addition to other CD34+ and/or CD45+ hematopoietic and stem/progenitor cells, infiltrated the cornea and were present in the stroma in high numbers by one day after PTK. The fibrocytes and other bone marrow-derived cells progressively decreased at four days and seven days after surgery. At four days after PTK, 5% of the GFP+ cells expressed activated caspase-3. At 14 days after PTK, more than 50% of GFP+ CD45+ cells were also αSMA+ myofibroblasts. At 21 days after PTK, few GFP+ αSMA+ cells persisted in the stroma and more than 95% of those remaining expressed activated caspase-3, indicating they were undergoing apoptosis. GFP+ CD45+ SMA+ cells that developed from 4 to 21 days after irregular PTK were likely developed from fibrocytes. After irregular PTK in the strain of C57BL/6-C57/BL/6-Tg(UBC-GFP)30Scha/J chimeric mice, however, more than 95% of fibrocytes and other hematopoietic cells underwent apoptosis prior to the development of mature αSMA+ myofibroblasts. Most GFP+ CD45+ αSMA+ myofibroblasts that did develop subsequently underwent apoptosis-likely due to epithelial basement membrane regeneration and deprivation of epithelium-derived TGFβ requisite for myofibroblast survival. Copyright © 2018 Elsevier Ltd. All rights reserved.

Apoptotic depletion of CD4+ T cells in idiopathic CD4+ T lymphocytopenia.

PubMed Central

Laurence, J; Mitra, D; Steiner, M; Lynch, D H; Siegal, F P; Staiano-Coico, L

1996-01-01

Progressive loss of CD4+ T lymphocytes, accompanied by opportunistic infections characteristic of the acquired immune deficiency syndrome, ahs been reported in the absence of any known etiology. The pathogenesis of this syndrome, a subset of idiopathic CD4+ T lymphocytopenia (ICL), is uncertain. We report that CD4+ T cells from seven of eight ICL patients underwent accelerated programmed cell death, a process facilitated by T cell receptor cross-linking. Apoptosis was associated with enhanced expression of Fas and Fas ligand in unstimulated cell populations, and partially inhibited by soluble anti-Fas mAb. In addition, apoptosis was suppressed by aurintricarboxylic acid, an inhibitor of calcium-dependent endonucleases and proteases, in cells from four of seven patients, The in vivo significance of these findings was supported by three factors: the absence of accelerated apoptosis in persons with stable, physiologic CD4 lymphopenia without clinical immune deficiency; detection of serum antihistone H2B autoantibodies, one consequence of DNA fragmentation, in some patients; and its selectivity, with apoptosis limited to the CD4 population in some, and occurring among CD8+ T cells predominantly in those individuals with marked depletion of both CD4+ T lymphocytes linked to clinical immune suppression have evidence for accelerated T cell apoptosis in vitro that may be pathophysiologic and amenable to therapy with apoptosis inhibitors. PMID:8609222

Embryo apoptosis identification: Oocyte grade or cleavage stage?

PubMed Central

Bakri, Noraina Mohd; Ibrahim, Siti Fatimah; Osman, Nurul Atikah; Hasan, Nurhaslina; Jaffar, Farah Hanan Fathihah; Rahman, Zulaiha Abdul; Osman, Khairul

2015-01-01

Apoptosis is a programed cell death that is vital for tissue homeostasis. However, embryo apoptosis had been known to be related to embryo fragmentation which should be avoided in in vitro fertilization (IVF). The purpose of this study was to evaluate the relationship of embryo apoptosis with the grade of immature oocytes and cleavage stage of in vitro produced (IVP) cattle embryos. This study consisted of 345 oocytes collected through ovary slicing. Immature oocytes were graded as A, B and C. This grading was based on cumulus cell thickness and compactness. All oocytes then underwent an in vitro maturation (IVM) procedure. An IVF was done 24 h after IVM culture. Prior to staining, stage of cleaved embryos was determined and classified as either 2, 4, 8 or >8-cell embryo stage. Apoptosis status of cleaved IVP embryos was determined by using annexin V-FITC staining technique at 48 and 72 h post insemination (hpi). Apoptosis status for each embryo was classified as either early or late. The result showed that there was no significant difference (p > 0.05) of apoptosis status among grade A, B and C embryos. All grades of oocytes showed embryo apoptosis where 1.5% late apoptosis for grade A, 4.5% and 10.4% of early and late apoptosis for grade B and grade C. Early apoptosis was not seen in grade A embryo. We also noted no significant difference (p > 0.05) of apoptosis status between 2, 4, 8 and >8-cell embryo stage. Early apoptosis was also not seen in >8-cell stage. Even though there were no differences in apoptosis expression between the three classes, the cleavage rate of grade A oocytes was significantly higher (p < 0.01) than grade B and grade C. In conclusion, the apoptosis expression in the embryo can occur regardless of the oocyte quality and the cleavage stage of the embryo produced. PMID:26858565

Inducing death in tumor cells: roles of the inhibitor of apoptosis proteins.

PubMed

Finlay, Darren; Teriete, Peter; Vamos, Mitchell; Cosford, Nicholas D P; Vuori, Kristiina

2017-01-01

The heterogeneous group of diseases collectively termed cancer results not just from aberrant cellular proliferation but also from a lack of accompanying homeostatic cell death. Indeed, cancer cells regularly acquire resistance to programmed cell death, or apoptosis, which not only supports cancer progression but also leads to resistance to therapeutic agents. Thus, various approaches have been undertaken in order to induce apoptosis in tumor cells for therapeutic purposes. Here, we will focus our discussion on agents that directly affect the apoptotic machinery itself rather than on drugs that induce apoptosis in tumor cells indirectly, such as by DNA damage or kinase dependency inhibition. As the roles of the Bcl-2 family have been extensively studied and reviewed recently, we will focus in this review specifically on the inhibitor of apoptosis protein (IAP) family. IAPs are a disparate group of proteins that all contain a baculovirus IAP repeat domain, which is important for the inhibition of apoptosis in some, but not all, family members. We describe each of the family members with respect to their structural and functional similarities and differences and their respective roles in cancer. Finally, we also review the current state of IAPs as targets for anti-cancer therapeutics and discuss the current clinical state of IAP antagonists.

[Roles of KLF5 in inhibition TNFα-induced SK-BR-3 breast cancer cell apoptosis].

PubMed

Shi, Jianhong; Liu, Caiyun; Zhang, Anyi; Cui, Naipeng; Wang, Bing; Chen, Baoping; Ma, Zhenfeng

2014-07-08

To explore the expression levels and roles of Krüpple-like factor 5 (KLF5) in tumor necrosis factor α (TNFα)-induced SK-BR-3 breast cancer cells. SK-BR-3 breast cancer cells were stimulated by TNFα at different concentrations (0, 1, 5, 10, 20 µg/L) for specified durations (0, 6, 12, 24, 36 h). Western blot was performed to detect KLF5 protein levels. Then Western blot and quantitative real-time PCR (qRT-PCR) were used to detect the expression levels of apoptosis genes. Flow cytometry and qRT-PCR were used to observe the effects of exogenous KLF5 on TNFα-induced apoptosis of SK-BR-3 breast cancer cell. KLF5 expression levels significantly decreased in TNFα-stimulated SK-BR-3 breast cancer cells in a concentration- and time-dependent manner. Quantitative RT-PCR results showed that TNFα up-regulate apoptosis gene caspase 3, caspase 9 and bax expression levels and down-regulate bcl-1 level in SK-BR-3 cells. Adenovirus expression vectors of pAd-GFP and pAd-GFP-KLF5 were constructed and used to infect SK-BR-3 breast cancer cells. Over-expression of GFP-KLF5 inhibited apoptosis in TNFα-stimulated SK-BR-3 breast cancer cells. TNFα reduces KLF5 expression in SK-BR-3 breast cancer cells and KLF5 participates in TNFα-induced SK-BR-3 cell apoptosis.

Protein Kinase G facilitates EGFR-mediated cell death in MDA-MB-468 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jackson, Nicole M.; Ceresa, Brian P., E-mail: brian.ceresa@louisville.edu

The Epidermal Growth Factor Receptor (EGFR) is a transmembrane receptor tyrosine kinase with critical implications in cell proliferation, migration, wound healing and the regulation of apoptosis. However, the EGFR has been shown to be hyper-expressed in a number of human malignancies. The MDA-MB-468 metastatic breast cell line is one example of this. This particular cell line hyper-expresses the EGFR and undergoes EGFR-mediated apoptosis in response to EGF ligand. The goal of this study was to identify the kinases that could be potential intermediates for the EGFR-mediated induction of apoptosis intracellularly. After identifying Cyclic GMP-dependent Protein Kinase G (PKG) as amore » plausible intermediate, we wanted to determine the temporal relationship of these two proteins in the induction of apoptosis. We observed a dose-dependent decrease in MDA-MB-468 cell viability, which was co-incident with increased PKG activity as measured by VASPSer239 phosphorylation. In addition, we observed a dose dependent decrease in cell viability, as well as an increase in apoptosis, in response to two different PKG agonists, 8-Bromo-cGMP and 8-pCPT-cGMP. MDA-MB-468 cells with reduced PKG activity had attenuated EGFR-mediated apoptosis. These findings indicate that PKG does not induce cell death via transphosphorylation of the EGFR. Instead, PKG activity occurs following EGFR activation. Together, these data indicate PKG as an intermediary in EGFR-mediated cell death, likely via apoptotic pathway.« less

MST1 activation by curcumin mediates JNK activation, Foxo3a nuclear translocation and apoptosis in melanoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu, Teng, E-mail: tengyu33@yahoo.com; Ji, Jiang; Guo, Yong-li

2013-11-08

Highlights: •Curcumin activates MST1 in melanoma cells. •MST1 mediates curcumin-induced apoptosis of melanoma cells. •ROS production is involved in curcumin-induced MST1 activation. •MST1 mediates curcumin-induced JNK activation in melanoma cells. •MST1 mediates curcumin-induced Foxo3a nuclear translocation and Bim expression. -- Abstract: Different groups including ours have shown that curcumin induces melanoma cell apoptosis, here we focused the role of mammalian Sterile 20-like kinase 1 (MST1) in it. We observed that curcumin activated MST1-dependent apoptosis in cultured melanoma cells. MST1 silencing by RNA interference (RNAi) suppressed curcumin-induced cell apoptosis, while MST1 over-expressing increased curcumin sensitivity. Meanwhile, curcumin induced reactive oxygen speciesmore » (ROS) production in melanoma cells, and the ROS scavenger, N-acetyl-cysteine (NAC), almost blocked MST1 activation to suggest that ROS might be required for MST1 activation by curcumin. c-Jun N-terminal protein kinase (JNK) activation by curcumin was dependent on MST1, since MST1 inhibition by RNAi or NAC largely inhibited curcumin-induced JNK activation. Further, curcumin induced Foxo3 nuclear translocation and Bim-1 (Foxo3 target gene) expression in melanoma cells, such an effect by curcumin was inhibited by MST1 RNAi. In conclusion, we suggested that MST1 activation by curcumin mediates JNK activation, Foxo3a nuclear translocation and apoptosis in melanoma cells.« less

Radiation Dose-effects on Cell Cycle, Apoptosis, and Marker Expression of Ataxia Telangiectasia-Heterozygous Human Breast Epithelial Cells

NASA Technical Reports Server (NTRS)

Cruz, A.; Bors, K.; Jansen, H.; Richmond, R.

2003-01-01

Ataxia-telangiectasia (A-T) is a radiation-sensitive genetic condition. AT-heterozygous human mammary epithelial cells (HMEC) were irradiated using a Cs137 source in order to compare cell cycle, apoptosis, and marker expression responses across 3 radiation doses. No differences in cell cycle and apoptosis were found with any of the radiation doses used (30, 60, and 90 rads) compared with the unirradiated control (0 rad). At the same doses, however, differences were found in marker expression, such as keratin 18 (kl8), keratin 14 (k14), insulin-like growth factor I receptor (IGF-IR), and connexin 43 (cx43). This may indicate that radiation sensitivity in the heterozygous state may be initiated through signal transduction responses.

Construction and analysis of a modular model of caspase activation in apoptosis

PubMed Central

Harrington, Heather A; Ho, Kenneth L; Ghosh, Samik; Tung, KC

2008-01-01

Background A key physiological mechanism employed by multicellular organisms is apoptosis, or programmed cell death. Apoptosis is triggered by the activation of caspases in response to both extracellular (extrinsic) and intracellular (intrinsic) signals. The extrinsic and intrinsic pathways are characterized by the formation of the death-inducing signaling complex (DISC) and the apoptosome, respectively; both the DISC and the apoptosome are oligomers with complex formation dynamics. Additionally, the extrinsic and intrinsic pathways are coupled through the mitochondrial apoptosis-induced channel via the Bcl-2 family of proteins. Results A model of caspase activation is constructed and analyzed. The apoptosis signaling network is simplified through modularization methodologies and equilibrium abstractions for three functional modules. The mathematical model is composed of a system of ordinary differential equations which is numerically solved. Multiple linear regression analysis investigates the role of each module and reduced models are constructed to identify key contributions of the extrinsic and intrinsic pathways in triggering apoptosis for different cell lines. Conclusion Through linear regression techniques, we identified the feedbacks, dissociation of complexes, and negative regulators as the key components in apoptosis. The analysis and reduced models for our model formulation reveal that the chosen cell lines predominately exhibit strong extrinsic caspase, typical of type I cell, behavior. Furthermore, under the simplified model framework, the selected cells lines exhibit different modes by which caspase activation may occur. Finally the proposed modularized model of apoptosis may generalize behavior for additional cells and tissues, specifically identifying and predicting components responsible for the transition from type I to type II cell behavior. PMID:19077196

Apoptosis in muscle-to-meat aging process: The omic witness.

PubMed

Longo, Valentina; Lana, Alessandro; Bottero, Maria Teresa; Zolla, Lello

2015-07-01

Meat derives from a muscle that undergoes a great number of biochemical and physiological changes. The anoxic condition established from the moment of animal sacrifice forces muscle cells to a sort of reaction, resulting in methodical programmed cell death to avoid necrosis. The duality autophagy and/or apoptosis is at the center of the scientific debate about the biological processes driving the muscle to meat conversion. Here we report an omic time course overview carried on proteome, phosphoproteome and metabolome of Piedmontese longissimus thoracis muscle searching for clues helping us to extricate through the dilemma. The survey depicts a progressive physiological impairing and our evidences push towards the apoptotic behavior: the proteomic time course trend of annexin A2, RKIP, HSPB6, αB crystalline, adenylate kinase, DJ-1 and 31kDa actin fragment; the 0-1day increased phosphorylation of myosin 2 and synaptopodin and the metabolomic time course trend of key metabolic indicators, like GSH/GSSG ratio, taurine and nitrotyrosine. The employed techniques provide strong indications about the likely apoptotic behavior of aging meat in muscle-to-meat conversion process. Our work underlines compelling evidences of the apoptotic behavior of Piedmontese beef muscle cells undergoing the muscle-to-meat process, whereas no autophagic clues are inferred from this omic investigation. Copyright © 2015 Elsevier B.V. All rights reserved.

Stomatin-like protein 2 is overexpressed in cervical cancer and involved in tumor cell apoptosis

PubMed Central

Deng, Huan; Deng, Yongjian; Liu, Feiye; Chen, Jie; Li, Zheng; Zhao, Kelei; Guan, Xiaoqian; Liang, Weijiang

2017-01-01

Stomatin-like protein 2 (SLP-2) is overexpressed in numerous types of human cancer and previous studies revealed that SLP-2 may function in mitochondria. The purpose of the present study was to evaluate the expression of SLP-2 in cervical cancer and the association between SLP-2 expression and clinical features, in addition to investigating the role of SLP-2 in the apoptosis of cervical cancer cells. The expression profile of SLP-2 was determined by quantitative polymerase chain reaction, western blotting and immunohistochemical staining. The effect of SLP-2 on cell apoptosis induced by chemotherapeutics in cervical cancer cells was evaluated using Annexin V staining and terminal deoxynucleotidyl-transferase-mediated dUTP nick end labeling (TUNEL) assays. The results indicated that SLP-2 expression in cervical cancer was significantly upregulated at the mRNA and protein levels, compared with that in normal cervical tissues. Immunohistochemical analysis revealed significant correlation between SLP-2 protein expression and clinical characteristics, including the squamous cell carcinoma antigen (P=0.003), deep stromal invasion (P=0.021), lymphovascular space involvement (P=0.044) and pelvic lymph node metastasis (P<0.001), which served as independent prognostic factors for predicting the shortening of overall survival time in patients with early-stage cervical cancer. In addition, TUNEL and Annexin V binding assays revealed that silencing SLP-2 expression significantly enhanced the sensitivity of cervical cancer cells to apoptosis induced by chemotherapeutics. Taken together, the results of the present study suggest that SLP-2 may be a progressive gene in the development of cervical cancer. Overexpression of SLP-2 serves an important role in the apoptosis of human cervical cancer cells. PMID:29181097

The necrosome promotes pancreatic oncogenesis via CXCL1 and Mincle-induced immune suppression.

PubMed

Seifert, Lena; Werba, Gregor; Tiwari, Shaun; Giao Ly, Nancy Ngoc; Alothman, Sara; Alqunaibit, Dalia; Avanzi, Antonina; Barilla, Rocky; Daley, Donnele; Greco, Stephanie H; Torres-Hernandez, Alejandro; Pergamo, Matthew; Ochi, Atsuo; Zambirinis, Constantinos P; Pansari, Mridul; Rendon, Mauricio; Tippens, Daniel; Hundeyin, Mautin; Mani, Vishnu R; Hajdu, Cristina; Engle, Dannielle; Miller, George

2016-04-14

Neoplastic pancreatic epithelial cells are believed to die through caspase 8-dependent apoptotic cell death, and chemotherapy is thought to promote tumour apoptosis. Conversely, cancer cells often disrupt apoptosis to survive. Another type of programmed cell death is necroptosis (programmed necrosis), but its role in pancreatic ductal adenocarcinoma (PDA) is unclear. There are many potential inducers of necroptosis in PDA, including ligation of tumour necrosis factor receptor 1 (TNFR1), CD95, TNF-related apoptosis-inducing ligand (TRAIL) receptors, Toll-like receptors, reactive oxygen species, and chemotherapeutic drugs. Here we report that the principal components of the necrosome, receptor-interacting protein (RIP)1 and RIP3, are highly expressed in PDA and are further upregulated by the chemotherapy drug gemcitabine. Blockade of the necrosome in vitro promoted cancer cell proliferation and induced an aggressive oncogenic phenotype. By contrast, in vivo deletion of RIP3 or inhibition of RIP1 protected against oncogenic progression in mice and was associated with the development of a highly immunogenic myeloid and T cell infiltrate. The immune-suppressive tumour microenvironment associated with intact RIP1/RIP3 signalling depended in part on necroptosis-induced expression of the chemokine attractant CXCL1, and CXCL1 blockade protected against PDA. Moreover, cytoplasmic SAP130 (a subunit of the histone deacetylase complex) was expressed in PDA in a RIP1/RIP3-dependent manner, and Mincle--its cognate receptor--was upregulated in tumour-infiltrating myeloid cells. Ligation of Mincle by SAP130 promoted oncogenesis, whereas deletion of Mincle protected against oncogenesis and phenocopied the immunogenic reprogramming of the tumour microenvironment that was induced by RIP3 deletion. Cellular depletion suggested that whereas inhibitory macrophages promote tumorigenesis in PDA, they lose their immune-suppressive effects when RIP3 or Mincle is deleted. Accordingly, T cells, which are not protective against PDA progression in mice with intact RIP3 or Mincle signalling, are reprogrammed into indispensable mediators of anti-tumour immunity in the absence of RIP3 or Mincle. Our work describes parallel networks of necroptosis-induced CXCL1 and Mincle signalling that promote macrophage-induced adaptive immune suppression and thereby enable PDA progression.

Epstein-Barr Virus nuclear antigen 1 (EBNA1) confers resistance to apoptosis in EBV-positive B-lymphoma cells through up-regulation of survivin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu Jie; Murakami, Masanao; Verma, Subhash C.

Resistance to apoptosis is an important component of the overall mechanism which drives the tumorigenic process. EBV is a ubiquitous human gamma-herpesvirus which preferentially establishes latent infection in viral infected B-lymphocytes. EBNA1 is typically expressed in most forms of EBV-positive malignancies and is important for replication of the latent episome in concert with replication of the host cells. Here, we investigate the effects of EBNA1 on survivin up-regulation in EBV-infected human B-lymphoma cells. We present evidence which demonstrates that EBNA1 forms a complex with Sp1 or Sp1-like proteins bound to their cis-element at the survivin promoter. This enhances the activitymore » of the complex and up-regulates survivin. Knockdown of survivin and EBNA1 showed enhanced apoptosis in infected cells and thus supports a role for EBNA1 in suppressing apoptosis in EBV-infected cells. Here, we suggest that EBV encoded EBNA1 can contribute to the oncogenic process by up-regulating the apoptosis suppressor protein, survivin in EBV-associated B-lymphoma cells.« less

Cell Cycle Deregulation in the Neurons of Alzheimer’s Disease

PubMed Central

Moh, Calvin; Kubiak, Jacek Z.; Bajic, Vladan P.; Zhu, Xiongwei; Smith, Mark A.

2018-01-01

The cell cycle consists of four main phases: G1, S, G2, and M. Most cells undergo these cycles up to 40–60 times in their life. However, neurons remain in a nondividing, nonreplicating phase, G0. Neurons initiate but do not complete cell division, eventually entering apoptosis. Research has suggested that like cancer, Alzheimer’s disease (AD) involves dysfunction in neuronal cell cycle reentry, leading to the development of the two-hit hypothesis of AD. The first hit is abnormal cell cycle reentry, which typically results in neuronal apoptosis and prevention of AD. However, with the second hit of chronic oxidative damage preventing apoptosis, neurons gain “immortality” analogous to tumor cells. Once both of these hits are activated, AD can develop and produce senile plaques and neurofibrillary tangles throughout brain tissue. In this review, we propose a mechanism for neuronal cell cycle reentry and the development of AD. PMID:21630160

A Novel Bcl-x Isoform Connected to the T Cell Receptor Regulates Apoptosis in T Cells

PubMed Central

Yang, Xiao-Feng; Weber, Georg F.

2014-01-01

Summary We define a novel Bcl-x isoform, Bcl-xγ, that is generated by alternative splicing and characterized by a unique 47 amino acid C-terminus. Bcl-xγ is expressed primarily in thymocytes, where it may depend on an interaction between the TCR and host MHC products, and in mature T cells, where its expression is associated with ligation of the T cell receptor. Overexpression of Bcl-xγ in T cells inhibits activation-induced apoptosis; inhibition of Bcl-xγ, after stable expression of Bcl-xγ antisense cDNA, enhances activation-induced apoptosis. In contrast to other Bcl-x isoforms, cells that fail to express Bcl-xγ after CD3 ligation undergo programmed cell death, while activated T cells that express Bcl-xγ are spared. Identification of Bcl-xγ helps provide amolecular explanation of T cell activation and death after antigen engagement. PMID:9390687

Identification of the key pathway of oxazolinoanthracyclines mechanism of action in cells derived from human solid tumors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Denel-Bobrowska, Marta, E-mail: mdenel@biol.uni.lo

Oxazolinodoxorubicin (O-DOX) and oxazolinodaunorubicin (O-DAU) are novel anthracycline derivatives with a modified daunosamine moiety. In the present study, we evaluated the cytotoxicities, genotoxicities and abilities of O-DOX and O-DAU to induce apoptosis in cancer cell lines (SKOV-3; A549; HepG2), and compared the results with their parent drugs. We assessed antiproliferative activity by MTT assay. We evaluated apoptosis-inducing ability by double-staining with fluorescent probes (Hoechst 33258/propidium iodide), and by determining expression levels of genes involved in programmed cell death by reverse transcription-polymerase chain reaction. Genotoxicities of the compounds were tested by comet assays. Oxazolinoanthracyclines demonstrated high anti-tumor activity. O-DOX had significantlymore » higher cytotoxicity, apoptosis-inducing ability, and genotoxicity compared with parental doxorubicin (DOX) in all tested conditions, while O-DAU activity differed among cell lines. The mechanism of oxazoline analog action appeared to involve the mitochondrial pathway of programmed cell death. These results provide further information about oxazoline derivatives of commonly used anthracycline chemotherapy agents. O-DOX and O-DAU have the ability to induce apoptosis in tumor cells. - Highlights: • Substituted amino group increased the anticancer activity of anthracyclines. • Mitochondrial apoptotic pathway seems to be involved in the mechanism of action. • Favorable biological properties of oxazoline derivatives were confirmed.« less

ATF4 induction through an atypical integrated stress response to ONC201 triggers p53-independent apoptosis in hematological malignancies

PubMed Central

Ishizawa, Jo; Kojima, Kensuke; Chachad, Dhruv; Ruvolo, Peter; Ruvolo, Vivian; Jacamo, Rodrigo O.; Borthakur, Gautam; Mu, Hong; Zeng, Zhihong; Tabe, Yoko; Allen, Joshua E.; Wang, Zhiqiang; Ma, Wencai; Lee, Hans C.; Orlowski, Robert; Sarbassov, Dos D.; Lorenzi, Philip L.; Huang, Xuelin; Neelapu, Sattva S.; McDonnell, Timothy; Miranda, Roberto N.; Wang, Michael; Kantarjian, Hagop; Konopleva, Marina; Davis, R. Eric.; Andreeff, Michael

2016-01-01

The clinical challenge posed by p53 abnormalities in hematological malignancies requires therapeutic strategies other than standard genotoxic chemotherapies. ONC201 is a first-in-class small molecule that activates p53-independent apoptosis, has a benign safety profile, and is in early clinical trials. We found that ONC201 caused p53-independent apoptosis and cell cycle arrest in cell lines and in mantle cell lymphoma (MCL) and acute myeloid leukemia (AML) samples from patients; these included samples from patients with genetic abnormalities associated with poor prognosis or cells that had developed resistance to the nongenotoxic agents ibrutinib and bortezomib. Moreover, ONC201 caused apoptosis in stem and progenitor AML cells and abrogated the engraftment of leukemic stem cells in mice while sparing normal bone marrow cells. ONC201 caused changes in gene expression similar to those caused by the unfolded protein response (UPR) and integrated stress responses (ISRs), which increase the translation of the transcription factor ATF4 through an increase in the phosphorylation of the translation initiation factor eIF2α. However, unlike the UPR and ISR, the increase in ATF4 abundance in ONC201-treated hematopoietic cells promoted apoptosis and did not depend on increased phosphorylation of eIF2α. ONC201 also inhibited mammalian target of rapamycin complex 1 (mTORC1) signaling, likely through ATF4-mediated induction of the mTORC1 inhibitor DDIT4. Overexpression of BCL-2 protected against ONC201-induced apoptosis, and the combination of ONC201 and the BCL-2 antagonist ABT-199 synergistically increased apoptosis. Thus, our results suggest that by inducing an atypical ISR and p53-independent apoptosis, ONC201 has clinical potential in hematological malignancies. PMID:26884599

Apoptosis and necrosis in the liver.

PubMed

Guicciardi, Maria Eugenia; Malhi, Harmeet; Mott, Justin L; Gores, Gregory J

2013-04-01

Because of its unique function and anatomical location, the liver is exposed to a multitude of toxins and xenobiotics, including medications and alcohol, as well as to infection by hepatotropic viruses, and therefore, is highly susceptible to tissue injury. Cell death in the liver occurs mainly by apoptosis or necrosis, with apoptosis also being the physiologic route to eliminate damaged or infected cells and to maintain tissue homeostasis. Liver cells, especially hepatocytes and cholangiocytes, are particularly susceptible to death receptor-mediated apoptosis, given the ubiquitous expression of the death receptors in the organ. In a quite unique way, death receptor-induced apoptosis in these cells is mediated by both mitochondrial and lysosomal permeabilization. Signaling between the endoplasmic reticulum and the mitochondria promotes hepatocyte apoptosis in response to excessive free fatty acid generation during the metabolic syndrome. These cell death pathways are partially regulated by microRNAs. Necrosis in the liver is generally associated with acute injury (i.e., ischemia/reperfusion injury) and has been long considered an unregulated process. Recently, a new form of "programmed" necrosis (named necroptosis) has been described: the role of necroptosis in the liver has yet to be explored. However, the minimal expression of a key player in this process in the liver suggests this form of cell death may be uncommon in liver diseases. Because apoptosis is a key feature of so many diseases of the liver, therapeutic modulation of liver cell death holds promise. An updated overview of these concepts is given in this article.

Herbal medicine as inducers of apoptosis in cancer treatment.

PubMed

Safarzadeh, Elham; Sandoghchian Shotorbani, Siamak; Baradaran, Behzad

2014-10-01

Cancer is uncontrolled growth of abnormal cells in the body. Nowadays, cancer is considered as a human tragedy and one of the most prevalent diseases in the wide, and its mortality resulting from cancer is being increased. It seems necessary to identify new strategies to prevent and treat such a deadly disease. Control survival and death of cancerous cell are important strategies in the management and therapy of cancer. Anticancer agents should kill the cancerous cell with the minimal side effect on normal cells that is possible through the induction of apoptosis. Apoptosis is known as programmed cell death in both normal and damaged tissues. This process includes some morphologically changes in cells such as rapid condensation and budding of the cell, formation of membrane-enclosed apoptotic bodies with well-preserved organelles. Induction of apoptosis is one of the most important markers of cytotoxic antitumor agents. Some natural compounds including plants induce apoptotic pathways that are blocked in cancer cells through various mechanisms in cancer cells. Multiple surveys reported that people with cancer commonly use herbs or herbal products. Vinca Alkaloids, Texans, podo phyllotoxin, Camptothecins have been clinically used as Plant derived anticancer agents. The present review summarizes the literature published so far regarding herbal medicine used as inducers of apoptosis in cancer.

Cultured Chinese hamster cells undergo apoptosis after exposure to cold but nonfreezing temperatures.

PubMed

Nagle, W A; Soloff, B L; Moss, A J; Henle, K J

1990-08-01

Cultured Chinese hamster V79 fibroblast cells at the transition from logarithmic to stationary growth have been shown to undergo apoptosis (programmed cell death) after cold shock [B. L. Soloff, W. A. Nagle, A. J. Moss, Jr., K. J. Henle, and J. T. Crawford, Biochem. Biophys. Res. Commun. 145, 876-883 (1987)]. In this report, we show that about 95% of the cell population was susceptible to cold-induced apoptosis, and the amount of cell killing was dependent on the duration of hypothermia. Cells treated for 0-90 min at 0 degrees C exhibited an exponential survival curve with a D0 of 32 min; thus, even short exposures to the cold (e.g., 5 min) produced measurable cell killing. The cold-induced injury was not produced by freezing, because similar results were observed at 6 degrees C, and cell killing was not influenced by the cryoprotective agent dimethyl sulfoxide. Cold-induced apoptosis was inhibited by rewarming at 23 degrees C, compared to 37 degrees C, by inhibitors of macromolecular synthesis, such as cycloheximide, and by 0.8 mM zinc sulfate. The results suggest that apoptosis represents a new manifestation of cell injury after brief exposure to 0-6 degrees C hypothermia.

Microbiota promotes systemic T-cell survival through suppression of an apoptotic factor

PubMed Central

Petersen, Charisse; Novis, Camille L.; Kubinak, Jason L.; Bell, Rickesha; Stephens, W. Zac; Lane, Thomas E.; Fujinami, Robert S.; Bosque, Alberto; O’Connell, Ryan M.; Round, June L.

2017-01-01

Symbiotic microbes impact the severity of a variety of diseases through regulation of T-cell development. However, little is known regarding the molecular mechanisms by which this is accomplished. Here we report that a secreted factor, Erdr1, is regulated by the microbiota to control T-cell apoptosis. Erdr1 expression was identified by transcriptome analysis to be elevated in splenic T cells from germfree and antibiotic-treated mice. Suppression of Erdr1 depends on detection of circulating microbial products by Toll-like receptors on T cells, and this regulation is conserved in human T cells. Erdr1 was found to function as an autocrine factor to induce apoptosis through caspase 3. Consistent with elevated levels of Erdr1, germfree mice have increased splenic T-cell apoptosis. RNA sequencing of Erdr1-overexpressing cells identified the up-regulation of genes involved in Fas-mediated cell death, and Erdr1 fails to induce apoptosis in Fas-deficient cells. Importantly, forced changes in Erdr1 expression levels dictate the survival of auto-reactive T cells and the clinical outcome of neuro-inflammatory autoimmune disease. Cellular survival is a fundamental feature regulating appropriate immune responses. We have identified a mechanism whereby the host integrates signals from the microbiota to control T-cell apoptosis, making regulation of Erdr1 a potential therapeutic target for autoimmune disease. PMID:28487480

The Selective Progesterone Receptor Modulator CDB4124 Inhibits Proliferation and Induces Apoptosis in Uterine Leiomyoma Cells

PubMed Central

Luo, Xia; Yin, Ping; Coon V., John S.; Cheng, You-Hong; Wiehle, Ronald D.; Bulun, Serdar E.

2009-01-01

Objective To evaluate the effects of selective progesterone receptor modulator CDB4124 on cell proliferation and apoptosis in cultured human uterine leiomyoma smooth muscle (LSM) cells and control myometrial smooth muscle (MSM) cells in matched uteri. Design Laboratory research. Setting Academic medical center. Patient(s) Premenopausal women (n=12) undergoing hysterectomy for leiomyoma-related symptoms. Intervention(s) Treatment of primary LSM and MSM cells with CDB4124 (10-8-10-6M) or vehicle for 24, 48 or 72 hours. Main Outcome Measure(s) Western blot for protein expression of proliferating cell nuclear antigen (PCNA), cleaved poly-adenosine 5’-diphosphate-ribose polymerase (PARP), Bcl-2 and Krüppel-like transcription factor 11 (KLF11); MTT assay to evaluate viable cell numbers; and real-time polymerase chain reaction to quantify mRNA levels. Result(s) Treatment with CDB4124 significantly decreased levels of the proliferation marker PCNA, the number of viable LSM cells, and the anti-apoptotic protein Bcl-2. On the other hand, treatment with CDB4124 increased levels of the apoptosis marker cleaved PARP and the tumor suppressor KLF11 in a dose- and time-dependent manner in LSM cells. In matched MSM cells, however, CDB4124 did not affect cell proliferation or apoptosis. Conclusion(s) CDB4124 selectively inhibits proliferation and induces apoptosis in LSM but not in MSM cells. PMID:20056218

Silibinin-induced glioma cell apoptosis by PI3K-mediated but Akt-independent downregulation of FoxM1 expression.

PubMed

Zhang, Mingjie; Liu, Yunhui; Gao, Yun; Li, Shaoyi

2015-10-15

The oncogenic transcription factor Forkhead box M1 (FoxM1) is overexpressed in many human tumors, including glioma. As a critical regulator of the cell cycle and apoptosis-related genes, FoxM1 is a potential therapeutic target against human malignant glioma. Silibinin, a flavonoid isolated from Silybum marianum, dose-dependently reduced glioma cell proliferation, promoted apoptosis, and downregulated FoxM1 expression. Knockdown of FoxM1 by small hairpin RNA (shRNA) transfection also promoted glioma cell apoptosis and augmented the antiproliferative and pro-apoptotic properties of silibinin. Moreover, silibinin increased caspase-3 activation, upregulated pro-apoptotic Bax, and suppressed anti-apoptotic Bcl-2 expression, effects enhanced by FoxM1 knockdown. Silibinin treatment suppressed U87 cell PI3K phospho-activation, and simultaneous silibinin exposure, FoxM1 knockdown, and PI3K inhibition additively increased U87 cell apoptosis. Furthermore, PI3K inhibition reduced FoxM1 expression. Akt activity was also suppressed by FoxM1 downregulation but Akt inhibition did not alter FoxM1 expression. Thus, silibinin likely inhibited glioma cell proliferation and induced apoptosis through inactivation of PI3K and FoxM1, leading to activation of the mitochondrial apoptotic pathway. FoxM1 may be a novel target for chemotherapy against human glioma. Copyright © 2015 Elsevier B.V. All rights reserved.

Gene Expression in Pre-MBT Embryos and Activation of Maternally-Inherited Program of Apoptosis to be Executed at around MBT as a Fail-Safe Mechanism in Xenopus Early Embryogenesis

PubMed Central

Shiokawa, Koichiro; Aso, Mai; Kondo, Takeshi; Uchiyama, Hiroaki; Kuroyanagi, Shinsaku; Takai, Jun-Ichi; Takahashi, Senji; Kajitani, Masayuki; Kaito, Chikara; Sekimizu, Kazuhisa; Takayama, Eiji; Igarashi, Kazuei; Hara, Hiroshi

2008-01-01

S-adenosylmethionine decarboxylase (SAMDC) is an enzyme which converts S-adenosylmethione (SAM), a methyl donor, to decarboxylated SAM (dcSAM), an aminopropyl donor for polyamine biosynthesis. In our studies on gene expression control in Xenopus early embryogenesis, we cloned the mRNA for Xenopus SAMDC, and overexpressed the enzyme by microinjecting its mRNA into Xenopus fertilized eggs. In the mRNA-injected embryos, the level of SAMDC was enormously increased, the SAM was exhausted, and protein synthesis was greatly inhibited, but cellular polyamine content did not change appreciably. SAMDC-overexpressed embryos cleaved and developed normally up to the early blastula stage, but at the midblastula stage, or the stage of midblastula transition (MBT), all the embryos were dissociated into cells, and destroyed due to execution of apoptosis. During cleavage SAMDC-overexpressed embryos transcribed caspase-8 gene, and this was followed by activation of caspase-9. When we overexpressed p53 mRNA in fertilized eggs, similar apoptosis took place at MBT, but in this case, transcription of caspase-8 did not occur, however activation of caspase-9 took place. Apoptosis induced by SAMDC-overexpression was completely suppressed by Bcl-2, whereas apoptosis induced by p53 overexpression or treatments with other toxic agents was only partially rescued. When we injected SAMDC mRNA into only one blastomere of 8- to 32-celled embryos, descendant cells of the mRNA-injected blastomere were segregated into the blastocoel and underwent apoptosis within the blastocoel, although such embryos continued to develop and became tadpoles with various extents of anomaly, reflecting the developmental fate of the eliminated cells. Thus, embryonic cells appear to check themselves at MBT and if physiologically severely-damaged cells occur, they are eliminated from the embryo by activation and execution of the maternally-inherited program of apoptosis. We assume that the apoptosis executed at MBT is a “fail-safe” mechanism of early development to save the embryo from accidental damages that take place during cleavage. PMID:19787085

The suppression of apoptosis by α-herpesvirus

PubMed Central

You, Yu; Cheng, An-Chun; Wang, Ming-Shu; Jia, Ren-Yong; Sun, Kun-Feng; Yang, Qiao; Wu, Ying; Zhu, Dekang; Chen, Shun; Liu, Ma-Feng; Zhao, Xin-Xin; Chen, Xiao-Yue

2017-01-01

Apoptosis, an important innate immune mechanism that eliminates pathogen-infected cells, is primarily triggered by two signalling pathways: the death receptor pathway and the mitochondria-mediated pathway. However, many viruses have evolved various strategies to suppress apoptosis by encoding anti-apoptotic factors or regulating apoptotic signalling pathways, which promote viral propagation and evasion of the host defence. During its life cycle, α-herpesvirus utilizes an elegant multifarious anti-apoptotic strategy to suppress programmed cell death. This progress article primarily focuses on the current understanding of the apoptosis-inhibition mechanisms of α-herpesvirus anti-apoptotic genes and their expression products and discusses future directions, including how the anti-apoptotic function of herpesvirus could be targeted therapeutically. PMID:28406478

Primary hepatocytes and their cultures in liver apoptosis research

PubMed Central

Vinken, Mathieu; Maes, Michaël; Oliveira, André G.; Cogliati, Bruno; Marques, Pedro E.; Menezes, Gustavo B.; Dagli, Maria Lúcia Zaidan; Vanhaecke, Tamara; Rogiers, Vera

2014-01-01

Apoptosis not only plays a key role in physiological demise of defunct hepatocytes, but is also associated with a plethora of acute and chronic liver diseases as well as with hepatotoxicity. The present paper focuses on the modelling of this mode of programmed cell death in primary hepatocyte cultures. Particular attention is paid to the activation of spontaneous apoptosis during the isolation of hepatocytes from the liver, its progressive manifestation upon the subsequent establishment of cell cultures and simultaneously to strategies to counteract this deleterious process. In addition, currently applied approaches to experimentally induce controlled apoptosis in this in vitro setting for mechanistic research purposes and thereby its detection using relevant biomarkers are reviewed. PMID:24013573

TNF-induced necroptosis requires the plasma membrane localization of the MLKL protein | Center for Cancer Research

Cancer.gov

The cell signaling protein tumor necrosis factor (TNF), produced by white blood cells, promotes inflammation and immunity processes such as fever and is involved in tumorigenesis and apoptosis (programmed cell death). However, dysregulation of TNF can also lead to another form of programmed cell death called necroptosis, which is characterized by a rise in intracellular Ca2+, generation of reactive oxygen species (ROS), intracellular acidity, depletion of ATP, and, eventually, plasma membrane rupture. TNF-induced necroptosis has been associated with a wide variety of diseases including neurodegenerative diseases, major depression, rheumatoid arthritis, and cancer. Whereas the signaling mechanisms underlying TNF-induced apoptosis have largely been determined, the events precipitating in TNF-initiated necroptosis are still unknown.

Dimethyloxalylglycine may be enhance the capacity of neural-like cells in treatment of Alzheimer disease.

PubMed

Ghasemi Moravej, Fahimeh; Vahabian, Mehrangiz; Soleimani Asl, Sara

2016-06-01

Although using differentiated stem cells is the best proposed option for the treatment of Alzheimer disease (AD), an efficient differentiation and cell therapy require enhanced cell survival and homing and decreased apoptosis. It seems that hypoxia preconditioning via Dimethyloxalylglycine (DMOG) may increase the capacity of MSC to induce neural like stem cells (NSCs). Furthermore, it can likely improve the viability of NSCs when transplanted into the brain of AD rats. © 2016 International Federation for Cell Biology.

2-O-Methylmagnolol upregulates the long non-coding RNA, GAS5, and enhances apoptosis in skin cancer cells.

