RESUMEN
Selenoproteins are rare proteins among all kingdoms of life containing the 21st amino acid, selenocysteine. Selenocysteine resembles cysteine, differing only by the substitution of selenium for sulfur. Yet the actual advantage of selenolate- versus thiolate-based catalysis has remained enigmatic, as most of the known selenoproteins also exist as cysteine-containing homologs. Here, we demonstrate that selenolate-based catalysis of the essential mammalian selenoprotein GPX4 is unexpectedly dispensable for normal embryogenesis. Yet the survival of a specific type of interneurons emerges to exclusively depend on selenocysteine-containing GPX4, thereby preventing fatal epileptic seizures. Mechanistically, selenocysteine utilization by GPX4 confers exquisite resistance to irreversible overoxidation as cells expressing a cysteine variant are highly sensitive toward peroxide-induced ferroptosis. Remarkably, concomitant deletion of all selenoproteins in Gpx4cys/cys cells revealed that selenoproteins are dispensable for cell viability provided partial GPX4 activity is retained. Conclusively, 200 years after its discovery, a specific and indispensable role for selenium is provided.
Asunto(s)
Apoptosis , Glutatión Peroxidasa/metabolismo , Convulsiones/metabolismo , Selenio/metabolismo , Animales , Supervivencia Celular , Células Cultivadas , Femenino , Glutatión Peroxidasa/genética , Células HEK293 , Humanos , Peróxido de Hidrógeno/toxicidad , Interneuronas/metabolismo , Peroxidación de Lípido , Masculino , Ratones , Ratones Endogámicos C57BL , Fosfolípido Hidroperóxido Glutatión Peroxidasa , Convulsiones/etiologíaRESUMEN
Regulatory myeloid immune cells, such as myeloid-derived suppressor cells (MDSCs), populate inflamed or cancerous tissue and block immune cell effector functions. The lack of mechanistic insight into MDSC suppressive activity and a marker for their identification has hampered attempts to overcome T cell inhibition and unleash anti-cancer immunity. Here, we report that human MDSCs were characterized by strongly reduced metabolism and conferred this compromised metabolic state to CD8+ T cells, thereby paralyzing their effector functions. We identified accumulation of the dicarbonyl radical methylglyoxal, generated by semicarbazide-sensitive amine oxidase, to cause the metabolic phenotype of MDSCs and MDSC-mediated paralysis of CD8+ T cells. In a murine cancer model, neutralization of dicarbonyl activity overcame MDSC-mediated T cell suppression and, together with checkpoint inhibition, improved the efficacy of cancer immune therapy. Our results identify the dicarbonyl methylglyoxal as a marker metabolite for MDSCs that mediates T cell paralysis and can serve as a target to improve cancer immune therapy.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Inmunoterapia/métodos , Melanoma/inmunología , Células Supresoras de Origen Mieloide/inmunología , Piruvaldehído/metabolismo , Amina Oxidasa (conteniendo Cobre)/metabolismo , Animales , Linfocitos T CD8-positivos/trasplante , Comunicación Celular , Proliferación Celular , Humanos , Tolerancia Inmunológica , Activación de Linfocitos , Melanoma Experimental , Ratones , Ratones Transgénicos , Neoplasias Experimentales , Receptor de Muerte Celular Programada 1/metabolismoRESUMEN
Mitochondrial fidelity is tightly linked to overall cellular homeostasis and is compromised in ageing and various pathologies1-3. Mitochondrial malfunction needs to be relayed to the cytosol, where an integrated stress response is triggered by the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) in mammalian cells4,5. eIF2α phosphorylation is mediated by the four eIF2α kinases GCN2, HRI, PERK and PKR, which are activated by diverse types of cellular stress6. However, the machinery that communicates mitochondrial perturbation to the cytosol to trigger the integrated stress response remains unknown1,2,7. Here we combine genome engineering and haploid genetics to unbiasedly identify genes that affect the induction of C/EBP homologous protein (CHOP), a key factor in the integrated stress response. We show that the mitochondrial protease OMA1 and the poorly characterized protein DELE1, together with HRI, constitute the missing pathway that is triggered by mitochondrial stress. Mechanistically, stress-induced activation of OMA1 causes DELE1 to be cleaved into a short form that accumulates in the cytosol, where it binds to and activates HRI via its C-terminal portion. Obstruction of this pathway can be beneficial or adverse depending on the type of mitochondrial perturbation. In addition to the core pathway components, our comparative genetic screening strategy identifies a suite of additional regulators. Together, these findings could be used to inform future strategies to modulate the cellular response to mitochondrial dysfunction in the context of human disease.
