RESUMO
Activation of the pseudokinase mixed lineage kinase domain-like (MLKL) upon its phosphorylation by the protein kinase RIPK3 triggers necroptosis, a form of programmed cell death in which rupture of cellular membranes yields release of intracellular components. We report that MLKL also associated with endosomes and controlled the transport of endocytosed proteins, thereby enhancing degradation of receptors and ligands, modulating their induced signaling and facilitating the generation of extracellular vesicles. This role was exerted on two quantitative grades: a constitutive one independent of RIPK3, and an enhanced one, triggered by RIPK3, where the association of MLKL with the endosomes was enhanced, and it was found to bind endosomal sorting complexes required for transport (ESCRT) proteins and the flotillins and to be excluded, together with them, from cells within vesicles. We suggest that release of phosphorylated MLKL within extracellular vesicles serves as a mechanism for self-restricting the necroptotic activity of this protein.
Assuntos
Apoptose/imunologia , Endossomos/metabolismo , Vesículas Extracelulares/metabolismo , Necrose/imunologia , Proteínas Quinases/metabolismo , Linhagem Celular , Humanos , Mutação/genética , Fosforilação , Engenharia de Proteínas , Proteínas Quinases/genética , Transporte Proteico , Proteômica , RNA Interferente Pequeno/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Transdução de SinaisRESUMO
OBJECTIVE: The aim: To provide best practices of disease management to improve treatment outcomes for this group of patients. PATIENTS AND METHODS: Materials and methods: The paper is based on first-hand experience in observing and treating 117 children aged from 6 months to 13 years with bilateral congenital cryptorchidism during a ten-year follow-up period, who were referred for surgical treatment, and 3 newborn boys with undescended testicles and testicular torsion. A complex of clinical and laboratory, instrumental, endocrinological, and genetic research methods was used for the survey of all patients. RESULTS: Results: Recognizing the action of a common causative factor for bilateral cryptorchidism, which is a consequence of primary endocrine disorders, makes it possible to predict bilateral identity of the location of testicles in this pathology, which we observed in 81 patients: bilateral inguinal cryptorchidism was registered in 49 (41.88%) children, bilateral abdominal cryptorchidism - in 32 (27.35%) children, a combination of inguinal and abdominal cryptorchidism - in 24 (20.51%) children. The following types of treatment were used in the studied group of children: 1 - primary surgical intervention - 4 children, representing 3.42%. 2 - observation and non-surgical treatment by an endocrinologist - 113 (96.58%) children. 3 - comprehensive treatment (surgical correction after hormone treatment) - 67 (59.29%) children. According to the research, hormone therapy had a positive effect on descent of the testicles in 89 (78.76%) patients: the testicles descended into the scrotum - in 22 (24.72%) children; the testicles descended in the inguinal canal - in 32 (35.95%) children; the testicles descended to the level of the opening to the inguinal canal - in 35 (39.33%) children. CONCLUSION: Conclusions: 1. All doctors, starting from the maternity hospital, polyclinic, children's unit, should identify children with bilateral cryptorchidism. All children diagnosed with bilateral cryptorchidism are referred to a surgeon or endocrinologist. The parents of a child with bilateral cryptorchidism should immediately consult a doctor. The study of the reasons for late admission of children to the surgical hospital revealed that 76.92% of patients sought medical advice late, after 1 year of life. 2. At the stage of diagnosis and determination of treatment tactics, an examination by an endocrinologist and a geneticist is necessary; ignoring them is considered an error in diagnostic and therapeutic tactics, since the process of descent of the testicles into the scrotum is hormone-dependent. 3. The indications for primary surgical treatment of a child with bilateral cryptorchidism involve a combination of cryptorchidism with inguinal hernia and pain syndrome, which might be caused by testicular torsion. 4. Hormone therapy provides better results of surgical intervention in bilateral cryptorchidism. The ineffectiveness of two courses of hormone therapy necessitates surgical treatment. 5. Comprehensive treatment of children with bilateral cryptorchidism (non-surgical hormone therapy and surgical correction) has led to good postoperative results in 71.64% of patients, satisfactory results - in 22.39% of children, recurrences - in 5.97% of patients. 7. A long-term follow-up observation should be carried out by a surgeon and endocrinologist until patients reach their reproductive years.