PubMed

Wang, Tong-Hong; Chan, Chieh-Wen; Fang, Jia-You; Shih, Ya-Min; Liu, Yi-Wen; Wang, Tzu-Chien V; Chen, Chi-Yuan

2017-03-02

Magnolol, a hydroxylated biphenol compound isolated from the bark of Magnolia officinalis, has been shown to exhibit anti-proliferative effect in various cancer cells, including skin cancer cells. Methoxylation of magnolol appears to improve its anti-inflammatory activity, yet the effect of this modification on the agent's antitumor activity remains unknown. In this work, we report that 2-O-methylmagnolol (MM1) displays improved antitumor activity against skin cancer cells compared to magnolol both in vitro and in vivo. The increased antitumor activity of MM1 appears to correlate with its increased ability to induce apoptosis. DNA microarray and network pathway analyses suggest that MM1 affects certain key factors involved in regulating apoptosis and programmed cell death. Interestingly, the level of the long non-coding (lnc) RNA of growth arrest-specific 5 (GAS5) was increased in MM1-treated cells, and inhibition of lncRNA GAS5 inhibited MM1-induced apoptosis. Conversely, overexpression of lncRNA GAS5 inhibited cell proliferation and promoted cell apoptosis in skin cancer cells. The expression of lncRNA GAS5 in the skin cancer tissues was found to be lower than that in the adjacent normal tissues in a majority of patients. Taken together, our findings suggest that MM1 has improved antitumor activity in skin cancer cells, and that this is due, at least in part, to the upregulation of lncRNA GAS5 and the enhancement of apoptosis.

Down-regulation of PKHD1 induces cell apoptosis through PI3K and NF-{kappa}B pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sun, Liping; Wang, Shixuan; Hu, Chaofeng

2011-04-15

Mutations in PKHD1 (polycystic kidney and hepatic disease gene 1) gene cause the autosomal recessive polycystic kidney disease (ARPKD). Fibrocystin/polyductin (FPC), encoded by PKHD1, is a membrane-associated receptor-like protein. Although it is widely accepted that cystogenesis is mostly due to aberrant cell proliferation and apoptosis, it is still unclear how apoptosis is regulated. The aim of this study is to analyze the relationship among apoptosis, phosphatidylinositol 3-kinase (PI3K)/Akt and nuclear factor {kappa}B (NF-{kappa}B) in FPC knockdown kidney cells. We show that PKHD1-silenced HEK293 cells demonstrate a higher PI3K/Akt activity. Selective inhibition of PI3K/Akt using LY294002 or wortmannin in these cellsmore » increases serum starvation-induced HEK293 cell apoptosis with a concomitant decrease in cell proliferation and higher caspase-3 activity. PI3K/Akt inhibition also leads to increased NF-{kappa}B activity in these cells. We conclude that the PI3K/Akt pathway is involved in apoptotic function in PKHD1-silenced cells, and PI3K/Akt inhibition correlates with upregulation of NF-{kappa}B activity. These observations provide a potential platform for determining FPC function and therapeutic investigation of ARPKD.« less

Inhibiting Polo-like kinase 1 causes growth reduction and apoptosis in pediatric acute lymphoblastic leukemia cells

PubMed Central

Hartsink-Segers, Stefanie A.; Exalto, Carla; Allen, Matthew; Williamson, Daniel; Clifford, Steven C.; Horstmann, Martin; Caron, Huib N.; Pieters, Rob; Den Boer, Monique L.

2013-01-01

This study investigated Polo-like kinase 1, a mitotic regulator often over-expressed in solid tumors and adult hematopoietic malignancies, as a potential new target in the treatment of pediatric acute lymphoblastic leukemia. Polo-like kinase 1 protein and Thr210 phosphorylation levels were higher in pediatric acute lymphoblastic leukemia (n=172) than in normal bone marrow mononuclear cells (n=10) (P<0.0001). High Polo-like kinase 1 protein phosphorylation, but not expression, was associated with a lower probability of event-free survival (P=0.042) and was a borderline significant prognostic factor (P=0.065) in a multivariate analysis including age and initial white blood cell count. Polo-like kinase 1 was necessary for leukemic cell survival, since short hairpin-mediated Polo-like kinase 1 knockdown in acute lymphoblastic leukemia cell lines inhibited cell proliferation by G2/M cell cycle arrest and induced apoptosis through caspase-3 and poly (ADP-ribose) polymerase cleavage. Primary patient cells with a high Polo-like kinase 1 protein expression were sensitive to the Polo-like kinase 1-specific inhibitor NMS-P937 in vitro, whereas cells with a low expression and normal bone marrow cells were resistant. This sensitivity was likely not caused by Polo-like kinase 1 mutations, since only one new mutation (Ser335Arg) was found by 454-sequencing of 38 pediatric acute lymphoblastic leukemia cases. This mutation did not affect Polo-like kinase 1 expression or NMS-P937 sensitivity. Together, these results indicate a pivotal role for Polo-like kinase 1 in pediatric acute lymphoblastic leukemia and show potential for Polo-like kinase 1-inhibiting drugs as an addition to current treatment strategies for cases expressing high Polo-like kinase 1 levels. PMID:23753023

Expression of insulin-like growth factor-1 and insulin-like growth factor-1 receptors in EL4 lymphoma cells overexpressing growth hormone.

PubMed

Weigent, Douglas A; Arnold, Robyn E

2005-03-01

Almost all of the previous studies with growth hormone (GH) have been done with exogenously supplied GH and, therefore, involve actions of the hormone through its receptor. However, the actions of endogenous or lymphocyte GH are still unclear. In a previous study, we showed that overexpression of GH (GHo) in a lymphoid cell line resulted in protection of the cells to apoptosis mediated by nitric oxide (NO). In the present study, we show that the protection from apoptosis could be transferred to control cells with culture fluids obtained from GHo cells and blocked by antibodies to the insulin-like growth factor-1 (IGF-1) or antibodies to the IGF-1-receptor (IGF-1R). Northern and Western blot analysis detected significantly higher levels of IGF-1 in cells overexpressing GH. An increase in the expression of the IGF-1R in GHo cells was also detected by Western blot analysis, (125)I-IGF-1 binding and analysis of IGF-1R promoter luciferase constructs. Transfection of GHo cells with a dominant negative IGF-1R mutant construct blocked the generation of NO and activation of Akt seen in GHo cells compared to vector alone control EL4 cells. The results suggest that one of the consequences of the overexpression of GH, in cells lacking the GH receptor, is an increase in the expression of IGF-1 and the IGF-1R which mediate the protection of EL4 lymphoma cells from apoptosis.

Novel TRAIL sensitizer Taraxacum officinale F.H. Wigg enhances TRAIL-induced apoptosis in Huh7 cells.

PubMed

Yoon, Ji-Yong; Cho, Hyun-Soo; Lee, Jeong-Ju; Lee, Hyo-Jung; Jun, Soo Young; Lee, Jae-Hye; Song, Hyuk-Hwan; Choi, SangHo; Saloura, Vassiliki; Park, Choon Gil; Kim, Cheol-Hee; Kim, Nam-Soon

2016-04-01

TRAIL (TNF-related apoptosis inducing ligand) is a promising anti-cancer drug target that selectively induces apoptosis in cancer cells. However, many cancer cells are resistant to TRAIL-induced apoptosis. Therefore, reversing TRAIL resistance is an important step for the development of effective TRAIL-based anti-cancer therapies. We previously reported that knockdown of the TOR signaling pathway regulator-like (TIPRL) protein caused TRAIL-induced apoptosis by activation of the MKK7-c-Jun N-terminal Kinase (JNK) pathway through disruption of the MKK7-TIPRL interaction. Here, we identified Taraxacum officinale F.H. Wigg (TO) as a novel TRAIL sensitizer from a set of 500 natural products using an ELISA system and validated its activity by GST pull-down analysis. Furthermore, combination treatment of Huh7 cells with TRAIL and TO resulted in TRAIL-induced apoptosis mediated through inhibition of the MKK7-TIPRL interaction and subsequent activation of MKK7-JNK phosphorylation. Interestingly, HPLC analysis identified chicoric acid as a major component of the TO extract, and combination treatment with chicoric acid and TRAIL induced TRAIL-induced cell apoptosis via JNK activation due to inhibition of the MKK7-TIPRL interaction. Our results suggest that TO plays an important role in TRAIL-induced apoptosis, and further functional studies are warranted to confirm the importance of TO as a novel TRAIL sensitizer for cancer therapy. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

Functional evaluation of the role of inhibitor of apoptosis proteins in chronic lymphocytic leukemia.

PubMed

Schliep, Stefan; Decker, Thomas; Schneller, Folker; Wagner, Hermann; Häcker, Georg

2004-06-01

The slow accumulation of malignant cells in chronic lymphocytic leukemia (CLL) suggests a defect in the induction of apoptosis in these cells. Previous studies have found sporadic alterations in the apoptotic apparatus in CLL cells, but a widespread defect has not been detected until now. A crucial checkpoint in the progression of apoptosis is the activity of inhibitor of apoptosis proteins (IAP) that control the activity of caspases upon the release of cytochrome c from mitochondria. The aim of this study was to evaluate the role of IAP in the regulation of apoptosis in CLL cells. Lysates from CLL cells were prepared, and the regular function of components of the cytochrome c-dependent caspase-activating machinery (the apoptosome) was investigated. The effect of IAP in caspase-inhibition was tested using a peptide derived from the mitochondrial IAP antagonist Smac/DIABLO. Regulation of expression as well as inhibitory function of the X-linked IAP (XIAP) by cytokines was analyzed. The apoptosome was found to be structurally and functionally competent in CLL. XIAP expression was enhanced by culture in the presence of cytokines. Smac/DIABLO was easily detectable in CLL cells and was released into the cytosol during apoptosis. No inhibitory effect of IAP was detected in CLL, irrespective of XIAP levels and culture conditions. Although XIAP is present in CLL cells and is up-regulated in conditions where apoptosis is prevented, no caspase-inhibiting anti-apoptotic effect of IAP is detectable. This is likely due to the high expression of Smac/DIABLO in CLL cells that is sufficient to overcome the caspase-inhibiting effect of IAP.

Regulation of Tumor Progression by Programmed Necrosis

PubMed Central

Jeon, Hyun Min; Jeong, Eui Kyong; Lee, Yig Ji; Kim, Cho Hee; Park, Hye Gyeong

2018-01-01

Rapidly growing malignant tumors frequently encounter hypoxia and nutrient (e.g., glucose) deprivation, which occurs because of insufficient blood supply. This results in necrotic cell death in the core region of solid tumors. Necrotic cells release their cellular cytoplasmic contents into the extracellular space, such as high mobility group box 1 (HMGB1), which is a nonhistone nuclear protein, but acts as a proinflammatory and tumor-promoting cytokine when released by necrotic cells. These released molecules recruit immune and inflammatory cells, which exert tumor-promoting activity by inducing angiogenesis, proliferation, and invasion. Development of a necrotic core in cancer patients is also associated with poor prognosis. Conventionally, necrosis has been thought of as an unregulated process, unlike programmed cell death processes like apoptosis and autophagy. Recently, necrosis has been recognized as a programmed cell death, encompassing processes such as oncosis, necroptosis, and others. Metabolic stress-induced necrosis and its regulatory mechanisms have been poorly investigated until recently. Snail and Dlx-2, EMT-inducing transcription factors, are responsible for metabolic stress-induced necrosis in tumors. Snail and Dlx-2 contribute to tumor progression by promoting necrosis and inducing EMT and oncogenic metabolism. Oncogenic metabolism has been shown to play a role(s) in initiating necrosis. Here, we discuss the molecular mechanisms underlying metabolic stress-induced programmed necrosis that promote tumor progression and aggressiveness. PMID:29636841

Expression of Progesterone Receptor Membrane Component-2 Within the Immature Rat Ovary and Its Role in Regulating Mitosis and Apoptosis of Spontaneously Immortalized Granulosa Cells1

PubMed Central

Griffin, Daniel; Liu, Xiufang; Pru, Cindy; Pru, James K.; Peluso, John J.

2014-01-01

ABSTRACT Progesterone receptor membrane component 2 (Pgrmc2) mRNA was detected in the immature rat ovary. By 48 h after eCG, Pgrmc2 mRNA levels decreased by 40% and were maintained at 48 h post-hCG. Immunohistochemical studies detected PGRMC2 in oocytes and ovarian surface epithelial, interstitial, thecal, granulosa, and luteal cells. PGRMC2 was also present in spontaneously immortalized granulosa cells, localizing to the cytoplasm of interphase cells and apparently to the mitotic spindle of cells in metaphase. Interestingly, PGRMC2 levels appeared to decrease during the G1 stage of the cell cycle. Moreover, overexpression of PGRMC2 suppressed entry into the cell cycle, possibly by binding the p58 form of cyclin dependent kinase 11b. Conversely, Pgrmc2 small interfering RNA (siRNA) treatment increased the percentage of cells in G1 and M stage but did not increase the number of cells, which was likely due to an increase in apoptosis. Depleting PGRMC2 did not inhibit cellular 3H-progesterone binding, but attenuated the ability of progesterone to suppress mitosis and apoptosis. Taken together these studies suggest that PGRMC2 affects granulosa cell mitosis by acting at two specific stages of the cell cycle. First, PGRMC2 regulates the progression from the G0 into the G1 stage of the cell cycle. Second, PGRMC2 appears to localize to the mitotic spindle, where it likely promotes the final stages of mitosis. Finally, siRNA knockdown studies indicate that PGRMC2 is required for progesterone to slow the rate of granulosa cell mitosis and apoptosis. These findings support a role for PGRMC2 in ovarian follicle development. PMID:24990806

Chk2 mediates RITA-induced apoptosis.

PubMed

de Lange, J; Verlaan-de Vries, M; Teunisse, A F A S; Jochemsen, A G

2012-06-01

Reactivation of the p53 tumor-suppressor protein by small molecules like Nutlin-3 and RITA (reactivation of p53 and induction of tumor cell apoptosis) is a promising strategy for cancer therapy. The molecular mechanisms involved in the responses to RITA remain enigmatic. Several groups reported the induction of a p53-dependent DNA damage response. Furthermore, the existence of a p53-dependent S-phase checkpoint has been suggested, involving the checkpoint kinase Chk1. We have recently shown synergistic induction of apoptosis by RITA in combination with Nutlin-3, and we observed concomitant Chk2 phosphorylation. Therefore, we investigated whether Chk2 contributes to the cellular responses to RITA. Strikingly, the induction of apoptosis seemed entirely Chk2 dependent. Transcriptional activity of p53 in response to RITA required the presence of Chk2. A partial rescue of apoptosis observed in Noxa knockdown cells emphasized the relevance of p53 transcriptional activity for RITA-induced apoptosis. In addition, we observed an early p53- and Chk2-dependent block of DNA replication upon RITA treatment. Replicating cells seemed more prone to entering RITA-induced apoptosis. Furthermore, the RITA-induced DNA damage response, which was not a secondary effect of apoptosis induction, was strongly attenuated in cells lacking p53 or Chk2. In conclusion, we identified Chk2 as an essential mediator of the cellular responses to RITA.

Chk2 mediates RITA-induced apoptosis

PubMed Central

de Lange, J; Verlaan-de Vries, M; Teunisse, A F A S; Jochemsen, A G

2012-01-01

Reactivation of the p53 tumor-suppressor protein by small molecules like Nutlin-3 and RITA (reactivation of p53 and induction of tumor cell apoptosis) is a promising strategy for cancer therapy. The molecular mechanisms involved in the responses to RITA remain enigmatic. Several groups reported the induction of a p53-dependent DNA damage response. Furthermore, the existence of a p53-dependent S-phase checkpoint has been suggested, involving the checkpoint kinase Chk1. We have recently shown synergistic induction of apoptosis by RITA in combination with Nutlin-3, and we observed concomitant Chk2 phosphorylation. Therefore, we investigated whether Chk2 contributes to the cellular responses to RITA. Strikingly, the induction of apoptosis seemed entirely Chk2 dependent. Transcriptional activity of p53 in response to RITA required the presence of Chk2. A partial rescue of apoptosis observed in Noxa knockdown cells emphasized the relevance of p53 transcriptional activity for RITA-induced apoptosis. In addition, we observed an early p53- and Chk2-dependent block of DNA replication upon RITA treatment. Replicating cells seemed more prone to entering RITA-induced apoptosis. Furthermore, the RITA-induced DNA damage response, which was not a secondary effect of apoptosis induction, was strongly attenuated in cells lacking p53 or Chk2. In conclusion, we identified Chk2 as an essential mediator of the cellular responses to RITA. PMID:22158418

Calpain Determines the Propensity of Adult Hippocampal Neural Stem Cells to Autophagic Cell Death Following Insulin Withdrawal.

PubMed

Chung, Kyung Min; Park, Hyunhee; Jung, Seonghee; Ha, Shinwon; Yoo, Seung-Jun; Woo, Hanwoong; Lee, Hyang Ju; Kim, Seong Who; Kim, Eun-Kyoung; Moon, Cheil; Yu, Seong-Woon

2015-10-01

Programmed cell death (PCD) has significant effects on the function of neural stem cells (NSCs) during brain development and degeneration. We have previously reported that adult rat hippocampal neural stem (HCN) cells underwent autophagic cell death (ACD) rather than apoptosis following insulin withdrawal despite their intact apoptotic capabilities. Here, we report a switch in the mode of cell death in HCN cells with calpain as a critical determinant. In HCN cells, calpain 1 expression was barely detectable while calpain 2 was predominant. Inhibition of calpain in insulin-deprived HCN cells further augmented ACD. In contrast, expression of calpain 1 switched ACD to apoptosis. The proteasome inhibitor lactacystin blocked calpain 2 degradation and elevated the intracellular Ca(2+) concentration. In combination, these effects potentiated calpain activity and converted the mode of cell death to apoptosis. Our results indicate that low calpain activity, due to absence of calpain 1 and degradation of calpain 2, results in a preference for ACD over apoptosis in insulin-deprived HCN cells. On the other hand, conditions leading to high calpain activity completely switch the mode of cell death to apoptosis. This is the first report on the PCD mode switching mechanism in NSCs. The dynamic change in calpain activity through the proteasome-mediated modulation of the calpain and intracellular Ca(2+) levels may be the critical contributor to the demise of NSCs. Our findings provide a novel insight into the complex mechanisms interconnecting autophagy and apoptosis and their roles in the regulation of NSC death. © 2015 AlphaMed Press.

A mechanism of apigenin-induced apoptosis is potentially related to anti-angiogenesis and anti-migration in human hepatocellular carcinoma cells.

PubMed

Kim, Bo Ra; Jeon, Young Keul; Nam, Myeong Jin

2011-07-01

Apigenin (APG) has been shown to have a strong anti-cancer effect on various cancer models via a programmed cell death, apoptosis. However, the fundamental mechanisms of these effects are still unclear. In the present study, we examined the question of whether or not APG can inhibit proliferation of hepatocellular carcinoma (HCC), huh-7 cells, resulting in apoptosis. In APG-treated cells, we observed typical features of apoptosis. To identify the proteins related to APG-induced apoptosis, we performed two-dimensional electrophoresis analysis and identified differentially expressed proteins. Among these proteins, we focused on vimentin, which plays a physiological role, such as cell migration and adhesion. We validated expression of vimentin in both mRNA and protein levels, verifying its decrease. In addition, we observed that APG down-regulated the expression levels of type I collagen, which collaborated with vimentin in cell migration and decreased the releasing amounts of VEGF and MMP-8, which are closely relevant to angiogenic activity. Finally, we confirmed the decreased capacity of cell migration due to down-regulation of vimentin, type I collagen, VEGF, and MMP-8 induced by APG. Based on the overall results, we suggested that vimentin was potentially associated with APG-induced apoptosis, as a key regulator in angiogenesis and migration. Copyright © 2011 Elsevier Ltd. All rights reserved.

RIP1 and RIP3 complex regulates radiation-induced programmed necrosis in glioblastoma.

PubMed

Das, Arabinda; McDonald, Daniel G; Dixon-Mah, Yaenette N; Jacqmin, Dustin J; Samant, Vikram N; Vandergrift, William A; Lindhorst, Scott M; Cachia, David; Varma, Abhay K; Vanek, Kenneth N; Banik, Naren L; Jenrette, Joseph M; Raizer, Jeffery J; Giglio, Pierre; Patel, Sunil J

2016-06-01

Radiation-induced necrosis (RN) is a relatively common side effect of radiation therapy for glioblastoma. However, the molecular mechanisms involved and the ways RN mechanisms differ from regulated cell death (apoptosis) are not well understood. Here, we compare the molecular mechanism of cell death (apoptosis or necrosis) of C6 glioma cells in both in vitro and in vivo (C6 othotopically allograft) models in response to low and high doses of X-ray radiation. Lower radiation doses were used to induce apoptosis, while high-dose levels were chosen to induce radiation necrosis. Our results demonstrate that active caspase-8 in this complex I induces apoptosis in response to low-dose radiation and inhibits necrosis by cleaving RIP1 and RI. When activation of caspase-8 was reduced at high doses of X-ray radiation, the RIP1/RIP3 necrosome complex II is formed. These complexes induce necrosis through the caspase-3-independent pathway mediated by calpain, cathepsin B/D, and apoptosis-inducing factor (AIF). AIF has a dual role in apoptosis and necrosis. At high doses, AIF promotes chromatinolysis and necrosis by interacting with histone H2AX. In addition, NF-κB, STAT-3, and HIF-1 play a crucial role in radiation-induced inflammatory responses embedded in a complex inflammatory network. Analysis of inflammatory markers in matched plasma and cerebrospinal fluid (CSF) isolated from in vivo specimens demonstrated the upregulation of chemokines and cytokines during the necrosis phase. Using RIP1/RIP3 kinase specific inhibitors (Nec-1, GSK'872), we also establish that the RIP1-RIP3 complex regulates programmed necrosis after either high-dose radiation or TNF-α-induced necrosis requires RIP1 and RIP3 kinases. Overall, our data shed new light on the relationship between RIP1/RIP3-mediated programmed necrosis and AIF-mediated caspase-independent programmed necrosis in glioblastoma.

Rapid simultaneous determination of apoptosis, necrosis, and viability in sulfur mustard exposed HaCaT cell cultures.

PubMed

Heinrich, A; Balszuweit, F; Thiermann, H; Kehe, K

2009-12-15

Sulfur mustard (SM; bis(2-chloroethyl)sulphide; HD) is a blister inducing agent causing DNA damage and subsequently, cell death, mostly by apoptosis in basal keratinocytes. Despite intensive investigations on the cellular mechanism, there are, as of now, no causal therapeutics to prevent or antagonize SM-related damage to cells and tissues. In order to develop treatment strategies against vesication, it is important to distinguish apoptosis from necrosis in SM treated human keratinocytes. DNA fragmentation is a hallmark of apoptosis and regulated by a cascade of enzymes (endonucleases, DNase I, NUC 18), which finally cut the chromatin into specific formations of 180-200 base pairs, the nucleosomes. A feasible way to monitor apoptosis is the detection of nucleosomes by means of the Cell Death Detection ELISA(plus) (CDDE). In contrast, during necrosis DNA fragmentation is at random and delivers larger fragments, which therefore are significantly less in number and predominantly occur in cell culture supernatant. To monitor necrosis, we measured the release of intracellular adenylate kinase (AK) into cell culture supernatant by means of the ToxiLight Bioluminescence Assay (TL). With combination of the Cell Death Detection ELISA(plus) and the ToxiLight Bioluminescence Assay, we acquired more comprehensive information on cell survival and mechanisms of cell death, following an SM exposure. To validate the assay we tested common apoptosis- and necrosis-inducing agents like SM 300 microM for 30 min, Lewisite (L) 60 microM for 5 min and Triton X-100 0.1%. The results show that it is possible to differentiate between the two modes of cell death and to quantify their extent. This assay is highly effective in quantifying apoptosis and necrosis caused by cytotoxic agents and in estimating protective effects of potential active pharmaceutical ingredients.

Prosurvival AMBRA1 turns into a proapoptotic BH3-like protein during mitochondrial apoptosis

PubMed Central

Strappazzon, Flavie; Di Rita, Anthea; Cianfanelli, Valentina; D'Orazio, Melania; Nazio, Francesca; Fimia, Gian Maria; Cecconi, Francesco

2016-01-01

ABSTRACT Autophagy and apoptosis are 2 stress-response mechanisms that are closely interconnected. However, the molecular interplays between these 2 pathways remain to be clarified. Here we report that the crucial proautophagic factor AMBRA1 can act as a positive mediator of mitochondrial apoptosis. Indeed, we show that, in a proapoptotic positive feedback loop, the C-terminal part of AMBRA1, generated by CASP/CASPASE cleavage upon apoptosis induction, inhibits the antiapoptotic factor BCL2 by a direct binding through its BH3-like domain. The mitochondrial AMBRA1-BCL2 complex is thus at the crossroad between autophagy and cell death and may represent a novel target in development of therapeutic approaches in clinical diseases. PMID:27123694

Erufosine simultaneously induces apoptosis and autophagy by modulating the Akt-mTOR signaling pathway in oral squamous cell carcinoma.

PubMed

Kapoor, Vaishali; Zaharieva, Maya M; Das, Satya N; Berger, Martin R

2012-06-01

We investigated the anticancer activity of erufosine in oral squamous carcinoma cell lines in terms of cell proliferation, colony formation, induction of autophagy/apoptosis, cell cycle and mTOR signaling pathway. Erufosine showed dose-dependent cytotoxicity in all cell lines, it induced autophagy as well as apoptosis, G2 cell cycle arrest and modulation of cyclin D1 expression. Further erufosine downregulated the phosphorylation of major components of mTOR pathway, like p-Akt at Ser473 and Thr308 residues, p-Raptor, p-mTOR, p-PRAS40 and its downstream substrates p-p70S6K and p-4EBP1 in a dose-dependent manner. The pre-treatment of tumor cells with p-mTOR siRNA increased cytotoxic effects of erufosine comparable to cisplatin but higher than rapamycin. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Beyond apoptosis: the mechanism and function of phosphatidylserine asymmetry in the membrane of activating mast cells.

PubMed

Rysavy, Noel M; Shimoda, Lori M N; Dixon, Alyssa M; Speck, Mark; Stokes, Alexander J; Turner, Helen; Umemoto, Eric Y

2014-01-01

Loss of plasma membrane asymmetry is a hallmark of apoptosis, but lipid bilayer asymmetry and loss of asymmetry can contribute to numerous cellular functions and responses that are independent of programmed cell death. Exofacial exposure of phosphatidylserine occurs in lymphocytes and mast cells after antigenic stimulation and in the absence of apoptosis, suggesting that there is a functional requirement for phosphatidylserine exposure in immunocytes. In this review we examine current ideas as to the nature of this functional role in mast cell activation. Mechanistically, there is controversy as to the candidate proteins responsible for phosphatidylserine translocation from the internal to external leaflet, and here we review the candidacies of mast cell PLSCR1 and TMEM16F. Finally we examine the potential relationship between functionally important mast cell membrane perturbations and phosphatidylserine exposure during activation.

Pyrrolidine dithiocarbamate-zinc(II) and -copper(II) complexes induce apoptosis in tumor cells by inhibiting the proteasomal activity☆

PubMed Central

Milacic, Vesna; Chen, Di; Giovagnini, Lorena; Diez, Alejandro; Fregona, Dolores; Dou, Q. Ping

2013-01-01

Zinc and copper are trace elements essential for proper folding, stabilization and catalytic activity of many metalloenzymes in living organisms. However, disturbed zinc and copper homeostasis is reported in many types of cancer. We have previously demonstrated that copper complexes induced proteasome inhibition and apoptosis in cultured human cancer cells. In the current study we hypothesized that zinc complexes could also inhibit the proteasomal chymotrypsin-like activity responsible for subsequent apoptosis induction. We first showed that zinc(II) chloride was able to inhibit the chymotrypsin-like activity of a purified 20S proteasome with an IC50 value of 13.8 μM, which was less potent than copper(II) chloride (IC50 5.3 μM). We then compared the potencies of a pyrrolidine dithiocarbamate (PyDT)-zinc(II) complex and a PyDT-copper(II) complex to inhibit cellular proteasomal activity, suppress proliferation and induce apoptosis in various human breast and prostate cancer cell lines. Consistently, zinc complex was less potent than copper complex in inhibiting the proteasome and inducing apoptosis. Additionally, zinc and copper complexes appear to use somewhat different mechanisms to kill tumor cells. Zinc complexes were able to activate calpain-, but not caspase-3-dependent pathway, while copper complexes were able to induce activation of both proteases. Furthermore, the potencies of these PyDT-metal complexes depend on the nature of metals and also on the ratio of PyDT to the metal ion within the complex, which probably affects their stability and availability for interacting with and inhibiting the proteasome in tumor cells. PMID:18501397

Elucidation of Altered Pathways in Tumor-Initiating Cells of Triple-Negative Breast Cancer: A Useful Cell Model System for Drug Screening.

PubMed

Christensen, Anne G; Ehmsen, Sidse; Terp, Mikkel G; Batra, Richa; Alcaraz, Nicolas; Baumbach, Jan; Noer, Julie B; Moreira, José; Leth-Larsen, Rikke; Larsen, Martin R; Ditzel, Henrik J

2017-08-01

A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics. Within these pairs, one, but not the other, resulted in tumors in immunodeficient NOD/Shi-scid/IL-2 Rγ null mice and efficiently formed mammospheres. Quantitative proteomics and phosphoproteomics were used to map signaling pathways associated with the tumor-initiating ability. Signaling associated with apoptosis was suppressed in tumor-initiating versus nontumorigenic counterparts with pro-apoptotic proteins, such as Bcl2-associated agonist of cell death (BAD), FAS-associated death domain protein (FADD), and myeloid differentiation primary response protein (MYD88), downregulated in tumor-initiating epithelial-like cells. Functional studies confirmed significantly lower apoptosis in tumor-initiating versus nontumorigenic cells. Moreover, central pathways, including β-catenin and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signaling, exhibited increased activation in the tumor-initiating cells. To evaluate the CSC model as a tool for drug screening, we assessed the effect of separately blocking NF-κB and Wnt/β-catenin signaling and found markedly reduced mammosphere formation, particularly for tumor-initiating cells. Similar reduction was also observed using patient-derived primary cancer cells. Furthermore, blocking NF-κB signaling in mice transplanted with tumor-initiating cells significantly reduced tumor outgrowth. Our study demonstrates that suppressed apoptosis, activation of pathways associated with cell viability, and CSCs are the major differences between tumor-initiating and nontumorigenic cells independent of their epithelial-like/mesenchymal-like phenotype. These altered pathways may provide targets for future drug development to eliminate CSCs, and the cell model may be a useful tool in such drug screenings. Stem Cells 2017;35:1898-1912. © 2017 AlphaMed Press.

Human replication protein Cdc6 is selectively cleaved by caspase 3 during apoptosis

PubMed Central

Pelizon, Cristina; d’Adda di Fagagna, Fabrizio; Farrace, Lorena; Laskey, Ronald A.

2002-01-01

In eukaryotes, the initiation of DNA replication involves the ordered assembly on chromatin of pre-replicative complexes (pre-RCs), including the origin recognition complex (ORC), Cdc6, Cdt1 and the minichromosome maintenance proteins (MCMs). In light of its indispensable role in the formation of pre-RCs, Cdc6 binding to chromatin represents a key step in the regulation of DNA replication and cell proliferation. Here, we study the human Cdc6 (HuCdc6) protein during programmed cell death (apoptosis). We find that HuCdc6, but not HuOrc2 (a member of the ORC) or HuMcm5 (one of the MCMs), is specifically cleaved in several human cell lines induced to undergo apoptosis by a variety of stimuli. Expression of caspase-uncleavable mutant HuCdc6 attenuates apoptosis, delaying cell death. Therefore, an important function for cleavage of HuCdc6 is to prevent a wounded cell from replicating and to facilitate death. PMID:12151338

Akt-phosphorylated Mitogen-activated Kinase-activating Death Domain Protein (MADD) Inhibits TRAIL-induced Apoptosis by Blocking Fas-associated Death Domain (FADD) Association with Death Receptor 4*

PubMed Central

Li, Peifeng; Jayarama, Shankar; Ganesh, Lakshmy; Mordi, David; Carr, Ryan; Kanteti, Prasad; Hay, Nissim; Prabhakar, Bellur S.

2010-01-01

MADD plays an essential role in cancer cell survival. Abrogation of endogenous MADD expression results in significant spontaneous apoptosis and enhanced susceptibility to tumor necrosis factor α-related apoptosis-inducing ligand (TRAIL)-induced apoptosis. However, the regulation of MADD function is largely unknown. Here, we demonstrate that endogenous MADD is phosphorylated at three highly conserved sites by Akt, and only the phosphorylated MADD can directly interact with the TRAIL receptor DR4 thereby preventing Fas-associated death domain recruitment. However, in cells susceptible to TRAIL treatment, TRAIL induces a reduction in MADD phosphorylation levels resulting in MADD dissociation from, and Fas-associated death domain association with DR4, which allows death-inducing signaling complex (DISC) formation leading to apoptosis. Thus, the pro-survival function of MADD is dependent upon its phosphorylation by Akt. Because Akt is active in most cancer cells and phosphorylated MADD confers resistance to TRAIL-induced apoptosis, co-targeting Akt-MADD axis is likely to increase efficacy of TRAIL-based therapies. PMID:20484047

Involvement of the MAPK and PI3K pathways in chitinase 3-like 1-regulated hyperoxia-induced airway epithelial cell death

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Mi Na; Lee, Kyung Eun; Hong, Jung Yeon

Highlights: Black-Right-Pointing-Pointer Hyperoxia induces apoptosis and chitinase 3-like 1 expression in human airway epithelial cells. Black-Right-Pointing-Pointer Presence of chitinase 3-like 1 affects airway epithelial cell death after hyperoxic exposure. Black-Right-Pointing-Pointer Silencing chitinase 3-like 1 manipulate the phosphorylation of ERK, p38 and Akt. -- Abstract: Background: Exposure to 100% oxygen causes hyperoxic acute lung injury characterized by cell death and injury of alveolar epithelial cells. Recently, the role of chitinase 3-like 1 (CHI3L1), a member of the glycosyl hydrolase 18 family that lacks chitinase activity, in oxidative stress was demonstrated in murine models. High levels of serum CHI3L1 have been associatedmore » with various diseases of the lung, such as asthma, chronic obstructive pulmonary disease, and cancer. However, the role of CHI3L1 in human airway epithelial cells undergoing oxidative stress remains unknown. In addition, the signaling pathways associated with CHI3L1 in this process are poorly understood. Purpose: In this study, we demonstrate the role of CHI3L1, along with the MAPK and PI3K signaling pathways, in hyperoxia-exposed airway epithelial cells. Method: The human airway epithelial cell line, BEAS-2B, was exposed to >95% oxygen (hyperoxia) for up to 72 h. Hyperoxia-induced cell death was determined by assessing cell viability, Annexin-V FITC staining, caspase-3 and -7 expression, and electron microscopy. CHI3L1 knockdown and overexpression studies were conducted in BEAS-2B cells to examine the role of CHI3L1 in hyperoxia-induced apoptosis. Activation of the MAPK and PI3K pathways was also investigated to determine the role of these signaling cascades in this process. Results: Hyperoxia exposure increased CHI3L1 expression and apoptosis in a time-dependent manner. CHI3L1 knockdown protected cells from hyperoxia-induced apoptosis. In contrast, CHI3L1 overexpression promoted cell death after hyperoxia exposure. Finally, phosphorylation of ERK1/2, p38, and Akt were affected by CHI3L1 knockdown. Conclusion: This study indicates that CHI3L1 is involved in hyperoxia-induced cell death, suggesting that CHI3L1 may be one of several cell death regulators influencing the MAPK and PI3K pathways during oxidative stress in human airway epithelial cells.« less

[The relationship between necroptosis and blinding eye diseases].

PubMed

Xie, L L; Jiang, B

2018-03-11

As a programmed cell death manner which is distinguished from apoptosis and autophagy, necroptosis is a newly discovered pathway of regulated necrosis that requires the protein receptor interacting protein kinases 1 and 3 and mixed lineage kinase domain-like protein. Necroptosis is mediated by death receptors, toll-like receptors and probably other mediators. Emerging evidences have delineated that necroptosis plays an important role in the occurrence and development of various blinding eye diseases. In this review, the related mechanism of necroptosis, the relationship between necroptosis and multiple blinding eye diseases, such as age-related macular degeneration, retinitis pigmentosa and glaucoma, and the potential therapeutic targets of necroptosis are discussed. (Chin J Ophthalmol, 2018, 54: 234-240) .

The flavonoid tangeretin activates the unfolded protein response and synergizes with imatinib in the erythroleukemia cell line K562.

PubMed

Lust, Sofie; Vanhoecke, Barbara; Van Gele, Mireille; Philippé, Jan; Bracke, Marc; Offner, Fritz

2010-06-01

We explored the mechanism of cell death of the polymethoxyflavone tangeretin (TAN) in K562 breakpoint cluster region-abelson murine leukemia (Bcr-Abl+) cells. Flow cytometric analysis showed that TAN arrested the cells in the G(2)/M phase and stimulated an accumulation of the cells in the sub-G(0) phase. TAN-induced cell death was evidenced by poly(ADP)-ribose polymerase cleavage, DNA laddering fragmentation, activation of the caspase cascade and downregulation of the antiapoptotic proteins Mcl-1 and Bcl-x(L). Pretreatment with the pancaspase inhibitor Z-VAD-FMK_blocked caspase activation and cell cycle arrest but did not inhibit apoptosis which suggest that other cell killing mechanisms like endoplasmic reticulum (ER)-associated cell death pathways could be involved. We demonstrated that TAN-induced apoptosis was preceded by a rapid activation of the proapoptotic arm of the unfolded protein response, namely PKR-like ER kinase. This was accompanied by enhanced levels of glucose-regulated protein of 78 kDa and of spliced X-box binding protein 1. Furthermore, TAN sensitized K562 cells to the cell killing effects of imatinib via an apoptotic mechanism. In conclusion, our results suggest that TAN is able to induce apoptosis in Bcr-Abl+ cells via cell cycle arrest and the induction of the unfolded protein response, and has synergistic cytotoxicity with imatinib.

Pathway-specific effect of caffeine on protection against UV irradiation-induced apoptosis in corneal epithelial cells.

PubMed

Wang, Ling; Lu, Luo

2007-02-01

To define the role of molecular interaction between the UV-induced JNK (c-Jun N-terminal kinase) cascade and corneal epithelial cell apoptosis and protection against apoptosis by caffeine. Rabbit and human corneal epithelial cells were cultured in DMEM/F12 medium containing 10% FBS and 5 microg/mL insulin at 37 degrees C in 5% CO(2). DNA fragmentation and ethidium bromide/acridine orange (EB/AO) nuclear staining were performed to detect cell death. Western blot, immunoprecipitation, and kinase assays were used to measure UV-induced mitogen-activated protein (MAP) kinase activity. UV irradiation-induced apoptosis through apoptosis signal-regulating kinase 1 (ASK1) and MAKK4 (SEK1) upstream from JNK was caffeine sensitive. Caffeine (1,3,7-trimethylxanthine), an agent that is one of the most popular additions to food consumed in the world and a potential enhancer of chemotherapy, effectively protected corneal epithelial cells against apoptosis by its specific effect on the JNK cascade. Theophylline (1,3-dimethylxanthine) exhibited an effect similar to that of caffeine on prevention of UV irradiation-induced apoptosis. However, alterations of either intracellular cAMP or Ca(2+) levels did not alter the effect of caffeine on the JNK signaling pathway. In addition, the blockade of PI3K-like kinases by wortmannin had no impact on the protective effect of caffeine against UV irradiation-induced apoptosis, suggesting that the protective effect of caffeine acts through a specific mechanism involving UV irradiation-induced activation of ASK1 and SEK1. In contrast, caffeine had no effects on melphalan-, hyperosmotic stress-, or IL-1beta-induced activation of the JNK signaling pathway in these cells. UV irradiation stress-induced activation of the ASK1-SEK1-JNK signaling pathway leading to apoptosis is a caffeine-sensitive process, and caffeine, as a multifunctional agent in cells, can specifically interact with the pathway to protect against apoptosis.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Yuan; Luo, Fangjun; Li, Hui

During inflammation in the glomerulus, the proliferation of myofiroblast-like mesangial cells is commonly associated with the pathological process. Macrophages play an important role in regulating the growth of resident mesangial cells in the glomeruli. Alternatively activated macrophage (M2 macrophage) is a subset of macrophages induced by IL-13/IL-4, which is shown to play a repair role in glomerulonephritis. Prompted by studies of development, we performed bone marrow derived macrophage and rat mesangial cell co-culture study. Conditioned medium from IL-4 primed M2 macrophages induced rat mesangial cell apoptosis. The pro-apoptotic effect of M2 macrophages was demonstrated by condensed nuclei stained with Hoechstmore » 33258, increased apoptosis rates by flow cytometry analysis and enhanced caspase-3 activation by western blot. Fas protein was up-regulated in rat mesangial cells, and its neutralizing antibody ZB4 partly inhibited M2 macrophage-induced apoptosis. The up-regulated arginase-1 expression in M2 macrophage also contributed to this apoptotic effect. These results indicated that the process of apoptosis triggered by conditioned medium from M2 macrophages, at least is partly conducted through Fas in rat mesangial cells. Our findings provide compelling evidence that M2 macrophages control the growth of mesangial cells in renal inflammatory conditions. - Highlights: • Conditioned-medium from M2 macrophages induces rat mesangial cell (MsC) apoptosis. • M2 macrophage conditioned medium exerts its pro-apoptotic effects via Fas ligand. • Arginase-1 activity in M2 macrophages plays a role in inducing apoptosis in rat MsC.« less

Effects of high glucose and advanced glycation end products on the expressions of sclerostin and RANKL as well as apoptosis in osteocyte-like MLO-Y4-A2 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tanaka, Ken-ichiro, E-mail: ken1nai@med.shimane-u.ac.jp; Yamaguchi, Toru, E-mail: yamaguch@med.shimane-u.ac.jp; Kanazawa, Ippei, E-mail: ippei.k@med.shimane-u.ac.jp

In diabetes mellitus (DM), high glucose (HG) and advanced glycation end products (AGEs) are involved in bone quality deterioration. Osteocytes produce sclerostin and receptor activator of nuclear factor-kB ligand (RANKL) and regulate osteoblast and osteoclast function. However, whether HG or AGEs directly affect osteocytes and regulate sclerostin and RANKL production is unknown. Here, we examined the effects of HG, AGE2, and AGE3 on the expression of sclerostin and RANKL and on apoptosis in osteocyte-like MLO-Y4-A2 cells. Treatment of the cells with 22 mM glucose, 100 μg/mL either AGE2 or AGE3 significantly increased the expression of sclerostin protein and mRNA; however, both AGEs,more » but not glucose, significantly decreased the expression of RANKL protein and mRNA. Moreover, treatment of the cells with HG, AGE2, or AGE3 for 72 h induced significant apoptosis. These detrimental effects of HG, AGE2, and AGE3 on sclerostin and RANKL expressions and on apoptosis were antagonized by pretreatment of the cells with 10{sup −8} M human parathyroid hormone (PTH)-(1–34). Thus, HG and AGEs likely suppress bone formation by increasing sclerostin expression in osteocytes, whereas AGEs suppress bone resorption by decreasing RANKL expression. Together, these processes may cause low bone turnover in DM. In addition, HG and AGEs may cause cortical bone deterioration by inducing osteocyte apoptosis. PTH may effectively treat these pathological processes and improve osteocyte function. - Highlights: • AGEs are involved in bone quality deterioration in diabetes mellitus (DM). • AGEs increased sclerostin as well as apoptosis, and decreased RANKL in osteocytes. • The effects of AGEs on osteocyte function were antagonized by human PTH-(1–34). • AGEs may cause low bone turnover and cortical porosity in DM. • PTH may be effective in bone quality deterioration by improving osteocyte function.« less

Modeled Microgravity Inhibits Apoptosis in Peripheral Blood Lymphocytes

NASA Technical Reports Server (NTRS)

Risin, Diana; Pellis, Neal R.