Asunto(s)
Citosol/metabolismo , Citosol/patología , Mitocondrias/metabolismo , Mitocondrias/patología , Proteínas Mitocondriales/metabolismo , Activación Enzimática , Factor 2 Eucariótico de Iniciación/metabolismo , Genoma Humano/genética , Humanos , Metaloendopeptidasas/metabolismo , Mitocondrias/enzimología , Fosforilación , Unión Proteica , Estrés Fisiológico , Factor de Transcripción CHOP/metabolismo , eIF-2 Quinasa/metabolismoRESUMEN
War and crises affect mental health, social attitudes, and cultural norms, which can exacerbate the state of long-term insecurity. With decades of armed conflict, the Democratic Republic of Congo is one example, and violence has become normalized in civilian settings. In this study, we tested the effectiveness of the NETfacts health system, an integrated model of evidence-based individual trauma treatment (Narrative Exposure Therapy [NET]) and a trauma-informed community-based intervention (NETfacts). Alongside changes in mental health outcomes (posttraumatic stress disorder, depression, social disapproval, and shame) we also investigated change in attitudes, including rape myth acceptance, stigmatization of survivors of sexual violence, and skepticism about the reintegration of former combatants. To test whether the additional community intervention is superior to individual NET alone, we implemented a randomized controlled design with six villages and interviewed a sample of 1,066 community members. Our results demonstrate that the NETfacts health system in comparison with NET alone more effectively reduced rape myth acceptance and with it ongoing victimization and perpetration. Community members of the NETfacts group also presented with less stigmatizing attitudes against survivors of sexual violence. Skepticism about the reintegration of former combatants declined in both groups. NETfacts appears to have increased motivation to engage in individual treatment. Synergizing the healing effects of individual and collective trauma exposure, the NETfacts health system appears to be an effective and scalable approach to correct degrading or ignominious norms and restore functioning and mental health in postconflict communities.
Asunto(s)
Violación , Delitos Sexuales , Trastornos por Estrés Postraumático , Humanos , Violación/psicología , Trastornos por Estrés Postraumático/terapia , Trastornos por Estrés Postraumático/psicología , ViolenciaRESUMEN
The Fe(II) and 2-oxoglutarate-dependent oxygenase Alkb homologue 1 (Alkbh1) has been shown to act on a wide range of substrates, like DNA, tRNA and histones. Thereby different enzymatic activities have been identified including, among others, demethylation of N3-methylcytosine (m3C) in RNA- and single-stranded DNA oligonucleotides, demethylation of N1-methyladenosine (m1A) in tRNA or formation of 5-formyl cytosine (f5C) in tRNA. In accordance with the different substrates, Alkbh1 has also been proposed to reside in distinct cellular compartments in human and mouse cells, including the nucleus, cytoplasm and mitochondria. Here, we describe further evidence for a role of human Alkbh1 in regulation of mitochondrial protein biogenesis, including visualizing localization of Alkbh1 into mitochondrial RNA granules with super-resolution 3D SIM microscopy. Electron microscopy and high-resolution respirometry analyses revealed an impact of Alkbh1 level on mitochondrial respiration, but not on mitochondrial structure. Downregulation of Alkbh1 impacts cell growth in HeLa cells and delays development in Caenorhabditis elegans, where the mitochondrial role of Alkbh1 seems to be conserved. Alkbh1 knockdown, but not Alkbh7 knockdown, triggers the mitochondrial unfolded protein response (UPRmt) in C. elegans.