Assuntos
Criptorquidismo , Torção do Cordão Espermático , Cirurgiões , Gravidez , Masculino , Recém-Nascido , Criança , Humanos , Feminino , Lactente , Criptorquidismo/cirurgia , Criptorquidismo/diagnóstico , Torção do Cordão Espermático/tratamento farmacológico , Gonadotropina Coriônica/uso terapêuticoRESUMO
Activation of antiviral responses by RNA sensors RIG-I and MDA5 must be stringently controlled. In this issue of Immunity, Wies et al. (2013) show that a requirement for activation-induced dephosphorylation of these proteins reinforces this restriction.
RESUMO
Caspase-8 deficiency in certain cells prompts chronic inflammation. One mechanism suggested to account for this inflammation is enhanced signaling for necrotic cell death, mediated by the protein kinases RIPK1 and RIPK3 that caspase-8 can cleave. We describe an activity of caspase-8 in dendritic cells that controls the initiation of inflammation in another way. Caspase-8 deficiency in these cells facilitated lipopolysaccharide-induced assembly and function of the NLRP3 inflammasome. This effect depended on the functions of RIPK1 and RIPK3, as well as of MLKL and PGAM5, two signaling proteins recently shown to contribute to RIPK3-mediated induction of necrosis. However, although enhancement of inflammasome assembly in the caspase-8-deficient cells shares proximal signaling events with the induction of necrosis, it occurred independently of cell death. These findings provide new insight into potentially pathological inflammatory processes to which RIPK1- and RIPK3-mediated signaling contributes.
Assuntos
Proteínas de Transporte/genética , Caspase 8/metabolismo , Inflamassomos/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Proteínas de Transporte/imunologia , Caspase 8/genética , Células Cultivadas , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Deleção de Genes , Inflamassomos/imunologia , Interleucina-1beta/metabolismo , Lipopolissacarídeos/imunologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Transdução de Sinais , Ativação TranscricionalRESUMO
STING, a protein mediating signaling activation by cytosolic DNA and known to act downstream of DNA sensor proteins, is shown also to bind DNA directly, raising the need to clarify the relative contributions of these distinct binding modes to DNA sensing.
RESUMO
Excessive responses to pattern-recognition receptors are prevented by regulatory mechanisms that affect the amounts and activities of the downstream signaling proteins. We report that activation of the transcription factor IRF3 by the ribonucleic acid sensor RIG-I was restricted by caspase-8-mediated cleavage of the RIP1 protein, which resulted in conversion of RIP1 from a signaling enhancer to a signaling inhibitor. The proteins RIP1 and caspase-8 were recruited to the RIG-I complex after viral infection and served antagonistic regulatory roles. Conjugation of ubiquitin chains to RIP1 facilitated assembly of the RIG-I complex, resulting in enhanced phosphorylation of IRF3. However, the ubiquitination of RIP1 also rendered it susceptible to caspase-8-mediated cleavage that yielded an inhibitory RIP1 fragment. The dependence of RIP1 cleavage on the same molecular change as that facilitating RIG-I signaling allows for RIG-I signaling to be restricted in its duration without compromising its initial activation.
Assuntos
Caspase 8/imunologia , Regulação da Expressão Gênica , Fator Regulador 3 de Interferon/imunologia , Complexo de Proteínas Formadoras de Poros Nucleares/imunologia , RNA Helicases/imunologia , Proteínas de Ligação a RNA/imunologia , Receptores do Ácido Retinoico/imunologia , Animais , Caspase 8/genética , Linhagem Celular Tumoral , Técnicas de Silenciamento de Genes , Células HEK293 , Células HeLa , Humanos , Immunoblotting , Camundongos , Análise em Microsséries , RNA Helicases/metabolismo , Proteínas Repressoras/imunologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
We present the generalized experimental results of performance degradation of hydrogen sensors based on metal-insulator-semiconductor field effect transistor (MISFET)with the structure Pd-Ta2O5-SiO2-Si. The n-channel MISFET elements were fabricated on silicon single chips together with temperature sensors and heater-resistors by means of conventional -technology. Two hundred cycles of responses to different hydrogen concentrations were measured during eight weeks using special measuring and temperature stabilization circuitries with a feedback loop based on the chip's thermo-sensor and heater. We show how the response parameters change during long-term tests of sensors under repeated hydrogen impacts. There were two stages of time-dependent response instability, the degradation of which depends on operating conditions, hydrogen concentrations, and time. To interpret results, we proposed the models, parameters of which were calculated using experimental data. These models can be used to predict performances of MISFET-based gas analysis devices for long-term operation.