2000-01-01

Microgravity interferes with numerous lymphocyte functions (expression of cell surface molecules, locomotion, polyclonal and antigen-specific activation, and the protein kinase C activity in signal transduction). The latter suggests that gravity may also affect programmed cell death (PCD) in lymphocyte populations. To test this hypothesis, we investigated spontaneous, activation- and radiation-induced PCD in peripheral blood mononuclear cells (PBMC) exposed to modeled microgravity using a rotating cell culture system. The results showed significant inhibition of radiation- and activation-induced apoptosis in modeled microgravity and provide insights into the potential mechanisms of this phenomenon.

Modeled microgravity inhibits apoptosis in peripheral blood lymphocytes

NASA Technical Reports Server (NTRS)

Risin, D.; Pellis, N. R.; McIntire, L. V. (Principal Investigator)

2001-01-01

Microgravity interferes with numerous lymphocyte functions (expression of cell surface molecules, locomotion, polyclonal and antigen-specific activation, and the protein kinase C activity in signal transduction). The latter suggests that gravity may also affect programmed cell death (PCD) in lymphocyte populations. To test this hypothesis, we investigated spontaneous, activation- and radiation-induced PCD in peripheral blood mononuclear cells exposed to modeled microgravity (MMG) using a rotating cell culture system. The results showed significant inhibition of radiation- and activation-induced apoptosis in MMG and provide insights into the potential mechanisms of this phenomenon.

Polo-like kinase 1 expression is suppressed by CCAAT/enhancer-binding protein α to mediate colon carcinoma cell differentiation and apoptosis.

PubMed

Dasgupta, Nirmalya; Thakur, Bhupesh Kumar; Ta, Atri; Das, Sayan; Banik, George; Das, Santasabuj

2017-07-01

Human polo-like kinase 1 (PLK1), a highly conserved serine/threonine kinase is a key player in several essential cell-cycle events. PLK1 is considered an oncogene and its overexpression often correlates with poor prognosis of cancers, including colorectal cancer (CRC). However, regulation of PLK1 expression in colorectal cells was never studied earlier and it is currently unknown if PLK1 regulates differentiation and apoptosis of CRC. PLK1 expression was analyzed by real-time PCR and western blotting. Transcriptional regulation was studied by reporter assay, gene knock-down, EMSA and ChIP. PLK1 expression was down-regulated during butyrate-induced differentiation of HT-29 and other CRC cells. Also, PLK1 down-regulation mediated the role of butyrate in CRC differentiation and apoptosis. We report here a novel transcriptional regulation of PLK1 by butyrate. Transcription factors CCAAT/enhancer-binding protein α (C/EBPα) and Oct-1 share an overlapping binding site over the PLK1 promoter. Elevated levels of C/EBPα by butyrate treatment of CRC cells competed out the activator protein Oct-1 from binding to the PLK1 promoter and sequestered it. Binding of C/EBPα was associated with increased deacetylation near the transcription start site (TSS) of the PLK1 promoter, which abrogated transcription through reduced recruitment of RNA polymerase II. We also found a synergistic role between the synthetic PLK1-inhibitor SBE13 and butyrate on the apoptosis of CRC cells. This study offered a novel p53-independent regulation of PLK1 during CRC differentiation and apoptosis. Down-regulation of PLK1 is one of the mechanisms underlying the anti-cancer role of dietary fibre-derived butyrate in CRC. Copyright © 2017 Elsevier B.V. All rights reserved.

cAMP prevents TNF-induced apoptosis through inhibiting DISC complex formation in rat hepatocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bhattacharjee, Rajesh; Xiang, Wenpei; Family Planning Research Institute, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, People's Republic of China

2012-06-22

Highlights: Black-Right-Pointing-Pointer cAMP blocks cell death induced by TNF and actinomycin D in cultured hepatocytes. Black-Right-Pointing-Pointer cAMP blocks NF-{kappa}B activation induced by TNF and actinomycin D. Black-Right-Pointing-Pointer cAMP blocks DISC formation following TNF and actinomycin D exposure. Black-Right-Pointing-Pointer cAMP blocks TNF signaling at a proximal step. -- Abstract: Tumor necrosis factor {alpha} (TNF) is a pleiotropic proinflammatory cytokine that plays a role in immunity and the control of cell proliferation, cell differentiation, and apoptosis. The pleiotropic nature of TNF is due to the formation of different signaling complexes upon the binding of TNF to its receptor, TNF receptor type 1more » (TNFR1). TNF induces apoptosis in various mammalian cells when the cells are co-treated with a transcription inhibitor like actinomycin D (ActD). When TNFR1 is activated, it recruits an adaptor protein, TNF receptor-associated protein with death domain (TRADD), through its cytoplasmic death effector domain (DED). TRADD, in turn, recruits other signaling proteins, including TNF receptor-associated protein 2 (TRAF2) and receptor-associated protein kinase (RIPK) 1, to form a complex. Subsequently, this complex combines with FADD and procaspase-8, converts into a death-inducing signaling complex (DISC) to induce apoptosis. Cyclic AMP (cAMP) is a second messenger that regulates various cellular processes such as cell proliferation, gene expression, and apoptosis. cAMP analogues are reported to act as anti-apoptotic agents in various cell types, including hepatocytes. We found that a cAMP analogue, dibutyryl cAMP (db-cAMP), inhibits TNF + ActD-induced apoptosis in rat hepatocytes. The protein kinase A (PKA) inhibitor KT-5720 reverses this inhibitory effect of cAMP on apoptosis. Cytoprotection by cAMP involves down-regulation of various apoptotic signal regulators like TRADD and FADD and inhibition of caspase-8 and caspase-3 cleavage. We also found that cAMP exerts its affect at the proximal level of TNF signaling by inhibiting the formation of the DISC complex upon the binding of TNF to TNFR1. In conclusion, our study shows that cAMP prevents TNF + ActD-induced apoptosis in rat hepatocytes by inhibiting DISC complex formation.« less

The role of necroptosis in the treatment of diseases.

PubMed

Cho, Young Sik

2018-04-11

Necroptosis is an emerging form of programmed cell death occurring via active and well-regulated necrosis, distinct from apoptosis morphologically, and biochemically. Necroptosis is mainly unmasked when apoptosis is compromised in response to cell stress. Unlike apoptotic cells, which are cleared by macrophages or neighboring cells, necrotic cells release danger signals, triggering inflammation, and exacerbating tissue damage. Evidence increasingly suggests that programmed necrosis is not only associated with pathophysiology of disease, but also induces innate immune response to viral infection. Therefore, necroptotic cell death plays both physiological and pathological roles. Physiologically, necroptosis induce an innate immune response as well as premature assembly of viral particles in cells infected with virus that abrogates host apoptotic machinery. On the other hand, necroptosis per se is detrimental, causing various diseases such as sepsis, neurodegenerative diseases and ischemic reperfusion injury. This review discusses the signaling pathways leading to necroptosis, associated necroptotic proteins with target-specific inhibitors and diseases involved. Several studies currently focus on protective approaches to inhibit necroptotic cell death. In cancer biology, however, anticancer drug resistance severely hampers the efficacy of chemotherapy based on apoptosis. Pharmacological switch of cell death finds therapeutic application in drug-resistant cancers. Therefore, the possible clinical role of necroptosis in cancer control will be discussed in brief.

[Biochemical changes in apoptosis and methods for their determination (review)].

PubMed

Sedláková, A; Kohút, A; Kalina, I

1999-08-01

Apoptosis or programmed cell death is a physiological process which occurs at different biological states as well as at disease process. Morphologically it is characterized by the chromatine condensation and other changes with preserved integrity of plasmatic membrane. The major and most frequently studied biochemical characteristic of apoptosis is a DNA fragmentation. In our paper attention is directed to the early biochemical changes in cell membranes, i.g., the externalization of phosphatidylserine, hydrolysis of sphingomyeline on the ceramide and activation of phospholipases especially phospholipase A2. In one part we described the changes of cysteine proteases (caspases), which play a key role in the execution of apoptosis. These biochemical changes are associated with ceramide signalization of apoptosis. Briefly are presented also some dates about apoptosis induction with reactive oxygen radicals and the role of the arachidonic acid metabolites in this process. We consider the investigation and determination of these changes as important parameters of apoptosis at some diseases, e.g., cancer or degenerative diseases, and of their treatment.

atRA-induced apoptosis of mouse embryonic palate mesenchymal cells involves activation of MAPK pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu Zengli; Xing Ying

2006-08-15

Our previous studies have shown that atRA treatment resulted in cell-cycle block and growth inhibition in mouse embryonic palatal mesenchymal (MEPM). In the current study, gestation day (GD) 13 MEPM cells were used to test the hypothesis that the growth inhibition by atRA is due to apoptosis. The effects of atRA on apoptosis were assessed by performing MTT assay, Cell Death Detection ELISA and flow cytometry, respectively. Data analysis confirmed that atRA treatment induced apoptosis-like cell death, as shown by decreased cell viability and increased fragmented DNA and sub-G1 fraction. atRA-induced apoptosis was associated with upregulation of bcl-2, translocation ofmore » bax protein to the mitochondria from the cytosol, activation of caspase-3 and cytochrome c release into cytosol. atRA-induced apoptosis was abrogated by z-DEVD-fmk, a caspase-3 specific inhibitor, and z-VAD-fmk, a general caspase inhibitor, suggesting that the atRA-induced cell death of MEPM cells occurs through the cytochrome c- and caspase-3-dependent pathways. In addition, atRA treatment caused a strong and sustained activation of c-Jun N-terminal kinase (JNK) and p38 kinase (p38), as well as an early but transient activation of extracellular signal-regulated kinase (ERK). Importantly, atRA-induced DNA fragmentation and capase-3 activation were prevented by pretreatment with the JNK inhibitor (SP600125) and the p38 MAPK inhibitor (SB202190), but not by pretreatment with MEK inhibitor (U0126). From these results, we suggest that mitogen-activated protein kinase-dependent pathways is involved in the atRA-induced apoptosis of MEPM cells.« less

Multiple, disparate roles for calcium signaling in apoptosis of human prostate and cervical cancer cells exposed to diindolylmethane.

PubMed

Savino, John A; Evans, Jodi F; Rabinowitz, Dorianne; Auborn, Karen J; Carter, Timothy H

2006-03-01

Diindolylmethane (DIM), derived from indole-3-carbinol in cruciferous vegetables, causes growth arrest and apoptosis of cancer cells in vitro. DIM also induces endoplasmic reticulum (ER) stress, and thapsigargin, a specific inhibitor of the sarcoplasmic reticulum/ER calcium-dependent ATPase, enhances this effect. We asked whether elevated cytosolic free calcium [Ca2+]i is required for cytotoxicity of DIM and thapsigargin in two cancer cells lines (C33A, from cervix, and DU145, from prostate). [Ca2+]i was measured in real-time by FURA-2 fluorescence. We tested whether DIM, thapsigargin, and DIM + thapsigargin cause apoptosis, measured by nucleosome release, under conditions that prevented elevation of [Ca2+]i, using both cell-permeable and cell-impermeable forms of the specific calcium chelator BAPTA. DIM, like thapsigargin, rapidly mobilized ER calcium. C33A and DU145 responded differently to perturbations in Ca2+ homeostasis, suggesting that DIM induces apoptosis by different mechanisms in these two cell lines and/or that calcium mobilization also activates different survival pathways in C33A and DU145. Apoptosis in C33A was independent of increased [Ca2+]i, suggesting that depletion of ER Ca2+ stores may be sufficient for cell killing, whereas apoptosis in DU145 required elevated [Ca2+]i for full response. Inhibitor studies using cyclosporin A and KN93 showed that Ca2+ signaling is important for cell survival but the characteristics of this response also differed in the two cell lines. Our results underscore the complex and variable nature of cellular responses to disrupted Ca2+ homeostasis and suggest that alteration Ca2+ homeostasis in the ER can induce cellular apoptosis by both calcium-dependent and calcium-independent mechanisms.

FLICE-like inhibitory protein (FLIP) protects against apoptosis and suppresses NF-kappaB activation induced by bacterial lipopolysaccharide.

PubMed

Bannerman, Douglas D; Eiting, Kristine T; Winn, Robert K; Harlan, John M

2004-10-01

Bacterial lipopolysaccharide (LPS) via its activation of Toll-like receptor-4 contributes to much of the vascular injury/dysfunction associated with gram-negative sepsis. Inhibition of de novo gene expression has been shown to sensitize endothelial cells (EC) to LPS-induced apoptosis, the onset of which correlates with decreased expression of FLICE-like inhibitory protein (FLIP). We now have data that conclusively establish a role for FLIP in protecting EC against LPS-induced apoptosis. Overexpression of FLIP protected against LPS-induced apoptosis, whereas down-regulation of FLIP using antisense oligonucleotides sensitized EC to direct LPS killing. Interestingly, FLIP overexpression suppressed NF-kappaB activation induced by LPS, but not by phorbol ester, suggesting a specific role for FLIP in mediating LPS activation. Conversely, mouse embryo fibroblasts (MEF) obtained from FLIP -/- mice showed enhanced LPS-induced NF-kappaB activation relative to those obtained from wild-type mice. Reconstitution of FLIP-/- MEF with full-length FLIP reversed the enhanced NF-kappaB activity elicited by LPS in the FLIP -/- cells. Changes in the expression of FLIP had no demonstrable effect on other known LPS/Tlr-4-activated signaling pathways including the p38, Akt, and Jnk pathways. Together, these data support a dual role for FLIP in mediating LPS-induced apoptosis and NF-kappaB activation.

Selective inhibition of FLICE-like inhibitory protein expression with small interfering RNA oligonucleotides is sufficient to sensitize tumor cells for TRAIL-induced apoptosis.

PubMed Central

Siegmund, Daniela; Hadwiger, Philipp; Pfizenmaier, Klaus; Vornlocher, Hans-Peter; Wajant, Harald

2002-01-01

BACKGROUND: Most tumors express death receptors and their activation represents a potential selective approach in cancer treatment. The most promising candidate for tumor selective death receptor-activation is tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/Apo2L, which activates the death receptors TRAIL-R1 and TRAIL-R2, and induces apoptosis preferentially in tumor cells but not in normal tissues. However, many cancer cells are not or only moderately sensitive towards TRAIL and require cotreatment with irradiation or chemotherapy to yield a therapeutically reasonable apoptotic response. Because chemotherapy can have a broad range of unwanted side effects, more specific means for sensitizing tumor cells for TRAIL are desirable. The expression of the cellular FLICE-like inhibitory protein (cFLIP) is regarded as a major cause of TRAIL resistance. We therefore analyzed the usefulness of targeting FLIP to sensitize tumor cells for TRAIL-induced apoptosis. MATERIALS AND METHODS: To selectively interfere with expression of cFLIP short double-stranded RNA oligonucleotides (small interfering RNAs [siRNAs]) were introduced in the human cell lines SV80 and KB by electroporation. Effects of siRNA on FLIP expression were analyzed by Western blotting and RNase protection assay and correlated with TRAIL sensitivity upon stimulation with recombinant soluble TRAIL and TRAIL-R1- and TRAIL-R2-specific agonistic antibodies. RESULTS: FLIP expression can be inhibited by RNA interference using siRNAs, evident from reduced levels of FLIP-mRNA and FLIP protein. Inhibition of cFLIP expression sensitizes cells for apoptosis induction by TRAIL and other death ligands. In accordance with the presumed function of FLIP as an inhibitor of death receptor-induced caspase-8 activation, down-regulation of FLIP by siRNAs enhanced TRAIL-induced caspase-8 activation. CONCLUSION: Inhibition of FLIP expression was sufficient to sensitize tumor cells for TRAIL-induced apoptosis. The combination of TRAIL and FLIP-targeting siRNA could therefore be a useful strategy to attack cancer cells, which are resistant to TRAIL alone. PMID:12520089

Platelet-Derived Growth Factor-BB Protects Mesenchymal Stem Cells (MSCs) Derived From Immune Thrombocytopenia Patients Against Apoptosis and Senescence and Maintains MSC-Mediated Immunosuppression

PubMed Central

Zhang, Jia-min; Feng, Fei-er; Wang, Qian-ming; Zhu, Xiao-lu; Fu, Hai-xia; Xu, Lan-ping; Liu, Kai-yan

2016-01-01

Immune thrombocytopenia (ITP) is characterized by platelet destruction and megakaryocyte dysfunction. Mesenchymal stem cells (MSCs) from ITP patients (MSC-ITP) do not exhibit conventional proliferative abilities and thus exhibit defects in immunoregulation, suggesting that MSC impairment might be a mechanism involved in ITP. Platelet-derived growth factor (PDGF) improves growth and survival in various cell types. Moreover, PDGF promotes MSC proliferation. The aim of the present study was to analyze the effects of PDGF-BB on MSC-ITP. We showed that MSC-ITP expanded more slowly and appeared flattened and larger. MSC-ITP exhibited increased apoptosis and senescence compared with controls. Both the intrinsic and extrinsic pathways account for the enhanced apoptosis. P53 and p21 expression were upregulated in MSC-ITP, but inhibition of p53 with pifithrin-α markedly inhibited apoptosis and senescence. Furthermore, MSCs from ITP patients showed a lower capacity for inhibiting the proliferation of activated T cells inducing regulatory T cells (Tregs) and suppressing the synthesis of anti-glycoprotein (GP)IIb-IIIa antibodies. PDGF-BB treatment significantly decreased the expression of p53 and p21 and increased survivin expression in MSC-ITP. In addition, the apoptotic rate and number of senescent cells in ITP MSCs were reduced. Their impaired ability for inhibiting activated T cells, inducing Tregs, and suppressing the synthesis of anti-GPIIb-IIIa antibodies was restored after PDGF-BB treatment. In conclusion, we have demonstrated that PDGF-BB protects MSCs derived from ITP patients against apoptosis, senescence, and immunomodulatory defects. This protective effect of PDGF-BB is likely mediated via the p53/p21 pathway, thus potentially providing a new therapeutic approach for ITP. Significance Immune thrombocytopenia (ITP) is characterized by platelet destruction and megakaryocyte dysfunction. Platelet-derived growth factor (PDGF) improves growth and survival in various cell types and promotes mesenchymal stem cell (MSC) proliferation. PDGF-BB protects MSCs derived from ITP patients against apoptosis, senescence, and immunomodulatory defects. This protective effect of PDGF-BB is likely mediated via the p53/p21 pathway, thus potentially providing a new therapeutic approach for ITP. PMID:27471307

N-acetylcysteine protects against cadmium-induced germ cell apoptosis by inhibiting endoplasmic reticulum stress in testes.

PubMed

Ji, Yan-Li; Wang, Hua; Zhang, Cheng; Zhang, Ying; Zhao, Mei; Chen, Yuan-Hua; Xu, De-Xiang

2013-03-01

Cadmium (Cd) is a reproductive toxicant that induces germ cell apoptosis in the testes. Previous studies have demonstrated that endoplasmic reticulum (ER) stress is involved in Cd-induced germ cell apoptosis. The aim of the present study was to investigate the effects of N-acetylcysteine (NAC), an antioxidant, on Cd-induced ER stress and germ cell apoptosis in the testes. Male CD-1 mice were intraperitoneally injected with CdCl2 (2.0 mg kg(-1)). As expected, acute Cd exposure induced germ cell apoptosis in the testes, as determined by terminal dUTP nick-end labelling (TUNEL). However, the administration of NAC alleviated Cd-induced germ cell apoptosis in the testes. Further analysis showed that NAC attenuated the Cd-induced upregulation of testicular glucose-regulated protein 78 (GRP78), an important ER molecular chaperone. Moreover, NAC inhibited the Cd-induced phosphorylation of testicular eukaryotic translation initiation factor 2α (eIF2α), a downstream target of the double-stranded RNA-activated kinase-like ER kinase (PERK) pathway. In addition, NAC blocked the Cd-induced activation of testicular X binding protein (XBP)-1, indicating that NAC attenuates the Cd-induced ER stress and the unfolded protein response (UPR). Interestingly, NAC almost completely prevented the Cd-induced elevation of C/EBP homologous protein (CHOP) and phosphorylation of c-Jun N-terminal kinase (JNK), two components of the ER stress-mediated apoptotic pathway. In conclusion, NAC protects against Cd-induced germ cell apoptosis by inhibiting endoplasmic reticulum stress in the testes.

N-acetylcysteine protects against cadmium-induced germ cell apoptosis by inhibiting endoplasmic reticulum stress in testes

PubMed Central

Ji, Yan-Li; Wang, Hua; Zhang, Cheng; Zhang, Ying; Zhao, Mei; Chen, Yuan-Hua; Xu, De-Xiang

2013-01-01

Cadmium (Cd) is a reproductive toxicant that induces germ cell apoptosis in the testes. Previous studies have demonstrated that endoplasmic reticulum (ER) stress is involved in Cd-induced germ cell apoptosis. The aim of the present study was to investigate the effects of N-acetylcysteine (NAC), an antioxidant, on Cd-induced ER stress and germ cell apoptosis in the testes. Male CD-1 mice were intraperitoneally injected with CdCl2 (2.0 mg kg−1). As expected, acute Cd exposure induced germ cell apoptosis in the testes, as determined by terminal dUTP nick-end labelling (TUNEL). However, the administration of NAC alleviated Cd-induced germ cell apoptosis in the testes. Further analysis showed that NAC attenuated the Cd-induced upregulation of testicular glucose-regulated protein 78 (GRP78), an important ER molecular chaperone. Moreover, NAC inhibited the Cd-induced phosphorylation of testicular eukaryotic translation initiation factor 2α (eIF2α), a downstream target of the double-stranded RNA-activated kinase-like ER kinase (PERK) pathway. In addition, NAC blocked the Cd-induced activation of testicular X binding protein (XBP)-1, indicating that NAC attenuates the Cd-induced ER stress and the unfolded protein response (UPR). Interestingly, NAC almost completely prevented the Cd-induced elevation of C/EBP homologous protein (CHOP) and phosphorylation of c-Jun N-terminal kinase (JNK), two components of the ER stress-mediated apoptotic pathway. In conclusion, NAC protects against Cd-induced germ cell apoptosis by inhibiting endoplasmic reticulum stress in the testes. PMID:23353715

Grape seed extract triggers apoptosis in Caco-2 human colon cancer cells through reactive oxygen species and calcium increase: extracellular signal-regulated kinase involvement.

PubMed

Dinicola, Simona; Mariggiò, Maria Addolorata; Morabito, Caterina; Guarnieri, Simone; Cucina, Alessandra; Pasqualato, Alessia; D'Anselmi, Fabrizio; Proietti, Sara; Coluccia, Pierpaolo; Bizzarri, Mariano

2013-09-14

Grape seed extract (GSE) from Italia, Palieri and Red Globe cultivars inhibits cell growth and induces apoptosis in Caco-2 human colon cancer cells in a dose-dependent manner. In order to investigate the mechanism(s) supporting the apoptotic process, we analysed reactive oxygen species (ROS) production, intracellular Ca2+ handling and extracellular signal-regulated kinase (ERK) activation. Upon exposure to GSE, ROS and intracellular Ca2+ levels increased in Caco-2 cells, concomitantly with ERK inactivation. As ERK activity is thought to be essential for promoting survival pathways, inhibition of this kinase is likely to play a relevant role in GSE-mediated anticancer effects. Indeed, pretreatment with N-acetyl cysteine, a ROS scavenger, reversed GSE-induced apoptosis, and promoted ERK phosphorylation. This effect was strengthened by ethylene glycol tetraacetic acid-mediated inhibition of extracellular Ca2+ influx. ROS and Ca2+ influx inhibition, in turn, increased ERK phosphorylation, and hence almost entirely suppressed GSE-mediated apoptosis. These data suggested that GSE triggers a previously unrecognised ERK-based mechanism, involving both ROS production and intracellular Ca2+ increase, eventually leading to apoptosis in cancer cells.

The timecourse of apoptotic cell death during postnatal remodeling of the mouse cochlea and its premature onset by triiodothyronine (T3).

PubMed

Peeters, R P; Ng, L; Ma, M; Forrest, D

2015-05-15

Apoptosis underlies various forms of tissue remodeling during development. Prior to the onset of hearing, thyroid hormone (T3) promotes cochlear remodeling, which involves regression of the greater epithelial ridge (GER), a transient structure of columnar cells adjacent to the mechanosensory hair cells. We investigated the timecourse of apoptosis in the GER and the influence of ectopic T3 on apoptosis. In saline-treated mice, activated caspase 3-positive cells were detected in the GER between postnatal days 7 and 13 and appeared progressively along the cochlear duct from base to apex over developmental time. T3 given on P0 and P1 advanced the overall program of apoptosis and remodeling by ~4 days. Thyroid hormone receptor β was required for these actions, suggesting a receptor-mediated process of initiation of apoptosis. Finally, T3 given only at P0 or P1 resulted in deafness in adult mice, thus revealing a transient period of susceptibility to long-term damage in the neonatal auditory system. Published by Elsevier Ireland Ltd.

Does mechanism matter? Unrelated neurotoxicants converge on cell cycle and apoptosis during neurodifferentiation.

PubMed

Slotkin, Theodore A; Seidler, Frederic J

2012-07-01

Mechanistically unrelated developmental neurotoxicants often produce neural cell loss culminating in similar functional and behavioral outcomes. We compared an organophosphate pesticide (diazinon), an organochlorine pesticide (dieldrin) and a metal (Ni(2+)) for effects on the genes regulating cell cycle and apoptosis in differentiating PC12 cells, an in vitro model of neuronal development. Each agent was introduced at 30μM for 24 or 72h, treatments devoid of cytotoxicity. Using microarrays, we examined the mRNAs encoding nearly 400 genes involved in each of the biological processes. All three agents targeted both the cell cycle and apoptosis pathways, evidenced by significant transcriptional changes in 40-45% of the cell cycle-related genes and 30-40% of the apoptosis-related genes. There was also a high degree of overlap as to which specific genes were affected by the diverse agents, with 80 cell cycle genes and 56 apoptosis genes common to all three. Concordance analysis, which assesses stringent matching of the direction, magnitude and timing of the transcriptional changes, showed highly significant correlations for pairwise comparisons of all the agents, for both cell cycle and apoptosis. Our results show that otherwise disparate developmental neurotoxicants converge on common cellular pathways governing the acquisition and programmed death of neural cells, providing a specific link to cell deficits. Our studies suggest that identifying the initial mechanism of action of a developmental neurotoxicant may be strategically less important than focusing on the pathways that converge on common final outcomes such as cell loss. Copyright © 2012 Elsevier Inc. All rights reserved.

DOES MECHANISM MATTER? UNRELATED NEUROTOXICANTS CONVERGE ON CELL CYCLE AND APOPTOSIS DURING NEURODIFFERENTIATION

PubMed Central

Slotkin, Theodore A.; Seidler, Frederic J.

2012-01-01

Mechanistically unrelated developmental neurotoxicants often produce neural cell loss culminating in similar functional and behavioral outcomes. We compared an organophosphate pesticide (diazinon), an organochlorine pesticide (dieldrin) and a metal (Ni2+) for effects on the genes regulating cell cycle and apoptosis in differentiating PC12 cells, an in vitro model of neuronal development. Each agent was introduced at 30 μM for 24 or 72 hr, treatments devoid of cytotoxicity. Using microarrays, we examined the mRNAs encoding nearly 400 genes involved in each of the biological processes. All three agents targeted both the cell cycle and apoptosis pathways, evidenced by significant transcriptional changes in 40–45% of the cell cycle-related genes and 30–40% of the apoptosis-related genes. There was also a high degree of overlap as to which specific genes were affected by the diverse agents, with 80 cell cycle genes and 56 apoptosis genes common to all three. Concordance analysis, which assesses stringent matching of the direction, magnitude and timing of the transcriptional changes, showed highly significant correlations for pairwise comparisons of all the agents, for both cell cycle and apoptosis. Our results show that otherwise disparate developmental neurotoxicants converge on common cellular pathways governing the acquisition and programmed death of neural cells, providing a specific link to cell deficits. Our studies suggest that identifying the initial mechanism of action of a developmental neurotoxicant may be strategically less important than focusing on the pathways that converge on common final outcomes such as cell loss. PMID:22546817

Interrogating the relevance of mitochondrial apoptosis for vertebrate development and postnatal tissue homeostasis

PubMed Central

Kaufmann, Thomas; Villunger, Andreas

2016-01-01

“Programmed cell death or ‘apoptosis’ is critical for organogenesis during embryonic development and tissue homeostasis in the adult. Its deregulation can contribute to a broad range of human pathologies, including neurodegeneration, cancer, or autoimmunity…” These or similar phrases have become generic opening statements in many reviews and textbooks describing the physiological relevance of apoptotic cell death. However, while the role in disease has been documented beyond doubt, facilitating innovative drug discovery, we wonder whether the former is really true. What goes wrong in vertebrate development or in adult tissue when the main route to apoptotic cell death, controlled by the BCL2 family, is impaired? Such scenarios have been mimicked by deletion of one or more prodeath genes within the BCL2 family, and gene targeting studies in mice exploring the consequences have been manifold. Many of these studies were geared toward understanding the role of BCL2 family proteins and mitochondrial apoptosis in disease, whereas fewer focused in detail on their role during normal development or tissue homeostasis, perhaps also due to an irritating lack of phenotype. Looking at these studies, the relevance of classical programmed cell death by apoptosis for development appears rather limited. Together, these many studies suggest either highly selective and context-dependent contributions of mitochondrial apoptosis or significant redundancy with alternative cell death mechanisms, as summarized and discussed here. PMID:27798841

Grape seed procyanidin extract modulates proliferation and apoptosis of pancreatic beta-cells.

PubMed

Cedó, Lídia; Castell-Auví, Anna; Pallarès, Victor; Blay, Mayte; Ardévol, Anna; Arola, Lluís; Pinent, Montserrat

2013-05-01

Grape seed procyanidin extract (GSPE) modulates glucose homeostasis and insulinemia in several animal models. Under pathological conditions, insulin levels are dependent on pancreatic beta-cell functionality, as well as on the beta-cell mass expansion or apoptosis in the pancreas. In this study, we analysed the effects of GSPE on modulating apoptosis and proliferation in beta-cells. We tested the effects of GSPE in the INS-1E pancreatic beta-cell line, either under basal or altered conditions with high glucose, insulin or palmitate levels. GSPE enhanced the pro-apoptotic effect of high glucose and showed clear antiproliferative effects under high glucose, insulin and palmitate conditions. These antiproliferative effects are likely due to high molecular weight compounds contained in the extract. GSPE also modulated pro- and anti-apoptotic markers in the pancreas of rats fed a cafeteria diet, with the effect depending on the dose of GSPE and duration of treatment. Thus, GSPE is able to modulate apoptosis and proliferation of beta-cells under altered, but not basal, conditions. Copyright © 2012 Elsevier Ltd. All rights reserved.

The Neuroprotective Effects of Decursin Isolated from Angelica gigas Nakai Against Amyloid β-Protein-Induced Apoptosis in PC 12 Cells via a Mitochondria-Related Caspase Pathway.

PubMed

Li, Li; Du, Jikun; Zou, Liyi; Xia, Haishan; Wu, Tie; Kim, Yongho; Lee, Yongwoo

2015-08-01

Decursin, purified from Angelica gigas Nakai, has been proven to exert neuroprotective property. Previous study revealed decursin protected the PC12 cells from Aβ25-35-induced oxidative cytotoxicity. The present study aimed to investigate whether decursin could protect PC12 cells from apoptosis caused by Aβ. Our results indicated that pretreatment of PC12 cells with decursin significantly inhibited Aβ25-35-induced cytotoxicity and apoptosis. The mechanism of action is likely to reverse Aβ25-35-induced mitochondrial dysfunction, including the reduction of mitochondrial membrane potential, the inhibition of reactive oxygen species production, and the decrease of mitochondrial release of cytochrome c in PC12 cells. In addition, decursin significantly suppressed the activity of caspase-3 and moderated the ratio of Bcl-2/Bax induced by Aβ25-35. These findings indicate that decursin exerts a neuroprotective effect against Aβ25-35-induced neurotoxicity in PC12 cells, at least in part, via suppressing the mitochondrial pathway of cellular apoptosis.

Proliferation of epithelial cell rests, formation of apical cysts, and regression of apical cysts after periapical wound healing.

PubMed

Lin, Louis M; Huang, George T-J; Rosenberg, Paul A

2007-08-01

There is continuing controversy regarding the potential for inflammatory apical cysts to heal after nonsurgical endodontic therapy. Molecular cell biology may provide answers to a series of related questions. How are the epithelial cell rests of Malassez stimulated to proliferate? How are the apical cysts formed? How does the lining epithelium of apical cysts regress after endodontic therapy? Epithelial cell rests are induced to divide and proliferate by inflammatory mediators, proinflammatory cytokines, and growth factors released from host cells during periradicular inflammation. Quiescent epithelial cell rests can behave like restricted-potential stem cells if stimulated to proliferate. Formation of apical cysts is most likely caused by the merging of proliferating epithelial strands from all directions to form a three-dimensional ball mass. After endodontic therapy, epithelial cells in epithelial strands of periapical granulomas and the lining epithelium of apical cysts may stop proliferating because of a reduction in inflammatory mediators, proinflammatory cytokines, and growth factors. Epithelial cells will also regress because of activation of apoptosis or programmed cell death through deprivation of survival factors or by receiving death signals during periapical wound healing.

The effects of arctigenin on human rheumatoid arthritis fibroblast-like synoviocytes.

PubMed

Liu, Hongbin; Yang, Yang; Cai, Xiaosong; Gao, Yunlong; Du, Jun; Chen, Shuo

2015-08-01

Rheumatoid arthritis fibroblast-like synoviocytes (RAFLSs) play an important role in the initiation and progression of RA, which are resistant to apoptosis and proliferate in an anchorage-independent manner. The effects of arctigenin on the proliferation and apoptosis of RAFLSs were explored. Arctigenin (0-160 µM) was used to treat RAFLSs for 48 h. Cell viability and apoptosis were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide assay and annexin V/propidium iodide staining. Western blot analysis was performed to detect the changes in apoptosis-related genes. Arctigenin decreased cell viability by 23, 30, and 38% at the dose of 10, 20, and 30 µM, respectively. The half maximal inhibitory concentration (IC50) of arctignein on RAFLSs was about 38 µM. Moreover, 9, 15, and 21% of RAFLSs are induced apoptosis by 10, 20, and 30 µM of arctigenin. The apoptotic response was due to the loss of mitochondrial membrane potential, coupled with the release of cytochrome C into cytoplasm, the up-regulation of pro-apoptotic protein, Bax, and down-regulation of antiapoptotic protein, B cell lymphoma 2 (Bcl-2). The activation of mitochondrial pathway in arctigenin-treated RAFLSs induced the cleavage of caspase-9, caspase-3, and poly (ADP-ribose) polymerase (PARP). Additionally, arctigenin inhibited the nuclear translocation of p65, decreased the degradation of inhibitor of kappa B alpha (IκBα), and attenuated the phosphorylation of Akt. Our results reveal that arctigenin inhibits cell proliferation and induces mitochondrial apoptosis of RAFLSs, which is associated with the modulation of NF-κB and Akt signaling pathways.

Relationship between intracellular pH, metabolic co-factors and caspase-3 activation in cancer cells during apoptosis.

PubMed

Sergeeva, Tatiana F; Shirmanova, Marina V; Zlobovskaya, Olga A; Gavrina, Alena I; Dudenkova, Varvara V; Lukina, Maria M; Lukyanov, Konstantin A; Zagaynova, Elena V

2017-03-01

A complex cascade of molecular events occurs in apoptotic cells but cell-to-cell variability significantly complicates determination of the order and interconnections between different processes. For better understanding of the mechanisms of programmed cell death, dynamic simultaneous registration of several parameters is required. In this paper we used multiparameter fluorescence microscopy to analyze energy metabolism, intracellular pH and caspase-3 activation in living cancer cells in vitro during staurosporine-induced apoptosis. We performed metabolic imaging of two co-factors, NAD(P)H and FAD, and used the genetically encoded pH-indicator SypHer1 and the FRET-based sensor for caspase-3 activity, mKate2-DEVD-iRFP, to visualize these parameters by confocal fluorescence microscopy and two-photon fluorescence lifetime imaging microscopy. The correlation between energy metabolism, intracellular pH and caspase-3 activation and their dynamic changes were studied in CT26 cancer cells during apoptosis. Induction of apoptosis was accompanied by a switch to oxidative phosphorylation, cytosol acidification and caspase-3 activation. We showed that alterations in cytosolic pH and the activation of oxidative phosphorylation are relatively early events associated with the induction of apoptosis. Copyright © 2017 Elsevier B.V. All rights reserved.

HAMLET kills tumor cells by apoptosis: structure, cellular mechanisms, and therapy.

PubMed

Gustafsson, Lotta; Hallgren, Oskar; Mossberg, Ann-Kristin; Pettersson, Jenny; Fischer, Walter; Aronsson, Annika; Svanborg, Catharina

2005-05-01

New cancer treatments should aim to destroy tumor cells without disturbing normal tissue. HAMLET (human alpha-lactalbumin made lethal to tumor cells) offers a new molecular approach to solving this problem, because it induces apoptosis in tumor cells but leaves normal differentiated cells unaffected. After partial unfolding and binding to oleic acid, alpha-lactalbumin forms the HAMLET complex, which enters tumor cells and freezes their metabolic machinery. The cells proceed to fragment their DNA, and they disintegrate with apoptosis-like characteristics. HAMLET kills a wide range of malignant cells in vitro and maintains this activity in vivo in patients with skin papillomas. In addition, HAMLET has striking effects on human glioblastomas in a rat xenograft model. After convection-enhanced delivery, HAMLET diffuses throughout the brain, selectively killing tumor cells and controlling tumor progression without apparent tissue toxicity. HAMLET thus shows great promise as a new therapeutic with the advantage of selectivity for tumor cells and lack of toxicity.

Imiquimod activates p53-dependent apoptosis in a human basal cell carcinoma cell line.

PubMed

Huang, Shi-Wei; Chang, Shu-Hao; Mu, Szu-Wei; Jiang, Hsin-Yi; Wang, Sin-Ting; Kao, Jun-Kai; Huang, Jau-Ling; Wu, Chun-Ying; Chen, Yi-Ju; Shieh, Jeng-Jer

2016-03-01

The tumor suppressor p53 controls DNA repair, cell cycle, apoptosis, autophagy and numerous other cellular processes. Imiquimod (IMQ), a synthetic toll-like receptor (TLR) 7 ligand for the treatment of superficial basal cell carcinoma (BCC), eliminates cancer cells by activating cell-mediated immunity and directly inducing apoptosis and autophagy in cancer cells. To evaluate the role of p53 in IMQ-induced cell death in skin cancer cells. The expression, phosphorylation and subcellular localization of p53 were detected by real-time PCR, luciferase reporter assay, cycloheximide chase analysis, immunoblotting and immunocytochemistry. Using BCC/KMC1 cell line as a model, the upstream signaling of p53 activation was dissected by over-expression of TLR7/8, the addition of ROS scavenger, ATM/ATR inhibitors and pan-caspase inhibitor. The role of p53 in IMQ-induced apoptosis and autophagy was assessed by genetically silencing p53 and evaluated by a DNA content assay, immunoblotting, LC3 puncta detection and acridine orange staining. IMQ induced p53 mRNA expression and protein accumulation, increased Ser15 phosphorylation, promoted nuclear translocation and up-regulated its target genes in skin cancer cells in a TLR7/8-independent manner. In BCC/KMC1 cells, the induction of p53 by IMQ was achieved through increased ROS production to stimulate the ATM/ATR-Chk1/Chk2 axis but was not mediated by inducing DNA damage. The pharmacological inhibition of ATM/ATR significantly suppressed IMQ-induced p53 activation and apoptosis. Silencing of p53 significantly decreased the IMQ-induced caspase cascade activation and apoptosis but enhanced autophagy. Mutant p53 skin cancer cell lines were more resistant to IMQ-induced apoptosis than wildtype p53 skin cancer cell lines. IMQ induced ROS production to stimulate ATM/ATR pathways and contributed to p53-dependent apoptosis in a skin basal cell carcinoma cell line BCC/KMC1. Copyright © 2015 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

Fluid shear stress inhibits TNF-alpha-induced apoptosis in osteoblasts: a role for fluid shear stress-induced activation of PI3-kinase and inhibition of caspase-3

NASA Technical Reports Server (NTRS)

Pavalko, Fredrick M.; Gerard, Rita L.; Ponik, Suzanne M.; Gallagher, Patricia J.; Jin, Yijun; Norvell, Suzanne M.

2003-01-01

In bone, a large proportion of osteoblasts, the cells responsible for deposition of new bone, normally undergo programmed cell death (apoptosis). Because mechanical loading of bone increases the rate of new bone formation, we hypothesized that mechanical stimulation of osteoblasts might increase their survival. To test this hypothesis, we investigated the effects of fluid shear stress (FSS) on osteoblast apoptosis using three osteoblast cell types: primary rat calvarial osteoblasts (RCOB), MC3T3-E1 osteoblastic cells, and UMR106 osteosarcoma cells. Cells were treated with TNF-alpha in the presence of cyclohexamide (CHX) to rapidly induce apoptosis. Osteoblasts showed significant signs of apoptosis within 4-6 h of exposure to TNF-alpha and CHX, and application of FSS (12 dyne/cm(2)) significantly attenuated this TNF-alpha-induced apoptosis. FSS activated PI3-kinase signaling, induced phosphorylation of Akt, and inhibited TNF-alpha-induced activation of caspase-3. Inhibition of PI3-kinase, using LY294002, blocked the ability of FSS to rescue osteoblasts from TNF-alpha-induced apoptosis and blocked FSS-induced inhibition of caspase-3 activation in osteoblasts treated with TNF-alpha. LY294002 did not, however, prevent FSS-induced phosphorylation of Akt suggesting that activation of Akt alone is not sufficient to rescue cells from apoptosis. This result also suggests that FSS can activate Akt via a PI3-kinase-independent pathway. These studies demonstrate for the first time that application of FSS to osteoblasts in vitro results in inhibition of TNF-alpha-induced apoptosis through a mechanism involving activation of PI3-kinase signaling and inhibition of caspases. FSS-induced activation of PI3-kinase may promote cell survival through a mechanism that is distinct from the Akt-mediated survival pathway. Copyright 2002 Wiley-Liss, Inc.

