Showing papers by "Guido Kroemer published in 2005"

Classification of cell death: recommendations of the Nomenclature Committee on Cell Death

Abstract: Different types of cell death are often defined by morphological criteria, without a clear reference to precise biochemical mechanisms. The Nomenclature Committee on Cell Death (NCCD) proposes unified criteria for the definition of cell death and of its different morphologies, while formulating several caveats against the misuse of words and concepts that slow down progress in the area of cell death research. Authors, reviewers and editors of scientific periodicals are invited to abandon expressions like 'percentage apoptosis' and to replace them with more accurate descriptions of the biochemical and cellular parameters that are actually measured. Moreover, at the present stage, it should be accepted that caspase-independent mechanisms can cooperate with (or substitute for) caspases in the execution of lethal signaling pathways and that 'autophagic cell death' is a type of cell death occurring together with (but not necessarily by) autophagic vacuolization. This study details the 2009 recommendations of the NCCD on the use of cell death-related terminology including 'entosis', 'mitotic catastrophe', 'necrosis', 'necroptosis' and 'pyroptosis'.

Inhibition of macroautophagy triggers apoptosis.

Abstract: Mammalian cells were observed to die under conditions in which nutrients were depleted and, simultaneously, macroautophagy was inhibited either genetically (by a small interfering RNA targeting Atg5, Atg6/Beclin 1-1, Atg10, or Atg12) or pharmacologically (by 3-methyladenine, hydroxychloroquine, bafilomycin A1, or monensin). Cell death occurred through apoptosis (type 1 cell death), since it was reduced by stabilization of mitochondrial membranes (with Bcl-2 or vMIA, a cytomegalovirus-derived gene) or by caspase inhibition. Under conditions in which the fusion between lysosomes and autophagosomes was inhibited, the formation of autophagic vacuoles was enhanced at a preapoptotic stage, as indicated by accumulation of LC3-II protein, ultrastructural studies, and an increase in the acidic vacuolar compartment. Cells exhibiting a morphology reminiscent of (autophagic) type 2 cell death, however, recovered, and only cells with a disrupted mitochondrial transmembrane potential were beyond the point of no return and inexorably died even under optimal culture conditions. All together, these data indicate that autophagy may be cytoprotective, at least under conditions of nutrient depletion, and point to an important cross talk between type 1 and type 2 cell death pathways.

Caspase-dependent immunogenicity of doxorubicin-induced tumor cell death.

Abstract: Systemic anticancer chemotherapy is immunosuppressive and mostly induces nonimmunogenic tumor cell death. Here, we show that even in the absence of any adjuvant, tumor cells dying in response to anthracyclins can elicit an effective antitumor immune response that suppresses the growth of inoculated tumors or leads to the regression of established neoplasia. Although both antracyclins and mitomycin C induced apoptosis with caspase activation, only anthracyclin-induced immunogenic cell death was immunogenic. Caspase inhibition by Z-VAD-fmk or transfection with the baculovirus inhibitor p35 did not inhibit doxorubicin (DX)-induced cell death, yet suppressed the immunogenicity of dying tumor cells in several rodent models of neoplasia. Depletion of dendritic cells (DCs) or CD8 + T cells abolished the immune response against DX-treated apoptotic tumor cells in vivo. Caspase inhibition suppressed the capacity of DX-killed cells to be phagocytosed by DCs, yet had no effect on their capacity to elicit DC maturation. Freshly excised tumors became immunogenic upon DX treatment in vitro, and intratumoral inoculation of DX could trigger the regression of established tumors in immunocompetent mice. These results delineate a procedure for the generation of cancer vaccines and the stimulation of anti-neoplastic immune responses in vivo.

Lysosomes and autophagy in cell death control.