RESUMO
Necrosis, a form of death characterized by rupture of the cell membrane, is closely interlinked with inflammation. Cellular components released during necrotic death can trigger inflammation. Conversely, inflammation often yields tissue damage and, as a consequence, cell death. Which occurs first--necrosis or inflammation--in specific in vivo situations is currently difficult to tell. A way out of this 'chicken-and-egg' conundrum may be found via the recent finding that both necrotic cell death and inflammation can be initiated by a distinct set of signaling proteins, the 'necrosome', that includes receptor-interacting protein (RIP)1, RIP3 and caspase-8. Further clarifying the function of these signaling proteins should make it possible to establish when they induce inflammation directly and when inflammation is caused by necrotic cell death.
Assuntos
Inflamação/imunologia , Animais , Humanos , Inflamação/metabolismo , Inflamação/patologia , Necrose/imunologia , Necrose/metabolismo , Transdução de SinaisRESUMO
Nociceptive axons undergo remodeling as they innervate their targets during development and in response to environmental insults and pathological conditions. How is nociceptive morphogenesis regulated? Here, we show that the microtubule destabilizer kinesin family member 2A (Kif2a) is a key regulator of nociceptive terminal structures and pain sensitivity. Ablation of Kif2a in sensory neurons causes hyperinnervation and hypersensitivity to noxious stimuli in young adult mice, whereas touch sensitivity and proprioception remain unaffected. Computational modeling predicts that structural remodeling is sufficient to explain the phenotypes. Furthermore, Kif2a deficiency triggers a transcriptional response comprising sustained upregulation of injury-related genes and homeostatic downregulation of highly specific channels and receptors at the late stage. The latter effect can be predicted to relieve the hyperexcitability of nociceptive neurons, despite persisting morphological aberrations, and indeed correlates with the resolution of pain hypersensitivity. Overall, we reveal a critical control node defining nociceptive terminal structure, which is regulating nociception.
Assuntos
Cinesinas , Nociceptividade , Proteínas Repressoras , Animais , Camundongos , Cinesinas/genética , Neurônios/fisiologia , Dor , Proteínas Repressoras/genéticaRESUMO
How transcription factors orchestrate the combinatorial expression of cell-surface proteins that, in turn, specify the wiring of the nervous system is an open question. In this issue of Neuron, Xie et al. reveal a new, unexpected layer of complexity.
Assuntos
Neurônios , Fatores de Transcrição , Sistema Nervoso , Neurônios/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Cells in vivo do not act in isolation. Therefore, when attempting to predict the results of pharmaceutical modulation of the function of a protein, we must also take into account the non-cell-autonomous consequences of such modulation. Studies of caspase-8 initially indicated that it serves as the proximal enzyme in cellular self-destruction dictated through the extrinsic cell-death pathway. Later studies revealed that it also participates in mechanisms affecting cell growth and survival. This essay presents a brief account of a study indicating that, apart from functional changes that are cell autonomous, tissue-specific deletion of caspase-8 in mice also has non-cell-autonomous effects with consequences that might even be the opposite of the cell-autonomous ones.