Asunto(s)
Histona H2a Dioxigenasa, Homólogo 1 de AlkB/metabolismo , Mitocondrias/metabolismo , ARN Mitocondrial/metabolismo , Células A549 , Enzimas AlkB/genética , Enzimas AlkB/metabolismo , Histona H2a Dioxigenasa, Homólogo 1 de AlkB/genética , Animales , Caenorhabditis elegans , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Electroforesis en Gel de Poliacrilamida , Células HEK293 , Células HT29 , Células HeLa , Humanos , Ratones , Microscopía Electrónica , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Consumo de Oxígeno/fisiología , Factor Tu de Elongación Peptídica/genética , Factor Tu de Elongación Peptídica/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Respuesta de Proteína Desplegada/genética , Respuesta de Proteína Desplegada/fisiologíaRESUMEN
Sorafenib represents the current standard of care for patients with advanced-stage hepatocellular carcinoma (HCC). However, acquired drug resistance occurs frequently during therapy and is accompanied by rapid tumor regrowth after sorafenib therapy termination. To identify the mechanism of this therapy-limiting growth resumption, we established robust sorafenib resistance HCC cell models that exhibited mitochondrial dysfunction and chemotherapeutic crossresistance. We found a rapid relapse of tumor cell proliferation after sorafenib withdrawal, which was caused by renewal of mitochondrial structures alongside a metabolic switch toward high electron transport system (ETS) activity. The translation-inhibiting antibiotic tigecycline impaired the biogenesis of mitochondrial DNA-encoded ETS subunits and limited the electron acceptor turnover required for glutamine oxidation. Thereby, tigecycline prevented the tumor relapse in vitro and in murine xenografts in vivo. These results offer a promising second-line therapeutic approach for advanced-stage HCC patients with progressive disease undergoing sorafenib therapy or treatment interruption due to severe adverse events.
Asunto(s)
Carcinoma Hepatocelular/tratamiento farmacológico , Resistencia a Antineoplásicos/efectos de los fármacos , Neoplasias Hepáticas/tratamiento farmacológico , Sorafenib/farmacología , Tigeciclina/farmacología , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Femenino , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Ratones SCID , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Recurrencia Local de Neoplasia/prevención & control , Inhibidores de la Síntesis de la Proteína/farmacologíaRESUMEN
BACKGROUND: The understanding of lysosomes has been expanded in recent research way beyond their view as cellular trash can. Lysosomes are pivotal in regulating metabolism, endocytosis and autophagy and are implicated in cancer. Recently it was discovered that the lysosomal V-ATPase, which is known to induce apoptosis, interferes with lipid metabolism in cancer, yet the interplay between these organelles is poorly understood. METHODS: LC-MS/MS analysis was performed to investigate lipid distribution in cells. Cell survival and signaling pathways were analyzed by means of cell biological methods (qPCR, Western Blot, flow cytometry, CellTiter-Blue). Mitochondrial structure was analyzed by confocal imaging and electron microscopy, their function was determined by flow cytometry and seahorse measurements. RESULTS: Our data reveal that interfering with lysosomal function changes composition and subcellular localization of triacylglycerids accompanied by an upregulation of PGC1α and PPARα expression, master regulators of energy and lipid metabolism. Furthermore, cardiolipin content is reduced driving mitochondria into fission, accompanied by a loss of membrane potential and reduction in oxidative capacity, which leads to a deregulation in cellular ROS and induction of mitochondria-driven apoptosis. Additionally, cells undergo a metabolic shift to glutamine dependency, correlated with the fission phenotype and sensitivity to lysosomal inhibition, most prominent in Ras mutated cells. CONCLUSION: This study sheds mechanistic light on a largely uninvestigated triangle between lysosomes, lipid metabolism and mitochondrial function. Insight into this organelle crosstalk increases our understanding of mitochondria-driven cell death. Our findings furthermore provide a first hint on a connection of Ras pathway mutations and sensitivity towards lysosomal inhibitors.
Asunto(s)
Muerte Celular , Metabolismo de los Lípidos , Mitocondrias/metabolismo , Muramidasa/metabolismo , Línea Celular Tumoral , Humanos , Estrés Oxidativo , ATPasas de Translocación de Protón Vacuolares/antagonistas & inhibidoresRESUMEN
The mechanisms responsible for the generation of a mature B1 and B2 cell compartment are still poorly understood. In this study, we demonstrated that absence of Dickkopf-3 (DKK3) led to changes in the composition of the B cell compartment, which were due to an altered development and maintenance program of B cells. Development of B2 cells was impaired at the pre- and immature B cell stage, resulting in decreased numbers of follicular B cells in adult DKK3-deficient mice. Furthermore, DKK3 limited B1 cell self-maintenance in the periphery, by decreasing the survival and proliferation behavior of B1 cells. DKK3 may act via the BCR signaling pathway, as Ca(2+) influx upon BCR stimulation was increased and SiglecG, a molecule shown to inhibit Calcium signaling, was downregulated in the absence of DKK3. DKK3-deficient mice exhibited altered Ab responses and an increased secretion of the cytokine IL-10. Additionally, DKK3 limited autoimmunity in a model of systemic lupus erythematosus. In summary, we identified DKK3 as a novel modulator interfering with B cell fate as well as the maintenance program of B cells, leading to changes in B cell immune responses.