Abstract: Lysosomal hydrolases participate in the digestion of endocytosed and autophagocytosed material inside the lysosomal/autolysosomal compartment in acute cell death when released into the cytosol and in cancer progression following their release into the extracellular space. Lysosomal alterations are common in cancer cells. The increased expression and altered trafficking of lysosomal enzymes participates in tissue invasion, angiogenesis and sensitization to the lysosomal death pathway. But lysosomal heat-shock protein 70 locally prevents lysosomal-membrane permeabilization. Similarly, alterations in the autophagic compartment are linked to carcinogenesis and resistance to chemotherapy. Targeting these pathways might constitute a novel approach to cancer therapy.

Caspase-independent cell death.

Abstract: Caspase activation has been frequently viewed as synonymous with apoptotic cell death; however, caspases can also contribute to processes that do not culminate in cell demise. Moreover, inhibition of caspases can have cytoprotective effects. In a number of different models, caspase inhibition does not maintain cellular viability and instead shifts the morphology of death from apoptosis to nonapoptotic pathways. Here, we explore the contribution of caspases to cell death, either as upstream signals or as downstream effectors contributing to apoptotic morphology, as well as alternative strategies for cell death inhibition. Such alternative strategies may either target catabolic hydrolases or be aimed at preventing mitochondrial membrane permeabilization and its upstream triggers.

Tumor cells convert immature myeloid dendritic cells into TGF-β–secreting cells inducing CD4+CD25+ regulatory T cell proliferation

Abstract: The mechanisms through which regulatory T cells accumulate in lymphoid organs of tumor-bearing hosts remain elusive. Our experiments indicate that the accumulation of CD4+CD25+ regulatory T cells (T reg cells) expressing FoxP3 and exhibiting immunosuppressive function originates from the proliferation of naturally occurring CD25+ T cells and requires signaling through transforming growth factor (TGF)-beta receptor II. During tumor progression, a subset of dendritic cells (DCs) exhibiting a myeloid immature phenotype is recruited to draining lymph nodes. This DC subset selectively promotes the proliferation of T reg cells in a TGF-beta-dependent manner in mice and rats. Tumor cells are necessary and sufficient to convert DCs into regulatory cells that secrete bioactive TGF-beta and stimulate T reg cell proliferation. In conclusion, tumor expansion can stimulate T reg cells via a specific DC subset.

The apoptosis/autophagy paradox: autophagic vacuolization before apoptotic death

Abstract: Autophagic cell death is morphologically characterized by an accumulation of autophagic vacuoles. Here, we show that inactivation of LAMP2 by RNA interference or by homologous recombination leads to autophagic vacuolization in nutrient-depleted cells. Cells that lack LAMP2 expression showed an enhanced accumulation of vacuoles carrying the marker LC3, yet a decreased colocalization of LC3 and lysosomes, suggesting that the fusion between autophagic vacuoles and lysosomes was inhibited. While a fraction of mitochondria from starved LAMP2-expressing cells colocalized with lysosomal markers, within autophagolysosomes, no such colocalization was found on removal of LAMP2 from the experimental system. Of note, LAMP1 depletion had no such effects and did not aggravate the phenotype induced by LAMP2-specific small interfering RNA. Serum and amino acid-starved LAMP2-negative cells exhibited an accumulation of autophagic vacuoles and then succumbed to cell death with hallmarks of apoptosis such as loss of the mitochondrial transmembrane potential, caspase activation and chromatin condensation. While caspase inhibition retarded cell death, it had no protective effect on mitochondria. Stabilization of mitochondria by overexpression of Bcl-2 or the mitochondrion-targeted cytomegalovirus protein vMIA, however, blocked all signs of apoptosis. Neither caspase inhibition nor mitochondrial stabilization antagonized autophagic vacuolization in LAMP2-deficient cells. Altogether, these data indicate that accumulation of autophagic vacuoles can precede apoptotic cell death. These findings argue against the clear-cut distinction between type 1 (apoptotic) and type 2 (autophagic) cell death.