Proliferate and survive: cell division cycle and apoptosis in human neuroblastoma.

PubMed

Borriello, Adriana; Roberto, Roberta; Della Ragione, Fulvio; Iolascon, Achille

2002-02-01

Neuroblastoma is one of the most frequent childhood cancers and a major cause of death from neoplasias of infancy. Although a wealth of studies on its molecular bases have been carried out, little conclusive information about its origin and evolution is available. Some intriguing findings have correlated neuroblastoma development with aberrations of two pivotal cellular processes generally altered in human cancers, namely cell division cycle and apoptosis. Indeed, it has been reported that neuroblastoma cell lines show accumulation of Id2 protein, a factor which is able to hamper the pRb protein antiproliferative activity. The increased Id2 is due to N-myc gene amplification and overexpression, a phenomenon frequently observed in neuroblastoma and an important independent negative marker. Moreover, neuroblastoma cells are frequently characterized by increased levels of survivin, an inhibitor of the apoptotic response, and by a deficiency of procaspase 8, a key intermediate of the programmed cell death cascade. These two events, probably, make neuroblastomas more resistant to programmed cell death. These recent findings might suggest that neuroblastoma cells have acquired the capability to proliferate easily and die difficultly. The mechanistic meaning of these data will be discussed in the present review. Moreover, we will suggest new therapeutic scenarios opened up by the described alterations of cell cycle and apoptosis engines.

‘Hints' in the killer protein gasdermin D: unveiling the secrets of gasdermins driving cell death

PubMed Central

Qiu, Shiqiao; Liu, Jing; Xing, Feiyue

2017-01-01

Pyroptosis is a lytic form of cell death distinguished from apoptosis, ferroptosis, necrosis, necroptosis, NETosis, oncosis, pyronecrosis and autophagy. Proinflammatory caspases cleave a gasdermin D (GSDMD) protein to generate a 31 kDa N-terminal domain. The cleavage relieves the intramolecular inhibition on the gasdermin-N domain, which then moves to the plasma membrane to exhibit pore-forming activity. Thus, GSDMD acts as the final and direct executor of pyroptotic cell death. Owing to the selective targeting of the inner leaflet of the plasma membrane with the pore-forming that determines pyroptotic cell death, GSDMD could be a potential target to control cell death or extracellular bacterial infections. Intriguingly, other gasdermin family members also share similar N-terminal domains, but they present different cell death programs. Herein, we summarize features and functions of the novel player proteins in cell death, including GSDMD triggering pyroptosis, Gsdma3/GSDMA initiating autophagy/apoptosis and DFNA5 inducing apoptosis/secondary necrosis. The gasdermin N terminus appears to be a novel pore-forming protein. This provides novel insight into the underlying roles and mechanisms of lytic or nonlytic forms of programmed cell death, as well as their potential applications in inflammation-associated diseases. PMID:28362726

Neutrophil Apoptosis: Relevance to the Innate Immune Response and Inflammatory Disease

PubMed Central

Fox, Sarah; Leitch, Andrew E.; Duffin, Rodger; Haslett, Christopher; Rossi, Adriano G.

2010-01-01

Neutrophils are the most abundant cell type involved in the innate immune response. They are rapidly recruited to sites of injury or infection where they engulf and kill invading microorganisms. Neutrophil apoptosis, the process of programmed cell death that prevents the release of neutrophil histotoxic contents, is tightly regulated and limits the destructive capacity of neutrophil products to surrounding tissue. The subsequent recognition and phagocytosis of apoptotic cells by phagocytic cells such as macrophages is central to the successful resolution of an inflammatory response and it is increasingly apparent that the dying neutrophil itself exerts an anti-inflammatory effect through modulation of surrounding cell responses, particularly macrophage inflammatory cytokine release. Apoptosis may be delayed, induced or enhanced by micro-organisms dependent on their immune evasion strategies and the health of the host they encounter. There is now an established field of research aimed at understanding the regulation of apoptosis and its potential as a target for therapeutic intervention in inflammatory and infective diseases. This review focuses on the physiological regulation of neutrophil apoptosis with respect to the innate immune system and highlights recent advances in mechanistic understanding of apoptotic pathways and their therapeutic manipulation in appropriate and excessive innate immune responses. PMID:20375550

Receptor Interactive Protein Kinase 3 Promotes Cisplatin-Triggered Necrosis in Apoptosis-Resistant Esophageal Squamous Cell Carcinoma Cells

PubMed Central

Zhao, Nan; Zhou, Lanping; Liu, Fang; Cichacz, Zbigniew; Zhang, Lin; Zhan, Qimin; Zhao, Xiaohang

2014-01-01

Cisplatin-based chemotherapy is currently the standard treatment for locally advanced esophageal cancer. Cisplatin has been shown to induce both apoptosis and necrosis in cancer cells, but the mechanism by which programmed necrosis is induced remains unknown. In this study, we provide evidence that cisplatin induces necrotic cell death in apoptosis-resistant esophageal cancer cells. This cell death is dependent on RIPK3 and on necrosome formation via autocrine production of TNFα. More importantly, we demonstrate that RIPK3 is necessary for cisplatin-induced killing of esophageal cancer cells because inhibition of RIPK1 activity by necrostatin or knockdown of RIPK3 significantly attenuates necrosis and leads to cisplatin resistance. Moreover, microarray analysis confirmed an anti-apoptotic molecular expression pattern in esophageal cancer cells in response to cisplatin. Taken together, our data indicate that RIPK3 and autocrine production of TNFα contribute to cisplatin sensitivity by initiating necrosis when the apoptotic pathway is suppressed or absent in esophageal cancer cells. These data provide new insight into the molecular mechanisms underlying cisplatin-induced necrosis and suggest that RIPK3 is a potential marker for predicting cisplatin sensitivity in apoptosis-resistant and advanced esophageal cancer. PMID:24959694

Inactivation of the small GTP binding protein Rho induces multinucleate cell formation and apoptosis in murine T lymphoma EL4.

PubMed

Moorman, J P; Bobak, D A; Hahn, C S

1996-06-01

The small G-protein Rho regulates the actin microfilament-dependent cytoskeleton. Exoenzyme C3 of Clostridium botulinum ADP-ribosylates Rho at Asn41, a modification that functionally inactivates Rho. Using a Sindbis virus-based transient gene expression system, we studied the role of Rho in murine EL4 T lymphoma cells. We generated a double subgenomic infectious Sindbis virus (dsSIN:C3) recombinant which expressed C3 in >95% of EL4 cells. This intracellular C3 resulted in modification and inactivation of virtually all endogenous Rho. dsSIN:C3 infection led to the formation of multinucleate cells, likely by inhibiting the actin microfilament-dependent step of cytokinesis. Intriguingly, in spite of the inhibition of cytokinesis, karyokinesis continued, with the result that cells containing a nuclear DNA content as high as 16N (eight nuclei) were observed. In addition, dsSIN:C3-mediated inactivation of Rho was a potent activator of apoptosis in EL4 cells. To discern whether the formation of multinucleate cells was responsible for the activation of apoptosis, 5-fluorouracil (5-FUra) was used to induce cell cycle arrest. As expected, EL4 cells treated with 5-FUra were prevented from forming multinucleate cells upon infection with dsSIN:C3. dsSIN:C3 infection, however, still caused marked apoptosis in 5-FUra-treated cells, indicating that this activation of apoptosis was independent of multinucleate cell formation.

Detecting contaminant-induced apoptosis and necrosis in lake trout thymocytes via flow cytometry.

USGS Publications Warehouse

Sweet, Leonard I.; Passino-Reader, Dora R.; Meier, Peter G.; Omann, Geneva M.; Stolen, J.S.; Fletcher, T.C.; Rowley, A.F.; Zelikoff, J.T.; Kaattari, S.L.; Smith, S.A.

1997-01-01

This chapter details the cytofluorometric techniques employed to assess levels of active (apoptosis) and passive (necrotic) cell death in untreated and contaminant-treated fish thymocytes. The thymus is believed to be a central component of hematopoiesis and immune function in teleosts (Abelli et al., 1996). Hence, chemically-elicited adverse effects to the thymus may result in immunomodulation and organ dysfunction. However, it is not well documented that environmental contaminants induce apoptosis, or programmed cell death. There is some evidence suggesting that low level exposure to waterborne contaminants can specifically induce cell death in the olfactory epithelium of rainbow trout (Julliard et al., 1996). Presently, only limited information is available in the literature regarding apoptotic death in piscine immune cells (Alford et al., 1994; Greenlee et al., 1991).

Mitochondrial membrane permeabilization and cell death during myocardial infarction: roles of calcium and reactive oxygen species

PubMed Central

Webster, Keith A

2013-01-01

Excess generation of reactive oxygen species (ROS) and cytosolic calcium accumulation play major roles in the initiation of programmed cell death during acute myocardial infarction. Cell death may include necrosis, apoptosis and autophagy, and combinations thereof. During ischemia, calcium handling between the sarcoplasmic reticulum and myofilament is disrupted and calcium is diverted to the mitochondria causing swelling. Reperfusion, while essential for survival, reactivates energy transduction and contractility and causes the release of ROS and additional ionic imbalance. During acute ischemia–reperfusion, the principal death pathways are programmed necrosis and apoptosis through the intrinsic pathway, initiated by the opening of the mitochondrial permeability transition pore and outer mitochondrial membrane permeabilization, respectively. Despite intense investigation, the mechanisms of action and modes of regulation of mitochondrial membrane permeabilization are incompletely understood. Extrinsic apoptosis, necroptosis and autophagy may also contribute to ischemia–reperfusion injury. In this review, the roles of dysregulated calcium and ROS and the contributions of Bcl-2 proteins, as well as mitochondrial morphology in promoting mitochondrial membrane permeability change and the ensuing cell death during myocardial infarction are discussed. PMID:23176689

The role of apoptosis in LDL transport through cultured endothelial cell monolayers

PubMed Central

Cancel, Limary M.; Tarbell, John M.

2009-01-01

We have previously shown that leaky junctions associated with dying or dividing cells are the dominant pathway for LDL transport under convective conditions, accounting for more than 90% of the transport [1]. To explore the role of apoptosis in the leaky junction pathway, TNFα and cycloheximide (TNFα/CHX) were used to induce an elevated rate of apoptosis in cultured bovine aortic endothelial cell (BAEC) monolayers and the convective fluxes of LDL and water were measured. Treatment with TNFα/CHX induced a 18.3-fold increase in apoptosis and a 4.4-fold increase in LDL permeability. Increases in apoptosis and permeability were attenuated by treatment with the caspase inhibitor Z-VAD-FMK. Water flux increased by 2.7-fold after treatment with TNFα/CHX, and this increase was not attenuated by treatment with Z-VAD-FMK. Immunostaining of the tight junction protein ZO-1 showed that TNFα/CHX treatment disrupts the tight junction in addition to inducing apoptosis. This disruption is present even when Z-VAD-FMK is used to inhibit apoptosis, and likely accounts for the increase in water flux. We found a strong correlation between the rate of apoptosis and the permeability of BAEC monolayers to LDL. These results demonstrate the potential of manipulating endothelial monolayer permeability by altering the rate of apoptosis pharmacollogicaly. This has implications for the treatment of atherosclerosis. PMID:19709659

A new lactoferrin- and iron-dependent lysosomal death pathway is induced by benzo[a]pyrene in hepatic epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gorria, Morgane; Tekpli, Xavier; Rissel, Mary

2008-04-15

While lysosomal disruption seems to be a late step of necrosis, a moderate lysosomal destabilization has been suggested to participate early in the apoptotic cascade. The origin of lysosomal dysfunction and its precise role in apoptosis or apoptosis-like process still needs to be clarified, especially upon carcinogen exposure. In this study, we focused on the implication of lysosomes in cell death induced by the prototype carcinogen benzo[a]pyrene (B[a]P; 50 nM) in rat hepatic epithelial F258 cells. We first demonstrated that B[a]P affected lysosomal morphology (increase in size) and pH (alkalinization), and that these changes were involved in caspase-3 activation andmore » cell death. Subsequently, we showed that lysosomal modifications were partly dependent on mitochondrial dysfunction, and that lysosomes together with mitochondria participate in B[a]P-induced oxidative stress. Using two iron chelators (desferrioxamine and deferiprone) and siRNA targeting the lysosomal iron-binding protease lactoferrin, we further demonstrated that both lysosomal iron content and lactoferrin were required for caspase-3 activation and apoptosis-like cell death.« less

Concanavalin A: A potential anti-neoplastic agent targeting apoptosis, autophagy and anti-angiogenesis for cancer therapeutics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Wen-wen; Yu, Jia-ying; Xu, Huai-long

2011-10-22

Highlights: {yields} ConA induces cancer cell death targeting apoptosis and autophagy. {yields} ConA inhibits cancer cell angiogenesis. {yields} ConA is utilized in pre-clinical and clinical trials. -- Abstract: Concanavalin A (ConA), a Ca{sup 2+}/Mn{sup 2+}-dependent and mannose/glucose-binding legume lectin, has drawn a rising attention for its remarkable anti-proliferative and anti-tumor activities to a variety of cancer cells. ConA induces programmed cell death via mitochondria-mediated, P73-Foxo1a-Bim apoptosis and BNIP3-mediated mitochondrial autophagy. Through IKK-NF-{kappa}B-COX-2, SHP-2-MEK-1-ERK, and SHP-2-Ras-ERK anti-angiogenic pathways, ConA would inhibit cancer cell survival. In addition, ConA stimulates cell immunity and generates an immune memory, resisting to the same genotypic tumor.more » These biological findings shed light on new perspectives of ConA as a potential anti-neoplastic agent targeting apoptosis, autophagy and anti-angiogenesis in pre-clinical or clinical trials for cancer therapeutics.« less

Microplasma Induced Cell Morphological Changes and Apoptosis of Ex Vivo Cultured Human Anterior Lens Epithelial Cells – Relevance to Capsular Opacification

PubMed Central

Hojnik, Nataša; Filipič, Gregor; Lazović, Saša; Vesel, Alenka; Primc, Gregor; Mozetič, Miran; Hawlina, Marko; Petrovski, Goran; Cvelbar, Uroš

2016-01-01

Inducing selective or targeted cell apoptosis without affecting large number of neighbouring cells remains a challenge. A plausible method for treatment of posterior capsular opacification (PCO) due to remaining lens epithelial cells (LECs) by reactive chemistry induced by localized single electrode microplasma discharge at top of a needle-like glass electrode with spot size ~3 μm is hereby presented. The focused and highly-localized atmospheric pressure microplasma jet with electrode discharge could induce a dose-dependent apoptosis in selected and targeted individual LECs, which could be confirmed by real-time monitoring of the morphological and structural changes at cellular level. Direct cell treatment with microplasma inside the medium appeared more effective in inducing apoptosis (caspase 8 positivity and DNA fragmentation) at a highly targeted cell level compared to treatment on top of the medium (indirect treatment). Our results show that single cell specific micropipette plasma can be used to selectively induce demise in LECs which remain in the capsular bag after cataract surgery and thus prevent their migration (CXCR4 positivity) to the posterior lens capsule and PCO formation. PMID:27832099

Effects of S-adenosylmethionine decarboxylase, polyamines, amino acids, and weak bases (amines and ammonia) on development and ribosomal RNA synthesis in Xenopus embryos.

PubMed

Shiokawa, Koichiro; Aso, Mai; Kondo, Takeshi; Takai, Jun-Ichi; Yoshida, Junki; Mishina, Takamichi; Fuchimukai, Kota; Ogasawara, Tsukasa; Kariya, Taro; Tashiro, Kosuke; Igarashi, Kazuei

2010-02-01

We have been studying control mechanisms of gene expression in early embryogenesis in a South African clawed toad Xenopus laevis, especially during the period of midblastula transition (MBT), or the transition from the phase of active cell division (cleavage stage) to the phase of extensive morphogenesis (post-blastular stages). We first found that ribosomal RNA synthesis is initiated shortly after MBT in Xenopus embryos and those weak bases, such as amines and ammonium ion, selectively inhibit the initiation and subsequent activation of rRNA synthesis. We then found that rapidly labeled heterogeneous mRNA-like RNA is synthesized in embryos at pre-MBT stage. We then performed cloning and expression studies of several genes, such as those for activin receptors, follistatin and aldolases, and then reached the studies of S-adenosylmethionine decarboxylase (SAMDC), a key enzyme in polyamine metabolism. Here, we cloned a Xenopus SAMDC cDNA and performed experiments to overexpress the in vitro-synthesized SAMDC mRNA in Xenopus early embryos, and found that the maternally preset program of apoptosis occurs in cleavage stage embryos, which is executed when embryos reach the stage of MBT. In the present article, we first summarize results on SAMDC and the maternal program of apoptosis, and then describe our studies on small-molecular-weight substances like polyamines, amino acids, and amines in Xenopus embryos. Finally, we summarize our studies on weak bases, especially on ammonium ion, as the specific inhibitor of ribosomal RNA synthesis in Xenopus embryonic cells.

Amarogentin Induces Apoptosis of Liver Cancer Cells via Upregulation of p53 and Downregulation of Human Telomerase Reverse Transcriptase in Mice.

PubMed

Huang, Chun; Li, Runqin; Zhang, Yinglin; Gong, Jianping

2017-10-01

Amarogentin has been reported to have a preventive effect on liver cancer via inducing cancer cell apoptosis. We attempted to elucidate the roles of p53-associated apoptosis pathways in the chemopreventive mechanism of amarogentin. The findings of this study will facilitate the development of a novel supplementary strategy for the treatment of liver cancer. The purity of amarogentin was assessed by high-performance liquid chromatography. The inhibitory ratios of the liver cell lines were determined using a Cell Counting Kit-8 following treatment with a gradient concentration of amarogentin. Cell apoptosis was detected by flow cytometry using annexin V-fluorescein isothiocyanate/propidium iodide kits. The gene and protein expression of p53-associated molecules, such as Akt, human telomerase reverse transcriptase, RelA, and p38, was detected by real-time quantitative polymerase chain reaction, Western blotting, and immunohistochemical staining in liver cancer cells and mouse tumor tissues after treatment with amarogentin. The inhibitory effect of amarogentin on cell proliferation was more obvious in liver cancer cells, and amarogentin was more likely to induce the apoptosis of liver cancer cells than that of normal liver cells. The gene and protein expression levels of Akt, RelA, and human telomerase reverse transcriptase were markedly higher in the control group than in the preventive group and treatment groups. Only the expression of human telomerase reverse transcriptase was downregulated, accompanied by the upregulation of p53. The results of our study suggest that amarogentin promotes apoptosis of liver cancer cells by the upregulation of p53 and downregulation of human telomerase reverse transcriptase and prevents the malignant transformation of these cells.

Amarogentin Induces Apoptosis of Liver Cancer Cells via Upregulation of p53 and Downregulation of Human Telomerase Reverse Transcriptase in Mice

PubMed Central

Li, Runqin; Zhang, Yinglin

2016-01-01

Background and Objective: Amarogentin has been reported to have a preventive effect on liver cancer via inducing cancer cell apoptosis. We attempted to elucidate the roles of p53-associated apoptosis pathways in the chemopreventive mechanism of amarogentin. The findings of this study will facilitate the development of a novel supplementary strategy for the treatment of liver cancer. Materials and Methods: The purity of amarogentin was assessed by high-performance liquid chromatography. The inhibitory ratios of the liver cell lines were determined using a Cell Counting Kit-8 following treatment with a gradient concentration of amarogentin. Cell apoptosis was detected by flow cytometry using annexin V-fluorescein isothiocyanate/propidium iodide kits. The gene and protein expression of p53-associated molecules, such as Akt, human telomerase reverse transcriptase, RelA, and p38, was detected by real-time quantitative polymerase chain reaction, Western blotting, and immunohistochemical staining in liver cancer cells and mouse tumor tissues after treatment with amarogentin. Results: The inhibitory effect of amarogentin on cell proliferation was more obvious in liver cancer cells, and amarogentin was more likely to induce the apoptosis of liver cancer cells than that of normal liver cells. The gene and protein expression levels of Akt, RelA, and human telomerase reverse transcriptase were markedly higher in the control group than in the preventive group and treatment groups. Only the expression of human telomerase reverse transcriptase was downregulated, accompanied by the upregulation of p53. Conclusion: The results of our study suggest that amarogentin promotes apoptosis of liver cancer cells by the upregulation of p53 and downregulation of human telomerase reverse transcriptase and prevents the malignant transformation of these cells. PMID:27402632

Different ways to die: cell death modes of the unicellular chlorophyte Dunaliella viridis exposed to various environmental stresses are mediated by the caspase-like activity DEVDase.

PubMed

Jiménez, Carlos; Capasso, Juan M; Edelstein, Charles L; Rivard, Christopher J; Lucia, Scott; Breusegem, Sophia; Berl, Tomás; Segovia, María

2009-01-01

Programmed cell death is necessary for homeostasis in multicellular organisms and it is also widely recognized to occur in unicellular organisms. However, the mechanisms through which it occurs in unicells, and the enzymes involved within the final response is still the subject of heated debate. It is shown here that exposure of the unicellular microalga Dunaliella viridis to several environmental stresses, induced different cell death morphotypes, depending on the stimulus received. Senescent cells demonstrated classical and unambiguous apoptotic-like characteristics such as chromatin condensation, DNA fragmentation, intact organelles, and blebbing of the cell membrane. Acute heat shock caused general swelling and altered plasma membrane, but the presence of chromatin clusters and DNA strand breaks suggested a necrotic-like event. UV irradiated cells presented changes typical for necrosis, together with apoptotic characteristics resembling an intermediate cell-death phenotype termed aponecrosis-like. Cells subjected to hyperosmotic shock revealed chromatin spotting without DNA fragmentation, and extensive cytoplasmic swelling and vacuolization, comparable to a paraptotic-like cell death phenotype. Nitrogen-starved cells showed pyknosis, blebbing, and cytoplasmic consumption, indicating a similarity to autophagic/vacuolar-like cell death. The caspase-like activity DEVDase was measured by using the fluorescent substrate Ac-DEVD-AMC and antibodies against the human caspase-3 active enzyme cross-reacted with bands, the intensity of which paralleled the activity. All the environmental stresses tested produced a substantial increase in both DEVDase activity and protein levels. The irreversible caspase-3 inhibitor Z-DEVD-FMK completely inhibited the enzymatic activity whereas serine and aspartyl proteases inhibitors did not. These results show that cell death in D. viridis does not conform to a single pattern and that environmental stimuli may produce different types of cell death depending on the type and intensity of the stimulus, all of which help to understand the cell death-dependent and cell death-independent functions of caspase-like proteins. Hence, these data support the theory that alternative, non-apoptotic programmed cell death (PCDs), exist either in parallel or in an independent manner with apoptosis and were already present in single-celled organisms that evolved some 1.2-1.6 billion years ago.

Different ways to die: cell death modes of the unicellular chlorophyte Dunaliella viridis exposed to various environmental stresses are mediated by the caspase-like activity DEVDase

PubMed Central

Jiménez, Carlos; Capasso, Juan M.; Edelstein, Charles L.; Rivard, Christopher J.; Lucia, Scott; Breusegem, Sophia; Berl, Tomás; Segovia, María

2009-01-01

Programmed cell death is necessary for homeostasis in multicellular organisms and it is also widely recognized to occur in unicellular organisms. However, the mechanisms through which it occurs in unicells, and the enzymes involved within the final response is still the subject of heated debate. It is shown here that exposure of the unicellular microalga Dunaliella viridis to several environmental stresses, induced different cell death morphotypes, depending on the stimulus received. Senescent cells demonstrated classical and unambiguous apoptotic-like characteristics such as chromatin condensation, DNA fragmentation, intact organelles, and blebbing of the cell membrane. Acute heat shock caused general swelling and altered plasma membrane, but the presence of chromatin clusters and DNA strand breaks suggested a necrotic-like event. UV irradiated cells presented changes typical for necrosis, together with apoptotic characteristics resembling an intermediate cell-death phenotype termed aponecrosis-like. Cells subjected to hyperosmotic shock revealed chromatin spotting without DNA fragmentation, and extensive cytoplasmic swelling and vacuolization, comparable to a paraptotic-like cell death phenotype. Nitrogen-starved cells showed pyknosis, blebbing, and cytoplasmic consumption, indicating a similarity to autophagic/vacuolar-like cell death. The caspase-like activity DEVDase was measured by using the fluorescent substrate Ac-DEVD-AMC and antibodies against the human caspase-3 active enzyme cross-reacted with bands, the intensity of which paralleled the activity. All the environmental stresses tested produced a substantial increase in both DEVDase activity and protein levels. The irreversible caspase-3 inhibitor Z-DEVD-FMK completely inhibited the enzymatic activity whereas serine and aspartyl proteases inhibitors did not. These results show that cell death in D. viridis does not conform to a single pattern and that environmental stimuli may produce different types of cell death depending on the type and intensity of the stimulus, all of which help to understand the cell death-dependent and cell death-independent functions of caspase-like proteins. Hence, these data support the theory that alternative, non-apoptotic programmed cell death (PCDs), exist either in parallel or in an independent manner with apoptosis and were already present in single-celled organisms that evolved some 1.2-1.6 billion years ago. PMID:19251986

BAX to basics: How the BCL2 gene family controls the death of retinal ganglion cells

PubMed Central

Maes, Margaret E.; Schlamp, Cassandra L.; Nickells, Robert W.

2017-01-01

Retinal ganglion cell (RGC) death is the principal consequence of injury to the optic nerve. For several decades, we have understood that the RGC death process was executed by apoptosis, suggesting that there may be ways to therapeutically intervene in this cell death program and provide a more direct treatment to the cells and tissues affected in diseases like glaucoma. A major part of this endeavor has been to elucidate the molecular biological pathways active in RGCs from the point of axonal injury to the point of irreversible cell death. A major component of this process is the complex interaction of members of the BCL2 gene family. Three distinct family members of proteins orchestrate the most critical junction in the apoptotic program of RGCs, culminating in the activation of pro-apoptotic BAX. Once active, BAX causes irreparable damage to mitochondria, while precipitating downstream events that finish off a dying ganglion cell. This review is divided into two major parts. First, we summarize the extent of knowledge of how BCL2 gene family proteins interact to facilitate the activation and function of BAX. This area of investigation has rapidly changed over the last few years and has yielded a dramatically different mechanistic understanding of how the intrinsic apoptotic program is run in mammalian cells. Second, we provided a comprehensive analysis of nearly two decades of investigation of the role of BAX in the process of RGC death, much of which has provided many important insights into the overall pathophysiology of diseases like glaucoma. PMID:28064040

BAX to basics: How the BCL2 gene family controls the death of retinal ganglion cells.

PubMed

Maes, Margaret E; Schlamp, Cassandra L; Nickells, Robert W

2017-03-01

Retinal ganglion cell (RGC) death is the principal consequence of injury to the optic nerve. For several decades, we have understood that the RGC death process was executed by apoptosis, suggesting that there may be ways to therapeutically intervene in this cell death program and provide a more direct treatment to the cells and tissues affected in diseases like glaucoma. A major part of this endeavor has been to elucidate the molecular biological pathways active in RGCs from the point of axonal injury to the point of irreversible cell death. A major component of this process is the complex interaction of members of the BCL2 gene family. Three distinct family members of proteins orchestrate the most critical junction in the apoptotic program of RGCs, culminating in the activation of pro-apoptotic BAX. Once active, BAX causes irreparable damage to mitochondria, while precipitating downstream events that finish off a dying ganglion cell. This review is divided into two major parts. First, we summarize the extent of knowledge of how BCL2 gene family proteins interact to facilitate the activation and function of BAX. This area of investigation has rapidly changed over the last few years and has yielded a dramatically different mechanistic understanding of how the intrinsic apoptotic program is run in mammalian cells. Second, we provided a comprehensive analysis of nearly two decades of investigation of the role of BAX in the process of RGC death, much of which has provided many important insights into the overall pathophysiology of diseases like glaucoma. Copyright © 2017 Elsevier Ltd. All rights reserved.

Beyond apoptosis: The mechanism and function of phosphatidylserine asymmetry in the membrane of activating mast cells

PubMed Central

Rysavy, Noel M.; Shimoda, Lori M. N.; Dixon, Alyssa M.; Speck, Mark; Stokes, Alexander J.; Turner, Helen; Umemoto, Eric Y.

2014-01-01

Loss of plasma membrane asymmetry is a hallmark of apoptosis, but lipid bilayer asymmetry and loss of asymmetry can contribute to numerous cellular functions and responses that are independent of programmed cell death. Exofacial exposure of phosphatidylserine occurs in lymphocytes and mast cells after antigenic stimulation and in the absence of apoptosis, suggesting that there is a functional requirement for phosphatidylserine exposure in immunocytes. In this review we examine current ideas as to the nature of this functional role in mast cell activation. Mechanistically, there is controversy as to the candidate proteins responsible for phosphatidylserine translocation from the internal to external leaflet, and here we review the candidacies of mast cell PLSCR1 and TMEM16F. Finally we examine the potential relationship between functionally important mast cell membrane perturbations and phosphatidylserine exposure during activation. PMID:25759911

Lovastatin induces apoptosis of ovarian cancer cells and synergizes with doxorubicin: potential therapeutic relevance.

PubMed

Martirosyan, Anna; Clendening, James W; Goard, Carolyn A; Penn, Linda Z

2010-03-18

Ovarian carcinoma is a rarely curable disease, for which new treatment options are required. As agents that block HMG-CoA reductase and the mevalonate pathway, the statin family of drugs are used in the treatment of hypercholesterolemia and have been shown to trigger apoptosis in a tumor-specific manner. Recent clinical trials show that the addition of statins to traditional chemotherapeutic strategies can increase efficacy of targeting statin-sensitive tumors. Our goal was to assess statin-induced apoptosis of ovarian cancer cells, either alone or in combination with chemotherapeutics, and then determine these mechanisms of action. The effect of lovastatin on ovarian cancer cell lines was evaluated alone and in combination with cisplatin and doxorubicin using several assays (MTT, TUNEL, fixed PI, PARP cleavage) and synergy determined by evaluating the combination index. The mechanisms of action were evaluated using functional, molecular, and pharmacologic approaches. We demonstrate that lovastatin induces apoptosis of ovarian cancer cells in a p53-independent manner and synergizes with doxorubicin, a chemotherapeutic agent used to treat recurrent cases of ovarian cancer. Lovastatin drives ovarian tumor cell death by two mechanisms: first, by blocking HMG-CoA reductase activity, and second, by sensitizing multi-drug resistant cells to doxorubicin by a novel mevalonate-independent mechanism. This inhibition of drug transport, likely through inhibition of P-glycoprotein, potentiates both DNA damage and tumor cell apoptosis. The results of this research provide pre-clinical data to warrant further evaluation of statins as potential anti-cancer agents to treat ovarian carcinoma. Many statins are inexpensive, off-patent generic drugs that are immediately available for use as anti-cancer agents. We provide evidence that lovastatin triggers apoptosis of ovarian cancer cells as a single agent by a mevalonate-dependent mechanism. Moreover, we also show lovastatin synergizes with doxorubicin, an agent administered for recurrent disease. This synergy occurs by a novel mevalonate-independent mechanism that antagonizes drug resistance, likely by inhibiting P-glycoprotein. These data raise important issues that may impact how statins can best be included in chemotherapy regimens.

INF-γ sensitizes head and neck squamous cell carcinoma cells to chemotherapy-induced apoptosis and necroptosis through up-regulation of Egr-1.

PubMed

Xu, Bei; Shu, Yongqian; Liu, Peng

2014-11-01

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. Acquired resistance to standard chemotherapy accounts for most of treatment failure. Here we demonstrate that Interferon-γ (INF-γ) may up-regulate Egr-1 gene expression in HNSCC cell line SCC-25. Forced expression of Egr-1 sensitizes SCC-25 cells to chemotherapy-induced apoptosis and necroptosis, a novel form of programmed cell death. Egr-1 up-regulation also significantly increases the production of Thrombospondin-1 (TSP-1), a matricellular glycoprotein which has been described to induce cell death in HNSCC. Moreover, INF-γ-induced sensitization of cells to chemotherapy-mediated cell death and TSP-1 production could be markedly abolished by Egr-1 silencing. The present investigation provides the first evidence that INF-γ may sensitize HNSCC cells to chemotherapy-induced apoptosis and necroptosis through up-regulation of Egr-1. These data support the combination use of INF-γ and cytotoxic drugs for HNSCC Therapy.

EGCG inhibits Cd(2+)-induced apoptosis through scavenging ROS rather than chelating Cd(2+) in HL-7702 cells.

PubMed

An, Zhen; Qi, Yongmei; Huang, Dejun; Gu, Xueyan; Tian, Yihong; Li, Ping; Li, Hui; Zhang, Yingmei

2014-05-01

Epigallocatechin-3-gallat (EGCG), the major catechin in green tea, shows a potential protective effect against heavy metal toxicity to humans. Apoptosis is one of the key events in cadmium (Cd(2+))-induced cytotoxicity. Nevertheless, the study of EGCG on Cd(2+)-induced apoptosis is rarely reported. The objective of this study was to clarify the effect and detailed mechanism of EGCG on Cd(2+)-induced apoptosis. Normal human liver cells (HL-7702) were treated with Cd(2+) for 21 h, and then co-treated with EGCG for 3 h. Cell viability, apoptosis, intracellular reactive oxygen species (ROS), malondialdehyde (MDA), mitochondrial membrane potential (MMP) and caspase-3 activity were detected. On the other hand, the chelation of Cd(2+) with EGCG was tested by UV-Vis spectroscopy analysis and Nuclear Magnetic Resonance ((1)H NMR) spectroscopy under neutral condition (pH 7.2). Cd(2+) significantly decreased the cell viability and induced apoptosis in HL-7702 cells. Conversely, EGCG co-treatment resulted in significant inhibition of Cd(2+)-induced reduction of cell viability and apoptosis, implying a rescue effect of EGCG against Cd(2+) poisoning. The protective effect most likely arises from scavenging ROS and maintaining redox homeostasis, as the generation of intracellular ROS and MDA is significantly reduced by EGCG, which further prevents MMP collapse and suppresses caspase-3 activity. However, no evidence is observed for the chelation of EGCG with Cd(2+) under neutral condition. Therefore, a clear conclusion from this work can be made that EGCG could inhibit Cd(2+)-induced apoptosis by acting as a ROS scavenger rather than a metal chelating agent.

Interaction of AIM with insulin-like growth factor-binding protein-4.

PubMed

You, Qiang; Wu, Yan; Yao, Nannan; Shen, Guannan; Zhang, Ying; Xu, Liangguo; Li, Guiying; Ju, Cynthia

2015-09-01

Apoptosis inhibitor of macrophages (AIM/cluster of differentiation 5 antigen-like/soluble protein α) has been shown to inhibit cellular apoptosis; however, the underlying molecular mechanism has not been elucidated. Using yeast two‑hybrid screening, the present study uncovered that AIM binds to insulin‑like growth factor binding protein‑4 (IGFBP‑4). AIM interaction with IGFBP‑4, as well as IGFBP‑2 and ‑3, but not with IGFBP‑1, ‑5 and ‑6, was further confirmed by co‑immunoprecipitation (co‑IP) using 293 cells. The binding activity and affinity between AIM and IGFBP‑4 in vitro were analyzed by co‑IP and biolayer interferometry. Serum depletion‑induced cellular apoptosis was attenuated by insulin‑like growth factor‑I (IGF‑I), and this effect was abrogated by IGFBP‑4. Of note, in the presence of AIM, the inhibitory effect of IGFBP‑4 on the anti‑apoptosis function of IGF‑I was attenuated, possibly through binding of AIM with IGFBP‑4. In conclusion, to the best of our knowledge, the present study provides the first evidence that AIM binds to IGFBP‑2, ‑3 and ‑4. The data suggest that this interaction may contribute to the mechanism of AIM-mediated anti-apoptosis function.

Interaction of AIM with insulin-like growth factor-binding protein-4

PubMed Central

YOU, QIANG; WU, YAN; YAO, NANNAN; SHEN, GUANNAN; ZHANG, YING; XU, LIANGGUO; LI, GUIYING; JU, CYNTHIA

2015-01-01

Apoptosis inhibitor of macrophages (AIM/cluster of differentiation 5 antigen-like/soluble protein α) has been shown to inhibit cellular apoptosis; however, the underlying molecular mechanism has not been elucidated. Using yeast two-hybrid screening, the present study uncovered that AIM binds to insulin-like growth factor binding protein-4 (IGFBP-4). AIM interaction with IGFBP-4, as well as IGFBP-2 and -3, but not with IGFBP-1, -5 and -6, was further confirmed by co-immunoprecipitation (co-IP) using 293 cells. The binding activity and affinity between AIM and IGFBP-4 in vitro were analyzed by co-IP and biolayer interferometry. Serum depletion-induced cellular apoptosis was attenuated by insulin-like growth factor-I (IGF-I), and this effect was abrogated by IGFBP-4. Of note, in the presence of AIM, the inhibitory effect of IGFBP-4 on the anti-apoptosis function of IGF-I was attenuated, possibly through binding of AIM with IGFBP-4. In conclusion, to the best of our knowledge, the present study provides the first evidence that AIM binds to IGFBP-2, -3 and -4. The data suggest that this interaction may contribute to the mechanism of AIM-mediated anti-apoptosis function. PMID:26135353

Malathion-induced granulosa cell apoptosis in caprine antral follicles: an ultrastructural and flow cytometric analysis.

PubMed

Bhardwaj, Jitender K; Saraf, Priyanka

2014-12-01

Organophosphate pesticides (OPs) like malathion interfere with normal ovarian function resulting in an increased incidence of atresia and granulosa cell apoptosis that plays a consequential role in the loss of ovarian follicles or follicular atresia. The aim of present study was to assess malathion-induced (100 nM) reproductive stress, ultrastructural damage and changes in apoptosis frequency in ovarian granulosa cells of antral follicles. Transmission electron microscopy (TEM) was employed for ultrastructural characterization, oxidative stress was evaluated using thiobarbituric acid reactive substances (TBARS) assay to measure lipid peroxidation, and apoptosis was quantified via flow cytometry. By TEM, apoptosis was identified by the presence of an indented nuclear membrane with blebbing, pyknotic crescent-shaped fragmented nuclei, increased vacuolization, degenerating mitochondria, and lipid droplets. The results indicate a significant increase in malondialdehyde (MDA) level (nmols/g wet tissue) at a 100 nM dose of malathion i.e. 7.57±0.033*, 8.53±0.12*, and 12.87±0.78** at 4, 6, or 8 h, respectively, as compared with controls (6.07±0.033, p<0.01*, p<0.05**) showing a positive correlation between malathion-induced lipid peroxidation and percentage of granulosa cell apoptosis (r=1; p<0.01). The parallel use of these three methods enabled us to determine the role of malathion in inducing apoptosis as a consequence of cytogenetic damage and oxidative stress generated in granulosa cells of antral follicles.

Two programmed cell death systems in Escherichia coli: an apoptotic-like death is inhibited by the mazEF-mediated death pathway.

PubMed

Erental, Ariel; Sharon, Idith; Engelberg-Kulka, Hanna

2012-01-01

In eukaryotes, the classical form of programmed cell death (PCD) is apoptosis, which has as its specific characteristics DNA fragmentation and membrane depolarization. In Escherichia coli a different PCD system has been reported. It is mediated by the toxin-antitoxin system module mazEF. The E. coli mazEF module is one of the most thoroughly studied toxin-antitoxin systems. mazF encodes a stable toxin, MazF, and mazE encodes a labile antitoxin, MazE, which prevents the lethal effect of MazF. mazEF-mediated cell death is a population phenomenon requiring the quorum-sensing pentapeptide NNWNN designated Extracellular Death Factor (EDF). mazEF is triggered by several stressful conditions, including severe damage to the DNA. Here, using confocal microscopy and FACS analysis, we show that under conditions of severe DNA damage, the triggered mazEF-mediated cell death pathway leads to the inhibition of a second cell death pathway. The latter is an apoptotic-like death (ALD); ALD is mediated by recA and lexA. The mazEF-mediated pathway reduces recA mRNA levels. Based on these results, we offer a molecular model for the maintenance of an altruistic characteristic in cell populations. In our model, the ALD pathway is inhibited by the altruistic EDF-mazEF-mediated death pathway.