Asunto(s)
Linfocitos B/inmunología , Proliferación Celular , Péptidos y Proteínas de Señalización Intercelular/inmunología , Transducción de Señal/inmunología , Proteínas Adaptadoras Transductoras de Señales , Animales , Apoptosis/genética , Apoptosis/inmunología , Autoanticuerpos/inmunología , Autoanticuerpos/metabolismo , Linfocitos B/metabolismo , Calcio/inmunología , Calcio/metabolismo , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Células Cultivadas , Citocinas/inmunología , Citocinas/metabolismo , Citometría de Flujo , Expresión Génica/inmunología , Humanos , Péptidos y Proteínas de Señalización Intercelular/deficiencia , Péptidos y Proteínas de Señalización Intercelular/genética , Lectinas/genética , Lectinas/inmunología , Lectinas/metabolismo , Lupus Eritematoso Sistémico/genética , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/metabolismo , Recuento de Linfocitos , Ratones Endogámicos C57BL , Ratones Endogámicos MRL lpr , Ratones Noqueados , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/inmunología , Receptores de Antígenos de Linfocitos B/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico , Transducción de Señal/genéticaRESUMEN
In healthy individuals, T cells react against incoming pathogens, but remain tolerant to self-antigens, thereby preventing autoimmune reactions. CD4 regulatory T cells are major contributors in induction and maintenance of peripheral tolerance, but a regulatory role has been also reported for several subsets of CD8 T cells. To determine the molecular basis of peripheral CD8 T-cell tolerance, we exploited a double transgenic mouse model in which CD8 T cells are neonatally tolerized following interaction with a parenchymal self-antigen. These tolerant CD8 T cells have regulatory capacity and can suppress T cells in an antigen-specific manner during adulthood. Dickkopf-3 (DKK3) was found to be expressed in the tolerant CD8 T cells and to be essential for the observed CD8 T-cell tolerance. In vitro, genetic deletion of DKK3 or blocking with antibodies restored CD8 T-cell proliferation and IL-2 production in response to the tolerizing self-antigen. Moreover, exogenous DKK3 reduced CD8 T-cell reactivity. In vivo, abrogation of DKK3 function reversed tolerance, leading to eradication of tumors expressing the target antigen and to rejection of autologous skin grafts. Thus, our findings define DKK3 as a immune modulator with a crucial role for CD8 T-cell tolerance.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Tolerancia Inmunológica/fisiología , Péptidos y Proteínas de Señalización Intercelular/fisiología , Proteínas Adaptadoras Transductoras de Señales , Animales , Western Blotting , Linfocitos T CD8-positivos/citología , Proliferación Celular , Citotoxicidad Inmunológica , Citometría de Flujo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The cell-toxic bile salt glycochenodeoxycholic acid (GCDCA) and taurochenodeoxycholic acid (TCDCA) are responsible for hepatocyte demise in cholestatic liver diseases, while tauroursodeoxycholic acid (TUDCA) is regarded hepatoprotective. We demonstrate the direct mitochondrio-toxicity of bile salts which deplete the mitochondrial membrane potential and induce the mitochondrial permeability transition (MPT). The bile salt mediated mechanistic mode of destruction significantly differs from that of calcium, the prototype MPT inducer. Cell-toxic bile salts initially bind to the mitochondrial outer membrane. Subsequently, the structure of the inner boundary membrane disintegrates. And it is only thereafter that the MPT is induced. This progressive destruction occurs in a dose- and time-dependent way. We demonstrate that GCDCA and TCDCA, but not TUDCA, preferentially permeabilize liposomes containing the mitochondrial membrane protein ANT, a process resembling the MPT induction in whole mitochondria. This suggests that ANT is one decisive target for toxic bile salts. To our knowledge this is the first report unraveling the consecutive steps leading to mitochondrial destruction by cell-toxic bile salts.