Does autophagy contribute to cell death

Abstract: Autophagy (specifically macroautophagy) is an evolutionarily conserved catabolic process where the cytoplasmic contents of a cell are sequestered within double membrane vacuoles, called autophagosomes, and subsequently delivered to the lysosome for degradation. Autophagy can function as a survival mechanism in starving cells. At the same time, extensive autophagy is commonly observed in dying cells, leading to its classification as an alternative form of programmed cell death. The functional contribution of autophagy to cell death has been a subject of great controversy. However, several recent loss-of-function studies of autophagy (Atg) genes have begun to address the roles of autophagy in both cell death and survival. Here, we review the emerging evidence in favor of and against autophagic cell death, discuss the possible roles that autophagic degradation might play in dying cells, and identify salient issues for future investigation.

Vital functions for lethal caspases

Abstract: Caspases are a family of cysteine proteases expressed as inactive zymogens in virtually all animal cells. These enzymes play a central role in most cell death pathways leading to apoptosis but growing evidences implicate caspases also in nonapoptotic functions. Several of these enzymes, activated in molecular platforms referred to as inflammasomes, play a role in innate immune response by processing some of the cytokines involved in inflammatory response. Caspases are requested for terminal differentiation of specific cell types, whether this differentiation process leads to enucleation or not. These enzymes play also a role in T and B lymphocyte proliferation and, in some circumstances, appear to be cytoprotective rather than cytotoxic. These pleiotropic functions implicate caspases in the control of life and death but the fine regulation of their dual effect remains poorly understood. The nonapoptotic functions of caspases implicate that cells can restrict the proteolytic activity of these enzymes to selected substrates. Deregulation of the pathways in which caspases exert these nonapoptotic functions is suspected to play a role in the pathophysiology of several human diseases.

Pharmacological manipulation of cell death: clinical applications in sight?

Abstract: This series of Reviews on cell death explores the creation of new therapies for correcting excessive or deficient cell death in human disease. Signal transduction pathways controlling cell death and the molecular core machinery responsible for cellular self-destruction have been elucidated with unprecedented celerity during the last decade, leading to the design of novel strategies for blocking pathological cell loss or for killing unwanted cells. Thus, an increasing number of compounds targeting a diverse range of apoptosis-related molecules are being explored at the preclinical and clinical levels. Beyond the agents that are already FDA approved, a range of molecules targeting apoptosis-regulatory transcription factors, regulators of mitochondrial membrane permeabilization, and inhibitors or activators of cell death-related proteases are under close scrutiny for drug development.

Muscle-Specific Loss of Apoptosis-Inducing Factor Leads to Mitochondrial Dysfunction, Skeletal Muscle Atrophy, and Dilated Cardiomyopathy

Abstract: Cardiac and skeletal muscle critically depend on mitochondrial energy metabolism for their normal function. Recently, we showed that apoptosis-inducing factor (AIF), a mitochondrial protein implicated in programmed cell death, plays a role in mitochondrial respiration. However, the in vivo consequences of AIF-regulated mitochondrial respiration resulting from a loss-of-function mutation in Aif are not known. Here, we report tissue-specific deletion of Aif in the mouse. Mice in which Aif has been inactivated specifically in cardiac and skeletal muscle exhibit impaired activity and protein expression of respiratory chain complex I. Mutant animals develop severe dilated cardiomyopathy, heart failure, and skeletal muscle atrophy accompanied by lactic acidemia consistent with defects in the mitochondrial respiratory chain. Isolated hearts from mutant animals exhibit poor contractile performance in response to a respiratory chain-dependent energy substrate, but not in response to glucose, supporting the notion that impaired heart function in mutant animals results from defective mitochondrial energy metabolism. These data provide genetic proof that the previously defined cell death promoter AIF has a second essential function in mitochondrial respiration and aerobic energy metabolism required for normal heart function and skeletal muscle homeostasis.

Mitochondrial permeability transition in apoptosis and necrosis.