Inhibition of Bim Enhances Replication of Varicella-Zoster Virus and Delays Plaque Formation in Virus-Infected Cells

PubMed Central

Liu, XueQiao

2014-01-01

Programmed cell death (apoptosis) is an important host defense mechanism against intracellular pathogens, such as viruses. Accordingly, viruses have evolved multiple mechanisms to modulate apoptosis to enhance replication. Varicella-zoster virus (VZV) induces apoptosis in human fibroblasts and melanoma cells. We found that VZV triggered the phosphorylation of the proapoptotic proteins Bim and BAD but had little or no effect on other Bcl-2 family members. Since phosphorylation of Bim and BAD reduces their proapoptotic activity, this may prevent or delay apoptosis in VZV-infected cells. Phosphorylation of Bim but not BAD in VZV-infected cells was dependent on activation of the MEK/extracellular signal-regulated kinase (ERK) pathway. Cells knocked down for Bim showed delayed VZV plaque formation, resulting in longer survival of VZV-infected cells and increased replication of virus, compared with wild-type cells infected with virus. Conversely, overexpression of Bim resulted in earlier plaque formation, smaller plaques, reduced virus replication, and increased caspase 3 activity. Inhibition of caspase activity in VZV-infected cells overexpressing Bim restored levels of virus production similar to those seen with virus-infected wild-type cells. Previously we showed that VZV ORF12 activates ERK and inhibits apoptosis in virus-infected cells. Here we found that VZV ORF12 contributes to Bim and BAD phosphorylation. In summary, VZV triggers Bim phosphorylation; reduction of Bim levels results in longer survival of VZV-infected cells and increased VZV replication. PMID:24227856

Toll-like Receptor 3 (TLR3) Induces Apoptosis via Death Receptors and Mitochondria by Up-regulating the Transactivating p63 Isoform α (TAP63α)*

PubMed Central

Sun, Ruili; Zhang, Yu; Lv, Qingshan; Liu, Bei; Jin, Miao; Zhang, Weijia; He, Qing; Deng, Minjie; Liu, Xueting; Li, Guancheng; Li, Yuehui; Zhou, Guohua; Xie, Pingli; Xie, Xiumei; Hu, Jinyue; Duan, Zhaojun

2011-01-01

Toll-like receptor 3 (TLR3), a member of the pathogen recognition receptors, is widely expressed in various cells and has been shown to activate immune signaling pathways by recognizing viral double-stranded RNA. Recently, it was reported that the activation of TLR3 induced apoptosis in some cells, but the detailed molecular mechanism is not fully understood. In this study, we found that in endothelial cells polyinosinic-polycytidylic acid (poly(I-C)) induced dose- and time-dependent cell apoptosis, which was elicited by TLR3 activation, as TLR3 neutralization and down-regulation repressed the apoptosis. Poly(I-C) induced the activation of both caspases 8 and 9, indicating that TLR3 triggered the signaling of both the extrinsic and intrinsic apoptotic pathways. Poly(I-C) up-regulated tumor necrosis factor-related apoptosis-inducing ligand and its receptors, death receptors 4/5, resulting in initiating the extrinsic pathway. Furthermore, poly(I-C) down-regulated anti-apoptotic protein, B cell lymphoma 2 (Bcl-2), and up-regulated Noxa, a key Bcl-2 homology 3-only antagonist of Bcl-2, leading to the priming of the intrinsic pathway. A p53-related protein, the transactivating p63 isoform α (TAp63α), was induced by TLR3 activation and contributed to the activation of both the intrinsic and extrinsic apoptotic pathways. Both the cells deficient in p63 gene expression by RNA interference and cells that overexpressed the N-terminally truncated p63 isoform α (ΔNp63α), a dominant-negative variant of TAp63α, by gene transfection, survived TLR3 activation. Taken together, TAp63α is a crucial regulator downstream of TLR3 to induce cell death via death receptors and mitochondria. PMID:21367858

Fas Apoptosis Inhibitory Molecule (FAIM) Contains a Novel Beta Sandwich in Contact with a Partially Ordered Domain

PubMed Central

Hemond, Michael; Rothstein, Thomas L.; Wagner, Gerhard

2009-01-01

Summary Fas apoptosis inhibitory molecule (FAIM) is a soluble cytosolic protein inhibitor of programmed cell death and is found in organisms throughout the animal kingdom. A short isoform (FAIM-S) is expressed in all tissue types, while an alternatively spliced long isoform (FAIM-L) is specifically expressed in the brain. Here FAIM-S is shown to consist of two independently folding domains in contact with one another. The NMR solution structure of the C-terminal domain of murine FAIM is solved in isolation and revealed to be a novel protein fold, a noninterleaved seven-stranded beta sandwich. The structure and sequence reveal several residues that are likely to be involved in functionally significant interactions with the N-terminal domain or other binding partners. Chemical shift perturbation is used to elucidate contacts made between the N- and C-terminal domains. PMID:19168072

TNF-induced necroptosis requires the plasma membrane localization of the MLKL protein | Center for Cancer Research

Cancer.gov

The cell signaling protein tumor necrosis factor (TNF), produced by white blood cells, promotes inflammation and immunity processes such as fever and is involved in tumorigenesis and apoptosis (programmed cell death). However, dysregulation of TNF can also lead to another form of programmed cell death called necroptosis, which is characterized by a rise in intracellular Ca2+,

Linc00472 suppresses proliferation and promotes apoptosis through elevating PDCD4 expression by sponging miR-196a in colorectal cancer.

PubMed

Ye, Yafei; Yang, Shengnan; Han, Yanping; Sun, Jingjing; Xv, Lijuan; Wu, Lina; Wang, Yongfeng; Ming, Liang

2018-06-21

Long intergenic non-coding RNA Linc00472 has been considered as a tumor suppressor in some cancers. However, the function and mechanism of Linc00472 in colorectal cancer has not been well elucidated. In this study, we found that Linc00472 was down-regulated in colorectal cancer tissues and cells. Elevated Linc00472 expression suppressed proliferation and induced apoptosis in colorectal cancer cells. Moreover, Linc00472 acted as a competing endogenous RNA (ceRNA) of miR-196a to release programmed cell death 4 (PDCD4). Furthermore, miR-196a overexpression or PDCD4 knockdown reversed Linc00472-mediated proliferation inhibition and apoptosis induction in colorectal cancer cells. Ectopic Linc00472 expression hindered tumor growth in vivo . Our study demonstrated that Linc00472 suppressed proliferation and induced apoptosis through up-regulating PDCD4 by decoying miR-196a, which may be an effective therapeutic target for colorectal cancer.

Activation of a PGC-1-related Coactivator (PRC)-dependent Inflammatory Stress Program Linked to Apoptosis and Premature Senescence*

PubMed Central

Gleyzer, Natalie; Scarpulla, Richard C.

2013-01-01

PGC-1-related coactivator (PRC), a growth-regulated member of the PGC-1 coactivator family, contributes to the expression of the mitochondrial respiratory apparatus. PRC also orchestrates a robust response to metabolic stress by promoting the expression of multiple genes specifying inflammation, proliferation, and metabolic reprogramming. Here, we demonstrate that this PRC-dependent stress program is activated during apoptosis and senescence, two major protective mechanisms against cellular dysfunction. Both PRC and its targets (IL1α, SPRR2D, and SPRR2F) were rapidly induced by menadione, an agent that promotes apoptosis through the generation of intracellular oxidants. Menadione-induced apoptosis and the PRC stress program were blocked by the antioxidant N-acetylcysteine. The PRC stress response was also activated by the topoisomerase I inhibitor 7-ethyl-10-hydroxycamptothecin (SN-38), an inducer of premature senescence in tumor cells. Cells treated with SN-38 displayed morphological characteristics of senescence and express senescence-associated β-galactosidase activity. In contrast to menadione, the SN-38 induction of the PRC program occurred over an extended time course and was antioxidant-insensitive. The potential adaptive function of the PRC stress response was investigated by treating cells with meclizine, a drug that promotes glycolytic energy metabolism and has been linked to cardio- and neuroprotection against ischemia-reperfusion injury. Meclizine increased lactate production and was a potent inducer of the PRC stress program, suggesting that PRC may contribute to the protective effects of meclizine. Finally, c-MYC and PRC were coordinately induced under all conditions tested, implicating c-MYC in the biological response to metabolic stress. The results suggest a general role for PRC in the adaptive response to cellular dysfunction. PMID:23364789

Polo-like kinase 1, a new therapeutic target in hepatocellular carcinoma

PubMed Central

Mok, Wei Chuen; Wasser, Shanthi; Tan, Theresa; Lim, Seng Gee

2012-01-01

AIM: To investigate the role of polo-like kinase 1 (PLK1) as a therapeutic target for hepatocellular carcinoma (HCC). METHODS: PLK1 gene expression was evaluated in HCC tissue and HCC cell lines. Gene knockdown with short-interfering RNA (siRNA) was used to study PLK1 gene and protein expression using real-time reverse transcription polymerase chain reaction (RT-PCR) and Western blotting, and cell proliferation using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2(4-sulfophenyl)-2H-tetrazolium (MTS) and bromodeoxyuridine (BrdU) assays. Apoptosis was evaluated using the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and caspase-inhibition assay. Huh-7 cells were transplanted into nude mice and co-cultured with PLK1 siRNA or control siRNA, and tumor progression was compared with controls. RESULTS: RT-PCR showed that PLK1 was overexpressed 12-fold in tumor samples compared with controls, and also was overexpressed in Huh-7 cells. siRNA against PLK1 showed a reduction in PLK1 gene and protein expression of up to 96% in Huh-7 cells, and a reduction in cell proliferation by 68% and 92% in MTS and BrdU cell proliferation assays, respectively. There was a 3-fold increase in apoptosis events, and TUNEL staining and caspase-3 assays suggested that this was caspase-independent. The pan-caspase inhibitor Z-VAD-FMK was unable to rescue the apoptotic cells. Immnofluorescence co-localized endonuclease-G to fragmented chromosomes, implicating it in apoptosis. Huh-7 cells transplanted subcutaneously into nude mice showed tumor regression in siPLK1-treated mice, but not in controls. CONCLUSION: Knockdown of PLK1 overexpression in HCC was shown to be a potential therapeutic target, leading to apoptosis through the endonuclease-G pathway. PMID:22826617

Cytosolic Irradiation of Femtosecond Laser Induces Mitochondria-dependent Apoptosis-like Cell Death via Intrinsic Reactive Oxygen Cascades

PubMed Central

Yoon, Jonghee; Ryu, Seung-wook; Lee, Seunghee; Choi, Chulhee

2015-01-01

High-intensity femtosecond lasers have recently been used to irreversibly disrupt nanoscale structures, such as intracellular organelles, and to modify biological functions in a reversible manner: so-called nanosurgery and biophotomodulation. Femtosecond laser pulses above the threshold intensity sufficient for reversible biophotomodulation can cause irreversible changes in the irradiated cell, eventually leading to cell death. Here, we demonstrated that cytosolic irradiation with a femtosecond laser produced intrinsic cascades of reactive oxygen species (ROS), which led to rapid apoptosis-like cell death via a caspase and poly (ADP-ribose) polymerase 1 (PARP-1) signaling pathway. We further showed that cells with enhanced mitochondrial fusion activity are more resilient to laser-induced stress compared to those with enforced mitochondrial fission. Taken together, these findings provide fundamental insight into how optical stimulation intervenes in intrinsic cellular signaling pathways and functions. PMID:25648455

Cytosolic irradiation of femtosecond laser induces mitochondria-dependent apoptosis-like cell death via intrinsic reactive oxygen cascades.

PubMed

Yoon, Jonghee; Ryu, Seung-Wook; Lee, Seunghee; Choi, Chulhee

2015-02-04

High-intensity femtosecond lasers have recently been used to irreversibly disrupt nanoscale structures, such as intracellular organelles, and to modify biological functions in a reversible manner: so-called nanosurgery and biophotomodulation. Femtosecond laser pulses above the threshold intensity sufficient for reversible biophotomodulation can cause irreversible changes in the irradiated cell, eventually leading to cell death. Here, we demonstrated that cytosolic irradiation with a femtosecond laser produced intrinsic cascades of reactive oxygen species (ROS), which led to rapid apoptosis-like cell death via a caspase and poly (ADP-ribose) polymerase 1 (PARP-1) signaling pathway. We further showed that cells with enhanced mitochondrial fusion activity are more resilient to laser-induced stress compared to those with enforced mitochondrial fission. Taken together, these findings provide fundamental insight into how optical stimulation intervenes in intrinsic cellular signaling pathways and functions.

Design, synthesis, and biological evaluation of polo-like kinase 1/eukaryotic elongation factor 2 kinase (PLK1/EEF2K) dual inhibitors for regulating breast cancer cells apoptosis and autophagy.

PubMed

Pan, Zhaoping; Chen, Yujuan; Liu, Jingyan; Jiang, Qinglin; Yang, Shengyong; Guo, Li; He, Gu

2018-01-20

Both PLK1 and EEF2K are serine⁄threonine kinases that play important roles in the proliferation and programmed cell death of various types of cancer. They are highly expressed in breast cancer tissues. Based on the multiple-complexes generated pharmacophore models of PLK1 and homology models of EEF2K, the integrated virtual screening is performed to discover novel PLK1/EEF2K dual inhibitors. The top ten hit compounds are selected and tested in vitro, and five of them display PLK1 and EEF2K inhibition in vitro. Based on the docking modes of the most potent hit compound, a series of derivatives are synthesized, characterized and biological assayed on the PLK1, EEF2K as well as breast cancer cell proliferation models. Compound 18i with satisfied inhibitory potency are shifted to molecular mechanism studies contained molecular dynamics simulations, cell cycles, apoptosis and autophagy assays. Our results suggested that these novel PLK1/EEF2K dual inhibitors can be used as lead compounds for further development breast cancer chemotherapy. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Detection of Mitochondrial Caspase Activity in Real Time In Situ in Live Cells

NASA Astrophysics Data System (ADS)

Zhang, Yingpei; Haskins, Catherine; Lopez-Cruzan, Marisa; Zhang, Jianhua; Centonze, Victoria E.; Herman, Brian

2004-08-01

Apoptosis plays an important role in many physiological and pathological processes. The initiation and execution of the cell death program requires activation of multiple caspases in a stringently temporal order. Here we describe a method that allows real-time observation of caspase activation in situ in live cells based on fluorescent resonance energy transfer (FRET) measurement using the prism and reflector imaging spectroscopy system (PARISS). When a fusion protein consisting of CFP connected to YFP via an intervening caspase substrate that has been targeted to a specific subcellular location is excited with a light source whose wavelength matches the cyan fluorescent protein (CFP) excitation peak, the energy absorbed by the CFP fluorophore is not emitted as fluorescence. Instead, the excitation energy is absorbed by the nearby yellow fluorescent protein (YFP) fluorophore that is covalently linked to CFP through a short peptide containing the caspase substrate. Cleavage of the linker peptide by caspases results in loss of FRET due to the separation of CFP and YFP fluorophores. Using a mitochondrially targeted CFP caspase 3 substrate YFP construct (mC3Y), we demonstrate for the first time that there is caspase-3-like activity in the mitochondrial matrix of some cells at very late stage of apoptosis.

Inhibition of cell proliferation by nobiletin, a dietary phytochemical, associated with apoptosis and characteristic gene expression, but lack of effect on early rat hepatocarcinogenesis in vivo.

PubMed

Ohnishi, Hiroyuki; Asamoto, Makoto; Tujimura, Kazunari; Hokaiwado, Naomi; Takahashi, Satoru; Ogawa, Kumiko; Kuribayashi, Masanori; Ogiso, Tadashi; Okuyama, Harumi; Shirai, Tomoyuki

2004-12-01

Dietary phytochemicals can inhibit the development of certain types of tumors. We here investigated the effects of nobiletin (Nob), garcinol (Gar), auraptene (Aur), beta-cryptoxanthin- and hesperidine-rich pulp (CHRP) and 1,1'-acetoxychavicol acetate (ACA) on hepatocarcinogenesis in a rat medium-term liver bioassay, and also examined their influence on cell proliferation, cell cycle kinetics, apoptosis and cell invasion of rat and human hepatocellular carcinoma (HCC) cells, MH1C1 and HepG2, respectively. While there were no obvious suppressive effects on the development of putative preneoplastic liver lesions, inhibition of hepatocarcinoma cell proliferation was evident in the Nob group. Nob also caused G2/M cell cycle arrest and apoptosis. Microarray analysis identified a set of genes specifically regulated by Nob, and these are likely to be involved in the observed growth suppression of HCC cells. These results suggest that phytochemicals might have chemopreventive potential in late stages of hepatocarcinogenesis.

Exploiting cannabinoid-induced cytotoxic autophagy to drive melanoma cell death.

PubMed

Armstrong, Jane L; Hill, David S; McKee, Christopher S; Hernandez-Tiedra, Sonia; Lorente, Mar; Lopez-Valero, Israel; Eleni Anagnostou, Maria; Babatunde, Fiyinfoluwa; Corazzari, Marco; Redfern, Christopher P F; Velasco, Guillermo; Lovat, Penny E

2015-06-01

Although the global incidence of cutaneous melanoma is increasing, survival rates for patients with metastatic disease remain <10%. Novel treatment strategies are therefore urgently required, particularly for patients bearing BRAF/NRAS wild-type tumors. Targeting autophagy is a means to promote cancer cell death in chemotherapy-resistant tumors, and the aim of this study was to test the hypothesis that cannabinoids promote autophagy-dependent apoptosis in melanoma. Treatment with Δ(9)-Tetrahydrocannabinol (THC) resulted in the activation of autophagy, loss of cell viability, and activation of apoptosis, whereas cotreatment with chloroquine or knockdown of Atg7, but not Beclin-1 or Ambra1, prevented THC-induced autophagy and cell death in vitro. Administration of Sativex-like (a laboratory preparation comprising equal amounts of THC and cannabidiol (CBD)) to mice bearing BRAF wild-type melanoma xenografts substantially inhibited melanoma viability, proliferation, and tumor growth paralleled by an increase in autophagy and apoptosis compared with standard single-agent temozolomide. Collectively, our findings suggest that THC activates noncanonical autophagy-mediated apoptosis of melanoma cells, suggesting that cytotoxic autophagy induction with Sativex warrants clinical evaluation for metastatic disease.

Nestin is essential for zebrafish brain and eye development through control of progenitor cell apoptosis.

PubMed

Chen, Hua-Ling; Yuh, Chiou-Hwa; Wu, Kenneth K

2010-02-19

Nestin is expressed in neural progenitor cells (NPC) of developing brain. Despite its wide use as an NPC marker, the function of nestin in embryo development is unclear. As nestin is conserved in zebrafish and its predicted sequence is clustered with the mammalian nestin orthologue, we used zebrafish as a model to investigate its role in embryogenesis. Injection of nestin morpholino (MO) into fertilized eggs induced time- and dose-dependent brain and eye developmental defects. Nestin morphants exhibited characteristic morphological changes including small head, small eyes and hydrocephalus. Histological examinations show reduced hind- and mid-brain size, dilated ventricle, poorly organized retina and underdeveloped lens. Injection of control nestin MO did not induce brain or eye changes. Nestin MO injection reduced expression of ascl1b (achaete-scute complex-like 1b), a marker of NPCs, without affecting its distribution. Nestin MO did not influence Elavl3/4 (Embryonic lethal, abnormal vision, Drosophila-like 3/4) (a neuronal marker), or otx2 (a midbrain neuronal marker), but severely perturbed cranial motor nerve development and axon distribution. To determine whether the developmental defects are due to excessive NPC apoptosis and/or reduced NPC proliferation, we analyzed apoptosis by TUNEL assay and acridine orange staining and proliferation by BrdU incorporation, pcna and mcm5 expressions. Excessive apoptosis was noted in hindbrain and midbrain cells. Apoptotic signals were colocalized with ascl1b. Proliferation markers were not significantly altered by nestin MO. These results suggest that nestin is essential for zebrafish brain and eye development probably through control of progenitor cell apoptosis.

Withaferin-A induces apoptosis in osteosarcoma U2OS cell line via generation of ROS and disruption of mitochondrial membrane potential.

PubMed

Li, A-X; Sun, M; Li, X

2017-03-01

Withaferin-A (WF-A) is a well-known dietary compound isolated from Withania somnifera. It has marked pharmacological potential and has been shown to exhibit antiproliferative activity against several types of cancerous cells. Currently, the main focus of anti-cancer therapeutic development is to identify apoptosis-inducing drug-like molecules. Osteosarcoma is a rare type of bone cancer affecting humans. The objective of the present study was therefore to evaluate the antitumor potential of WF-A against several osteosarcoma cell lines. MTT assay was used to evaluate WF-A against osteosarcoma cell lines and to calculate the IC50. DAPI staining was used to confirm the apoptosis-inducing potential of WF-A. Mitochondrial membrane potential, reactive oxygen species (ROS) assay, and Western blotting were used to confirm the basis of apoptosis. The results of the present study revealed that WF-A exhibited strong antiproliferative activity against all the cells lines, with IC50 ranging from 0.32 to 7.6 µM. The lowest IC50 (0.32 µM) was observed against U2OS cell line and, therefore, it was selected for further analysis. DAPI staining indicated that WF-A exhibited antiproliferative activity via induction of apoptosis. Moreover, WF-A induced a ROS-mediated reduction in mitochondrial membrane potential in a dose-dependent manner and activation of caspase-3 in osteosarcoma cells. We suggest that WF-A may prove a potent therapeutic agent for inducing apoptosis in osteosarcoma cell lines via generation of ROS and disruption of mitochondrial membrane potential.

Tumor necrosis factor-α inhibits effects of aryl hydrocarbon receptor ligands on cell death in human lymphocytes.

PubMed

Ghatrehsamani, Mahdi; Soleimani, Masoud; Esfahani, Behjat A Moayedi; Shirzad, Hedayatollah; Hakemi, Mazdak G; Mossahebimohammadi, Majid; Eskandari, Nahid; Adib, Minoo

2015-01-01

Activation of aryl hydrocarbon receptor (AhR) leads to diverse outcome in various kinds of cells. AhR activation may induce apoptosis or prevent of apoptosis and cell death. Recent studies suggest that apoptosis effects of AhR can be modulated by inflammatory cytokine like tumor necrosis factor alpha (TNF-α). In this study, we try to investigate the possible interaction of TNF-α with the 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD), a ligand of AhR, on peripheral lymphocytes. Human peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood by discontinuous density gradient centrifugation on ficoll. Isolated PBMCs were divided into four groups: Control group, TNF-α administered group, TCDD administered group, co-administered group with TCDD and TNF-α. Cells were maintained for a week in lymphocyte culture condition. Then, TNF-α was added to group 2 and 4. Finally, apoptosis and necrosis were analyzed in all samples using flowcytometry. In group 4, the mean percent of necrosis and apoptosis in TCDD treatment groups was significantly larger than other groups; (P < 0.05). Furthermore, there was no significant difference between the mean percent of cell death in TNF-α administered group and TCDD administered group (P > 0.05). However, the mean percent of cell death in co-administered group with TCDD and TNF-α was significantly lower than other groups; (P < 0.05). TNF-α could significantly inhibit effects of TCDD on lymphocytes apoptosis. Combination effects of TNF-α and TCDD on lymphocyte increase cell survival.

Hypoxia preconditioning increases survival and decreases expression of Toll-like receptor 4 in pulmonary artery endothelial cells exposed to lipopolysaccharide

PubMed Central

Nanchal, Rahul; Audi, Said; Konduri, G. Ganesh; Medhora, Meetha

2013-01-01

Abstract Pulmonary or systemic infections and hypoxemic respiratory failure are among the leading causes of admission to intensive care units, and these conditions frequently exist in sequence or in tandem. Inflammatory responses to infections are reproduced by lipopolysaccharide (LPS) engaging Toll-like receptor 4 (TLR4). Apoptosis is a hallmark of lung injury in sepsis. This study was conducted to determine whether preexposure to LPS or hypoxia modulated the survival of pulmonary artery endothelial cells (PAECs). We also investigated the role TLR4 receptor expression plays in apoptosis due to these conditions. Bovine PAECs were cultured in hypoxic or normoxic environments and treated with LPS. TLR4 antagonist TAK-242 was used to probe the role played by TLR4 receptors in cell survival. Cell apoptosis and survival were measured by caspase 3 activity and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) incorporation. TLR4 expression and tumor necrosis factor α (TNF-α) production were also determined. LPS increased caspase 3 activity in a TAK-242-sensitive manner and decreased MTT incorporation. Apoptosis was decreased in PAECs preconditioned with hypoxia prior to LPS exposure. LPS increased TNF-α production, and hypoxic preconditioning blunted it. Hypoxic preconditioning reduced LPS-induced TLR4 messenger RNA and TLR4 protein. TAK-242 decreased to baseline the LPS-stimulated expression of TLR4 messenger RNA regardless of environmental conditions. In contrast, LPS followed by hypoxia substantially increased apoptosis and cell death. In conclusion, protection from LPS-stimulated PAEC apoptosis by hypoxic preconditioning is attributable in part to reduction in TLR4 expression. If these signaling pathways apply to septic patients, they may account for differing sensitivities of individuals to acute lung injury depending on oxygen tensions in PAECs in vivo. PMID:24618542

Apoptosis induced by microtubule disrupting drugs in cultured human lymphoma cells. Inhibitory effects of phorbol ester and zinc sulphate.

PubMed

Takano, Y; Okudaira, M; Harmon, B V

1993-03-01

The effects of the microtubule disrupting drugs (MDD) vinblastine, vincristine and colchicine on a human lymphoma cell line, BM 13674, were investigated. Twelve hours after administration of vinblastine (10(-3) mg/ml), vincristine (10(-2) mg/ml) or colchicine (10(-2) mg/ml), cell death with the characteristic morphology of apoptosis was observed in 71.6%, 82.2% and 76.9% of the cells respectively. The mode of death was confirmed as apoptotic by the occurrence of internucleosomal DNA cleavage, which was demonstrated by agarose gel electrophoresis. For the purpose of casting light on the mechanism involved, inhibition tests were performed on apoptosis induced by one of these drugs, vinblastine, using a phorbol ester (PDBu), zinc sulphate and cycloheximide. PDBu, an activator of protein kinase C, and zinc sulphate, a putative inhibitor of the endonuclease were thought to be responsible for internucleosomal DNA cleavage; both markedly reduced the induction of apoptosis. The protein synthesis inhibitor cycloheximide, on the other hand, had no inhibitory effect. Moreover, cycloheximide treatment per se enhanced apoptosis. This suggests that new protein synthesis is not required for the execution of vinblastine-induced apoptosis. Such a finding is in accord with recent reports suggesting that the "death program" within many cell types may be primed but unable to proceed due to concomitant production of specific "apoptotic inhibitors". It is suggested that phorbol esters prevent vinblastine-induced apoptosis in the BM 13674 cells by activating one or more of these specific "apoptotic inhibitors", possibly by means of PKC-mediated phosphorylation.

Synergistic Effect of Curcumin in Combination with Anticancer Agents in Human Retinoblastoma Cancer Cell Lines.

PubMed

Sreenivasan, Seethalakshmi; Krishnakumar, Subramanian

2015-01-01

Curcumin (diferuloylmethane), a phenolic compound obtained from the rhizome of the herb Curcuma longa, is known to have anti-proliferative and anti-tumor properties. In this study, we evaluated the cytotoxic effect of curcumin alone and in combination with individual drugs like carboplatin, etoposide, or vincristine in a human retinoblastoma (RB) cancer cell line. A drug-drug interaction was analyzed using the median effect/isobologram method and combination index values were used to characterize the interaction as synergistic or additive. We also performed the apoptosis and cell-cycle kinetics study with single drugs in combination with curcumin in a human RB cell lines (Y79 and Weri-Rb1). Curcumin caused concentration-dependent decrease in cell proliferation, cell kinetics, and also induced apoptosis in both the RB cell lines. When combination of curcumin with individual drugs like carboplatin or etoposide or vincristine was treated on to RB cells, both cell viability and cell cycling were reduced and increased apoptosis was noted, in comparison with single drug treatment. These effects were significant in both the cell lines, indicating the ability of curcumin to increase the sensitivity of RB cells to chemotherapy drugs. Our in vitro findings showed that the combination of curcumin with single drug treatment showed marked synergistic inhibitory effect against RB cell lines. These results suggest that curcumin can be used as a modulator which may have a potential therapeutic value for the treatment of RB cancer patients.

ARP2, a novel pro-apoptotic protein expressed in epithelial prostate cancer LNCaP cells and epithelial ovary CHO transformed cells.

PubMed

Mas-Oliva, Jaime; Navarro-Vidal, Enrique; Tapia-Vieyra, Juana Virginia

2014-01-01

Neoplastic epithelial cells generate the most aggressive types of cancers such as those located in the lung, breast, colon, prostate and ovary. During advanced stages of prostate cancer, epithelial cells are associated to the appearance of androgen-independent tumors, an apoptotic-resistant phenotype that ultimately overgrows and promotes metastatic events. We have previously identified and electrophysiologically characterized a novel Ca(2+)-permeable channel activated during apoptosis in the androgen-independent prostate epithelial cancer cell line, LNCaP. In addition, we reported for the first time the cloning and characterization of this channel-like molecule named apoptosis regulated protein 2 (ARP2) associated to a lethal influx of Ca(2+) in Xenopus oocytes. In the present study, LNCaP cells and Chinese hamster ovary cells (CHO cell line) transfected with arp2-cDNA are induced to undergo apoptosis showing an important impact on cell viability and activation of caspases 3 and 7 when compared to serum deprived grown cells and ionomycin treated cells. The subcellular localization of ARP2 in CHO cells undergoing apoptosis was studied using confocal microscopy. While apoptosis progresses, ARP2 initially localized in the peri-nuclear region of cells migrates with time towards the plasma membrane region. Based on the present results and those of our previous studies, the fact that ARP2 constitutes a novel cation channel is supported. Therefore, ARP2 becomes a valuable target to modulate the influx and concentration of calcium in the cytoplasm of epithelial cancer cells showing an apoptotic-resistant phenotype during the onset of an apoptotic event.

Geary autocorrelation and DCCA coefficient: Application to predict apoptosis protein subcellular localization via PSSM

NASA Astrophysics Data System (ADS)

Liang, Yunyun; Liu, Sanyang; Zhang, Shengli

2017-02-01

Apoptosis is a fundamental process controlling normal tissue homeostasis by regulating a balance between cell proliferation and death. Predicting subcellular location of apoptosis proteins is very helpful for understanding its mechanism of programmed cell death. Prediction of apoptosis protein subcellular location is still a challenging and complicated task, and existing methods mainly based on protein primary sequences. In this paper, we propose a new position-specific scoring matrix (PSSM)-based model by using Geary autocorrelation function and detrended cross-correlation coefficient (DCCA coefficient). Then a 270-dimensional (270D) feature vector is constructed on three widely used datasets: ZD98, ZW225 and CL317, and support vector machine is adopted as classifier. The overall prediction accuracies are significantly improved by rigorous jackknife test. The results show that our model offers a reliable and effective PSSM-based tool for prediction of apoptosis protein subcellular localization.

MicroRNA-214 Promotes Apoptosis in Canine Hemangiosarcoma by Targeting the COP1-p53 Axis.

PubMed

Heishima, Kazuki; Mori, Takashi; Sakai, Hiroki; Sugito, Nobuhiko; Murakami, Mami; Yamada, Nami; Akao, Yukihiro; Maruo, Kohji

2015-01-01

MicroRNA-214 regulates both angiogenic function in endothelial cells and apoptosis in various cancers. However, the regulation and function of miR-214 is unclear in canine hemangiosarcoma, which is a spontaneous model of human angiosarcoma. The expression and functional roles of miR-214 in canine hemangiosarcoma were presently explored by performing miRNA TaqMan qRT-PCR and transfecting cells with synthetic microRNA. Here, we report that miR-214 was significantly down-regulated in the cell lines used and in clinical samples of canine hemangiosarcoma. Restoration of miR-214 expression reduced cell growth and induced apoptosis in canine hemangiosarcoma cell lines through transcriptional activation of p53-regulated genes although miR-214 had a slight effect of growth inhibition on normal endothelial cells. We identified COP1, which is a critical negative regulator of p53, as a novel direct target of miR-214. COP1 was overexpressed and the specific COP1 knockdown induced apoptosis through transcriptional activation of p53-regulated genes as well as did miR-214-transfection in HSA cell lines. Furthermore, p53 knockdown abolished the miR-214-COP1-mediated apoptosis; thus, miR-214 and COP1 regulated apoptosis through controlling p53 in HSA. In conclusion, miR-214 functioned as a tumor suppressor in canine hemangiosarcoma by inducing apoptosis through recovering the function of p53. miR-214 down-regulation and COP1 overexpression is likely to contribute to tumorigenesis of HSA. Therefore, targeting miR-214-COP1-p53 axis would possibly be a novel effective strategy for treatment of canine hemangiosarcoma and capable of being applied to the development of novel therapeutics for human angiosarcoma.

Inhibitory effects of 3-bromopyruvate in human nasopharyngeal carcinoma cells.

PubMed

Zou, Xue; Zhang, Mengxiao; Sun, Yiming; Zhao, Surong; Wei, Yingmei; Zhang, Xudong; Jiang, Chenchen; Liu, Hao

2015-10-01

Tumor cells depend on aerobic glycolysis for adenosine triphosphate (ATP) production, which is therefore targeted by therapeutic agents. The compound 3-bromopyruvate (3-BrPA), a strong alkylating agent and hexokinase inhibitor, inhibits tumor cell glycolysis and the production of ATP, causing apoptosis. 3-BrPA induces apoptosis of nasopharyngeal carcinoma (NPC) cell lines HNE1 and CNE-2Z, which may be related to its molecular mechanisms. In the present study, we investigated the effects of 3-BrPA on the viability, reactive oxygen species (ROS), apoptosis and other types of programmed cell death in NPC cells in vitro and in vivo. PI staining showed significant apoptosis in NPC cells accompanied by the overproduction of ROS and downregulation of mitochondrial membrane potential (MMP, ΔΨm) by 3-BrPA. However, the ROS scavenger N-acetyl-L-cysteine (NAC) significantly reduced 3-BrPA-induced apoptosis by decreasing ROS and facilitating the recovery of MMP. We elucidated the molecular mechanisms underlying 3-BrPA activity and found that it caused mitochondrial dysfunction and ROS production, leading to necroptosis of NPC cells. We investigated the effects of the caspase inhibitor z-VAD-fmk, which inhibits apoptosis but promotes death domain receptor (DR)-induced NPC cell necrosis. Necrostatin-1 (Nec-1) inhibits necroptosis, apparently via a DR signaling pathway and thus abrogates the effects of z-VAD‑fmk. In addition, we demonstrated the effective attenuation of 3-BrPA-induced necrotic cell death by Nec-1. Finally, animal studies proved that 3-BrPA exhibited significant antitumor activity in nude mice. The present study is the first demonstration of 3-BrPA-induced non-apoptotic necroptosis and ROS generation in NPC cells and provides potential strategies for developing agents against apoptosis‑resistant cancers.

Autophagy and Apoptosis Act as Partners to Induce Germ Cell Death after Heat Stress in Mice

PubMed Central

Zhang, Mianqiu; Jiang, Min; Bi, Ye; Zhu, Hui; Zhou, Zuomin; Sha, Jiahao

2012-01-01

Testicular heating suppresses spermatogenesis which is marked by germ cell loss via apoptotic pathways. Recently, it is reported that autophagy also can be induced by heat treatment in somatic cells. In this study, the status of autophagy in germ cells after heat treatment, as well as the partnership between autophagy and apoptosis in these cells was investigated. The results demonstrated that besides initiating apoptotic pathways, heat also induced autophagic pathways in germ cells. Exposure of germ cells to hyperthermia resulted in several specific features of the autophagic process, including autophagosome formation and the conversion of LC3-I to LC3-II. Furthermore, the ubiquitin-like protein conjugation system was implicated as being likely responsible for heat-induced autophagy in germ cells since all genes involving this system were found to be expressed in the testes. In addition, the upstream protein in this system, Atg7 (Autophagy-related gene 7), was found to be expressed in all types of spermatogenic cells, and its expression level was positively correlated with the level of autophagy in germ cells. As a result, Atg7 was selected as the investigative target to further analyze the role of autophagy in heat-induced germ cell death. It was shown that down expression of Atg7 protein resulted in the notable decrease in the level of autophagy in heat-treated germ cells, and this down-regulation of autophagy caused by Atg7 knockdown further reduced the apoptotic rate of germ cells. These results suggest that autophagy plays a positive role in the process of germ cell apoptosis after heat treatment. In conclusion, this study demonstrates that heat triggers autophagy and apoptosis in germ cells. These two mechanisms might act as partners, not antagonist, to induce cell death and lead to eventual destruction of spermatogenesis. PMID:22848486

Endothelial necrosis at 1h post-burn predicts progression of tissue injury

PubMed Central

Hirth, Douglas; McClain, Steve A.; Singer, Adam J.; Clark, Richard A.F.

2013-01-01

Burn injury progression has not been well characterized at the cellular level. To define burn injury progression in terms of cell death, histopathologic spatiotemporal relationships of cellular necrosis and apoptosis were investigated in a validated porcine model of vertical burn injury progression. Cell necrosis was identified by High Mobility Group Box 1 protein and apoptosis by Caspase 3a staining of tissue samples taken 1h, 24h and 7 days post-burn. Level of endothelial cell necrosis at 1h was predictive of level of apoptosis at 24h (Pearson's r=0.87) and of level of tissue necrosis at 7 days (Pearson's r=0.87). Furthermore, endothelial cell necrosis was deeper than interstitial cell necrosis at 1h (p<0.001). Endothelial cell necrosis at 1h divided the zone of injury progression (Jackson's zone of stasis) into an upper subzone with necrotic endothelial cells and initially viable adnexal and interstitial cells at 1h that progressed to necrosis by 24h, and a lower zone with initially viable endothelial cells at 1h, but necrosis and apoptosis of all cell types by 24h. Importantly, this spatiotemporal series of events and rapid progression resembles myocardial infarction and stroke, and implicates mechanisms of these injuries, ischemia, ischemia reperfusion, and programmed cell death, in burn progression. PMID:23627744

Chlamydia Inhibit Host Cell Apoptosis by Degradation of Proapoptotic BH3-only Proteins

PubMed Central

Fischer, Silke F.; Vier, Juliane; Kirschnek, Susanne; Klos, Andreas; Hess, Simone; Ying, Songmin; Häcker, Georg

2004-01-01

Chlamydia are obligate intracellular bacteria that replicate in a vacuole inside a host cell. Chlamydial infection has been shown to protect the host cell against apoptotic stimuli. This is likely important for the ability of Chlamydia to reproduce in human cells. Here we show that resistance to apoptosis is conveyed by the destruction of the proapoptotic BH3-only proteins Bim/Bod, Puma, and Bad during infection. Apoptotic stimuli were blocked upstream of the mitochondrial activation of Bax/Bak. During infection with both species, Chlamydia trachomatis and Chlamydia pneumoniae, Bim protein gradually disappeared without noticeable changes in Bim mRNA. The disappearance was blocked by inhibitors of the proteasome. Infected cells retained sensitivity to Bim expressed by transfection, indicating functional relevance of the Bim disappearance. Fusion to Bim targeted the green fluorescent protein for destruction during infection. Analysis of truncation mutants showed that a short region of Bim containing the BH3 domain was sufficient for destruction during chlamydial infection. Like Bim, Puma and Bad proteins disappeared during infection. These results reveal a novel way by which microbes can interfere with the host cell's apoptotic machinery, and provide a molecular explanation of the cellular resistance to apoptosis during infection with Chlamydia. PMID:15452181

ATF4 induction through an atypical integrated stress response to ONC201 triggers p53-independent apoptosis in hematological malignancies.

PubMed

Ishizawa, Jo; Kojima, Kensuke; Chachad, Dhruv; Ruvolo, Peter; Ruvolo, Vivian; Jacamo, Rodrigo O; Borthakur, Gautam; Mu, Hong; Zeng, Zhihong; Tabe, Yoko; Allen, Joshua E; Wang, Zhiqiang; Ma, Wencai; Lee, Hans C; Orlowski, Robert; Sarbassov, Dos D; Lorenzi, Philip L; Huang, Xuelin; Neelapu, Sattva S; McDonnell, Timothy; Miranda, Roberto N; Wang, Michael; Kantarjian, Hagop; Konopleva, Marina; Davis, R Eric; Andreeff, Michael

2016-02-16

The clinical challenge posed by p53 abnormalities in hematological malignancies requires therapeutic strategies other than standard genotoxic chemotherapies. ONC201 is a first-in-class small molecule that activates p53-independent apoptosis, has a benign safety profile, and is in early clinical trials. We found that ONC201 caused p53-independent apoptosis and cell cycle arrest in cell lines and in mantle cell lymphoma (MCL) and acute myeloid leukemia (AML) samples from patients; these included samples from patients with genetic abnormalities associated with poor prognosis or cells that had developed resistance to the nongenotoxic agents ibrutinib and bortezomib. Moreover, ONC201 caused apoptosis in stem and progenitor AML cells and abrogated the engraftment of leukemic stem cells in mice while sparing normal bone marrow cells. ONC201 caused changes in gene expression similar to those caused by the unfolded protein response (UPR) and integrated stress responses (ISRs), which increase the translation of the transcription factor ATF4 through an increase in the phosphorylation of the translation initiation factor eIF2α. However, unlike the UPR and ISR, the increase in ATF4 abundance in ONC201-treated hematopoietic cells promoted apoptosis and did not depend on increased phosphorylation of eIF2α. ONC201 also inhibited mammalian target of rapamycin complex 1 (mTORC1) signaling, likely through ATF4-mediated induction of the mTORC1 inhibitor DDIT4. Overexpression of BCL-2 protected against ONC201-induced apoptosis, and the combination of ONC201 and the BCL-2 antagonist ABT-199 synergistically increased apoptosis. Thus, our results suggest that by inducing an atypical ISR and p53-independent apoptosis, ONC201 has clinical potential in hematological malignancies. Copyright © 2016, American Association for the Advancement of Science.

The Immunomodulatory Small Molecule Imiquimod Induces Apoptosis in Devil Facial Tumour Cell Lines.

PubMed

Patchett, Amanda L; Darby, Jocelyn M; Tovar, Cesar; Lyons, A Bruce; Woods, Gregory M

2016-01-01

The survival of the Tasmanian devil (Sarcophilus harrisii) is threatened by devil facial tumour disease (DFTD). This transmissible cancer is usually fatal, and no successful treatments have been developed. In human studies, the small immunomodulatory molecule imiquimod is a successful immunotherapy, activating anti-tumour immunity via stimulation of toll-like receptor-7 (TLR7) signaling pathways. In addition, imiquimod is a potent inducer of apoptosis in human tumour cell lines via TLR7 independent mechanisms. Here we investigate the potential of imiquimod as a DFTD therapy through analysis of treated DFTD cell lines and Tasmanian devil fibroblasts. WST-8 proliferation assays and annexin V apoptosis assays were performed to monitor apoptosis, and changes to the expression of pro- and anti-apoptotic genes were analysed using qRT-PCR. Our results show that DFTD cell lines, but not Tasmanian devil fibroblasts, are sensitive to imiquimod-induced apoptosis in a time and concentration dependent manner. Induction of apoptosis was accompanied by down-regulation of the anti-apoptotic BCL2 and BCLXL genes, and up-regulation of the pro-apoptotic BIM gene. Continuous imiquimod treatment was required for these effects to occur. These results demonstrate that imiquimod can deregulate DFTD cell growth and survival in direct and targeted manner. In vivo, this may increase DFTD vulnerability to imiquimod-induced TLR7-mediated immune responses. Our findings have improved the current knowledge of imiquimod action in tumour cells for application to both DFTD and human cancer therapy.

Virus-induced apoptosis and phosphorylation form of metacaspase in the marine coccolithophorid Emiliania huxleyi.

PubMed

Liu, Jingwen; Cai, Weicong; Fang, Xian; Wang, Xueting; Li, Guiling

2018-04-01

Lytic viral infection and programmed cell death (PCD) are thought to represent two distinct death mechanisms in phytoplankton, unicellular photoautotrophs that drift with ocean currents. PCD (apoptosis) is mainly brought about by the activation of caspases, a protease family with unique substrate selectivity. Here, we demonstrated that virus infection induced apoptosis of marine coccolithophorid Emiliania huxleyi BOF92 involving activation of metacaspase. E. huxleyi cells exhibited cell death process akin to that of apoptosis when exposed to virus infection. We observed typical hallmarks of apoptosis including cell shrinkage, associated nuclear morphological changes and DNA fragmentation. Immunoblotting revealed that antibody against human active-caspase-3 shared epitopes with a protein of ≈ 23 kDa; whose pattern of expression correlated with the onset of cell death. Moreover, analysis on two-dimensional gel electrophoresis revealed that two spots of active caspase-3 co-migrated with the different isoelectric points. Phosphatase treatment of cytosolic extracts containing active caspases-3 showed a mobility shift, suggesting that phosphorylated form of this enzyme might be present in the extracts. Computational prediction of phosphorylation sites based on the amino acid sequence of E. huxleyi metacaspase showed multiple phosphorylated sites for serine, threonine and tyrosine residues. This is the first report showing that phosphorylation modification of metacaspase in E. huxleyi might be required for certain biochemical and morphological changes during virus induced apoptosis.

Palmitate induces VSMC apoptosis via toll like receptor (TLR)4/ROS/p53 pathway.