Assuntos
Caspase 8/fisiologia , Animais , Apoptose , Caspase 8/genética , Inibidores Enzimáticos/farmacologia , Deleção de Genes , Hepatectomia , Hepatócitos/enzimologia , Camundongos , Camundongos TransgênicosRESUMO
Caspase-8, the proximal enzyme in the death-induction pathway of the TNF/nerve growth factor receptor family, is activated upon juxtaposition of its molecules within the receptor complexes and is then self-processed. Caspase-8 also contributes to the regulation of cell survival and growth, but little is known about the similarities or the differences between the mechanisms of these nonapoptotic functions and of the enzyme's apoptotic activity. In this study, we report that in bacterial artificial chromosome-transgenic mice, in which the aspartate residue upstream of the initial self-processing site in caspase-8 (D387) was replaced by alanine, induction of cell death by Fas is compromised. However, in contrast to caspase-8-deficient mice, which die in utero at mid-gestation, the mice mutated at D387 were born alive and seemed to develop normally. Moreover, mice with the D387A mutation showed normal in vitro growth responses of T lymphocytes to stimulation of their Ag receptor as well as of B lymphocytes to stimulation by LPS, normal differentiation of bone marrow macrophage precursors in response to M-CSF, and normal generation of myeloid colonies by the bone marrow hematopoietic progenitors, all of which are compromised in cells deficient in caspase-8. These finding indicated that self-processing of activated caspase-8 is differentially involved in the different functions of this enzyme: it is needed for the induction of cell death through the extrinsic cell death pathway but not for nonapoptotic functions of caspase-8.
Assuntos
Síndrome de Alstrom , Apoptose/genética , Caspase 8/genética , Cromossomos Artificiais Bacterianos/genética , Regulação Enzimológica da Expressão Gênica/imunologia , Mutação , Processamento Pós-Transcricional do RNA/imunologia , Animais , Formação de Anticorpos/genética , Apresentação de Antígeno/genética , Apoptose/imunologia , Caspase 8/metabolismo , Linhagem Celular , Proliferação de Células , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Células Clonais , Citotoxicidade Imunológica/genética , Citotoxicidade Imunológica/imunologia , Ativação Enzimática/genética , Ativação Enzimática/imunologia , Proteína Ligante Fas/toxicidade , Humanos , Ativação Linfocitária/genética , Vírus da Coriomeningite Linfocítica/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos TransgênicosRESUMO
NF-kappaB transcription factors have key roles in inflammation, immune response, oncogenesis and protection against apoptosis. In most cells, these factors are kept inactive in the cytoplasm through association with IkappaB inhibitors. After stimulation by various reagents, IkappaB is phosphorylated by the IkappaB kinase (IKK) complex and degraded by the proteasome, allowing NF-kappaB to translocate to the nucleus and activate its target genes. Here we report that CYLD, a tumour suppressor that is mutated in familial cylindromatosis, interacts with NEMO, the regulatory subunit of IKK. CYLD also interacts directly with tumour-necrosis factor receptor (TNFR)-associated factor 2 (TRAF2), an adaptor molecule involved in signalling by members of the family of TNF/nerve growth factor receptors. CYLD has deubiquitinating activity that is directed towards non-K48-linked polyubiquitin chains, and negatively modulates TRAF-mediated activation of IKK, strengthening the notion that ubiquitination is involved in IKK activation by TRAFs and suggesting that CYLD functions in this process. Truncations of CYLD found in cylindromatosis result in reduced enzymatic activity, indicating a link between impaired deubiquitination of CYLD substrates and human pathophysiology.
Assuntos
NF-kappa B/antagonistas & inibidores , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina/metabolismo , Sequência de Aminoácidos , Catálise , Linhagem Celular , Enzima Desubiquitinante CYLD , Humanos , Quinase I-kappa B , Dados de Sequência Molecular , Mutação , NF-kappa B/metabolismo , Ligação Proteica , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Especificidade por Substrato , Fator 2 Associado a Receptor de TNF , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/genéticaRESUMO
Deletion of the Casp8 gene in epithelial tissues of mice results in severe inflammatory pathologies. Its ubiquitous deletion, or its specific deletion in endothelial cells, results in intrauterine death associated with capillary damage. These pathologies are all preventable by co-deletion of Casp8 and the genes encoding either the RIPK1 or the RIPK3 protein kinase. Since activation of RIPK3 in Caspase-8-deficient cells can trigger necroptotic cell death, and since RIPK1 can activate RIPK3, it is widely assumed that the inflammatory states resulting from Caspase-8 deficiency occur as a consequence of RIPK3-induced necroptosis. Here, we report that although on a Ripk3-null background Casp8 deletion in mice does not result in outright pathological changes, it triggers enhanced expression of a variety of inflammatory genes in utero, which gradually subsides after birth. Deletion of Ripk1, or even of only one of its two alleles, obliterates this activation. Resembling the embryonic pathology observed in RIPK3-expressing cells, the activation of inflammatory genes observed on a Ripk3-null background seems to be initiated in endothelial cells. Analysis of endothelial cells isolated from livers of Caspase-8-deficient embryos revealed neither an increase in the amount of RIPK1 in these cells after Casp8 deletion, nor triggering of RIPK1 phosphorylation. These findings indicate that the triggering of inflammation by Casp8 deletion in mice occurs, in part, independently of necroptosis or other functions of RIPK3, and rather reflects enhanced RIPK1-dependent signaling for activation of inflammatory genes.