Abstract: Apoptosis has classically been viewed as a process not involving mitochondria, whereas the implication of mitochondrial dysfunction in necrosis has been recognized for several decades. Recently, it has become clear that apoptosis implies a disruption of mitochondrial membrane intregrity that is decisive for the cell death process. Cytofluorometric methods assessing the mitochondrial membrane function and structure can be employed to demonstrate that, at least in most models of apoptosis, mitochondrial changes precede caspase and nuclease activation. Moreover, pharmacological and genetic experiments suggest that the loss of mitochondrial membrane integrity is a critical event of the apoptotic process, beyond or at the point of no return of programmed cell death. Inhibitors of the mitochondrial megachannel (= permeability transition pore) can prevent both the mitochondrial and the post-mitochondrial manifestations of apoptosis.

Mechanisms of apoptosis induction by the HIV-1 envelope

Abstract: The envelope glycoprotein complex (Env) of human immunodeficiency virus-1 (HIV-1) can induce apoptosis by a cornucopia of distinct mechanisms. A soluble Env derivative, gp120, can kill cells through signals that are transmitted by chemokine receptors such as CXCR4. Cell surface-bound Env (gp120/gp41), as present on the plasma membrane of HIV-1-infected cells, can kill uninfected bystander cells expressing CD4 and CXCR4 (or similar chemokine receptors, depending on the Env variant) by at least three different mechanisms. First, a transient interaction involving the exchange of lipids between the two interacting cells (‘the kiss of death’) may lead to the selective death of single CD4-expressing target cells. Second, fusion of the interacting cells may lead to the formation of syncytia which then succumb to apoptosis in a complex pathway involving the activation of several kinases (cyclin-dependent kinase-1, Cdk1; checkpoint kinase-2, Chk2; mammalian target of rapamycin, mTOR; p38 mitogen-activated protein kinase, p38 MAPK; inhibitor of NF-κB kinase, IKK), as well as the activation of several transcription factors (NF-κB, p53), finally resulting in the activation of the mitochondrial pathway of apoptosis. Third, if the Env-expressing cell is at an early stage of imminent apoptosis, its fusion with a CD4-expressing target cell can precipitate the death of both cells, through a process that may be considered as contagious apoptosis and which does not involve Cdk1, mTOR, p38 nor p53, yet does involve mitochondria. Activation of some of the above- mentioned lethal signal transducers have been detected in patients’ tissues, suggesting that HIV-1 may indeed trigger apoptosis through molecules whose implication in Env-induced killing has initially been discovered in vitro.

Essential role of p53 phosphorylation by p38 MAPK in apoptosis induction by the HIV-1 envelope

Abstract: The proapoptotic activity of the transcription factor p53 critically depends on the phosphorylation of serine 46 (p53S46P). Here, we show that syncytia containing p53S46P could be detected in lymph node biopsies from human immunodeficiency virus (HIV)-1 carriers, in the brain of patients with HIV-1–associated dementia and in cocultures of HeLa expressing the HIV-1 envelope glycoprotein complex (Env) with HeLa cells expressing CD4. In this latter model, cell death was the result of a sequential process involving cell fusion, nuclear fusion (karyogamy), phosphorylation of serine 15 (p53S15P), later on serine 46 (p53S46P), and transcription of p53 target genes. Cytoplasmic p38 mitogen-activated protein kinase (MAPK) was found to undergo an activating phosphorylation (p38T180/Y182P [p38 with phosphorylated threonine 180 and tyrosine 182]) before karyogamy and to translocate into karyogamic nuclei. p38T180/Y182P colocalized and coimmunoprecipitated with p53S46P. Recombinant p38 phosphorylated recombinant p53 on serine 46 in vitro. Inhibition of p38 MAPK by pharmacological inhibitors, dominant-negative p38, or small interfering RNA, suppressed p53S46P (but not p53S15P), the expression of p53-inducible genes, the conformational activation of proapoptotic Bax and Bak, the release of cytochrome c from mitochondria, and consequent apoptosis. p38T180/Y182P was also detected in HIV-1–induced syncytia, in vivo, in patients' lymph nodes and brains. Dominant-negative MKK3 or MKK6 inhibited syncytial activation of p38, p53S46P, and apoptosis. Altogether, these findings indicate that p38 MAPK-mediated p53 phosphorylation constitutes a critical step of Env-induced apoptosis.