PubMed

Zhang, Yuanjun; Xia, Guanghao; Zhang, Yaqiong; Liu, Juxiang; Liu, Xiaowei; Li, Weihua; Lv, Yaya; Wei, Suhong; Liu, Jing; Quan, Jinxing

2017-08-01

Toll-like receptor 4 (TLR4) has been implicated in vascular inflammation, as well as in the pathogenesis of atherosclerosis and diabetes. Vascular smooth muscle cell (VSMC) apoptosis has been shown to induce plaque vulnerability in atherosclerosis. Previous studies reported that palmitate induced apoptosis in VSMCs; however, the role of TLR4 in palmitate-induced apoptosis in VSMCs has not yet been defined. In this study, we investigated whether or not palmitate-induced apoptosis depended on the activation of the TLR4 pathway. VSMCs were treated with or without palmitate, CRISPR/Cas9z-mediated genome editing methods were used to deplete TLR4 expression, while NADPH oxidase inhibitors were used to inhibit reactive oxygen species (ROS) generation. Cell apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, ROS was measured using the 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) method, the mRNA and protein expression levels of caspase 3, caspase 9, BCL-2 and p53 were studied by real-time polymerase chain reaction (RT-PCR) and ELISA. Palmitate significantly promotes VSMC apoptosis, ROS generation, and expression of caspase 3, caspase 9 and p53; while NADPH oxidase inhibitor pretreatment markedly attenuated these effects. Moreover, knockdown of TLR4 significantly blocked palmitate-induced ROS generation and VSMC apoptosis accompanied by inhibition of caspase 3, caspase 9, p53 expression and restoration of BCL-2 expression. Our results suggest that palmitate-induced apoptosis depends on the activation of the TLR4/ROS/p53 signaling pathway, and that TLR4 may be a potential therapeutic target for the prevention and treatment of atherosclerosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Prototheca zopfii Induced Ultrastructural Features Associated with Apoptosis in Bovine Mammary Epithelial Cells

PubMed Central

Shahid, Muhammad; Wang, Jianfang; Gu, Xiaolong; Chen, Wei; Ali, Tariq; Gao, Jian; Han, Dandan; Yang, Rui; Fanning, Séamus; Han, Bo

2017-01-01

Prototheca zopfii infections are becoming global concerns in humans and animals. Bovine protothecal mastitis is characterized by deteriorating milk quality and quantity, thus imparting huge economic losses to dairy industry. Previous published studies mostly focused on the prevalence and characterization of P. zopfii from mastitis. However, the ultrastructural pathomorphological changes associated with apoptosis in bovine mammary epithelial cells (bMECs) are not studied yet. Therefore, in this study we aimed to evaluate the in vitro comparative apoptotic potential of P. zopfii genotype-I and -II on bMECs using flow cytometry, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The results showed fast growth rate and higher adhesion capability of genotype-II in bMECs as compared with genotype-I. The viability of bMECs infected with P. zopfii genotype-II was significantly decreased after 12 h (p < 0.05) and 24 h (p < 0.01) in comparison with control cells. Contrary, genotype-I couldn't show any significant effects on cell viability. Moreover, after infection of bMECs with genotype-II, the apoptosis increased significantly at 12 h (p < 0.05) and 24 h (p < 0.01) as compared with control group. Genotype-I couldn't display any significant effects on cell apoptosis. The host specificity of P. zopfii was also tested in mouse osteoblast cells, and the results suggest that genotype-I and -II could not cause any significant apoptosis in these cell lines. SEM interpreted the pathomorphological alterations in bMECs after infection. Adhesion of P. zopfii with cells and further disruption of cytomembrane validated the apoptosis caused by genotype-II under SEM. While genotype-1 couldn't cause any significant apoptosis in bMECs. Furthermore, genotype-II induced apoptotic manifested specific ultrastructure features, like cytoplasmic cavitation, swollen mitochondria, pyknosis, cytomembrane disruption, and appearance of apoptotic bodies under TEM. The findings of the current study revealed that genotype-II has the capability to invade and survive within the bMECs, thus imparting significant damages to the mammary cells which result in apoptosis. This study represents the first insights into the pathomorphological and ultrastructure features of apoptosis in bMECs induced by P. zopfii genotype-II. PMID:28752077

Simultaneous induction of apoptosis and necroptosis by Tanshinone IIA in human hepatocellular carcinoma HepG2 cells.

PubMed

Lin, C-Y; Chang, T-W; Hsieh, W-H; Hung, M-C; Lin, I-H; Lai, S-C; Tzeng, Y-J

Tanshinone IIA (Tan IIA), a constituent of the traditional medicinal plant Salvia miltiorrhiza BUNGE, has been reported to possess anticancer activity through induction of apoptosis in many cancer cells. Surprisingly, the present study finds that Tan IIA simultaneously causes apoptosis and necroptosis in human hepatocellular carcinoma HepG2 cells. We further find that apoptosis can be converted to necroptosis by pan-caspase inhibitor Z-VAD-fmk, and the two death modes can be blocked by necroptotic inhibitor necrostatin-1. The underlying mechanisms are revealed by analysis of the signaling molecules using western blotting. In control cells, FLICE inhibitory protein in short form (FLIP S ) is expressed in relatively high levels and binds to caspase 8 in ripoptosome, which supposedly sustains cell survival. However, in Tan IIA-treated cells, FLIP S is down-regulated and may thus cause homodimer formation of cleaved caspase 8, cleavage of receptor-interacting serine/threonine-protein kinases 1, 3 (RIP1, RIP3), and mixed-lineage kinase domain-like (MLKL), in turn leads to cell apoptosis. In parallel, Tan IIA causes necroptosis by forming a suggested necrosomal complex composed of RIP1/RIP3. Regarding the inhibitors, z-VAD-fmk diminishes the cleaved caspase 8, RIP1, RIP3, and MLKL induced by Tan IIA, and reconstructs the ripoptosome complex, which marks cells moving from apoptosis to necroptosis. Nec-1 recovers the Tan IIA down-regulated FLIP S , consequently causes FLIP S to form heterodimer with caspase 8 and thus block apoptosis. Meanwhile, cleaved forms of RIP1 and RIP3 were observed preventing necroptosis. Intriguingly, the cytotoxicity of tumor necrosis factor-related apoptosis-inducing ligand to HepG2 cells is enhanced by Tan IIA in a pilot study, which may be attributed to low FLIP S levels induced by Tan IIA. In short, Tan IIA simultaneously induces both Nec-1 inhibition and FLIP S regulation-mediated apoptosis/necroptosis, which has not been previously documented. Moreover, the involvement of the cleavage type of MLKL in executing necroptosis warrants further investigation.

Density-dependent induction of apoptosis by transforming growth factor-beta 1 in a human ovarian carcinoma cell line.

PubMed

Mathieu, C; Jozan, S; Mazars, P; Côme, M G; Moisand, A; Valette, A

1995-01-01

Transforming growth factor-beta 1 inhibited proliferation of a human ovarian carcinoma cell line (NIH-OVCAR-3). The inhibition of NIH-OVCAR-3 cell proliferation was accompanied by a decrease in clonogenic potential, evidenced by the reduced ability of TGF-beta 1-treated NIH-OVCAR-3 cells to form colonies on a plastic substratum. This rapid decrease of clonogenic potential, which was detected 6 h after addition of TGF-beta 1 was dose-dependent (IC50 = 4 pM). Fluorescence microscopy of DAPI-stained cells supported by electron-microscopic examination showed that TGF-beta 1 induced chromatin condensation and nuclear fragmentation. In addition, oligonucleosomal-sized fragments were detected in the TGF-beta 1-treated cells. These features indicated that TGF-beta 1 induced NIH-OVCAR-3 cell death by an apoptosis-like mechanism. This TGF-beta 1 apoptotic effect was subject to modulation by cell density. It was observed that an increase in cell density (up to 20 x 10(3) cells/cm2) protected NIH-OVCAR-3 cells against apoptosis induced by TGF-beta 1. Conditioned medium from high-density cultures of NIH-OVCAR-3 cells did not inhibit apoptosis induced by TGF-beta 1 on NIH-OVCAR-3 cells cultured at low density, suggesting that the protective effect of cell density was not related to the cell secretion of a soluble survival factor.

Hsp70- and p53-reponses after heat treatment and/or X-irradiation mediate the susceptibility of hematopoietic cells to undergo apoptosis.

PubMed

Nijhuis, E H A; Poot, A A; Feijen, J; Vermes, I

2008-02-01

The effect of heat treatment in combination with X-irradiation was examined with regard to expression of p53, a tumor suppressor gene product, and Hsp70, a heat-shock protein, in association with the occurrence of programmed cell death (apoptosis). Three hematopoietic cell lines (HSB2, HL60 and Kasumi-1), which differ in p53 status, were exposed to 42.5 degrees C during one hour and/or X-radiation (total dose 8 Gy). After exposure, both mRNA and protein expression levels of Hsp70 and p53 were investigated by real-time PCR (polymerase chain reaction) and Western blotting. Apoptosis was simultaneously analyzed by observation of cell morphology as well as flowcytometric determination of Annexin V binding to phosphatidylserine and propidium iodide exclusion. Both HL60 and HSB2 cell lines with a low p53 status and a quick response to heat treatment with Hsp70 over-expression are less susceptible to heat-induced apoptosis compared to Kasumi-1 cells with wild-type p53 protein and no Hsp70 response. The combination of first applying X-irradiation followed by heat treatment resulted in the most effective induction of apoptosis due to impairment of the Hsp70 response in all three cell lines. These results indicate that the Hsp70 response and p53 status mediate the susceptibility of hematopoietic cells to undergo heat-induced apoptosis. Therefore, these parameters can be used as markers to predict the effectiveness of hyperthermia in cancer treatment.

Ap4A induces apoptosis in human cultured cells.

PubMed

Vartanian, A; Alexandrov, I; Prudowski, I; McLennan, A; Kisselev, L

1999-07-30

Diadenosine oligophosphates (Ap(n)A) have been proposed as intracellular and extracellular signaling molecules in animal cells. The ratio of diadenosine 5',5'''-P1,P3-triphosphate to diadenosine 5',5'''-P1,P4-tetraphosphate (Ap3A/Ap4A) is sensitive to the cellular status and alters when cultured cells undergo differentiation or are treated with interferons. In cells undergoing apoptosis induced by DNA topoisomerase II inhibitor VP16, the concentration of Ap3A decreases significantly while that of Ap4A increases. Here, we have examined the effects of exogenously added Ap3A and Ap4A on apoptosis and morphological differentiation. Penetration of Ap(n)A into cells was achieved by cold shock. Ap4A at 10 microM induced programmed cell death in human HL60, U937 and Jurkat cells and mouse VMRO cells and this effect appeared to require Ap4A breakdown as hydrolysis-resistant analogues of Ap4A were inactive. On its own, Ap3A induced neither apoptosis nor cell differentiation but did display strong synergism with the protein kinase C activators 12-deoxyphorbol-13-O-phenylacetate and 12-deoxyphorbol-13-O-phenylacetate-20-acetate in inducing differentiation of HL60 cells. We propose that Ap4A and Ap3A are physiological antagonists in determination of the cellular status: Ap4A induces apoptosis whereas Ap3A is a co-inductor of differentiation. In both cases, the mechanism of signal transduction remains unknown.

The targeted inhibition of mitochondrial Hsp90 overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yan, Chunlan; Department of Physiology, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310058; Oh, Joon Seok

Previous studies have reported that a Gamitrinib variant containing triphenylphosphonium (G-TPP) binds to mitochondrial Hsp90 and rapidly inhibits its activity, thus inducing the apoptotic pathway in the cells. Accordingly, G-TPP shows a potential as a promising drug for the treatment of cancer. A cell can die from different types of cell death such as apoptosis, necrosis, necroptosis, and autophagic cell death. In this study, we further investigated the mechanisms and modes of cell death in the G-TPP-treated Hep3B and U937 cell lines. We discovered that G-TPP kills the U937 cells through the apoptotic pathway and the overexpression of Bcl-2 significantlymore » inhibits U937 cell death to G-TPP. We further discovered that G-TPP kills the Hep3B cells by activating necroptosis in combination with the partial activation of caspase-dependent apoptosis. Importantly, G-TPP overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis. We also observed that G-TPP induces compensatory autophagy in the Hep3B cell line. We further found that whereas there is a Bcl-2-Beclin 1 interaction in response to G-TPP, silencing the beclin 1 gene failed to block LC3-II accumulation in the Hep3B cells, indicating that G-TPP triggers Beclin 1-independent protective autophagy in Hep3B cells. Taken together, these data reveal that G-TPP induces cell death through a combination of death pathways, including necroptosis and apoptosis, and overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis. These findings are important for the therapeutic exploitation of necroptosis as an alternative cell death program to bypass the resistance to apoptosis. Highlights: ► G-TPP binds to mitochondrial Hsp90. ► G-TPP induces apoptosis in U937 human leukemia cancer cells. ► G-TPP induces combination of death pathways in Hep3B cell. ► G-TPP overcomes the resistance conferred by Bcl-2 in Hep3B cells via necroptosis. ► G-TPP triggers Beclin 1-independent protective autophagy in Hep3B cells.« less

HSF-1 activates the ubiquitin proteasome system to promote non-apoptotic developmental cell death in C. elegans.

PubMed

Kinet, Maxime J; Malin, Jennifer A; Abraham, Mary C; Blum, Elyse S; Silverman, Melanie R; Lu, Yun; Shaham, Shai

2016-03-08

Apoptosis is a prominent metazoan cell death form. Yet, mutations in apoptosis regulators cause only minor defects in vertebrate development, suggesting that another developmental cell death mechanism exists. While some non-apoptotic programs have been molecularly characterized, none appear to control developmental cell culling. Linker-cell-type death (LCD) is a morphologically conserved non-apoptotic cell death process operating in Caenorhabditis elegans and vertebrate development, and is therefore a compelling candidate process complementing apoptosis. However, the details of LCD execution are not known. Here we delineate a molecular-genetic pathway governing LCD in C. elegans. Redundant activities of antagonistic Wnt signals, a temporal control pathway, and mitogen-activated protein kinase kinase signaling control heat shock factor 1 (HSF-1), a conserved stress-activated transcription factor. Rather than protecting cells, HSF-1 promotes their demise by activating components of the ubiquitin proteasome system, including the E2 ligase LET-70/UBE2D2 functioning with E3 components CUL-3, RBX-1, BTBD-2, and SIAH-1. Our studies uncover design similarities between LCD and developmental apoptosis, and provide testable predictions for analyzing LCD in vertebrates.

Deficiency of base excision repair enzyme NEIL3 drives increased predisposition to autoimmunity

PubMed Central

Massaad, Michel J.; Zhou, Jia; Tsuchimoto, Daisuke; Chou, Janet; Jabara, Haifa; Janssen, Erin; Glauzy, Salomé; Olson, Brennan G.; Morbach, Henner; Ohsumi, Toshiro K.; Schmitz, Klaus; Kane, Jennifer; Torisu, Kumiko; Chouery, Eliane; Megarbane, Andre; Kang, Peter B.; Al-Idrissi, Eman; Aldhekri, Hasan; Meffre, Eric; Mizui, Masayuki; Manis, John P.; Al-Herz, Waleed; Wallace, Susan S.; Geha, Raif S.

2016-01-01

Alterations in the apoptosis of immune cells have been associated with autoimmunity. Here, we have identified a homozygous missense mutation in the gene encoding the base excision repair enzyme Nei endonuclease VIII-like 3 (NEIL3) that abolished enzymatic activity in 3 siblings from a consanguineous family. The NEIL3 mutation was associated with fatal recurrent infections, severe autoimmunity, hypogammaglobulinemia, and impaired B cell function in these individuals. The same homozygous NEIL3 mutation was also identified in an asymptomatic individual who exhibited elevated levels of serum autoantibodies and defective peripheral B cell tolerance, but normal B cell function. Further analysis of the patients revealed an absence of LPS-responsive beige-like anchor (LRBA) protein expression, a known cause of immunodeficiency. We next examined the contribution of NEIL3 to the maintenance of self-tolerance in Neil3–/– mice. Although Neil3–/– mice displayed normal B cell function, they exhibited elevated serum levels of autoantibodies and developed nephritis following treatment with poly(I:C) to mimic microbial stimulation. In Neil3–/– mice, splenic T and B cells as well as germinal center B cells from Peyer’s patches showed marked increases in apoptosis and cell death, indicating the potential release of self-antigens that favor autoimmunity. These findings demonstrate that deficiency in NEIL3 is associated with increased lymphocyte apoptosis, autoantibodies, and predisposition to autoimmunity. PMID:27760045

Untangling the Roles of Anti-Apoptosis in Regulating Programmed Cell Death using Humanized Yeast Cells

PubMed Central

Clapp, Caitlin; Portt, Liam; Khoury, Chamel; Sheibani, Sara; Eid, Rawan; Greenwood, Matthew; Vali, Hojatollah; Mandato, Craig A.; Greenwood, Michael T.

2012-01-01

Genetically programmed cell death (PCD) mechanisms, including apoptosis, are important for the survival of metazoans since it allows, among things, the removal of damaged cells that interfere with normal function. Cell death due to PCD is observed in normal processes such as aging and in a number of pathophysiologies including hypoxia (common causes of heart attacks and strokes) and subsequent tissue reperfusion. Conversely, the loss of normal apoptotic responses is associated with the development of tumors. So far, limited success in preventing unwanted PCD has been reported with current therapeutic approaches despite the fact that inhibitors of key apoptotic inducers such as caspases have been developed. Alternative approaches have focused on mimicking anti-apoptotic processes observed in cells displaying increased resistance to apoptotic stimuli. Hormesis and pre-conditioning are commonly observed cellular strategies where sub-lethal levels of pro-apoptotic stimuli lead to increased resistance to higher or lethal levels of stress. Increased expression of anti-apoptotic sequences is a common mechanism mediating these protective effects. The relevance of the latter observation is exemplified by the observation that transgenic mice overexpressing anti-apoptotic genes show significant reductions in tissue damage following ischemia. Thus strategies aimed at increasing the levels of anti-apoptotic proteins, using gene therapy or cell penetrating recombinant proteins are being evaluated as novel therapeutics to decrease cell death following acute periods of cell death inducing stress. In spite of its functional and therapeutic importance, more is known regarding the processes involved in apoptosis than anti-apoptosis. The genetically tractable yeast Saccharomyces cerevisiae has emerged as an exceptional model to study multiple aspects of PCD including the mitochondrial mediated apoptosis observed in metazoans. To increase our knowledge of the process of anti-apoptosis, we screened a human heart cDNA expression library in yeast cells undergoing PCD due to the conditional expression of a mammalian pro-apoptotic Bax cDNA. Analysis of the multiple Bax suppressors identified revealed several previously known as well as a large number of clones representing potential novel anti-apoptotic sequences. The focus of this review is to report on recent achievements in the use of humanized yeast in genetic screens to identify novel stress-induced PCD suppressors, supporting the use of yeast as a unicellular model organism to elucidate anti-apoptotic and cell survival mechanisms. PMID:22708116

Knockdown of CAVEOLIN-1 Sensitizes Human Basal-Like Triple-Negative Breast Cancer Cells to Radiation.

PubMed

Zou, Man; Li, Yanhui; Xia, Shu; Chu, Qian; Xiao, Xiaoguang; Qiu, Hong; Chen, Yu; Zheng, Zu'an; Liu, Fei; Zhuang, Liang; Yu, Shiying

2017-01-01

Triple-negative breast cancer (TNBC) is a high-risk breast cancer phenotype without specific targeted therapy options and is significantly associated with increased local recurrence in patients treated with radiotherapy. CAVEOLIN-1 (CAV-1)-mediated epidermal growth factor receptor (EGFR) nuclear translocation following irradiation promotes DNA repair and thus induces radiation resistance. In this study, we aimed to determine whether knockdown of CAV-1 enhances the radiosensitivity of basal-like TNBC cell lines and to explore the possible mechanisms. Western blotting was used to compare protein expression in a panel of breast cancer cell lines. Nuclear accumulation of EGFR as well as DNA repair and damage at multiple time points following irradiation with or without CAV-1 siRNA pretreatment were investigated using western blotting and confocal microscopy. The radiosensitizing effect of CAV-1 siRNA was evaluated using a clonogenic assay. Flowcytometry was performed to analyse cell apoptosis and cell cycle alteration. We found that CAV-1 is over-expressed in basal-like TNBC cell lines and barely expressed in HER-2-positive cells; additionally, we observed that HER-2-positive cell lines are more sensitive to irradiation than basal-like TNBC cells. Our findings revealed that radiation-induced EGFR nuclear translocation was impaired by knockdown of CAV-1. In parallel, radiation-induced elevation of DNA repair proteins was also hampered by pretreatment with CAV-1 siRNA before irradiation. Silencing of CAV-1 also promoted DNA damage 24 h after irradiation. Colony formation assays verified that cells could be radiosensitized after knockdown of CAV-1. Furthermore, G2/M cell cycle arrest and apoptosis enhancement may also contribute to the radiosensitizing effect of CAV-1 siRNA. Our results support the hypothesis that CAV-1 knockdown by siRNA causes increased radiosensitivity in basal-like TNBC cells. The mechanisms associated with this effect are reduced DNA repair through delayed CAV-1-associated EGFR nuclear accumulation and induction of G2/M arrest and apoptosis through the combined effects of CAV-1 siRNA and radiation. © 2017 The Author(s). Published by S. Karger AG, Basel.

A selective plasmin inhibitor, trans-aminomethylcyclohexanecarbonyl-L-(O-picolyl)tyrosine-octylamide (YO-2), induces thymocyte apoptosis.

PubMed

Lee, Eibai; Enomoto, Riyo; Takemura, Kazu; Tsuda, Yuko; Okada, Yoshio

2002-04-01

The treatment of rat thymocytes with YO-2, a novel inhibitor of plasmin, resulted in an increase in DNA fragmentation. DNA fragmentation was also induced by another YO compounds such as YO-0, -3, -4 and -5. These YO compounds are the inhibitor of plasmin activity. On the other hand, YO-1, -6 and -8 that hardly inhibit plasmin activity had no effect on DNA fragmentation. Analysis of fragmented DNA from thymocytes treated with YO-2 by agarose gel electrophoresis revealed that the compound caused internucleosomal DNA fragmentation. In addition, judging from a laser scanning microscopy, annexin V-positive and propidium iodide-negative cells were increased by the treatment of the cells with the compound. Moreover, chromatin condensation was observed in thymocytes treated with the compound. These results demonstrated that YO-2 induces thymocyte apoptosis. There seemed to be some correlation between the apoptosis induced by YO compounds and their plasmin inhibitory effect. However, because the other protease inhibitors including pepstatin A, leupeptin, AEBSF, DFP and E-64-d did not affect DNA fragmentation, YO compounds are likely to have unique mechanism on plasmin or to show the effect on the other plasmin-like proteases. The plasmin inhibitory activity may have an important role in YO-2-induced apoptosis. Furthermore, the stimulations of caspase-8, -9 and -3-like activities were observed in thymocytes treated with YO-2. These results suggest that YO-2 induces thymocyte apoptosis via activation of caspase cascade.

Cell death in the thymus--it' s all a matter of contacts.

PubMed

Minter, Lisa M; Osborne, Barbara A

2003-06-01

Apoptosis, or programmed cell death, plays a critical role in shaping the T cell repertoire, deleting unproductive as well as potentially autoreactive T cells. Our understanding of how thymocyte apoptosis is regulated is continually evolving, as new essential modulators of this process are discovered. A conundrum that remains, however, is how signaling through essentially the same receptors and cascades evokes distinct biological responses: death by neglect, positive or negative selection. We hypothesize that the immunological synapse (IS) may be critical to transducing survival signals during thymocyte development, and suggest that factors affecting IS assembly may also influence T cell selection.

Astragaloside IV attenuates injury caused by myocardial ischemia/reperfusion in rats via regulation of toll-like receptor 4/nuclear factor-κB signaling pathway.

PubMed

Lu, Meili; Tang, Futian; Zhang, Jing; Luan, Aina; Mei, Meng; Xu, Chonghua; Zhang, Suping; Wang, Hongxin; Maslov, Leonid N

2015-04-01

Myocardial ischemia/reperfusion (MI/R) injury, in which inflammatory response and cell apoptosis play a vital role, is frequently encountered in clinical practice. Astragaloside IV (AsIV), a small molecular saponin of Astragalus membranaceus, has been shown to confer protective effects against many cardiovascular diseases. The present study was aimed to investigate the antiinflammatory and antiapoptotic effects and the possible mechanism of AsIV on MI/R injury in rats. Rats were randomly divided into sham operation group, MI/R group and groups with combinations of MI/R and different doses of AsIV. The results showed that the expressions of myocardial toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB) were significantly increased, and apoptosis of cardiomyocytes was induced in MI/R group compared with that in sham operation group. Administration of AsIV attenuated MI/R injury, downregulated the expressions of TLR4 and NF-κB and inhibited cell apoptosis as evidenced by decreased terminal deoxynucleotidyl transferase dUTP nick end labeling positive cells, B-cell lymphoma-2 associated X protein and caspase-3 expressions and increased B-cell lymphoma-2 expression compared with that in MI/R group. In addition, AsIV treatment reduced levels of inflammatory cytokines induced by MI/R injury. In conclusion, our results demonstrated that AsIV downregulates TLR4/NF-κB signaling pathway and inhibits cell apoptosis, subsequently attenuating MI/R injury in rats. Copyright © 2015 John Wiley & Sons, Ltd.

Graphene oxide and reduced graphene oxide induced neural pheochromocytoma-derived PC12 cell lines apoptosis and cell cycle alterations via the ERK signaling pathways.

PubMed

Kang, Yiyuan; Liu, Jia; Wu, Junrong; Yin, Qian; Liang, Huimin; Chen, Aijie; Shao, Longquan

2017-01-01

Given the novel applications of graphene materials in biomedical and electronics industry, the health hazards of these particles have attracted extensive worldwide attention. Although many studies have been performed on graphene material-induced toxic effects, toxicological data for the effect of graphene materials on the nervous system are lacking. In this study, we focused on the biological effects of graphene oxide (GO) and reduced graphene oxide (rGO) materials on PC12 cells, a type of traditional neural cell line. We found that GO and rGO exerted significant toxic effects on PC12 cells in a dose- and time-dependent manner. Moreover, apoptosis appeared to be a response to toxicity. A potent increase in the number of PC12 cells at G0/G1 phase after GO and rGO exposure was detected by cell cycle analysis. We found that phosphorylation levels of ERK signaling molecules, which are related to cell cycle regulation and apoptosis, were significantly altered after GO and rGO exposure. In conclusion, our results show that GO has more potent toxic effects than rGO and that apoptosis and cell cycle arrest are the main toxicity responses to GO and rGO treatments, which are likely due to ERK pathway regulation.

Cell death proteomics database: consolidating proteomics data on cell death.

PubMed

Arntzen, Magnus Ø; Bull, Vibeke H; Thiede, Bernd

2013-05-03

Programmed cell death is a ubiquitous process of utmost importance for the development and maintenance of multicellular organisms. More than 10 different types of programmed cell death forms have been discovered. Several proteomics analyses have been performed to gain insight in proteins involved in the different forms of programmed cell death. To consolidate these studies, we have developed the cell death proteomics (CDP) database, which comprehends data from apoptosis, autophagy, cytotoxic granule-mediated cell death, excitotoxicity, mitotic catastrophe, paraptosis, pyroptosis, and Wallerian degeneration. The CDP database is available as a web-based database to compare protein identifications and quantitative information across different experimental setups. The proteomics data of 73 publications were integrated and unified with protein annotations from UniProt-KB and gene ontology (GO). Currently, more than 6,500 records of more than 3,700 proteins are included in the CDP. Comparing apoptosis and autophagy using overrepresentation analysis of GO terms, the majority of enriched processes were found in both, but also some clear differences were perceived. Furthermore, the analysis revealed differences and similarities of the proteome between autophagosomal and overall autophagy. The CDP database represents a useful tool to consolidate data from proteome analyses of programmed cell death and is available at http://celldeathproteomics.uio.no.

[Biological function of trophology and the pathogenesis of metabolic syndrome--syndrome of overeating. Phylogenetically theory of general pathology, role of leptin and adiponectin].

PubMed

Titov, V N

2014-01-01

Metabolic syndrome (overeating) is a phylogenetically-determined succession of symptoms with the same pathogenesis. There is only one etiological factor, namely, increased consumption of physiologically optimal food. Enterocytes and omental fat cells are a phylogenetically early paracrine-regulated cell community that realizes the biological reactions of exo- and endotrophy. Visceral obesity, high levels of unesterified fatty acids (FA), formation of a pool of micellar FA in the blood, integration of these FA into endothelial cell plasma membrane and enlargement of adipocytes are the causes of hydrodynamic pressure elevation. Toll-like receptors recognize the associates between albumin and greater than physiological number of FA as "foreing" and initiate inflammatory response. "Endoplasm stress" develops in lipid-overloaded cells, protein synthesis (folding) in them is impaired and apoptosis-like cell death is activated. Visceral fat is a phylogenetically early depot of FA to fulfill the biological function of homeostasis, trophology, endoecology and adaptation; it is regulated at the level of paracrine communities and is anatomically limited. The subcutaneous fat depot fulfills the phylogenetically late function of locomotion; the depot size is not anatomically limited. Visceral fat cells have no receptors for phylogenetically late insulin (INS); specialized adipocyes bearing INS and GLUT4 receptors are cells that form the subcutaneous depot. These cells are regulated by phylogenetically late humoral factors at the entire body level. Leptin is an initiator of humoral hypothalamic regulation of in vivo number of ontogenetically programmed number of visceral INS-insensitive fat cells. It prevents "endoplasm stress" and apoptosis, being designed to regulate the amount of consumed food. Leptin initiates storage of FA from visceral pool into subcutaneous pool. Adiponectin is a phylogenetically late humoral hypothalamic regulatory factor that controls optimal number of fat cells in vivo. Its biological role consists in regulation of the number (proliferation) of insulin-dependent adipocytes in subcutaneous fatty tissue.

Withaferin-A Induces Apoptosis in Osteosarcoma U2OS Cell Line via Generation of ROS and Disruption of Mitochondrial Membrane Potential.

PubMed

Zhang, Hui-Liang; Zhang, Hong

2017-01-01

Withaferin-A (WF-A) is a well-known dietary compound isolated from Withania sominifera . It has tremendous pharmacological potential and has been shown to exhibit antiproliferative activity against several types of cancerous cells. Currently, the main focus of anti-cancer therapeutic development is to identify apoptosis inducing drug-like molecules. Osteosarcoma is a rare type of osteocancer, affecting human. The present study therefore focused on the evaluation of antitumor potential of WF-A against several osteosarcoma cell lines. MTT assay was used to evaluate WF-A against osteosarcoma cell lines and to calculate the IC 50 . DAPI staining was used to confirm the apoptosis inducing potential of WF-A. Mitochondrial membrane potential, reactive oxygen species (ROS) assay, and Western blotting were used to confirm the basis of apoptosis. The results revealed that that WF-A exhibited strong antiproliferative activity against all the cells lines, with IC 50 ranging from 0.32 to 7.6 μM. The lowest IC 50 (0.32 μM) was observed against U2OS cell line and therefore it was selected for further analysis. DAPI staining indicated that WF-A exhibited antiproliferative activity via induction of apoptosis. Moreover, WF-A induced ROS-mediated reduction in mitochondrial membrane potential ΔΨm) in a dose-dependent manner and activation of caspase-3 in osteosarcoma cells. We propose that WF-A may prove a potent therapeutic agent for inducing apoptosis in osteosarcoma cell lines via generation of ROS and disruption of mitochondrial membrane potential. WF-A exhibits strong anticancer activity against osteosarcoma cell linesAntiproliferative activity of WF-A is via induction of apoptosisWF-A induced ROS-mediated reduction in mitochondrial membrane potentialWF-A induced expression of caspase-3 in osteosarcoma cells. Abbreviations used: WA: Withaferin A; ROS: Reactive oxygen species; OS: Osteosarcoma; MMP: Mitochondrial membrane potential.

Memantine Can Reduce Ethanol-Induced Caspase-3 Activity and Apoptosis in H4 Cells by Decreasing Intracellular Calcium.

PubMed

Wang, Xiaolong; Chen, Jiajun; Wang, Hongbo; Yu, Hao; Wang, Changliang; You, Jiabin; Wang, Pengfei; Feng, Chunmei; Xu, Guohui; Wu, Xu; Zhao, Rui; Zhang, Guohua

2017-08-01

Caspase-3 activation and apoptosis are associated with various neurodegenerative disorders. Calcium activation is an important factor in promoting apoptosis. We, therefore, assessed the role of intracellular calcium in ethanol-induced activation of caspase-3 in H4 human neuroglioma cells and the protective effect of the NMDA receptor antagonist, memantine, on ethanol-induced apoptosis in H4 cells. H4 cells were treated with 100 mM EtOH (in culture medium) for 2 days. For interaction studies, cells were treated with memantine (4 μM), EDTA (1 mM), or BAPTA-AM (10 μM) before treatment with EtOH. Knockdown of the gene encoding the NR1 subunit of the NMDA receptor was performed using RNAi. Apoptosis was detected by Annexin V-FITC/PI staining and flow cytometry. Cell viability was detected using an MTS cell proliferation kit. Fluorescence dual wavelength spectrophotometry was used to determine the intracellular calcium concentration. The levels of NR1, caspase-3, IP3R1, and SERCA1 proteins were detected by western blotting. NR1, IP3R1, and SERCA1 mRNA levels were detected by qPCR. We observed increased expression of NR1, IP3R1, SERCA1, and increased intracellular levels of calcium ions in H4 cells exposed to ethanol. In addition, the calcium chelators, EDTA and BAPTA, and RNAi disruption of the NMDA receptor reduced ethanol-induced caspase-3 activation in H4 cells. Memantine treatment reduced the ethanol-induced increase of intracellular calcium, caspase-3 activation, apoptosis, and the ethanol-induced decrease in cell viability. Our results indicate that ethanol-induced caspase-3 activation and apoptosis are likely to be dependent on cytosolic calcium levels and that they can be reduced by memantine treatment.

Reserve stem cells: Reprogramming of differentiated cells fuels repair, metaplasia, and neoplasia in the adult gastrointestinal tract

PubMed Central

Mills, Jason C.; Sansom, Owen J.

2016-01-01

It has long been known that differentiated cells can switch fates, especially in vitro, but only recently has there been a critical mass of publications describing the mechanisms adult, post-mitotic cells use in vivo to reverse their differentiation state. We propose that this sort of cellular reprogramming is a fundamental cellular process akin to apoptosis or mitosis. Because reprogramming can invoke regenerative cells from mature cells, it is critical to the longterm maintenance of tissues like the pancreas, which encounter large insults during adulthood but lack constitutively active adult stem cells to repair the damage. However, even in tissues with adult stem cells, like stomach and intestine, reprogramming may allow mature cells to serve as reserve (“quiescent”) stem cells when normal stem cells are compromised. We propose that the potential downside to reprogramming is that it increases risk for cancers that occur late in adulthood. Mature, long-lived cells may have years of exposure to mutagens. Mutations that affect the physiological function of differentiated, post-mitotic cells may lead to apoptosis, but mutations in genes that govern proliferation might not be selected against. Hence, reprogramming with reentry into the cell cycle might unmask those mutations, causing an irreversible progenitor-like, proliferative state. We review recent evidence showing that reprogramming fuels irreversible metaplastic and precancerous proliferations in stomach and pancreas. Finally, we illustrate how we think reprogrammed differentiated cells are likely candidates as cells of origin for cancers of the intestine. PMID:26175494

A microtubule inhibitor, ABT-751, induces autophagy and delays apoptosis in Huh-7 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wei, Ren-Jie

The objective was to investigate the upstream mechanisms of apoptosis which were triggered by a novel anti-microtubule drug, ABT-751, in hepatocellular carcinoma-derived Huh-7 cells. Effects of ABT-751 were evaluated by immunocytochemistry, flow cytometric, alkaline comet, soft agar, immunoblotting, CytoID, green fluorescent protein-microtubule associated protein 1 light chain 3 beta detection, plasmid transfection, nuclear/cytosol fractionation, coimmunoprecipitation, quantitative reverse transcription-polymerase chain reaction, small-hairpin RNA interference and mitochondria/cytosol fractionation assays. Results showed that ABT-751 caused dysregulation of microtubule, collapse of mitochondrial membrane potential, generation of reactive oxygen species (ROS), DNA damage, G{sub 2}/M cell cycle arrest, inhibition of anchorage-independent cell growth and apoptosismore » in Huh-7 cells. ABT-751 also induced early autophagy via upregulation of nuclear TP53 and downregulation of the AKT serine/threonine kinase (AKT)/mechanistic target of rapamycin (MTOR) pathway. Through modulation of the expression levels of DNA damage checkpoint proteins and G{sub 2}/M cell cycle regulators, ABT-751 induced G{sub 2}/M cell cycle arrest. Subsequently, ABT-751 triggered apoptosis with marked downregulation of B-cell CLL/lymphoma 2, upregulation of mitochondrial BCL2 antagonist/killer 1 and BCL2 like 11 protein levels, and cleavages of caspase 8 (CASP8), CASP9, CASP3 and DNA fragmentation factor subunit alpha proteins. Suppression of ROS significantly decreased ABT-751-induced autophagic and apoptotic cells. Pharmacological inhibition of autophagy significantly increased the percentages of ABT-751-induced apoptotic cells. The autophagy induced by ABT-751 plays a protective role to postpone apoptosis by exerting adaptive responses following microtubule damage, ROS and/or impaired mitochondria. - Highlights: • An anti-microtubule agent, ABT-751, induces autophagy and apoptosis in Huh-7 cells. • ABT-751 induces early autophagy and delays apoptosis. • ABT-751 induces autophagy via modulations of nuclear TP53 and AKT/MTOR pathways. • ABT-751-induced apoptosis is ROS-, mitochondria- and caspase-dependent. • Inhibition of autophagy enhances ABT-751-induced apoptosis.« less

Methamphetamine exposure triggers apoptosis and autophagy in neuronal cells by activating the C/EBPβ-related signaling pathway.

PubMed

Xu, Xiang; Huang, Enping; Luo, Baoying; Cai, Dunpeng; Zhao, Xu; Luo, Qin; Jin, Yili; Chen, Ling; Wang, Qi; Liu, Chao; Lin, Zhoumeng; Xie, Wei-Bing; Wang, Huijun

2018-06-25

Methamphetamine (Meth) is a widely abused psychoactive drug that primarily damages the nervous system, notably causing dopaminergic neuronal apoptosis. CCAAT-enhancer binding protein (C/EBPβ) is a transcription factor and an important regulator of cell apoptosis and autophagy. Insulin-like growth factor binding protein (IGFBP5) is a proapoptotic factor that mediates Meth-induced neuronal apoptosis, and Trib3 (tribbles pseudokinase 3) is an endoplasmic reticulum (ER) stress-inducible gene involved in autophagic cell death through the mammalian target of rapamycin (mTOR) signaling pathway. To test the hypothesis that C/EBPβ is involved in Meth-induced IGFBP5-mediated neuronal apoptosis and Trib3-mediated neuronal autophagy, we measured the protein expression of C/EBPβ after Meth exposure and evaluated the effects of silencing C/EBPβ, IGFBP5, or Trib3 on Meth-induced apoptosis and autophagy in neuronal cells and in the rat striatum after intrastriatal Meth injection. We found that, at relatively high doses, Meth exposure increased C/EBPβ protein expression, which was accompanied by increased neuronal apoptosis and autophagy; triggered the IGFBP5-mediated, p53-up-regulated modulator of apoptosis (PUMA)-related mitochondrial apoptotic signaling pathway; and stimulated the Trib3-mediated ER stress signaling pathway through the Akt-mTOR signaling axis. We also found that autophagy is an early response to Meth-induced stress upstream of apoptosis and plays a detrimental role in Meth-induced neuronal cell death. These results suggest that Meth exposure induces C/EBPβ expression, which plays an essential role in the neuronal apoptosis and autophagy induced by relatively high doses of Meth; however, relatively low concentrations of Meth did not change the expression of C/EBPβ in vitro. Further studies are needed to elucidate the role of C/EBPβ in low-dose Meth-induced neurotoxicity.-Xu, X., Huang, E., Luo, B., Cai, D., Zhao, X., Luo, Q., Jin, Y., Chen, L., Wang, Q., Liu, C., Lin, Z., Xie, W.-B., Wang, H. Methamphetamine exposure triggers apoptosis and autophagy in neuronal cells by activating the C/EBPβ-related signaling pathway.

Artemisinin Represses Telomerase Subunits and Induces Apoptosis in HPV-39 Infected Human Cervical Cancer Cells.

PubMed

Mondal, Anushree; Chatterji, Urmi

2015-09-01

Artemisinin, a plant-derived antimalarial drug with relatively low toxicity on normal cells in humans, has selective anticancer activities in various types of cancers, both in vitro and in vivo. In the present study, we have investigated the anticancer effects of artemisinin in human cervical cancer cells, with special emphasis on its role in inducing apoptosis and repressing cell proliferation by inhibiting the telomerase subunits, ERα which is essential for maintenance of the cervix, and downstream components like VEGF, which is known to activate angiogenesis. Effects of artemisinin on apoptosis of ME-180 cells were measured by flow cytometry, DAPI, and annexin V staining. Expression of genes and proteins related to cell proliferation and apoptosis was quantified both at the transcriptional and translational levels by semi-quantitative RT-PCR and western blot analysis, respectively. Our findings demonstrated that artemisinin significantly downregulated the expression of ERα and its downstream component, VEGF. Antiproliferative activity was also supported by decreased telomerase activity and reduced expression of hTR and hTERT subunits. Additionally, artemisinin reduced the expression of the HPV-39 viral E6 and E7 components. Artemisinin-induced apoptosis was confirmed by FACS, nuclear chromatin condensation, annexin V staining. Increased expression of p53 with concomitant decrease in expression of the p53 inhibitor Mdm2 further supported that artemisinin-induced apoptosis was p53-dependent. The results clearly indicate that artemisinin induces antiproliferative and proapoptotic effects in HPV-39-infected ME-180 cells, and warrants further trial as an effective anticancer drug. © 2015 Wiley Periodicals, Inc.

Effects of Different Zinc Species on Cellar Zinc Distribution, Cell Cycle, Apoptosis and Viability in MDAMB231 Cells.