Asunto(s)
Ácido Glicoquenodesoxicólico/toxicidad , Mitocondrias Hepáticas/efectos de los fármacos , Translocasas Mitocondriales de ADP y ATP/agonistas , Ácido Tauroquenodesoxicólico/farmacología , Animales , Permeabilidad de la Membrana Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Liposomas/química , Hígado/química , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias Cardíacas/química , Mitocondrias Hepáticas/metabolismo , Mitocondrias Hepáticas/patología , Translocasas Mitocondriales de ADP y ATP/aislamiento & purificación , Proteínas de Transporte de Membrana Mitocondrial/agonistas , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Membranas Mitocondriales/química , Membranas Mitocondriales/efectos de los fármacos , Poro de Transición de la Permeabilidad Mitocondrial , Miocardio/química , Ratas , Ácido Tauroquenodesoxicólico/toxicidad , Canales Aniónicos Dependientes del Voltaje/química , Canales Aniónicos Dependientes del Voltaje/aislamiento & purificaciónRESUMEN
Mitochondrial dysfunctions decisively contribute to the progression of human diseases, implying that functional tests of isolated mitochondria may furnish conclusive information for diagnosis and therapy. Classical mitochondrial isolation methods, however, lack precisely adjustable settings for cell rupture, which is the most critical step in this procedure, and this complicates subsequent analyses. Here, we present an efficient method to isolate functionally active, intact mitochondria from cultured or primary cells and minute tissue samples in a rapid, highly reproducible manner.
Asunto(s)
Hepatocitos/ultraestructura , Mitocondrias Hepáticas/ultraestructura , Neuronas/ultraestructura , Animales , Automatización de Laboratorios , Biomarcadores/metabolismo , Fraccionamiento Celular , Línea Celular Tumoral , Hepatocitos/metabolismo , Humanos , Ratones , Ratones Endogámicos C57BL , Mitocondrias Hepáticas/metabolismo , Neuronas/metabolismo , Cultivo Primario de Células , Ratas , Ratas Endogámicas BUF , Ratas Endogámicas WKYRESUMEN
BACKGROUND: Excessive formation of reactive oxygen species contributes to tissue injury and functional deterioration after myocardial ischemia/reperfusion. Especially, mitochondrial reactive oxygen species are capable of opening the mitochondrial permeability transition pore, a harmful event in cardiac ischemia/reperfusion. Thioredoxins are key players in the cardiac defense against oxidative stress. Mutations in the mitochondrial thioredoxin reductase (thioredoxin reductase-2, Txnrd2) gene have been recently identified to cause dilated cardiomyopathy in patients. Here, we investigated whether mitochondrial thioredoxin reductase is protective against myocardial ischemia/reperfusion injury. METHODS AND RESULTS: In mice, α-MHC-restricted Cre-mediated Txnrd2 deficiency, induced by tamoxifen (Txnrd2-/-ic), aggravated systolic dysfunction and cardiomyocyte cell death after ischemia (90 minutes) and reperfusion (24 hours). Txnrd2-/-ic was accompanied by a loss of mitochondrial integrity and function, which was resolved on pretreatment with the reactive oxygen species scavenger N-acetylcysteine and the mitochondrial permeability transition pore blocker cyclosporin A. Likewise, Txnrd2 deletion in embryonic endothelial precursor cells and embryonic stem cell-derived cardiomyocytes, as well as introduction of Txnrd2-shRNA into adult HL-1 cardiomyocytes, increased cell death on hypoxia and reoxygenation, unless N-acetylcysteine was coadministered. CONCLUSIONS: We report that Txnrd2 exerts a crucial function during postischemic reperfusion via thiol regeneration. The efficacy of cyclosporin A in cardiac Txnrd2 deficiency may indicate a role for Txnrd2 in reducing mitochondrial reactive oxygen species, thereby preventing opening of the mitochondrial permeability transition pore.