Dynamic changes in Mcl-1 expression regulate macrophage viability or commitment to apoptosis during bacterial clearance

Abstract: Macrophages are critical effectors of bacterial clearance and must retain viability, despite exposure to toxic bacterial products, until key antimicrobial functions are performed. Subsequently, host-mediated macrophage apoptosis aids resolution of infection. The ability of macrophages to make this transition from resistance to susceptibility to apoptosis is important for effective host innate immune responses. We investigated the role of Mcl-1, an essential regulator of macrophage lifespan, in this switch from viability to apoptosis, using the model of pneumococcal-associated macrophage apoptosis. Upon exposure to pneumococci, macrophages initially upregulate Mcl-1 protein and maintain viability for up to 14 hours. Subsequently, macrophages reduce expression of full-length Mcl-1 and upregulate a 34-kDa isoform of Mcl-1 corresponding to a novel BH3-only splice variant, Mcl-1(Exon-1). Change in expression of Mcl-1 protein is associated with mitochondrial membrane permeabilization, which is characterized by loss of mitochondrial inner transmembrane potential and translocation of cytochrome c and apoptosis-inducing factor. Following pneumococcal infection, macrophages expressing full-length human Mcl-1 as a transgene exhibit a delay in apoptosis and in bacterial killing. Mcl-1 transgenic mice clear pneumococci from the lung less efficiently than nontransgenic mice. Dynamic changes in Mcl-1 expression determine macrophage viability as well as antibacterial host defense.

Redundant cell death mechanisms as relics and backups.

Abstract: Here we review recent observations indicating the existence of redundant cell death mechanisms. We speculate that this redundancy reflects a particular evolutionary history for cellular demise. Autophagic or apoptotic elements might have been added to a primordial death mechanism, initially improving cell dismantling and later acquiring the ability to act themselves as death effectors. The resulting redundancy of cell death mechanisms has pathophysiological implications.

Inhibition of adenine nucleotide translocator pore function and protection against apoptosis in vivo by an HIV protease inhibitor

Abstract: Inhibitors of HIV protease have been shown to have antiapoptotic effects in vitro, yet whether these effects are seen in vivo remains controversial. In this study, we have evaluated the impact of the HIV protease inhibitor (PI) nelfinavir, boosted with ritonavir, in models of nonviral disease associated with excessive apoptosis. In mice with Fas-induced fatal hepatitis, Staphylococcal enterotoxin B-induced shock, and middle cerebral artery occlusion-induced stroke, we demonstrate that PIs significantly reduce apoptosis and improve histology, function, and/or behavioral recovery in each of these models. Further, we demonstrate that both in vitro and in vivo, PIs block apoptosis through the preservation of mitochondrial integrity and that in vitro PIs act to prevent pore function of the adenine nucleotide translocator (ANT) subunit of the mitochondrial permeability transition pore complex.

PK11195 potently sensitizes to apoptosis induction independently from the peripheral benzodiazepin receptor.

Abstract: 1-(2-Chlorophenyl-N-methylpropyl)-3-isoquinolinecarboxamide (PK11195) is a prototypic ligand of the peripheral benzodiazepine receptor (PBR), a mitochondrial outer membrane protein. PK11195 can be used to chemosensitize tumor cells to a variety of chemotherapeutic agents, both in vitro and in vivo. PK11195 has been suggested to exert this effect via inhibition of the multiple drug resistance (MDR) pump and by direct mitochondrial effects which could be mediated by the PBR. Here, we established a model system in which PK11195 and another PBR ligand, 7-chloro-5-(4-chlorophenyl)-1,3-dihydro-1-methyl-2H-1,4-benzodiazepin-2-one (Ro5-4864), sensitize to nutrient depletion-induced cell death. In this MDR-independent model, PK11195 and Ro5-4864 are fully active even when the PBR is knocked down by small interfering RNA. Cells that lack PBR possess low-affinity binding sites for PK11195 and Ro5-4864. The starvation-sensitizing effects of PK11195 are not due to a modulation of the adaptive response of starved cells, namely autophagy and NF-kappaB activation. Rather, it appears that the combination of PK11195 with autophagy or NF-kappaB inhibitors has a potent synergistic death-inducing effect. Starved cells treated with PK11195 exhibit characteristics of apoptosis, including loss of the mitochondrial transmembrane potential, mitochondrial cytochrome c release, caspase activation and chromatin condensation. Accordingly, stabilization of mitochondria by overexpression of Bcl-2 or expression of the viral mitochondrial inhibitor (vMIA) from cytomegalovirus inhibits cell death induced by PK11195 plus starvation. Thus, PK11195 potently sensitizes to apoptosis via a pathway that involves mitochondria, yet does not involve the PBR.