PubMed

Wang, Yan-hong; Zhao, Wen-jie; Zheng, Wei-juan; Mao, Li; Lian, Hong-zhen; Hu, Xin; Hua, Zi-chun

2016-03-01

Intracellular metal elements exist in mammalian cells with the concentration range from picomoles per litre to micromoles per litre and play a considerable role in various biological procedures. Element provided by different species can influence the availability and distribution of the element in a cell and could lead to different biological effects on the cell's growth and function. Zinc as an abundant and widely distributed essential trace element, is involved in numerous and relevant physiological functions. Zinc homeostasis in cells, which is regulated by metallothioneins, zinc transporter/SLC30A, Zrt-/Irt-like proteins/SLC39A and metal-response element-binding transcription factor-1 (MTF-1), is crucial for normal cellular functioning. In this study, we investigated the influences of different zinc species, zinc sulphate, zinc gluconate and bacitracin zinc, which represented inorganic, organic and biological zinc species, respectively, on cell cycle, viability and apoptosis in MDAMB231 cells. It was found that the responses of cell cycle, apoptosis and death to different zinc species in MDAMB231 cells are different. Western blot analysis of the expression of several key proteins in regulating zinc-related transcription, cell cycle, apoptosis, including MTF-1, cyclin B1, cyclin D1, caspase-8 and caspase-9 in treated cells further confirmed the observed results on cell level.

Role of insulin-like growth factor-1 (IGF-1) in regulating cell cycle progression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Qi-lin; Yang, Tian-lun; Yin, Ji-ye

2009-11-06

Aims: Insulin-like growth factor-1 (IGF-1) is a polypeptide protein hormone, similar in molecular structure to insulin, which plays an important role in cell migration, cell cycle progression, cell survival and proliferation. In this study, we investigated the possible mechanisms of IGF-1 mediated cell cycle redistribution and apoptosis of vascular endothelial cells. Method: Human umbilical vein endothelial cells (HUVECs) were pretreated with 0.1, 0.5, or 2.5 {mu}g/mL of IGF-1 for 30 min before the addition of Ang II. Cell cycle redistribution and apoptosis were examined by flow cytometry. Expression of Ang II type 1 (AT{sub 1}) mRNA and cyclin E proteinmore » were determined by RT-PCR and Western blot, respectively. Results: Ang II (1 {mu}mol/L) induced HUVECs arrested at G{sub 0}/G{sub 1}, enhanced the expression level of AT{sub 1} mRNA in a time-dependent manner, reduced the enzymatic activity of nitric oxide synthase (NOS) and nitric oxide (NO) content as well as the expression level of cyclin E protein. However, IGF-1 enhanced NOS activity, NO content, and the expression level of cyclin E protein, and reduced the expression level of AT{sub 1} mRNA. L-NAME significantly counteracted these effects of IGF-1. Conclusions: Our data suggests that IGF-1 can reverse vascular endothelial cells arrested at G{sub 0}/G{sub 1} and apoptosis induced by Ang II, which might be mediated via a NOS-NO signaling pathway and is likely associated with the expression levels of AT1 mRNA and cyclin E proteins.« less

Topological Transitions in Mitochondrial Membranes controlled by Apoptotic Proteins

NASA Astrophysics Data System (ADS)

Hwee Lai, Ghee; Sanders, Lori K.; Mishra, Abhijit; Schmidt, Nathan W.; Wong, Gerard C. L.; Ivashyna, Olena; Schlesinger, Paul H.

2010-03-01

The Bcl-2 family comprises pro-apoptotic proteins, capable of permeabilizing the mitochondrial membrane, and anti-apoptotic members interacting in an antagonistic fashion to regulate programmed cell death (apoptosis). They offer potential therapeutic targets to re-engage cellular suicide in tumor cells but the extensive network of implicated protein-protein interactions has impeded full understanding of the decision pathway. We show, using synchrotron x-ray diffraction, that pro-apoptotic proteins interact with mitochondrial-like model membranes to generate saddle-splay (negative Gaussian) curvature topologically required for pore formation, while anti-apoptotic proteins can deactivate curvature generation by molecules drastically different from Bcl-2 family members and offer evidence for membrane-curvature mediated interactions general enough to affect very disparate systems.

The essence of insect metamorphosis and aging: electrical rewiring of cells driven by the principles of juvenile hormone-dependent Ca(2+)-homeostasis.

PubMed

De Loof, Arnold; De Haes, Wouter; Janssen, Tom; Schoofs, Liliane

2014-04-01

In holometabolous insects the fall to zero of the titer of Juvenile Hormone ends its still poorly understood "status quo" mode of action in larvae. Concurrently it initiates metamorphosis of which the programmed cell death of all internal tissues that actively secrete proteins, such as the fat body, midgut, salivary glands, prothoracic glands, etc. is the most drastic aspect. These tissues have a very well developed rough endoplasmic reticulum, a known storage site of intracellular Ca(2+). A persistent high [Ca(2+)]i is toxic, lethal and causal to apoptosis. Metamorphosis becomes a logical phenomenon if analyzed from: (1) the causal link between calcium toxicity and apoptosis; (2) the largely overlooked fact that at least some isoforms of Ca(2+)-ATPases have a binding site for farnesol-like endogenous sesquiterpenoids (FRS). The Ca(2+)-ATPase blocker thapsigargin, like JH a sesquiterpenoid derivative, illustrates how absence of JH might work. The Ca(2+)-homeostasis system is concurrently extremely well conserved in evolution and highly variable, enabling tissue-, developmental-, and species specificity. As long as JH succeeds in keeping [Ca(2+)]i low by keeping the Ca(2+)-ATPases pumping, it acts as "the status quo" hormone. When it disappears, its various inhibitory effects are lifted. The electrical wiring system of cells, in particular in the regenerating tissues, is subject to change during metamorphosis. The possibility is discussed that in vertebrates an endogenous farnesol-like sesquiterpenoid, probably farnesol itself, acts as a functional, but hitherto completely overlooked Juvenile anti-aging "Inbrome", a novel concept in signaling. Copyright © 2014 Elsevier Inc. All rights reserved.

Expression of progesterone receptor membrane component-2 within the immature rat ovary and its role in regulating mitosis and apoptosis of spontaneously immortalized granulosa cells.

PubMed

Griffin, Daniel; Liu, Xiufang; Pru, Cindy; Pru, James K; Peluso, John J

2014-08-01

Progesterone receptor membrane component 2 (Pgrmc2) mRNA was detected in the immature rat ovary. By 48 h after eCG, Pgrmc2 mRNA levels decreased by 40% and were maintained at 48 h post-hCG. Immunohistochemical studies detected PGRMC2 in oocytes and ovarian surface epithelial, interstitial, thecal, granulosa, and luteal cells. PGRMC2 was also present in spontaneously immortalized granulosa cells, localizing to the cytoplasm of interphase cells and apparently to the mitotic spindle of cells in metaphase. Interestingly, PGRMC2 levels appeared to decrease during the G1 stage of the cell cycle. Moreover, overexpression of PGRMC2 suppressed entry into the cell cycle, possibly by binding the p58 form of cyclin dependent kinase 11b. Conversely, Pgrmc2 small interfering RNA (siRNA) treatment increased the percentage of cells in G1 and M stage but did not increase the number of cells, which was likely due to an increase in apoptosis. Depleting PGRMC2 did not inhibit cellular (3)H-progesterone binding, but attenuated the ability of progesterone to suppress mitosis and apoptosis. Taken together these studies suggest that PGRMC2 affects granulosa cell mitosis by acting at two specific stages of the cell cycle. First, PGRMC2 regulates the progression from the G0 into the G1 stage of the cell cycle. Second, PGRMC2 appears to localize to the mitotic spindle, where it likely promotes the final stages of mitosis. Finally, siRNA knockdown studies indicate that PGRMC2 is required for progesterone to slow the rate of granulosa cell mitosis and apoptosis. These findings support a role for PGRMC2 in ovarian follicle development. © 2014 by the Society for the Study of Reproduction, Inc.

Interactions between insulin-like growth factor-I, estrogen receptor-α (ERα) and ERβ in regulating growth/apoptosis of MCF-7 human breast cancer cells

PubMed Central

Mendoza, Rhone A.; Enriquez, Marlene I; Mejia, Sylvia M; Moody, Emily E; Thordarson, Gudmundur

2011-01-01

Understanding of the interactions between estradiol (E2) and insulin-like growth factor-I (IGF-I) is still incomplete. Cell lines derived from the MCF-7 breast cancer cells were generated with suppressed expression of the IGF-I receptor (IGF-IR), termed IGF-IR.low cells, by stable transfection using small interfering RNA (siRNA) expression vector. Vector for control cells carried sequence generating non-interfering RNA. Concomitant with reduction in the IGF-IR levels, the IGF-IR.low cells also showed a reduction in estrogen receptor α (ERα) and progesterone receptor expressions and an elevation in the expression of ERβ. The number of the IGF-IR.low cells was reduced in response to IGF-I and human growth hormone plus epidermal growth factor, but E2 did not cause increase in the number of the IGF-IR.low cells compared to controls. Proliferation rate of IGF-IR.low cells was only reduced in response to E2 compared to controls, whereas their basal and hormone stimulated apoptosis rate was increased. Phosphorylation of p38 mitogen activated protein kinase (p38 MAPK) was increased in the IGF-IR.low cells after treatment with E2, without affecting control cells. Further, phosphorylation of the tumor suppressor protein p53 was elevated in the IGF-IR.low cells compared to the controls. Summary, suppressing the IGF-IR expression decreased the level of ERα but increased the level of ERβ. Overall growth rate of the IGF-IR.low cells was reduced mostly through an increase in apoptosis without affecting proliferation substantially. We hypothesize that a decreased ERα:ERβ ratio triggered a rapid phosphorylation of p38 MAPK which in turn phosphorylated the p53 tumor suppressor and accelerated apoptosis rate. PMID:20974640

E1A enhances cellular sensitivity to DNA-damage-induced apoptosis through PIDD-dependent caspase-2 activation.

PubMed

Radke, Jay R; Siddiqui, Zeba K; Figueroa, Iris; Cook, James L

Expression of the adenoviral protein, E1A, sensitizes mammalian cells to a wide variety of apoptosis-inducing agents through multiple cellular pathways. For example, E1A sensitizes cells to apoptosis induced by TNF-superfamily members by inhibiting NF-kappa B (NF- κ B)-dependent gene expression. In contrast, E1A sensitization to nitric oxide, an inducer of the intrinsic apoptotic pathway, is not dependent upon repression of NF- κ B-dependent transcription but rather is dependent upon caspase-2 activation. The latter observation suggested that E1A-induced enhancement of caspase-2 activation might be a critical factor in cellular sensitization to other intrinsic apoptosis pathway-inducing agents. Etoposide and gemcitabine are two DNA damaging agents that induce intrinsic apoptosis. Here we report that E1A-induced sensitization to both of these agents, like NO, is independent of NF- κ B activation but dependent on caspase-2 activation. The results show that caspase-2 is a key mitochondrial-injuring caspase during etoposide and gemcitabine-induced apoptosis of E1A-positive cells, and that caspase-2 is required for induction of caspase-3 activity by both chemotherapeutic agents. Expression of PIDD was required for caspase-2 activation, mitochondrial injury and enhanced apoptotic cell death. Furthermore, E1A-enhanced sensitivity to injury-induced apoptosis required PIDD cleavage to PIDD-CC. These results define the PIDD/caspase-2 pathway as a key apical, mitochondrial-injuring mechanism in E1A-induced sensitivity of mammalian cells to chemotherapeutic agents.

Olive (Olea europaea) Leaf Extract Induces Apoptosis and Monocyte/Macrophage Differentiation in Human Chronic Myelogenous Leukemia K562 Cells: Insight into the Underlying Mechanism

PubMed Central

Han, Junkyu; Jlaiel, Lobna; Sayadi, Sami; Isoda, Hiroko

2014-01-01

Differentiation therapy is an attractive approach aiming at reversing malignancy and reactivating endogenous differentiation programs in cancer cells. Olive leaf extract, known for its antioxidant activity, has been demonstrated to induce apoptosis in several cancer cells. However, its differentiation inducing properties and the mechanisms involved are still poorly understood. In this study, we investigated the effect of Chemlali Olive Leaf Extract (COLE) for its potential differentiation inducing effect on multipotent leukemia K562 cells. Results showed that COLE inhibits K562 cells proliferation and arrests the cell cycle at G0/G1, and then at G2/M phase over treatment time. Further analysis revealed that COLE induces apoptosis and differentiation of K562 cells toward the monocyte lineage. Microarray analysis was conducted to investigate the underlying mechanism of COLE differentiation inducing effect. The differentially expressed genes such as IFI16, EGR1, NFYA, FOXP1, CXCL2, CXCL3, and CXCL8 confirmed the commitment of K562 cells to the monocyte/macrophage lineage. Thus our results provide evidence that, in addition to apoptosis, induction of differentiation is one of the possible therapeutic effects of olive leaf in cancer cells. PMID:24803988

Olive (Olea europaea) leaf extract induces apoptosis and monocyte/macrophage differentiation in human chronic myelogenous leukemia K562 cells: insight into the underlying mechanism.

PubMed

Samet, Imen; Han, Junkyu; Jlaiel, Lobna; Sayadi, Sami; Isoda, Hiroko

2014-01-01

Differentiation therapy is an attractive approach aiming at reversing malignancy and reactivating endogenous differentiation programs in cancer cells. Olive leaf extract, known for its antioxidant activity, has been demonstrated to induce apoptosis in several cancer cells. However, its differentiation inducing properties and the mechanisms involved are still poorly understood. In this study, we investigated the effect of Chemlali Olive Leaf Extract (COLE) for its potential differentiation inducing effect on multipotent leukemia K562 cells. Results showed that COLE inhibits K562 cells proliferation and arrests the cell cycle at G0/G1, and then at G2/M phase over treatment time. Further analysis revealed that COLE induces apoptosis and differentiation of K562 cells toward the monocyte lineage. Microarray analysis was conducted to investigate the underlying mechanism of COLE differentiation inducing effect. The differentially expressed genes such as IFI16, EGR1, NFYA, FOXP1, CXCL2, CXCL3, and CXCL8 confirmed the commitment of K562 cells to the monocyte/macrophage lineage. Thus our results provide evidence that, in addition to apoptosis, induction of differentiation is one of the possible therapeutic effects of olive leaf in cancer cells.

Zoledronate induces apoptosis in cells from fibro-cellular membrane of unicameral bone cyst (UBC).

PubMed

Yu, John; Chang, Seong-Sil; Suratwala, Sanjeev; Chung, Woo-Sik; Abdelmessieh, Peter; Lee, Hahn-Jun; Yang, Jay; Lee, Francis Young-In

2005-09-01

Unicameral bone cyst (UBC) is a benign cystic lesion in children which is prone to fracture. Various treatments are available, but recurrence after different types of percutaneous injection therapy can cause bone destruction and pathologic fracture. The potential therapeutic effects of anti-resorptive agents, such as bisphosphonates, have not been investigated for UBC. The objective of this study was to characterize the cells from the fibro-cellular membrane of unicameral bone cyst (UBC cells) and to determine whether zoledronate, a nitrogen-containing bisphosphonate, could induce apoptosis in UBC cells. Flow cytometry and immunoblotting were performed in order to determine whether zoledronate induced apoptosis. Cells derived from normal human trabecular bones were used as controls against UBC cells to compare the effect of zoledronate in inducing apoptosis. Immunohisto/cytochemistry (IHC/ICC) and mini-array analyses were performed on tissues and cultured cells. Isolated peripheral blood mononuclear cells were incubated with conditioned media from the UBC cells to determine whether they are capable of inducing osteoclastogenesis. UBC membrane is composed of cells staining positively with CD68, SDF-1, STRO-1 and RANKL, but in vitro cells showed no staining with antibodies to CD68 and STRO-1, suggesting that there was a clonal selection of stromal cells during cell culture. UBC cells also express RUNX2 (runt-related transcription factor-2, core binding factor-1), a key transcription factor for osteoblastic differentiation. In addition, media collected from UBC cells induced a generation of multi-nucleated osteoclast-like cells of peripheral blood mononuclear cells. Zoledronate induced apoptosis of UBC cells in a dose-dependent manner. Apoptosis was evidenced by induction of the active cleaved form of caspase-3. The baseline apoptotic fractions were similar in UBC cells and trabecular bone cells. However, in the overall apoptotic fractions in this study, trabecular bone cells showed 17.2% of apoptosis, significantly lower than 24.2% of UBC cells (p-value=0.007). With the various zoledronate concentrations, mean apoptotic fractions of trabecular bone cells was 19.2%, significantly lower than 27.8% of UBC cells (p-value=0.040). With GGOH co-treatment in various zoledronate concentrations, 15.1% apoptosis was shown in trabecular bone cells, which was not significantly lower than 20.6% of UBC cells (p-value=0.076). This data suggests that zoledronate causes apoptosis in both UBC and trabecular bone cells by inhibition of the mevalonate pathway. In addition to the known anti-osteoclastogenic effect of bisphosphonates, the GGOH inhibitory effects of zoledronate were more prominent in UBC cells than trabecular bone cells, indicating their potential therapeutic role in UBC.

Monitoring cell morphology during necrosis and apoptosis by quantitative phase imaging

NASA Astrophysics Data System (ADS)

Mugnano, Martina; Calabuig, Alejandro; Grilli, Simonetta; Miccio, Lisa; Ferraro, Pietro

2015-05-01

Cellular morphology changes and volume alterations play significant roles in many biological processes and they are mirrors of cell functions. In this paper, we propose the Digital Holographic microscope (DH) as a non-invasive imaging technique for a rapid and accurate extraction of morphological information related to cell death. In particular, we investigate the morphological variations that occur during necrosis and apoptosis. The study of necrosis is extremely important because it is often associated with unwarranted loss of cells in human pathologies such as ischemia, trauma, and some forms of neurodegeneration; therefore, a better elucidation in terms of cell morphological changes could pave the way for new treatments. Also, apoptosis is extremely important because it's involved in cancer, both in its formation and in medical treatments. Because the inability to initiate apoptosis enhances tumour formation, current cancer treatments target this pathway. Within this framework, we have developed a transmission off-axis DH apparatus integrated with a micro incubator for investigation of living cells in a temperature and CO2 controlled environment. We employ DH to analyse the necrosis cell death induced by laser light (wavelength 473 nm, light power 4 mW). We have chosen as cellular model NIH 3T3 mouse embryonic fibroblasts because their adhesive features such as morphological changes, and the time needed to adhere and spread have been well characterized in the literature. We have monitored cell volume changes and morphological alterations in real time in order to study the necrosis process accurately and quantitatively. Cell volume changes were evaluated from the measured phase changes of light transmitted through cells. Our digital holographic experiments showed that after exposure of cells to laser light for 90-120 min., they swell and then take on a balloon-like shape until the plasma membrane ruptures and finally the cell volume decreases. Furthermore, we present a preliminary study on the variation of morphological parameters in case of cell apoptosis induced by exposure to 10 μM cadmium chloride. We employ the same cell line, monitoring the process for 18 hours. In the vast group of environmental pollutants, the toxic heavy metal cadmium is considered a likely candidate as a causative agent of several types of cancers. Widely distributed and used in industry, and with a broad range of target organs and a long half-life (10-30 years) in the human body, this element has been long known for its multiple adverse effects on human health, through occupational or environmental exposure. In apoptosis, we measure cell volume decrease and cell shrinking. Both data of apoptosis and necrosis were analysed by means of a Sigmoidal Statistical Distribution function, which allows several quantitative data to be established, such as swelling and cell death time, flux of intracellular material from inside to outside the cell, initial and final volume versus time. In addition, we can quantitatively study the cytoplasmatic granularity that occurs during necrosis. As a future application, DH could be employed as a non-invasive and label-free method to distinguish between apoptosis and necrosis in terms of morphological parameters.

Effects of RNA interference-mediated silencing of toll-like receptor 4 gene on proliferation and apoptosis of human breast cancer MCF-7 and MDA-MB-231 cells: An in vitro study.

PubMed

Gao, Xiao-Ling; Yang, Jiao-Jiao; Wang, Shu-Juan; Chen, Yan; Wang, Bei; Cheng, Er-Jing; Gong, Jian-Nan; Dong, Yan-Ting; Liu, Dai; Wang, Xiang-Li; Huang, Ya-Qiong; An, Dong-Dong

2018-06-22

Breast cancer is known as the most prevalent cancer in women worldwide, and has an undeniable negative impact on public health, both physically, and mentally. This study aims to investigate the effects of toll-like receptor 4 (TLR4) gene silencing on proliferation and apoptosis of human breast cancer cells to explore for a new theoretical basis for its treatment. TLR4 small interference RNA (siRNA) fragment recombinant plasmids were constructed, including TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3. Human breast cancer MCF-7 and MDA-MB-231 cells were assigned into blank, negative control (NC), TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3 groups. MCF-7 and MDA-MB-231 cell growth was detected by MTT assay. Apoptosis and cell cycle were determined by flow cytometry. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis were conducted to determine the expression of TLR4, CDK4, cyclin D1, Livin, Bcl-2, p53, c-FLIP, and caspase-3. In comparison with the NC and blank groups, the TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3 groups showed decreased the expression of TLR4, inhibited proliferation of MCF-7 and MDA-MB-231 cells and promoted MCF-7 and MDA-MB-231 cell apoptosis, and the cells were blocked in G1 phase. In comparison with the NC and blank groups, in the TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3 groups, siRNA-TLR4 significantly increased expression of p53 and caspase-3 in MCF-7 and MDA-MB-231 cells, while it decreased the expressions of CDK4, cyclinD1, Livin, Bal-2, and c-FLIP. The study demonstrates that TLR4 gene silencing inhibits proliferation and induces apoptosis of MCF-7 and MDA-MB-231 cells. © 2018 Wiley Periodicals, Inc.

Induction of apoptosis in non-small cell lung carcinoma A549 cells by PGD₂ metabolite, 15d-PGJ₂.

PubMed

Wang, Jun-Jie; Mak, Oi-Tong

2011-11-01

PGD2 (prostaglandin D2) is a mediator in various pathophysiological processes, including inflammation and tumorigenesis. PGD2 can be converted into active metabolites and is known to activate two distinct receptors, DP (PGD2 receptor) and CRTH2/DP2 (chemoattractant receptor-homologous molecule expressed on Th2 cells). In the past, PGD2 was thought to be involved principally in the process of inflammation. However, in recent years, several studies have shown that PGD2 has anti-proliferative ability against tumorigenesis and can induce cellular apoptosis via activation of the caspase-dependent pathway in human colorectal cancer cells, leukaemia cells and eosinophils. In the lung, where PGD2 is highly released when sensitized mast cells are challenged with allergen, the mechanism of PGD2-induced apoptosis is unclear. In the present study, A549 cells, a type of NSCLC (non-small cell lung carcinoma), were treated with PGD2 under various conditions, including while blocking DP and CRTH2/DP2 with the selective antagonists BWA868C and ramatroban respectively. We report here that PGD2 induces A549 cell death through the intrinsic apoptotic pathway, although the process does not appear to involve either DP or CRTH2/DP2. Similar results were also found with H2199 cells, another type of NSCLC. We found that PGD2 metabolites induce apoptosis effectively and that 15d-PGJ2 (15-deoxy-Δ12,14-prostaglandin J2) is a likely candidate for the principal apoptotic inducer in PGD2-induced apoptosis in NSCLC A549 cells.

Combined radiation and p53 gene therapy of malignant glioma cells.

PubMed

Badie, B; Goh, C S; Klaver, J; Herweijer, H; Boothman, D A

1999-01-01

More than half of malignant gliomas reportedly have alterations in the p53 tumor suppressor gene. Because p53 plays a key role in the cellular response to DNA-damaging agents, we investigated the role of p53 gene therapy before ionizing radiation in cultured human glioma cells containing normal or mutated p53. Three established human glioma cell lines expressing the wild-type (U87 MG, p53wt) or mutant (A172 and U373 MG, p53mut) p53 gene were transduced by recombinant adenoviral vectors bearing human p53 (Adp53) and Escherichia coli beta-galactosidase genes (AdLacZ, control virus) before radiation (0-20 Gy). Changes in p53, p21, and Bax expression were studied by Western immunoblotting, whereas cell cycle alterations and apoptosis were investigated by flow cytometry and nuclear staining. Survival was assessed by clonogenic assays. Within 48 hours of Adp53 exposure, all three cell lines demonstrated p53 expression at a viral multiplicity of infection of 100. p21, which is a p53-inducible downstream effector gene, was overexpressed, and cells were arrested in the G1 phase. Bax expression, which is thought to play a role in p53-induced apoptosis, did not change with either radiation or Adp53. Apoptosis and survival after p53 gene therapy varied. U87 MG (p53wt) cells showed minimal apoptosis after Adp53, irradiation, or combined treatments. U373 MG (p53mut) cells underwent massive apoptosis and died within 48 hours of Adp53 treatment, independent of irradiation. Surprisingly, A172 (p53mut) cells demonstrated minimal apoptosis after Adp53 exposure; however, unlike U373 MG cells, apoptosis increased with radiation dose. Survival of all three cell lines was reduced dramatically after >10 Gy. Although Adp53 transduction significantly reduced the survival of U373 MG cells and inhibited A172 growth, it had no effect on the U87 MG cell line. Transduction with AdLacZ did not affect apoptosis or cell cycle progression and only minimally affected survival in all cell lines. We conclude that responses to p53 gene therapy are variable among gliomas and most likely depend upon both cellular p53 status and as yet ill-defined downstream pathways involving activation of cell cycle regulatory and apoptotic genes.

MST1 is a key regulator of beta cell apoptosis and dysfunction in diabetes.

PubMed

Ardestani, Amin; Paroni, Federico; Azizi, Zahra; Kaur, Supreet; Khobragade, Vrushali; Yuan, Ting; Frogne, Thomas; Tao, Wufan; Oberholzer, Jose; Pattou, Francois; Conte, Julie Kerr; Maedler, Kathrin

2014-04-01

Apoptotic cell death is a hallmark of the loss of insulin-producing beta cells in all forms of diabetes mellitus. Current treatments fail to halt the decline in functional beta cell mass, and strategies to prevent beta cell apoptosis and dysfunction are urgently needed. Here, we identified mammalian sterile 20-like kinase-1 (MST1) as a critical regulator of apoptotic beta cell death and function. Under diabetogenic conditions, MST1 was strongly activated in beta cells in human and mouse islets and specifically induced the mitochondrial-dependent pathway of apoptosis through upregulation of the BCL-2 homology-3 (BH3)-only protein BIM. MST1 directly phosphorylated the beta cell transcription factor PDX1 at T11, resulting in the latter's ubiquitination and degradation and thus in impaired insulin secretion. MST1 deficiency completely restored normoglycemia, beta cell function and survival in vitro and in vivo. We show MST1 as a proapoptotic kinase and key mediator of apoptotic signaling and beta cell dysfunction and suggest that it may serve as target for the development of new therapies for diabetes.

Validity of SW982 synovial cell line for studying the drugs against rheumatoid arthritis in fluvastatin-induced apoptosis signaling model

PubMed Central

Chang, Jae-Ho; Lee, Kyu-Jae; Kim, Soo-Ki; Yoo, Dae-Hyun; Kang, Tae-Young

2014-01-01

Background & objectives: To study effects of drugs against rheumatoid arthritis (RA) synoviocytes or fibroblast like synoviocytes (FLS) are used. To overcome the drawbacks of using FLS, this study was conducted to show the validity of SW982 synovial cell line in RA study. Methods: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Annexin V propidium iodide (PI) staining, mitochondrial membrane potential assay, Triton X-114 Phase partitioning, and immunolot for apoptosis signaling in SW982 human synovial cell line were performed. Results: Fluvastatin induced apoptosis in a dose- and time-dependent manner in TNFα -stimulated SW982 human synovial cells. A geranylgeranylpyrophosphate (GGPP) inhibitor, but not a farnesylpyrophosphate (FPP) inhibitor, induced apoptosis, and fluvastatin-induced apoptosis was associated with the translocation of isoprenylated RhoA and Rac1 proteins from the cell membrane to the cytosol. Fluvastatin-induced downstream apoptotic signals were associated with inhibition of the phosphoinositide 3-kinase (PI3K)/Akt pathway. Accordingly, 89 kDa apoptotic cleavage fragment of poly (ADP-ribose) polymerase (PARP) was detected. Interpretation & conclusions: Collectively, our data indicate that fluvastatin induces apoptotic cell death in TNFα-stimulated SW982 human synovial cells through the inactivation of the geranylgerenylated membrane fraction of RhoA and Rac1 proteins and the subsequent inhibition of the PI3K/Akt signaling pathway. This finding shows the validity of SW982 cell line for RA study. PMID:24604047

A Druggable TCF4 and BRD4 dependent Transcriptional Network Sustains Malignancy in Blastic Plasmacytoid Dendritic Cell Neoplasm

PubMed Central

Ceribelli, Michele; Hou, Zhiying Esther; Kelly, Priscilla N.; Huang, Da Wei; Wright, George; Ganapathi, Karthik; Evbuomwan, Moses O.; Pittaluga, Stefania; Shaffer, Arthur L.; Marcucci, Guido; Forman, Stephen J.; Xiao, Wenming; Guha, Rajarshi; Zhang, Xiaohu; Ferrer, Marc; Chaperot, Laurence; Plumas, Joel; Jaffe, Elaine S.; Thomas, Craig J.; Reizis, Boris; Staudt, Louis M.

2016-01-01

SUMMARY Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is an aggressive and largely incurable hematologic malignancy originating from plasmacytoid dendritic cells (pDCs). Using RNA interference screening, we identified the E-box transcription factor TCF4 as a master regulator of the BPDCN oncogenic program. TCF4 served as a faithful diagnostic marker of BPDCN, and its downregulation caused the loss of the BPDCN-specific gene expression program and apoptosis. High-throughput drug screening revealed that bromodomain and extra-terminal domain inhibitors (BETi’s) induced BPDCN apoptosis, which was attributable to disruption of a BPDCN-specific transcriptional network controlled by TCF4-dependent super-enhancers. BETi’s retarded the growth of BPDCN xenografts, supporting their clinical evaluation in this recalcitrant malignancy. PMID:27846392

Clinical significance of SLP-2 in hepatocellular carcinoma tissues and its regulation in cancer cell proliferation, migration, and EMT

PubMed Central

Lin, Xiaoqi; Lin, Qingjun; Han, Ming; Guo, Guohu

2017-01-01

Stomatin-like protein 2 (SLP-2) gene was significantly upregulated in a variety of tumor tissues and found to be involved in proliferation and metastasis. However, its functional role in hepatocellular carcinoma (HCC) remains unknown. Our study was to investigate the function of SLP-2 in cell proliferation, migration, invasion, cell apoptosis, and the process of epithelial–mesenchymal transition (EMT) in HCC. SLP-2 mRNA and protein expression in HCC were assessed by qRT-PCR and immunohistochemical staining. In vitro, we determined cell proliferation, migration, invasion, and cell apoptosis by CCK-8, transwell, and flow cytometry assays, respectively. SLP-2 was found to be upregulated at both mRNA and protein levels in HCC tissues, and its aberrant overexpression was linked with poor prognosis in patients with HCC. SLP-2 downregulation by siRNAs significantly suppressed cell proliferation, migration, invasion, anti-apoptosis abilities, and inhibited EMT process in vitro. In conclusion, the present study demonstrated the overexpression of SLP-2 in HCC tissues for the first time. As an effective regulator involved in cell proliferation, migration, invasion, cell apoptosis, and EMT, SLP-2 could be a novel therapeutic target for patients with HCC who express high levels of SLP-2. PMID:29033585

Clinical significance of SLP-2 in hepatocellular carcinoma tissues and its regulation in cancer cell proliferation, migration, and EMT.

PubMed

Huang, Yijie; Chen, Yexi; Lin, Xiaoqi; Lin, Qingjun; Han, Ming; Guo, Guohu

2017-01-01

Stomatin-like protein 2 ( SLP-2 ) gene was significantly upregulated in a variety of tumor tissues and found to be involved in proliferation and metastasis. However, its functional role in hepatocellular carcinoma (HCC) remains unknown. Our study was to investigate the function of SLP-2 in cell proliferation, migration, invasion, cell apoptosis, and the process of epithelial-mesenchymal transition (EMT) in HCC. SLP-2 mRNA and protein expression in HCC were assessed by qRT-PCR and immunohistochemical staining. In vitro, we determined cell proliferation, migration, invasion, and cell apoptosis by CCK-8, transwell, and flow cytometry assays, respectively. SLP-2 was found to be upregulated at both mRNA and protein levels in HCC tissues, and its aberrant overexpression was linked with poor prognosis in patients with HCC. SLP-2 downregulation by siRNAs significantly suppressed cell proliferation, migration, invasion, anti-apoptosis abilities, and inhibited EMT process in vitro. In conclusion, the present study demonstrated the overexpression of SLP-2 in HCC tissues for the first time. As an effective regulator involved in cell proliferation, migration, invasion, cell apoptosis, and EMT, SLP-2 could be a novel therapeutic target for patients with HCC who express high levels of SLP-2.

Measurement of subcellular texture by optical Gabor-like filtering with a digital micromirror device

PubMed Central

Pasternack, Robert M.; Qian, Zhen; Zheng, Jing-Yi; Metaxas, Dimitris N.; White, Eileen; Boustany, Nada N.

2010-01-01

We demonstrate an optical Fourier processing method to quantify object texture arising from subcellular feature orientation within unstained living cells. Using a digital micromirror device as a Fourier spatial filter, we measured cellular responses to two-dimensional optical Gabor-like filters optimized to sense orientation of nonspherical particles, such as mitochondria, with a width around 0.45 μm. Our method showed significantly rounder structures within apoptosis-defective cells lacking the proapoptotic mitochondrial effectors Bax and Bak, when compared with Bax/Bak expressing cells functional for apoptosis, consistent with reported differences in mitochondrial shape in these cells. By decoupling spatial frequency resolution from image resolution, this method enables rapid analysis of nonspherical submicrometer scatterers in an under-sampled large field of view and yields spatially localized morphometric parameters that improve the quantitative assessment of biological function. PMID:18830354

Placental Apoptosis in Health and Disease

PubMed Central

Sharp, Andrew N.; Heazell, Alexander E.P.; Crocker, Ian P.; Mor, Gil

2011-01-01

Apoptosis, programmed cell death, is an essential feature of normal placental development but is exaggerated in association with placental disease. Placental development relies upon effective implantation and invasion of the maternal decidua by the placental trophoblast. In normal pregnancy, trophoblast apoptosis increases with placental growth and advancing gestation. However, apoptosis is notably exaggerated in the pregnancy complications, hydatidiform mole, pre-eclampsia, and intra-uterine growth restriction (IUGR). Placental apoptosis may be initiated by a variety of stimuli, including hypoxia and oxidative stress. In common with other cell-types, trophoblast apoptosis follows the extrinsic or intrinsic pathways culminating in the activation of caspases. In contrast, the formation of apoptotic bodies is less clearly identified, but postulated by some to involve the clustering of apoptotic nuclei and liberation of this material into the maternal circulation. In addition to promoting a favorable maternal immune response, the release of this placental-derived material is thought to provoke the endothelial dysfunction of pre-eclampsia. Widespread apoptosis of the syncytiotrophoblast may also impair trophoblast function leading to the reduction in nutrient transport seen in IUGR. A clearer understanding of placental apoptosis and its regulation may provide new insights into placental pathologies, potentially suggesting therapeutic targets. PMID:20367628

Toll-Like Receptor-3 and Geographic Atrophy in Age-Related Macular Degeneration

PubMed Central

Yang, Zhenglin; Stratton, Charity; Francis, Peter J.; Kleinman, Mark E.; Tan, Perciliz L.; Gibbs, Daniel; Tong, Zongzhong; Chen, Haoyu; Constantine, Ryan; Yang, Xian; Chen, Yuhong; Zeng, Jiexi; Davey, Lisa; Ma, Xiang; Hau, Vincent S.; Wang, Chi; Harmon, Jennifer; Buehler, Jeanette; Pearson, Erik; Patel, Shrena; Kaminoh, Yuuki; Watkins, Scott; Luo, Ling; Zabriskie, Norman A.; Bernstein, Paul S.; Cho, Wongil; Schwager, Andrea; Hinton, David R; Klein, Michael L; Hamon, Sara C.; Simmons, Emily; Yu, Beifeng; Campochiaro, Betsy; Sunness, Janet S.; Campochiaro, Peter; Jorde, Lynn; Parmigiani, Giovanni; Zack, Donald J.; Katsanis, Nicholas; Ambati, Jayakrishna; Zhang, Kang

2008-01-01

BACKGROUND Age-related macular degeneration (AMD) is the most common cause of irreversible visual impairment in the developed world. Advanced AMD is comprised of geographic atrophy (GA) and choroidal neovascularization (CNV). Specific genetic variants that predispose for GA are largely unknown. METHODS We tested (i) for association between the functional toll-like receptor-3 (TLR3) variant rs3775291 (L412F) and AMD in European Americans and (ii) the effect of TLR3 L and F variants on the viability of human retinal pigment epithelium (RPE) cells in vitro and on RPE cell apoptosis in wildtype and Tlr3−/− mice. RESULTS The F variant (or T allele at single nucleotide polymorphism at rs3775291) was associated with protection against GA (P=0.005); this association was replicated in two independent GA case-control series (P=5.43×10−4 and P=0.002, respectively. We observed no association between TLR3 variants and CNV. The rs377291 variant is probably critical to the function of TLR3, because a prototypic TLR3 ligand induced cell death and apoptosis in human RPE cells with the LL genotype to a greater extent than it did RPE cells with the LF genotype. Moreover, the ligand induced more RPE cell death and apoptosis in wild-type than in Tlr3−/− mice. CONCLUSIONS The TLR3 412F variant confers protection against GA, probably by suppressing RPE cell death. Given that double stranded RNA can activate TLR3-mediated apoptosis, our results suggest a possible role for viral dsRNA transcripts in the development of GA and raise awareness of potential toxicity induced by short interfering RNA (siRNA) therapeutics in the eye. PMID:18753640

Role of the HTLV-1 viral factors in the induction of apoptosis.

PubMed

Karimi, Mohammad; Mohammadi, Hamed; Hemmatzadeh, Maryam; Mohammadi, Asadollah; Rafatpanah, Houshang; Baradaran, Behzad

2017-01-01

Adult T-cell leukemia (ATL) and HTLV-1-associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP) are the two main diseases that are caused by the HTLV-1 virus. One of the features of HTLV-1 infection is its resistance against programmed cell death, which maintains the survival of cells to oncogenic transformation and underlies the viruses' therapeutic resistance. Two main genes by which the virus develops cancer are Tax and HBZ; playing an essential role in angiogenesis in regulating viral transcription and modulating multiple host factors as well as apoptosis pathways. Here we have reviewed by prior research how the apoptosis pathways are suppressed by the Tax and HBZ and new drugs which have been designed to deal with this suppression. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Interplay between Ret and Fap-1 regulates CD95-mediated apoptosis in medullary thyroid cancer cells.

PubMed

Nicolini, Valentina; Cassinelli, Giuliana; Cuccuru, Giuditta; Bongarzone, Italia; Petrangolini, Giovanna; Tortoreto, Monica; Mondellini, Piera; Casalini, Patrizia; Favini, Enrica; Zaffaroni, Nadia; Zunino, Franco; Lanzi, Cinzia

2011-10-01

Emerging evidence suggests that Ret oncoproteins expressed in medullary thyroid cancer (MTC) might evade the pro-apoptotic function of the dependence receptor proto-Ret by directly impacting the apoptosis machinery. Identification of the molecular determinants of the interplay between Ret signaling and apoptosis might provide a relevant contribution to the optimization of Ret-targeted therapies. Here, we describe the cross-talk between Ret-M918T oncogenic mutant responsible for type 2B multiple endocrine syndrome (MEN2B), and components of death receptor-mediated extrinsic apoptosis pathway. In the human MEN2B-type MTC cell line MZ-CRC-1 expressing Ret-M918T, Ret was found associated with Fap-1, known as inhibitor of the CD95 death receptor trafficking to the cell membrane, and with procaspase-8, the initiator pro-form caspase in the extrinsic apoptosis pathway. Cell treatment with the anti-tumor Ret kinase inhibitor RPI-1 inhibited tyrosine phosphorylation of procaspase-8, likely inducing its local activation, followed by downregulation of both Ret and Fap-1, and translocation of CD95 into lipid rafts. According to the resulting increase of CD95 cell surface expression, the CD95 agonist antibody CH11 enhanced RPI-1-induced cell growth inhibition and apoptosis. RET RNA interference downregulated Fap-1 protein in MZ-CRC-1 cells, whereas exogenous RET-M918T upregulated Fap-1 in HEK293 cells. Overall, these data indicate that the Ret oncoprotein exerts opposing controls on Fap-1 and CD95, increasing Fap-1 expression and decreasing CD95 cell surface expression. The functional interplay of the Ret mutant with the extrinsic apoptosis pathway provides a mechanism possibly contributing to MTC malignant phenotype and a rational basis for novel therapeutic strategies combining Ret inhibitors and CD95 agonists. Copyright © 2011 Elsevier Inc. All rights reserved.

Induction of apoptosis in a flounder gill cell line by lymphocystis disease virus infection.

PubMed

Hu, G-B; Cong, R-S; Fan, T-J; Mei, X-G

2004-11-01

Lymphocystis disease virus (LCDV), a large icosahedral DNA virus classified to the iridovirus family, is the causative agent of lymphocystis, a disease which occurs in marine and freshwater fish species and is characterized by formation of papilloma-like lesions on the surface of the skin. In vitro, LCDV infection causes flounder gill cells, an adherent cell line, to exhibit an obvious cytopathic effect (CPE). In order to test whether apoptosis is responsible for the observed CPE, cells infected with LCDV at a multiplicity of infection (m.o.i.) of 5 PFU per cell were examined at various time intervals for the appearance of apoptotic signs. Nuclear fragmentation, DNA laddering and caspase activation were observed in the infected cells at the time (i.e. 10 days post-infection) when an intensive CPE was observed. These findings demonstrate that LCDV is capable of inducing apoptosis in vitro, which is different from the result of LCDV infection in vivo, and consequently suggest an intricate LCDV-host interaction.

Optical coherence tomography spectral analysis for detecting apoptosis in vitro and in vivo

NASA Astrophysics Data System (ADS)

Farhat, Golnaz; Giles, Anoja; Kolios, Michael C.; Czarnota, Gregory J.

2015-12-01

Apoptosis is a form of programmed cell death characterized by a series of predictable morphological changes at the subcellular level, which modify the light-scattering properties of cells. We present a spectroscopic optical coherence tomography (OCT) technique to detect changes in subcellular morphology related to apoptosis in vitro and in vivo. OCT data were acquired from acute myeloid leukemia (AML) cells treated with cisplatin over a 48-h period. The backscatter spectrum of the OCT signal acquired from the cell samples was characterized by calculating its in vitro integrated backscatter (IB) and spectral slope (SS). The IB increased with treatment duration, while the SS decreased, with the most significant changes occurring after 24 to 48 h of treatment. These changes coincided with striking morphological transformations in the cells and their nuclei. Similar trends in the spectral parameter values were observed in vivo in solid tumors grown from AML cells in mice, which were treated with chemotherapy and radiation. Our results provide a strong foundation from which future experiments may be designed to further understand the effect of cellular morphology and kinetics of apoptosis on the OCT signal and demonstrate the feasibility of using this technique in vivo.