Asunto(s)
Mitocondrias/enzimología , Daño por Reperfusión Miocárdica/metabolismo , Estrés Oxidativo/fisiología , Compuestos de Sulfhidrilo/metabolismo , Tiorredoxina Reductasa 2/metabolismo , Acetilcisteína/farmacología , Animales , Muerte Celular/efectos de los fármacos , Muerte Celular/fisiología , Células Cultivadas , Ciclosporina/farmacología , Células Madre Embrionarias/citología , Células Endoteliales/citología , Inhibidores Enzimáticos/farmacología , Depuradores de Radicales Libres/farmacología , Regulación Enzimológica de la Expresión Génica/fisiología , Células Madre Hematopoyéticas/citología , Ratones , Ratones Noqueados , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Daño por Reperfusión Miocárdica/patología , Daño por Reperfusión Miocárdica/fisiopatología , Miocitos Cardíacos/citología , Estrés Oxidativo/efectos de los fármacos , Tiorredoxina Reductasa 1/genética , Tiorredoxina Reductasa 1/metabolismo , Tiorredoxina Reductasa 2/genéticaRESUMEN
Thymic selection shapes the T cell repertoire to ensure maximal antigenic coverage against pathogens while preventing autoimmunity. Recognition of self-peptides in the context of peptide-MHC complexes by the TCR is central to this process, which remains partially understood at the molecular level. In this study we provide genetic evidence that the Nck adapter proteins are essential for thymic selection. In vivo Nck deletion resulted in a reduction of the thymic cellularity, defective positive selection of low-avidity T cells, and impaired deletion of thymocytes engaged by low-potency stimuli. Nck-deficient thymocytes were characterized by reduced ERK activation, particularly pronounced in mature single positive thymocytes. Taken together, our findings identify a crucial role for the Nck adapters in enhancing TCR signal strength, thereby fine-tuning the threshold of thymocyte selection and shaping the preimmune T cell repertoire.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/inmunología , Proteínas Oncogénicas/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Transducción de Señal/inmunología , Linfocitos T/inmunología , Timo/inmunología , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Activación Enzimática/genética , Activación Enzimática/inmunología , Quinasas MAP Reguladas por Señal Extracelular/genética , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Eliminación de Gen , Antígenos de Histocompatibilidad/genética , Antígenos de Histocompatibilidad/inmunología , Antígenos de Histocompatibilidad/metabolismo , Ratones , Ratones Noqueados , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Péptidos/genética , Péptidos/inmunología , Péptidos/metabolismo , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/metabolismo , Transducción de Señal/genética , Linfocitos T/citología , Linfocitos T/metabolismo , Timo/citología , Timo/metabolismoRESUMEN
BACKGROUND: Interpersonal violence damages mental health and frequently leads to aggressive defence strategies. If survivors are subsequently blamed for the events, both consequences worsen. Stigma flourishes, especially when survivors are silenced so that details of the trauma remain unknown. Breaking the secrecy both at the individual and collective level is key to enable the healing and reconciliation of individuals and communities living under continuous threat. METHOD: The NETfacts health system is a stepped care model with three components: (1) Narrative Exposure Therapy (NET), an evidence-based trauma therapy that includes survivor testimony (2) NET for Forensic Offender Rehabilitation (FORNET) acknowledges that perpetrators are frequently also victims and assists in reducing aggression and the attraction to violence, and (3) a community intervention disseminating and discussing Facts derived from NET treatment (NETfacts) to challenge the collective avoidance of atrocities and other traumatic material. The intervention was piloted in a community with 497 adult residents in Eastern Democratic Republic of Congo. The willingness of clients to consent to sharing their anonymised testimonies (with a focus on sexual violence survivors and ex-combatants) was investigated, together with other components of feasibility including security and clinical safety, extent of support of respected local authorities and participation rates. As secondary outcomes, clinical and social measures were assessed before and post NETfacts among 200 village residents of whom 160 self-enrolled and 40 had not participated in any form of treatment. RESULTS: Implementation was feasible with 248 clients from a partner project giving consent to use their testimonies and high support of respected local authorities and participation rates (56% of residents self-enrolled in NETfacts). Immediate beneficial effects were shown for posttraumatic stress and rejection of rape myths among NETfacts participants who experienced multiple traumatic events in their own past. Attitudes towards ex-combatants improved and the perceived lack of social acknowledgement after trauma increased independent from participation. No significant change was observed for depressive symptoms. CONCLUSION: NETfacts is a feasible and promising approach to challenge the culture of secrecy surrounding trauma, suppression and social exclusion. Long term effectiveness requires further evaluation.