Assuntos
Caspase 8/metabolismo , Embrião de Mamíferos/enzimologia , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Ativação Transcricional , Animais , Caspase 8/genética , Inflamação/embriologia , Inflamação/genética , Camundongos , Camundongos Knockout , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Transdução de Sinais/genéticaAssuntos
Anti-Inflamatórios/metabolismo , Caspase 8/metabolismo , Animais , RNA Helicases DEAD-box/metabolismo , Epiderme/enzimologia , Epiderme/patologia , Humanos , Fator Regulador 3 de Interferon/metabolismo , Camundongos , Fatores de Crescimento Neural/metabolismo , Transdução de Sinais , Fatores de Necrose Tumoral/metabolismoRESUMO
Cells dying by apoptosis can trigger an anti-inflammatory gene response in other cells by releasing a compound called adenosine monophosphate.
Assuntos
Monofosfato de Adenosina/fisiologia , Apoptose/fisiologia , Tolerância Imunológica/fisiologia , AnimaisRESUMO
Emerging evidence indicates that the molecular mechanisms of cell death have regulatory roles in inflammation and that the molecular changes that are associated with different forms of cell death affect the course of inflammation in different ways. In this Timeline article, we discuss how our understanding of the mechanisms and functional roles of tissue injury and cell death in inflammation has evolved on the basis of almost two centuries of study. We describe how such ideas have led to our current models of cell death and inflammation, and we highlight the remaining gaps in our knowledge of the subject.
Assuntos
Inflamação/história , Animais , Morte Celular/imunologia , História do Século XIX , História do Século XX , História do Século XXI , Humanos , Inflamação/imunologia , Inflamação/patologia , Modelos Imunológicos , Transdução de SinaisRESUMO
Emerging evidence indicates that necrotic cell death can be regulated by a specific set of signaling molecules. Studies showing that the same signaling molecules also trigger inflammation, and that when cells die necrotically some of the molecules they release facilitate inflammation, raised the possibility that the death induced by these signaling molecules ("necroptosis") serves to trigger inflammation. Here we briefly discuss the work done on the anti-inflammatory function of caspase-8 and its relation to the inhibitory effect of this enzyme on the induction of necroptosis. The studies imply that caspase-8 and the other proximal signaling proteins known to participate in the induction and regulation of necroptosis are too pleiotropic to serve as reliable molecular probes for determining the relative contribution of this death mode to in vivo processes.
Assuntos
Apoptose/imunologia , Caspase 8/imunologia , Necrose/imunologia , Animais , Anti-Inflamatórios/imunologia , Caspase 8/genética , Humanos , Inflamação/genética , Inflamação/imunologia , Camundongos , Transdução de Sinais/imunologiaRESUMO
Necroptosis-a form of programmed necrotic cell death-and its resulting release of damage-associated molecular patterns (DAMPs) are believed to participate in the triggering of inflammatory processes. To assess the relative contribution of this cell death mode to inflammation, we need to know what other cellular effects can be exerted by molecules shown to trigger necrotic death, and the extent to which those effects might themselves contribute to inflammation. Here, we describe the technical approaches that have been applied to assess the impact of the main signaling molecules known to mediate activation of necroptosis upon generation of inflammatory cytokines in LPS-treated mouse bone marrow-derived dendritic cells. The findings obtained by this assessment indicated that signaling molecules known to initiate necroptosis can also initiate activation of the NLRP3 inflammasome, thereby inducing inflammation independently of cell death by triggering the generation of proinflammatory cytokines such as IL-1ß.