Chemosensitization by erythropoietin through inhibition of the NF-kappaB rescue pathway.

Abstract: Two cell lines that exemplify erythropoietin (EPO) receptor-positive tumors, human renal carcinoma cell lines RCC and the myelomonocytic leukemia cell line U937, were investigated for the apoptosis-modulatory potential of EPO. Cells cultured in the presence of EPO exhibited an elevated apoptotic response to cancer chemotherapeutic agents such as daunorubicin (Dauno) and vinblastine (VBL). Chemosensitization by EPO did not involve an increase in p53 activation, yet correlated with enhanced Bax/Bak-dependent mitochondrial membrane perturbation and caspase maturation. In vitro monotherapy with Dauno or VBL induced the degradation of IκBα, provoked the translocation of NF-κB p65/50 to the nucleus and stimulated the expression of an NF-κB-activatable reporter gene. All these signs of NF-κB activation were perturbed in the presence of EPO. Inhibition of JAK2, one of the receptor-proximal elements of EPO-mediated signal transduction, greatly diminished the EPO-mediated chemosensitization and NF-κB inhibition. EPO lost its death-facilitating effects in the presence of an NF-κB inhibitor, underscoring the cause–effect relationship between EPO-mediated chemosensitization and NF-κB inhibition. Altogether, these results suggest that, at least in a specific subset of tumors, EPO receptor agonists can prevent activation of the NF-κB pathway, thereby enhancing the propensity of EPO receptor-positive tumor cells to undergo apoptosis.

Characterization of Cell Death Pathways in Human Immunodeficiency Virus-Associated Encephalitis

Abstract: Human immunodeficiency virus (HIV)-associated dementia is a neurodegenerative syndrome characterized by cognitive decline, personality change, and motor deficits. HIV-associated encephalitis (HAE), the neuropathology responsible for HIV-associated dementia, involves the formation of multinucleated giant cells or syncytia. In this article we describe the apoptotic pathways activated in the brains of HAE-affected patients. Approximately 50% of multinuclear giant cells exhibited apoptotic DNA fragmentation as detected by the terminal dUTP nick-end labeling technique. In addition, the presence of syncytia in the frontal cortex of ∼35% of HAE patients correlated with the number of cells expressing the HIV-1 protein p24. Histochemical and immunohistochemical analyses revealed that HAE-associated syncytia underwent apoptosis through a mitochondrial pathway previously delineated for HIV-1 envelope-elicited syncytia in vitro. We observed over-expression of the mammalian target of rapamycin (mTOR), a kinase that mediates activation of the pro-apoptotic transcription factor p53, and p53-dependent up-regulation of two effectors of mitochondrial apoptosis, namely the BH3-only proteins Puma and transglutaminase type 2 (TG2). Interestingly, although mTOR activation and Puma induction were observed in dying syncytia and neurons, IkB phosphorylation and TG2 up-regulation were only found in syncytia. These findings provide substantial new information on the cell death mechanisms that regulate HAE, suggesting an important pathogenetic role of syncytia in the disease.

Constitutive presence of cytochrome c in the cytosol of a chemoresistant leukemic cell line.