Mitoguazone induces apoptosis via a p53-independent mechanism.

PubMed

Davidson, K; Petit, T; Izbicka, E; Koester, S; Von Hoff, D D

1998-08-01

Mitoguazone (methylglyoxal bisguanylhydrazone, methyl-GAG or MGBG) is a synthetic polycarbonyl derivative with activity in patients with Hodgkin's and non-Hodgkin's lymphoma, head and neck cancer, prostate cancer, and esophageal cancer. Mitoguazone has also recently been documented to have activity in patients with AIDS-related lymphoma. Among anticancer drugs, mitoguazone has a unique mechanism of action via interference with the polyamine biosynthetic pathway. Polyamines stabilize DNA structure by non-covalent cross-bridging between phosphate groups on opposite strands. In addition, mitoguazone causes uncoupling of oxidative phosphorylation. In this study, the ability of mitoguazone to induce apoptosis by inhibiting the polyamine pathway was assessed in three Burkitt's lymphoma cell lines (Raji, Ramos and Daudi) and one prostate carcinoma cell line (MPC 3). Additional evaluations were performed in two human breast cancer cell lines (MCF7 with wild-type p53 and VM4K with mutated p53) to determine whether the p53 tumor suppressor gene was required for efficient apoptosis induction. The present study demonstrated that mitoguazone induces apoptosis in all the different human cancer cell lines tested in a concentration- and time-dependent way, and triggers a p53-independent programmed cell death in the human breast cancer MCF7 cell line.

Amplexicaule A exerts anti-tumor effects by inducing apoptosis in human breast cancer

PubMed Central

Shu, Guangwen; Wan, Dingrong; He, Feng; Loaec, Morgann; Ding, Yali; Li, Jun; Dovat, Sinisa; Yang, Gaungzhong; Song, Chunhua

2016-01-01

Chemotherapy is the main treatment for patients with breast cancer metastases, but natural alternatives have been receiving attention for their potential as novel anti-tumor reagents. Amplexicaule A (APA) is a flavonoid glucoside isolated from rhizomes of Polygonum amplexicaule D. Don var. sinense Forb (PADF). We found that APA has anti-tumor effects in a breast cancer xenograft mouse model and induces apoptosis in breast cancer cell lines. APA increased levels of cleaved caspase-3,-8,-9 and PARP, which resulted from suppression of MCL-1 and BCL-2 expression in the cells. APA also inactivated the Akt/mTOR pathway in breast cancer cells. Thus, APA exerts a strong anti-tumor effect on breast cancer cells, most likely through induction of apoptosis. Our study is the first to identify this novel anti-tumor compound and provides a new strategy for isolation and separation of single compounds from herbs. PMID:26943775

ASC Induces Apoptosis via Activation of Caspase-9 by Enhancing Gap Junction-Mediated Intercellular Communication

PubMed Central

Hida, Shigeaki; Fujii, Chifumi; Taniguchi, Shun’ichiro; Ito, Kensuke; Matsumura, Tomio; Okada, Nagisa; Sakaizawa, Takashi; Kobayashi, Akira; Takeoka, Michiko; Miyagawa, Shin-ichi

2017-01-01

ASC (apoptosis-associated speck-like protein containing a CARD) is a key adaptor molecule of inflammasomes that mediates inflammatory and apoptotic signals. Aberrant methylation-induced silencing of ASC has been observed in a variety of cancer cells, thus implicating ASC in tumor suppression, although this role remains incompletely defined especially in the context of closely neighboring cell proliferation. As ASC has been confirmed to be silenced by abnormal methylation in HT1080 fibrosarcoma cells as well, this cell line was investigated to characterize the precise role and mechanism of ASC in tumor progression. The effects of ASC were examined using in vitro cell cultures based on comparisons between low and high cell density conditions as well as in a xenograft murine model. ASC overexpression was established by insertion of the ASC gene into pcDNA3 and pMX-IRES-GFP vectors, the latter being packed into a retrovirus and subjected to reproducible competitive assays using parental cells as an internal control, for evaluation of cell viability. p21 and p53 were silenced using shRNA. Cell viability was suppressed in ASC-expressing transfectants as compared with control cells at high cell density conditions in in vitro culture and colony formation assays and in in vivo ectopic tumor formation trials. This suppression was not detected in low cell density conditions. Furthermore, remarkable progression of apoptosis was observed in ASC-introduced cells at a high cell density, but not at a low one. ASC-dependent apoptosis was mediated not by p21, p53, or caspase-1, but rather by cleavage of caspase-9 as well as by suppression of the NF-κB-related X-linked inhibitor-of-apoptosis protein. Caspase-9 cleavage was observed to be dependent on gap junction formation. The remarkable effect of ASC on the induction of apoptosis through caspase-9 and gap junctions revealed in this study may lead to promising new approaches in anticancer therapy. PMID:28056049

Cyclic mechanical stretch contributes to network development of osteocyte-like cells with morphological change and autophagy promotion but without preferential cell alignment in rat.

PubMed

Inaba, Nao; Kuroshima, Shinichiro; Uto, Yusuke; Sasaki, Muneteru; Sawase, Takashi

2017-09-01

Osteocytes play important roles in controlling bone quality as well as preferential alignment of biological apatite c -axis/collagen fibers. However, the relationship between osteocytes and mechanical stress remains unclear due to the difficulty of three-dimensional (3D) culture of osteocytes in vitro . The aim of this study was to investigate the effect of cyclic mechanical stretch on 3D-cultured osteocyte-like cells. Osteocyte-like cells were established using rat calvarial osteoblasts cultured in a 3D culture system. Cyclic mechanical stretch (8% amplitude at a rate of 2 cycles min -1 ) was applied for 24, 48 and 96 consecutive hours. Morphology, cell number and preferential cell alignment were evaluated. Apoptosis- and autophagy-related gene expression levels were measured using quantitative PCR. 3D-cultured osteoblasts became osteocyte-like cells that expressed osteocyte-specific genes such as Dmp1 , Cx43 , Sost , Fgf23 and RANKL , with morphological changes similar to osteocytes. Cell number was significantly decreased in a time-dependent manner under non-loaded conditions, whereas cyclic mechanical stretch significantly prevented decreased cell numbers with increased expression of anti-apoptosis-related genes. Moreover, cyclic mechanical stretch significantly decreased cell size and ellipticity with increased expression of autophagy-related genes, LC3b and atg7 . Interestingly, preferential cell alignment did not occur, irrespective of mechanical stretch. These findings suggest that an anti-apoptotic effect contributes to network development of osteocyte-like cells under loaded condition. Spherical change of osteocyte-like cells induced by mechanical stretch may be associated with autophagy upregulation. Preferential alignment of osteocytes induced by mechanical load in vivo may be partially predetermined before osteoblasts differentiate into osteocytes and embed into bone matrix.

Methylmercury causes neuronal cell death through the suppression of the TrkA pathway: In vitro and in vivo effects of TrkA pathway activators

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fujimura, Masatake, E-mail: fujimura@nimd.go.jp; Usuki, Fusako

Methylmercury (MeHg) is an environmental toxin which induces cell death specific for the nervous systems. Here we show that MeHg causes neuronal cell death through the suppression of the tropomyosin receptor kinase A (TrkA) pathway, and that compounds activating the TrkA pathway prevent MeHg-induced nerve damage in vitro and in vivo. We first investigated the mechanism of MeHg-induced neurotoxicity in differentiating neurons using PC12 cells. Exposure to 100 nM MeHg for 1 day induced apoptosis in differentiating PC12 cells. Further, MeHg-induced apoptosis was preceded by inhibition of neurite extension, as determined by ELISA analyses of the neurite-specific protein neurofilament tripletmore » H protein (NF-H). To determine the mechanism of MeHg-induced apoptosis, we evaluated the effects of MeHg on the TrkA pathway, which is known to regulate neuronal differentiation and viability. Western blot analysis demonstrated that, like the TrkA phosphorylation inhibitor K252a, MeHg inhibited phosphorylation of TrkA and its downstream effectors. Furthermore, GM1 ganglioside and its analog MCC-257, which enhance TrkA phosphorylation, overcame the effect of MeHg in neurons, supporting the involvement of the TrkA pathway in MeHg-induced nerve damage. Finally, we demonstrated that MCC-257 rescued the clinical sign and pathological changes in MeHg-exposed rats. These findings indicate that MeHg-induced apoptosis in neuron is triggered by inhibition of the TrkA pathway, and that GM1 ganglioside and MCC-257 effectively prevent MeHg-induced nerve damage. - Highlights: • Exposure to 100 nM MeHg for 1 day induced apoptosis in differentiating PC12 cells. • Inhibition of neurite extension was involved in MeHg-induced apoptosis. • Like the TrkA phosphorylation inhibitor, MeHg inhibited phosphorylation of TrkA. • GM1 ganglioside and its analog effectively prevented MeHg-induced nerve damage.« less

An Apoptotic 'Eat Me' Signal: Phosphatidylserine Exposure.

PubMed

Segawa, Katsumori; Nagata, Shigekazu

2015-11-01

Apoptosis and the clearance of apoptotic cells are essential processes in animal development and homeostasis. For apoptotic cells to be cleared, they must display an 'eat me' signal, most likely phosphatidylserine (PtdSer) exposure, which prompts phagocytes to engulf the cells. PtdSer, which is recognized by several different systems, is normally confined to the cytoplasmic leaflet of the plasma membrane by a 'flippase'; apoptosis activates a 'scramblase' that quickly exposes PtdSer on the cell surface. The molecules that flip and scramble phospholipids at the plasma membrane have recently been identified. Here we discuss recent findings regarding the molecular mechanisms of apoptotic PtdSer exposure and the clearance of apoptotic cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

Nrf2/Keap1 system regulates vascular smooth muscle cell apoptosis for vascular homeostasis: role in neointimal formation after vascular injury

PubMed Central

Ashino, Takashi; Yamamoto, Masayuki; Numazawa, Satoshi

2016-01-01

Abnormal increases in vascular smooth muscle cells (VSMCs) in the intimal region after a vascular injury is a key event in developing neointimal hyperplasia. To maintain vascular function, proliferation and apoptosis of VSMCs is tightly controlled during vascular remodeling. NF-E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) system, a key component of the oxidative stress response that acts in maintaining homeostasis, plays an important role in neointimal hyperplasia after a vascular injury; however, the role of Nrf2/Keap1 in VSMC apoptosis has not been clarified. Here we report that 14 days after arterial injury in mice, TUNEL-positive VSMCs are detected in both the neointimal and medial layers. These layers contain cells expressing high levels of Nrf2 but low Keap1 expression. In VSMCs, Keap1 depletion induces features of apoptosis, such as positive TUNEL staining and annexin V binding. These changes are associated with an increased expression of nuclear Nrf2. Simultaneous Nrf2 depletion inhibits Keap1 depletion-induced apoptosis. At 14 days after the vascular injury, Nrf2-deficient mice demonstrated fewer TUNEL-positive cells and increased neointimal formation in the neointimal and medial areas. The results suggest that the Nrf2/Keap1 system regulates VSMC apoptosis during neointimal formation, thereby inhibiting neointimal hyperplasia after a vascular injury. PMID:27198574

Role of TGF-β signaling in curcumin-mediated inhibition of tumorigenicity of human lung cancer cells

PubMed Central

Datta, Raktima; Halder, Sunil K.

2014-01-01

Purpose Curcumin has been shown to have potent anti-cancer activities like inhibition of cell proliferation, induction of apoptosis, and suppression of angiogenesis. Transforming growth factor-β (TGF-β) signaling plays a complex role in tumor suppression and promotion depending on the tumor type and stage. However, the effect of curcumin on TGF-β signaling in cancer cells and the role of TGF-β signaling in curcumin-induced anticancer activities have not been determined. Here, we investigate the role of curcumin on TGF-β signaling, and whether TGF-β signaling is involved in the antitumor activities of curcumin. Methods Human non-small cell lung cancer (NSCLC) cell lines, ACC-LC-176 (without TGF-β signaling), H358, and A549 (with TGF-β signaling) were treated with curcumin to determine cell growth, apoptosis, and tumorigenicity. Antitumor activities of curcumin were determined using these cell lines and an in vivo mouse model. We also tested the effect of curcumin on TGF-β/Smad signaling by western blotting and by luciferase assays. Results Curcumin inhibited cell growth and induced apoptosis of all three NSCLC cell lines in vitro and in vivo. It significantly reduced subcutaneous tumor growth by these three cell lines irrespective of TGF-β signaling status. Curcumin inhibited TGF-β-induced Smad2/3 phosphorylation and transcription in H358 and A549 cells, but not in ACC-LC-176 cells. Conclusions Curcumin reduces tumorigenicity of human lung cancer cells in vitro and in vivo by inhibiting cell proliferation and promoting apoptosis. These results suggest that TGF-β signaling is not directly involved in curcumin-mediated growth inhibition, induction of apoptosis, and inhibition of tumorigenicity. PMID:23224523

Differential roles of SS18-SSX fusion gene and insulin-like growth factor-1 receptor in synovial sarcoma cell growth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Toernkvist, Maria; Natalishvili, Natalia; Xie Yuntao

2008-04-11

Recently we demonstrated that the synovial sarcoma specific fusion gene SS18-SSX is crucial for cyclin D1 expression and is linked to cell proliferation. In this report we explore the role of SS18-SSX and IGF-1R for their potential functions in cellular proliferation and survival in cultured synovial sarcoma cells. We found that targeting of SS18-SSX mRNA by antisense oligonucleotide treatment drastically and rapidly decreased cell proliferation but caused only a slight increase of apoptosis. The synovial sarcoma cells were confirmed to express IGF-1R, and treatment with an IGF-1R inhibitor resulted in substantially reduced cell viability by inducing apoptosis in these cells.more » Conversely, inhibition of the IGF-1R resulted only in a slight to moderate decrease in DNA synthesis. In conclusion, SS18-SSX and IGF-1R seem to play important but different roles in maintaining malignant growth of synovial sarcoma cells. Whereas SS18-SSX maintains cyclin D1 and cell proliferation, IGF-1R protects from apoptosis.« less

Triggering the apoptosis of targeted human renal cancer cells by the vibration of anisotropic magnetic particles attached to the cell membrane.

PubMed

Leulmi, Selma; Chauchet, Xavier; Morcrette, Melissa; Ortiz, Guillermo; Joisten, Hélène; Sabon, Philippe; Livache, Thierry; Hou, Yanxia; Carrière, Marie; Lequien, Stéphane; Dieny, Bernard

2015-10-14

Cancer cells develop resistance to chemotherapy, and the side effects encountered seriously limit the effectiveness of treatments. For these reasons, the search for alternative therapies that target cancer cells without affecting healthy tissues is currently one of the most active areas of research on cancer. The present study focuses on a recently proposed approach for cancer cell destruction based on the targeted triggering of cancer cell spontaneous death through the mechanical vibration of anisotropic magnetic micro/nanoparticles attached to the cell membranes at low frequencies (∼20 Hz) and in weak magnetic fields (∼30 mT). The study was conducted in vitro, on human renal cancer cells with superparamagnetic-like particles. Three types of such particles made of NiFe or magnetite were prepared and characterized (either synthetic antiferromagnetic, vortex or polycrystalline with random grain anisotropy). The triggering of the apoptosis of these cancer cells was demonstrated with NiFe vortex particles and statistically characterized by flow-cytometry studies. The death pathway via apoptosis and not necrosis was identified by the clear observation of caspase activation.

Triggering the apoptosis of targeted human renal cancer cells by the vibration of anisotropic magnetic particles attached to the cell membrane

NASA Astrophysics Data System (ADS)

Leulmi, Selma; Chauchet, Xavier; Morcrette, Melissa; Ortiz, Guillermo; Joisten, Hélène; Sabon, Philippe; Livache, Thierry; Hou, Yanxia; Carrière, Marie; Lequien, Stéphane; Dieny, Bernard

2015-09-01

Cancer cells develop resistance to chemotherapy, and the side effects encountered seriously limit the effectiveness of treatments. For these reasons, the search for alternative therapies that target cancer cells without affecting healthy tissues is currently one of the most active areas of research on cancer. The present study focuses on a recently proposed approach for cancer cell destruction based on the targeted triggering of cancer cell spontaneous death through the mechanical vibration of anisotropic magnetic micro/nanoparticles attached to the cell membranes at low frequencies (~20 Hz) and in weak magnetic fields (~30 mT). The study was conducted in vitro, on human renal cancer cells with superparamagnetic-like particles. Three types of such particles made of NiFe or magnetite were prepared and characterized (either synthetic antiferromagnetic, vortex or polycrystalline with random grain anisotropy). The triggering of the apoptosis of these cancer cells was demonstrated with NiFe vortex particles and statistically characterized by flow-cytometry studies. The death pathway via apoptosis and not necrosis was identified by the clear observation of caspase activation.

Compound 49b Prevents Diabetes-Induced Apoptosis through Increased IGFBP-3 Levels

PubMed Central

Zhang, Qiuhua; Guy, Kimberly; Pagadala, Jayaprakash; Jiang, Youde; Walker, Robert J; Liu, Luhong; Soderland, Carl; Kern, Timothy S; Ferry, Robert; He, Hui; Yates, C. Ryan; Miller, Duane D; Steinle, Jena J

2012-01-01

Purpose. To determine whether Compound 49b, a novel PKA-activating drug, can prevent diabetic-like changes in the rat retina through increased insulin-like growth factor binding protein-3 (IGFBP-3) levels. Methods. For the cell culture studies, we used both human retinal endothelial cells (REC) and retinal Müller cells in either 5 mM (normal) or 25 mM (high) glucose. Cells were treated with 50 nM Compound 49b alone of following treatment with protein kinase A (PKA) siRNA or IGFBP-3 siRNA. Western blotting and ELISA analyses were done to verify PKA and IGFBP-3 knockdown, as well as to measure apoptotic markers. For animal studies, we used streptozotocin-treated rats after 2 and 8 months of diabetes. Some rats were treated topically with 1 mM Compound 49b. Analyses were done for retinal thickness, cell numbers in the ganglion cell layer, pericyte ghosts, and numbers of degenerate capillaries, as well as electroretinogram and heart morphology. Results. Compound 49b requires active PKA and IGFBP-3 to prevent apoptosis of REC. Compound 49b significantly reduced the numbers of degenerate capillaries and pericyte ghosts, while preventing the decreased retinal thickness and loss of cells in the ganglion cell layer. Compound 49b maintained a normal electroretinogram, with no changes in blood pressure, intraocular pressure, or heart morphological changes. Conclusions. Topical Compound 49b is able to prevent diabetic-like changes in the rat retina, without producing systemic changes. Compound 49b is able to prevent REC apoptosis through increasing IGFBP-3 levels, which are reduced in response to hyperglycemia. PMID:22467575

The modulation of apoptosis by oncogenic viruses

PubMed Central

2013-01-01

Transforming viruses can change a normal cell into a cancer cell during their normal life cycle. Persistent infections with these viruses have been recognized to cause some types of cancer. These viruses have been implicated in the modulation of various biological processes, such as proliferation, differentiation and apoptosis. The study of infections caused by oncogenic viruses had helped in our understanding of several mechanisms that regulate cell growth, as well as the molecular alterations leading to cancer. Therefore, transforming viruses provide models of study that have enabled the advances in cancer research. Viruses with transforming abilities, include different members of the Human Papillomavirus (HPV) family, Hepatitis C virus (HCV), Human T-cell Leukemia virus (HTLV-1), Epstein Barr virus (EBV) and Kaposi’s Sarcoma Herpesvirus (KSHV). Apoptosis, or programmed cell death, is a tightly regulated process that plays an important role in development and homeostasis. Additionally, it functions as an antiviral defense mechanism. The deregulation of apoptosis has been implicated in the etiology of diverse diseases, including cancer. Oncogenic viruses employ different mechanisms to inhibit the apoptotic process, allowing the propagation of infected and damaged cells. During this process, some viral proteins are able to evade the immune system, while others can directly interact with the caspases involved in apoptotic signaling. In some instances, viral proteins can also promote apoptosis, which may be necessary for an accurate regulation of the initial stages of infection. PMID:23741982

Transfected Poly(I:C) Activates Different dsRNA Receptors, Leading to Apoptosis or Immunoadjuvant Response in Androgen-independent Prostate Cancer Cells*

PubMed Central

Palchetti, Sara; Starace, Donatella; De Cesaris, Paola; Filippini, Antonio; Ziparo, Elio; Riccioli, Anna

2015-01-01

Despite the effectiveness of surgery or radiation therapy for the treatment of early-stage prostate cancer (PCa), there is currently no effective strategy for late-stage disease. New therapeutic targets are emerging; in particular, dsRNA receptors Toll-like receptor 3 (TLR3) and cytosolic helicases expressed by cancer cells, once activated, exert a pro-apoptotic effect in different tumors. We previously demonstrated that the synthetic analog of dsRNA poly(I:C) induces apoptosis in the androgen-dependent PCa cell line LNCaP in a TLR3-dependent fashion, whereas only a weak apoptotic effect is observed in the more aggressive and androgen-independent PCa cells PC3 and DU145. In this paper, we characterize the receptors and the signaling pathways involved in the remarkable apoptosis induced by poly(I:C) transfected by Lipofectamine (in-poly(I:C)) compared with the 12-fold higher free poly(I:C) concentration in PC3 and DU145 cells. By using genetic inhibition of different poly(I:C) receptors, we demonstrate the crucial role of TLR3 and Src in in-poly(I:C)-induced apoptosis. Therefore, we show that the increased in-poly(I:C) apoptotic efficacy is due to a higher binding of endosomal TLR3. On the other hand, we show that in-poly(I:C) binding to cytosolic receptors MDA5 and RIG-I triggers IRF3-mediated signaling, leading uniquely to the up-regulation of IFN-β, which likely in turn induces increased TLR3, MDA5, and RIG-I proteins. In summary, in-poly(I:C) activates two distinct antitumor pathways in PC3 and DU145 cells: one mediated by the TLR3/Src/STAT1 axis, leading to apoptosis, and the other one mediated by MDA5/RIG-I/IRF3, leading to immunoadjuvant IFN-β expression. PMID:25568326

Semaphorin 3A is a retrograde cell death signal in developing sympathetic neurons

PubMed Central

Wehner, Amanda B.; Abdesselem, Houari; Dickendesher, Travis L.; Imai, Fumiyasu; Yoshida, Yutaka; Giger, Roman J.; Pierchala, Brian A.

2016-01-01

ABSTRACT During development of the peripheral nervous system, excess neurons are generated, most of which will be lost by programmed cell death due to a limited supply of neurotrophic factors from their targets. Other environmental factors, such as ‘competition factors' produced by neurons themselves, and axon guidance molecules have also been implicated in developmental cell death. Semaphorin 3A (Sema3A), in addition to its function as a chemorepulsive guidance cue, can also induce death of sensory neurons in vitro. The extent to which Sema3A regulates developmental cell death in vivo, however, is debated. We show that in compartmentalized cultures of rat sympathetic neurons, a Sema3A-initiated apoptosis signal is retrogradely transported from axon terminals to cell bodies to induce cell death. Sema3A-mediated apoptosis utilizes the extrinsic pathway and requires both neuropilin 1 and plexin A3. Sema3A is not retrogradely transported in older, survival factor-independent sympathetic neurons, and is much less effective at inducing apoptosis in these neurons. Importantly, deletion of either neuropilin 1 or plexin A3 significantly reduces developmental cell death in the superior cervical ganglia. Taken together, a Sema3A-initiated apoptotic signaling complex regulates the apoptosis of sympathetic neurons during the period of naturally occurring cell death. PMID:27143756

Direct inhibition of interleukin-2 receptor alpha-mediated signaling pathway induces G1 arrest and apoptosis in human head-and-neck cancer cells.

PubMed

Kuhn, Deborah J; Dou, Q Ping

2005-05-15

Overexpression of the interleukin-2 receptor (IL-2R) alpha chain in tumor cells is associated with tumor progression and a poor patient prognosis. IL-2Ralpha is responsible for the high affinity binding of the receptor to IL-2, leading to activation of several proliferative and anti-apoptotic intracellular signaling pathways. We have previously shown that human squamous cell carcinoma of a head-and-neck line (PCI-13) genetically engineered to overexpress IL-2Ralpha exhibit increased transforming activity, proliferation, and drug resistance, compared to the vector control cells (J Cell Biochem 2003;89:824-836). In this study, we report that IL-2Ralpha(+) cells express high levels of total and phosphorylated Jak3 protein and are more resistant to apoptosis induced by a Jak3 inhibitor than the control LacZ cells. Furthermore, we used daclizumab, a monoclonal antibody specific to IL-2Ralpha, and determined the effects of IL-2Ralpha inhibition on cell cycle and apoptosis as well as the involvement of potential cell cycle and apoptosis regulatory proteins. We found that daclizumab induces G(1) arrest, associated with down-regulation of cyclin A protein, preferentially in IL-2Ralpha(+) cells, but not in LacZ cells. In addition, daclizumab activates apoptotic death program via Bcl-2 down-regulation preferentially in IL-2Ralpha(+) cells. Finally, daclizumab also sensitizes IL-2Ralpha(+) cells to other apoptotic stimuli, although the effect is moderate. These results indicate that daclizumab inhibits the proliferative potential of IL-2Ralpha(+) cells via inhibition of cell cycle progression and induction of apoptosis.

Apoptosis and tumor cell death in response to HAMLET (human alpha-lactalbumin made lethal to tumor cells).

PubMed

Hallgren, Oskar; Aits, Sonja; Brest, Patrick; Gustafsson, Lotta; Mossberg, Ann-Kristin; Wullt, Björn; Svanborg, Catharina

2008-01-01

HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a molecular complex derived from human milk that kills tumor cells by a process resembling programmed cell death. The complex consists of partially unfolded alpha-lactalbumin and oleic acid, and both the protein and the fatty acid are required for cell death. HAMLET has broad antitumor activity in vitro, and its therapeutic effect has been confirmed in vivo in a human glioblastoma rat xenograft model, in patients with skin papillomas and in patients with bladder cancer. The mechanisms of tumor cell death remain unclear, however. Immediately after the encounter with tumor cells, HAMLET invades the cells and causes mitochondrial membrane depolarization, cytochrome c release, phosphatidyl serine exposure, and a low caspase response. A fraction of the cells undergoes morphological changes characteristic of apoptosis, but caspase inhibition does not rescue the cells and Bcl-2 overexpression or altered p53 status does not influence the sensitivity of tumor cells to HAMLET. HAMLET also creates a state of unfolded protein overload and activates 20S proteasomes, which contributes to cell death. In parallel, HAMLET translocates to tumor cell nuclei, where high-affinity interactions with histones cause chromatin disruption, loss of transcription, and nuclear condensation. The dying cells also show morphological changes compatible with macroautophagy, and recent studies indicate that macroautophagy is involved in the cell death response to HAMLET. The results suggest that HAMLET, like a hydra with many heads, may interact with several crucial cellular organelles, thereby activating several forms of cell death, in parallel. This complexity might underlie the rapid death response of tumor cells and the broad antitumor activity of HAMLET.

Tributyltin induces Yca1p-dependent cell death of yeast Saccharomyces cerevisiae.

PubMed

Chahomchuen, Thippayarat; Akiyama, Koichi; Sekito, Takayuki; Sugimoto, Naoko; Okabe, Masaaki; Nishimoto, Sogo; Sugahara, Takuya; Kakinuma, Yoshimi

2009-10-01

Tributyltin chloride (TBT), an environmental pollutant, is toxic to a variety of eukaryotic and prokaryotic organisms. Although it has been reported that TBT induces apoptotic cell death in mammalian, the action of TBT on eukaryotic microorganisms has not yet been fully investigated. In this study we examined the mechanism involved in cell death caused by TBT exposure in Saccharomyces cerevisiae. The median lethal concentration of TBT was 10 microM for the parent strain BY4741 and 3 microM for the pdr5Delta mutant defective in a major multidrug transporter, respectively. Fluorescence microscopic observations revealed nuclear condensation and chromatin fragmentation in cells treated with TBT indicating that cells underwent an apoptosis-like cell dearth. TBT-induced cell death was suppressed by deletion of the yca1 gene encoding a homologue of the mammalian caspase. In parallel, reactive oxygen species (ROS) were produced by TBT. These results suggest that TBT induces apoptosis-like cell death in yeast via an Yca1p-dependent pathway possibly downstream of the ROS production. This is the first report on TBT-induced apoptotic cell death in yeast.

Targeting the programmed cell death 1: programmed cell death ligand 1 pathway reverses T cell exhaustion in patients with sepsis

PubMed Central

2014-01-01

Introduction A major pathophysiologic mechanism in sepsis is impaired host immunity which results in failure to eradicate invading pathogens and increased susceptibility to secondary infections. Although many immunosuppressive mechanisms exist, increased expression of the inhibitory receptor programmed cell death 1 (PD-1) and its ligand (PD-L1) are thought to play key roles. The newly recognized phenomenon of T cell exhaustion is mediated in part by PD-1 effects on T cells. This study tested the ability of anti-PD-1 and anti-PD-L1 antibodies to prevent apoptosis and improve lymphocyte function in septic patients. Methods Blood was obtained from 43 septic and 15 non-septic critically-ill patients. Effects of anti-PD-1, anti-PD-L1, or isotype-control antibody on lymphocyte apoptosis and interferon gamma (IFN-γ) and interleukin-2 (IL-2) production were quantitated by flow cytometry. Results Lymphocytes from septic patients produced decreased IFN-γ and IL-2 and had increased CD8 T cell expression of PD-1 and decreased PD-L1 expression compared to non-septic patients (P<0.05). Monocytes from septic patients had increased PD-L1 and decreased HLA-DR expression compared to non-septic patients (P<0.01). CD8 T cell expression of PD-1 increased over time in ICU as PD-L1, IFN-γ, and IL2 decreased. In addition, donors with the highest CD8 PD-1 expression together with the lowest CD8 PD-L1 expression also had lower levels of HLA-DR expression in monocytes, and an increased rate of secondary infections, suggestive of a more immune exhausted phenotype. Treatment of cells from septic patients with anti-PD-1 or anti-PD-L1 antibody decreased apoptosis and increased IFN-γ and IL-2 production in septic patients; (P<0.01). The percentage of CD4 T cells that were PD-1 positive correlated with the degree of cellular apoptosis (P<0.01). Conclusions In vitro blockade of the PD-1:PD-L1 pathway decreases apoptosis and improves immune cell function in septic patients. The current results together with multiple positive studies of anti-PD-1 and anti-PD-L1 in animal models of bacterial and fungal infections and the relative safety profile of anti-PD-1/anti-PD-L1 in human oncology trials to date strongly support the initiation of clinical trials testing these antibodies in sepsis, a disorder with a high mortality. PMID:24387680

Cholecystokinin-8-induced hypoplasia of the rat pancreas: influence of nitric oxide on cell proliferation and programmed cell death.

PubMed

Trulsson, Lena M; Gasslander, Thomas; Svanvik, Joar

2004-10-01

The background of cholecystokinin-8 (CCK-8)-induced hypoplasia in the pancreas is not known. In order to increase our understanding we studied the roles of nitric oxide and NF-kappaB in rats. CCK-8 was injected for 4 days, in a mode known to cause hypoplasia, and the nitric oxide formation was either decreased by means of N(omega)-nitro-L-arginine (L-NNA) or increased by S-nitroso-N-acetylpencillamine (SNAP). The activation of NF-kappaB was quantified by ELISA detection, apoptosis with caspase-3 and histone-associated DNA-fragmentation and mitotic activity in the acinar, centroacinar and ductal cells were visualized by the incorporation of [(3)H]-thymidine. Pancreatic histology and weight as well as protein- and DNA contents were also studied. Intermittent CCK injections reduced pancreatic weight, protein and DNA contents and increased apoptosis, acinar cell proliferation and nuclear factor kappaB (NF-kappaB) activation. It also caused vacuolisation of acinar cells. The inhibition of endogenous nitric oxide formation by L-NNA further increased apoptosis and NF-kappaB activation but blocked the increased proliferation and vacuolisation of acinar cells. The DNA content was not further reduced. SNAP given together with CCK-8 increased apoptosis and other pathways of cell death, raised proliferation of acinar cells and strongly reduced the DNA content in the pancreas. Histological examination showed no inflammation in any group. We conclude that during CCK-8-induced pancreatic hypoplasia, endogenously formed nitric oxide suppresses apoptosis but increases cell death along non-apoptotic pathways and stimulates regeneration of acinar cells. Exogenous nitric oxide enhances the acinar cell turnover by increasing both apoptotic and non-apoptotic cell death and cell renewal. In this situation NF-kappaB activation seems not to inhibit apoptosis nor promote cell proliferation.

Navigation to the graveyard-induction of various pathways of necrosis and their classification by flow cytometry.

PubMed

Janko, Christina; Munoz, Luis; Chaurio, Ricardo; Maueröder, Christian; Berens, Christian; Lauber, Kirsten; Herrmann, Martin

2013-01-01

Apoptosis and necrosis reflect the program of cell death employed by a dying cell and the final stage of death, respectively. Whereas apoptosis is defined as a physiological, highly organized cell death process, necrosis is commonly considered to be accidental and uncontrolled. Physiological and weak pathological death stimuli preferentially induce apoptosis, while harsh non-physiological insults often immediately instigate (primary) necrosis. If an apoptosing cell transits into a phase of plasma membrane disintegration, this stage of death is referred to as secondary or post-apoptotic necrosis.Here, we present several conditions that stimulate primary and/or secondary necrosis and show that necrosis displays considerably different time courses. For subclassification of necrotic phenotypes we employed a flow cytometric single-tube 4-color staining technique including annexin A5-FITC, propidium iodide, DiIC1(5), and Hoechst 33342.

Up-regulation of p53 and mitochondrial signaling pathway in apoptosis by a combination of COX-2 inhibitor, Celecoxib and Dolastatin 15, a marine mollusk linear peptide in experimental colon carcinogenesis.

PubMed

Piplani, Honit; Vaish, Vivek; Rana, Chandan; Sanyal, Sankar N

2013-11-01

Programmed cell death, also known as apoptosis, is an active process occurring in eukaryotic cells and it depends on various sets of pro and anti-apoptotic proteins. Chemoprevention of colorectal cancer can be achieved by inducing apoptosis using synthetic compound, Celecoxib and natural peptide, Dolastatin 15 in an effective manner. But the apoptotic signaling by these two drugs remain unclear. The present study was thus focused on the role of Bcl2 family of proteins and their interplay with p53 in rats during the chemoprevention by these two drugs. After treatment for 6 wk with 1, 2-dimethylhydrazine (DMH), animals showed a marked occurrence of multiple plaque lesions. However, a simultaneous treatment with Celecoxib and Dolastatin 15 decreases such number to a significant level. DMH treatment also decreases the number of apoptotic cells in the colonic enterocytes which were corrected to the normal level by Celecoxib and Dolastatin 15. An increased expression of Bcl2 while other proteins like Bax, Apaf-1, cyt c, and caspases in the apoptotic pathway, and the tumor suppressor proteins, p53 and p21 get down-regulated after DMH treatment which were reverted back to normal with Celecoxib and Dolastatin 15. Also, cells having high mitochondrial membrane potential had been seen to increase to significant levels which were reduced after the administration of these anti-inflammatory drugs. In silico molecular docking studies also showed that Dolastatin 15 and Celecoxib may bind to the active site pocket of Bcl2 , thus revealing the direct target of Dolastatin 15 and Celecoxib apart from binding to COX-2. © 2012 Wiley Periodicals, Inc.

Oxidative stress: a key regulator of leiomyoma cell survival.

PubMed

Fletcher, Nicole M; Abusamaan, Mohammed S; Memaj, Ira; Saed, Mohammed G; Al-Hendy, Ayman; Diamond, Michael P; Saed, Ghassan M

2017-06-01

To determine the effects of attenuating oxidative stress with the use of dichloroacetate (DCA) on the expression of key redox enzymes myeloperoxidase (MPO) and inducible nitric oxide synthase (iNOS) as well as on apoptosis. Prospective experimental study. University medical center. Cells established from myometrium and uterine fibroid from the same patients. Cells were exposed to normal (20% O 2 ) or hypoxic (2% O 2 ) conditions for 24 hours with or without DCA (20 μg/mL), a metabolic modulator that shifts anaerobic to aerobic metabolism. Nitrate/nitrite (iNOS activity indicator), iNOS, Bcl-2/Bax ratio, MPO, and caspase-3 activities and levels were determined by means of Greiss assay, real-time reverse-transcription polymerase chain reaction, and ELISA. Data were analyzed with the use of SPSS by means of one-way analysis of variance with Tukey post hoc analysis and independent t tests. MPO, iNOS, and nitrate/nitrite expression were higher in leiomyoma than in myometrial cells, and they were further enhanced by hypoxia in myometrial cells. Treatment with the use of DCA decreased MPO, iNOS, and nitrate/nitrite levels and negated the effect of hypoxia in both types of cells. Leiomyoma cells showed less apoptosis, as indicated by both caspase-3 activity and the Bcl-2/Bax ratio, than myometrial cells. Hypoxia further decreased apoptosis in myometrial cells with no further effect on leiomyoma cells. Treatment with DCA resulted in increased apoptosis in both types of cells, even in the presence of hypoxia. Shifting anaerobic to aerobic metabolism with the use of DCA resulted in an increase in apoptosis in leiomyoma cells and protected myometrial cells from the acquisition of the leiomyoma-like phenotype. Copyright © 2017 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Pulsed Electromagnetic Field Exposure Reduces Hypoxia and Inflammation Damage in Neuron-Like and Microglial Cells.

PubMed

Vincenzi, Fabrizio; Ravani, Annalisa; Pasquini, Silvia; Merighi, Stefania; Gessi, Stefania; Setti, Stefania; Cadossi, Ruggero; Borea, Pier Andrea; Varani, Katia

2017-05-01

In the present study, the effect of low-frequency, low-energy pulsed electromagnetic fields (PEMFs) has been investigated by using different cell lines derived from neuron-like cells and microglial cells. In particular, the primary aim was to evaluate the effect of PEMF exposure in inflammation- and hypoxia-induced injury in two different neuronal cell models, the human neuroblastoma-derived SH-SY5Y cells and rat pheochromocytoma PC12 cells and in N9 microglial cells. In neuron-like cells, live/dead and apoptosis assays were performed in hypoxia conditions from 2 to 48 h. Interestingly, PEMF exposure counteracted hypoxia damage significantly reducing cell death and apoptosis. In the same cell lines, PEMFs inhibited the activation of the hypoxia-inducible factor 1α (HIF-1α), the master transcriptional regulator of cellular response to hypoxia. The effect of PEMF exposure on reactive oxygen species (ROS) production in both neuron-like and microglial cells was investigated considering their key role in ischemic injury. PEMFs significantly decreased hypoxia-induced ROS generation in PC12, SH-SY5Y, and N9 cells after 24 or 48 h of incubation. Moreover, PEMFs were able to reduce some of the most well-known pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, and IL-8 release in N9 microglial cells stimulated with different concentrations of LPS for 24 or 48 h of incubation time. These results show a protective effect of PEMFs on hypoxia damage in neuron-like cells and an anti-inflammatory effect in microglial cells suggesting that PEMFs could represent a potential therapeutic approach in cerebral ischemic conditions. J. Cell. Physiol. 232: 1200-1208, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Cathepsin B Contributes to Autophagy-related 7 (Atg7)-induced Nod-like Receptor 3 (NLRP3)-dependent Proinflammatory Response and Aggravates Lipotoxicity in Rat Insulinoma Cell Line

PubMed Central

Li, Shali; Du, Leilei; Zhang, Lu; Hu, Yue; Xia, Wenchun; Wu, Jia; Zhu, Jing; Chen, Lingling; Zhu, Fengqi; Li, Chunxian; Yang, SiJun

2013-01-01

Impairment of glucose-stimulated insulin secretion caused by the lipotoxicity of palmitate was found in β-cells. Recent studies have indicated that defects in autophagy contribute to pathogenesis in type 2 diabetes. Here, we report that autophagy-related 7 (Atg7) induced excessive autophagic activation in INS-1(823/13) cells exposed to saturated fatty acids. Atg7-induced cathepsin B (CTSB) overexpression resulted in an unexpected significant increase in proinflammatory chemokine and cytokine production levels of IL-1β, monocyte chemotactic protein-1, IL-6, and TNF-α. Inhibition of receptor-interacting protein did not affect the inflammatory response, ruling out involvement of necrosis. CTSB siRNA suppressed the inflammatory response but did not affect apoptosis significantly, suggesting that CTSB was a molecular linker between autophagy and the proinflammatory response. Blocking caspase-3 suppressed apoptosis but did not affect the inflammatory response, suggesting that CTSB induced inflammatory effects independently of apoptosis. Silencing of Nod-like receptor 3 (NLRP3) completely abolished both IL-1β secretion and the down-regulation effects of Atg7-induced CTSB overexpression on glucose-stimulated insulin secretion impairment, thus identifying the NLRP3 inflammasome as an autophagy-responsive element in the pancreatic INS-1(823/13) cell line. Combined together, our results indicate that CTSB contributed to the Atg7-induced NLRP3-dependent proinflammatory response, resulting in aggravation of lipotoxicity, independently of apoptosis in the pancreatic INS-1(823/13) cell line. PMID:23986436

Akt-dependent glucose metabolism promotes Mcl-1 synthesis to maintain cell survival and resistance to Bcl-2 inhibition.

PubMed

Coloff, Jonathan L; Macintyre, Andrew N; Nichols, Amanda G; Liu, Tingyu; Gallo, Catherine A; Plas, David R; Rathmell, Jeffrey C

2011-08-01

Most cancer cells utilize aerobic glycolysis, and activation of the phosphoinositide 3-kinase/Akt/mTOR pathway can promote this metabolic program to render cells glucose dependent. Although manipulation of glucose metabolism may provide a means to specifically eliminate cancer cells, mechanistic links between cell metabolism and apoptosis remain poorly understood. Here, we examined the role and metabolic regulation of the antiapoptotic Bcl-2 family protein Mcl-1 in cell death upon inhibition of Akt-induced aerobic glycolysis. In the presence of adequate glucose, activated Akt prevented the loss of Mcl-1 expression and protected cells from growth factor deprivation-induced apoptosis. Mcl-1 associated with and inhibited the proapoptotic Bcl-2 family protein Bim, contributing to cell survival. However, suppression of glucose metabolism led to induction of Bim, decreased expression of Mcl-1, and apoptosis. The proapoptotic Bcl-2/Bcl-xL/Bcl-w inhibitor, ABT-737, shows clinical promise, but Mcl-1 upregulation can promote resistance. Importantly, inhibition of glucose metabolism or mTORC1 overcame Mcl-1-mediated resistance in diffuse large B cell leukemic cells. Together these data show that Mcl-1 protein synthesis is tightly controlled by metabolism and that manipulation of glucose metabolism may provide a mechanism to suppress Mcl-1 expression and sensitize cancer cells to apoptosis.