Asunto(s)
Violación , Trastornos por Estrés Postraumático , Adulto , Humanos , Estudios de Factibilidad , Trastornos por Estrés Postraumático/psicología , Violencia/psicología , Violación/psicología , Agresión/psicologíaRESUMEN
OBJECTIVE: Classical ATP-independent non-shivering thermogenesis enabled by uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) is activated, but not essential for survival, in the cold. It has long been suspected that futile ATP-consuming substrate cycles also contribute to thermogenesis and can partially compensate for the genetic ablation of UCP1 in mouse models. Futile ATP-dependent thermogenesis could thereby enable survival in the cold even when brown fat is less abundant or missing. METHODS: In this study, we explore different potential sources of UCP1-independent thermogenesis and identify a futile ATP-consuming triglyceride/fatty acid cycle as the main contributor to cellular heat production in brown adipocytes lacking UCP1. We uncover the mechanism on a molecular level and pinpoint the key enzymes involved using pharmacological and genetic interference. RESULTS: ATGL is the most important lipase in terms of releasing fatty acids from lipid droplets, while DGAT1 accounts for the majority of fatty acid re-esterification in UCP1-ablated brown adipocytes. Furthermore, we demonstrate that chronic cold exposure causes a pronounced remodeling of adipose tissues and leads to the recruitment of lipid cycling capacity specifically in BAT of UCP1-knockout mice, possibly fueled by fatty acids from white fat. Quantification of triglyceride/fatty acid cycling clearly shows that UCP1-ablated animals significantly increase turnover rates at room temperature and below. CONCLUSION: Our results suggest an important role for futile lipid cycling in adaptive thermogenesis and total energy expenditure.
Asunto(s)
Tejido Adiposo Pardo , Termogénesis , Adenosina Trifosfato/metabolismo , Tejido Adiposo Pardo/metabolismo , Animales , Ácidos Grasos/metabolismo , Ratones , Ratones Noqueados , Triglicéridos/metabolismo , Proteína Desacopladora 1/genética , Proteína Desacopladora 1/metabolismoRESUMEN
Reintegration of ex-combatants involves multiple challenges. In addition to the trauma-related psychological sequelae, social obstacles in the community can aggravate psychopathological aggressive tendencies and lead to the continuation of violence in civilian life. However, the association between others' negative attitudes and ex-combatants' ongoing perpetration of violence remains largely unexplored. Between September 2018 and May 2019, we assessed a representative community sample of adults in Eastern DR Congo (N = 1,058) and measured trauma exposure, perpetration, mental health problems (PTSD, depression, and appetitive aggression), perceived stigma (shame, perceived lack of social acknowledgement), experienced stigma, and skepticism toward reintegration with ex-combatants. Male ex-combatants (12%, n = 129) had more past trauma and violence perpetration than other community members and a greater number of recent conflicts (including both victimization and perpetration) within the community and with strangers/organized violence. They reported more experienced stigma, more severe PTSD symptoms but were less skeptical about reintegration. Ex-combatants' ongoing violence was predicted by an interplay of the community's skepticism toward reintegration and ex-combatants' perceived and recently experienced stigma (often attributed to the armed group history) and mental health problems, in addition to lifetime traumatization. These findings promote the need for combined interventions that address individual mental health problems including aggression and collective discriminatory attitudes and behaviors.
Asunto(s)
Estereotipo , Violencia , Agresión , República Democrática del Congo , Humanos , MasculinoRESUMEN
BACKGROUND: Survivors of sexual violence are frequently condemned and socially excluded. Myths about rape may translate into stigmatization, diminish disclosure, prevent help-seeking from support structures and worsen mental health. Areas of conflict or organized violence remain the evident hotspots of sexual victimization. However, little is known about prevalence and predictors of rape myths in these settings or their association with survivors' disclosure, stigmatization and psychopathology. METHOD: Between September 2018 and May 2019, we assessed in a representative sample of 1066 individuals from six communities in Eastern DRC traumatic exposure, sexual perpetration, threats to social integrity, perceived stigmatization (perceived lack of social acknowledgement, shame), stigmatizing attitudes towards survivors (negative attitudes and willingness to provide support, rape myths acceptance), and mental illness (PTSD, depression). RESULTS: Survivors of sexual violence (33%, n = 184 of women, 16%, n = 84 of men) reported more traumatic exposure, threats to social integrity, shame, perceived lack of social acknowledgement, PTSD symptoms and depression. Their social environment affirmed various stigmatizing attitudes (5-89% affirmations). Beliefs in rape myths were predicted by its average acceptance in the community, education, and witness of others' sexual victimization. The rates of cases whose history of sexual victimization was socially disclosed were higher in communities and among survivors with low rape myths acceptance and disclosure showed associations with perceived stigmatization. Rape myths acceptance among individuals without a history of sexual victimization was associated with survivors' recently experienced threats to social integrity which predicted their stigma perceptions and mental illness. CONCLUSION: Rape myths acceptance in the community is associated with stigma and trauma-related mental illness of sexual violence survivors. This adds up to the psychic burden of trauma.