Abstract: The release of holocytochrome c (cyt c) from mitochondria into the cytosol is reportedly a landmark of the execution phase of apoptosis. As shown here, the P-glycoprotein- (P-gp) expressing K562/ADR cell line (but not the parental K562 cell line) exhibits both cytosolic and mitochondrial cyt c in the absence of any signs of apoptosis. K562/ADR cells were found to be relatively resistant to a variety of different inducers of apoptosis, and blocking the P-gp did not reverse this resistance. The release of cyt c in non-apoptotic K562/ADR cells was not accompanied by that of any other mitochondrial apoptogenic protein, such as AIF or Smac/DIABLO, and was inhibited by Bcl-2 over expression. In addition, using a cell-free system, we show that mitochondria isolated from K562/ADR cells spontaneously released cyt c. These data suggest that cyt c release may be compatible with the preservation of mitochondrial integrity and function, as well as cell proliferation.

Cell death mechanisms in HIV-associated dementia: the involvement of syncytia.

Abstract: 1 National Institute for Infectious Diseases, ‘Lazzaro Spallanzani’, Via Portuense 292, 00149 Rome, Italy 2 CNRS-UMR8125, Institut Gustave Roussy, 39 rue Camille-Desmoulins, F-94805 Villejuif, France 3 Department of Biology, University of Rome ‘Tor Vergata’, Via della Ricerca Scientifica 1, 00173, Rome, Italy * Corresponding author: Professor M Piacentini, Laboratory of Cell Biology and E.M., INMI-IRCCS L. Spallanzani, via Portuense, 292 00149 Rome, Italy. Tel: þ 39 06 55170966; fax: þ 39 06 5582825; E-mail: mauro.piacentini@uniroma2.it

A cellular machine generating apoptosis-prone aneuploid cells.

Abstract: HeLa cells manipulated to express the human immunodeficiency virus (HIV-1)-encoded envelope glycoprotein complex (Env) can fuse with HeLa cells expressing the Env receptor CD4 as well as a chemokine coreceptor (e.g. CXCR4), thus forming syncytia. We have used such Env-elicited syncytia to dissect a lethal p53-dependent signal transduction cascade relevant to AIDS, as well as ‘mitotic catastrophe’, an apoptosis-like cell death that occurs during the metaphase, after fusion of nonsynchronized cells and inactivation of the cell cycle checkpoint kinase Chk2. One of the intrinsic advantages of a model of cell death affecting giant multinuclear cells is the ease with which the subcellular localization of apoptosis-regulatory proteins can be studied. In a coculture of Cell Tracker Green-labeled HeLa Env cells and Cell Tracker Red-labeled HeLa CD4 cells, syncytia can be easily detected as double-strained cells, the vast majority of which have a X4n DNA content. However, a fraction (B10%) of such bona fide syncytia have anB2n DNA content, as determined by simultaneous staining with Hoechst

Anthracyclin induced immunogenic dead or dying cells composition

Abstract: The present invention is aimed at a composition comprising immunogenic dead or dying cancer cells or infected cells and fragments or fractions thereof obtained by treatment with anthracyclins, to a process of manufacturing said composition and to its use for treating cancer, for preparing a vaccine or for treating infectious diseases. It also relates to a medicament comprising at least one anthracyclin to be injected into or at the vicinity of the tumor.

Cell death pathways in retroviral infection

Abstract: Viruses mercilessly exploit their host cells with the exclusive aim of assuring their own survival and propagation. In fact, viruses control all facets of the host cell biology, subverting its metabolism, influencing the cell division cycle and regulating the apoptotic machinery. Viruses target the cell death program at different levels, following a three-layered strategy. First, viruses must avoid the rapid death of freshly infected cells, which constitutes one of the most ancient antiviral defense mechanisms. For example, retroviruses can trigger the DNA damage response as a correlate of retroviral DNA integration, and obviously cell death induced at this stage would abort viral infection. Second, viruses can actively kill the infected target cell, at late stages of the viral life cycle, to ensure its liberation and propagation to adjacent cells. Induction of host cell death may also induce a tolerogenic type of cell death, apoptosis (as opposed to necrotic lysis), thus subverting the host’s immune response. Third, viruses can induce the death of noninfected inflammatory or immune cells. This ‘bystander killing’ is induced with the scope of paralyzing the host’s innate and cognate defense systems. As a result, viruses employ multiple strategies to manipulate (induce or inhibit) cell death. The present Special Issue of Cell Death and Differentiation specifically deals with the regulation of cellular demise by retroviruses, in particular by the human immunodeficiency virus (HIV), which has infected an estimated 60 million people worldwide. Owing to the widespread use of the highly active antiretroviral therapy (HAART), the incidence of AIDS is now declining in the developed West, although the number of HIV-infected individuals still increases. Since viral escape from HAART is increasingly frequent, HIV-1 infection thus will continue to constitute a major epidemiological, medical and scientific problem. HIV kills human cells of different types by a cornucopia of different mechanisms. HIV is particularly malicious since it primarily infects cells from the cognate and innate immune systems (in particular, CD4þ T cells and monocytes), in which it manipulates apoptosis through direct effects. In addition, HIV uses cell-exposed proteins (such as the Envelope glycoprotein complex, Env) or secreted proteins including Vpr to induce cell death in a variety of different cell types. To kill cells, HIV profoundly influences the cell biology, including the cytoskeleton, by acting on multiple cellular receptors, not only the classical HIV-1 receptor CD4 with its chemokine coreceptor. In addition, HIV-1 may induce subtle tactics to induce the death of HIV-1-specific immune effectors, for instance by inducing the expression of CD95//Fas on infected cells, which then engages the CD95/Fas death receptor on CD8þ T cells. Although the detailed mechanisms accounting for T-cell death in vivo are not known, HIV-1 infection profoundly influences the T-cell repertoire, both in CD4þ and in CD8þ T cells in vivo. Animal models such as the infection of macaques by simian immunodeficiency virus (SIV) may be important for the elucidation of the exact molecular mechanisms of the apoptotic pathways participating in the pathogenesis of AIDS. Importantly, HIV-1 provokes a neuropathological response involving all cell types in the brain. The incidence of the socalled HIV-1-associated dementia (HAD) is increasing, and its pathogenesis is likely to be complex, involving multiple direct and indirect apoptosis-inducing mechanisms culminating in neurodegeneration. Soluble viral products such as gp120 and Tat as well as inflammatory responses play a major role in HAD. Moreover, direct mechanisms leading to the formation of synctia may contribute to HAD. The apoptotic response of synctia is likely to involve the activation of p53, illustrating how host cell-intrinsic factors can contribute to viral pathogenesis. Human T-cell leukemia virus type 1 (HTLV-1) is responsible for adult T-cell leukemia/lymphoma, as well as progressive demyelinating neurodegenerative disease termed HTLVassociated myelopathy/tropica spastic paraparesis. HTLV-1 encodes the viral oncogene Tax, which can inhibit apoptosis through the activation of the NF-kB pathway. In addition, HTLV-1 encodes for a protein, p13II, which can sensitize for apoptosis induction, presumably through a direct effect on the mitochondrial inner membrane. How and to which extent p13/II contributes to the HTLV-1-mediated disease states remains on open conundrum. Antiretroviral agents used for the treatment of AIDS can modulate host cell apoptosis, even in the absence of retroviruses. Thus, HIV-1 protease inhibitors can both inhibit and induce apoptosis, depending on the dose and on the cell type investigated. It may thus be speculated that such drugs may become useful in clinical applications in which apoptosis modulation is a therapeutic goal. We do hope that the readers of Cell Death and Differentiation will appreciate the present compendium of articles on retroviral and antiretroviral modulation of cell death pathways. May the ideas shared by the authors stimulate further debate and experimentation. It appears reasonable to anticipate that a detailed comprehension of cell death modulation by retroviruses will open new therapeutic avenues that may reduce viral reproduction, boost antiretroviral immune responses or, alternatively, dampen deleterious host cell loss, for instance in the central nervous system.