RESUMEN
Isolation of mitochondria is a crucial method for examining molecular details of this organelle's manifold functions. Historically, mitochondrial isolations required large amounts of sample material which impeded their isolation from cultured cells. We have therefore developed a method allowing for controlled and reproducible isolation of intact and functional mitochondria from diverse cell types in culture. Here we provide a methodological update of this approach together with a protocol for the subsequent analysis of such isolated mitochondria by electron microscopy. Combining the isolation procedure with this powerful imaging method can reveal ultrastructural mitochondrial peculiarities in disease settings that might not be evident in intact cells and allows for assessment of mitochondrial membrane integrity and sample purity.
Asunto(s)
Neoplasias Hepáticas/patología , Microscopía Electrónica/métodos , Mitocondrias Hepáticas/patología , Línea Celular Tumoral , Separación Celular/instrumentación , Separación Celular/métodos , Diseño de Equipo , Células Hep G2 , Humanos , Procesamiento de Imagen Asistido por Computador/métodosRESUMEN
OBJECTIVE: Individuals who return from armed groups present with a history of traumatic events including perpetration. Subsequent severe mental stress and heightened levels of reactive and appetitive aggression may persist and if left untreated, frequently impede peacebuilding and societal stability. In this study, we tested a revised adaptation of Narrative Exposure Therapy (NET; Schauer et al., 2011) for Forensic Offender Rehabilitation (FORNET) implemented in a sample of male former combatants in war-torn regions of the DR Congo. METHOD: We applied a longitudinal parallel-group randomized controlled design with treatment as usual (TAU) as control condition and 3-5 and 6-9 months follow-up assessments. The effect of treatment over time on clinical and social outcomes was tested with GLMMs; appetitive aggression and current violent behavior (CVB) were specified as primary and posttraumatic stress as secondary outcomes. RESULTS: FORNET decreased appetitive aggression (within group Cohen's dz = 2.00), CVB (dz = .90) and posttraumatic stress (dz = 1.48) significantly more than treatment as usual. Clinical significance was obtained for all outcomes. Remarkably, NET clients also reduced their substance abuse (dz = .68) even though this was not targeted within the intervention. Depression, perceived social acknowledgement and subjective solidarity with (para)military life decreased. CONCLUSION: FORNET is a compact and scalable psychotherapeutic intervention that effectively reduces current aggressive behavior including physical abuse against children, intimate partner violence (IPV), and community violence. FORNET further decreases appetitive aggression, posttraumatic stress symptoms, and other clinical and social problems that commonly hinder demobilization, reintegration, and post-conflict peacebuilding. (PsycInfo Database Record (c) 2021 APA, all rights reserved).
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Agresión/psicología , Terapia Implosiva/métodos , Terapia Narrativa/métodos , Trastornos por Estrés Postraumático/psicología , Trastornos por Estrés Postraumático/terapia , Veteranos/psicología , Adolescente , Adulto , Anciano , República Democrática del Congo , Humanos , Masculino , Persona de Mediana Edad , Factores de Riesgo , Resultado del Tratamiento , Veteranos/estadística & datos numéricos , Violencia , Adulto JovenRESUMEN
Astrocyte-to-neuron conversion is a promising avenue for neuronal replacement therapy. Neurons are particularly dependent on mitochondrial function, but how well mitochondria adapt to the new fate is unknown. Here, we determined the comprehensive mitochondrial proteome of cortical astrocytes and neurons, identifying about 150 significantly enriched mitochondrial proteins for each cell type, including transporters, metabolic enzymes, and cell-type-specific antioxidants. Monitoring their transition during reprogramming revealed late and only partial adaptation to the neuronal identity. Early dCas9-mediated activation of genes encoding mitochondrial proteins significantly improved conversion efficiency, particularly for neuron-enriched but not astrocyte-enriched antioxidant proteins. For example, Sod1 not only improves the survival of the converted neurons but also elicits a faster conversion pace, indicating that mitochondrial proteins act as enablers and drivers in this process. Transcriptional engineering of mitochondrial proteins with other functions improved reprogramming as well, demonstrating a broader role of mitochondrial proteins during fate conversion.