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1.
PLoS Biol ; 18(8): e3000807, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32760056

RESUMO

Radiotherapy is a commonly used conditioning regimen for bone marrow transplantation (BMT). Cytotoxicity limits the use of this life-saving therapy, but the underlying mechanisms remain poorly defined. Here, we use the syngeneic mouse BMT model to test the hypothesis that lethal radiation damages tissues, thereby unleashing signals that indiscriminately activate the inflammasome pathways in host and transplanted cells. We find that a clinically relevant high dose of radiation causes severe damage to bones and the spleen through mechanisms involving the NLRP3 and AIM2 inflammasomes but not the NLRC4 inflammasome. Downstream, we demonstrate that gasdermin D (GSDMD), the common effector of the inflammasomes, is also activated by radiation. Remarkably, protection against the injury induced by deadly ionizing radiation occurs only when NLRP3, AIM2, or GSDMD is lost simultaneously in both the donor and host cell compartments. Thus, this study reveals a continuum of the actions of lethal radiation relayed by the inflammasome-GSDMD axis, initially affecting recipient cells and ultimately harming transplanted cells as they grow in the severely injured and toxic environment. This study also suggests that therapeutic targeting of inflammasome-GSDMD signaling has the potential to prevent the collateral effects of intense radiation regimens.


Assuntos
Células da Medula Óssea/efeitos da radiação , Transplante de Medula Óssea , Proteínas de Ligação a DNA/genética , Inflamassomos/efeitos da radiação , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteínas de Ligação a Fosfato/genética , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Proteínas de Ligação a DNA/deficiência , Feminino , Fêmur/citologia , Fêmur/metabolismo , Regulação da Expressão Gênica , Inflamassomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR/deficiência , Proteínas de Ligação a Fosfato/deficiência , Piroptose/genética , Piroptose/efeitos da radiação , Transdução de Sinais , Baço/metabolismo , Baço/patologia , Baço/efeitos da radiação , Transplante Isogênico , Irradiação Corporal Total , Raios X
2.
Leukemia ; 34(7): 1726-1729, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32483300

RESUMO

The scientific community faces an unexpected and urgent challenge related to the SARS-CoV-2 pandemic and is investigating the role of receptors involved in entry of this virus into cells as well as pathomechanisms leading to a cytokine "storm," which in many cases ends in severe acute respiratory syndrome, fulminant myocarditis and kidney injury. An important question is if it may also damage hematopoietic stem progenitor cells?


Assuntos
Infecções por Coronavirus/epidemiologia , Síndrome da Liberação de Citocina/epidemiologia , Células-Tronco Hematopoéticas/virologia , Inflamassomos/imunologia , Pandemias , Pneumonia Viral/epidemiologia , Síndrome Respiratória Aguda Grave/epidemiologia , Lesão Renal Aguda/epidemiologia , Lesão Renal Aguda/imunologia , Lesão Renal Aguda/prevenção & controle , Lesão Renal Aguda/virologia , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/imunologia , Betacoronavirus/patogenicidade , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Síndrome da Liberação de Citocina/imunologia , Síndrome da Liberação de Citocina/prevenção & controle , Síndrome da Liberação de Citocina/virologia , Citocinas/antagonistas & inibidores , Citocinas/genética , Citocinas/imunologia , Furanos/farmacologia , Regulação da Expressão Gênica , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/imunologia , Humanos , Imunidade Inata/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Inflamassomos/antagonistas & inibidores , Inflamassomos/genética , Miocardite/epidemiologia , Miocardite/imunologia , Miocardite/prevenção & controle , Miocardite/virologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/imunologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Piroptose/efeitos dos fármacos , Piroptose/genética , Piroptose/imunologia , Fatores de Risco , Síndrome Respiratória Aguda Grave/imunologia , Síndrome Respiratória Aguda Grave/prevenção & controle , Síndrome Respiratória Aguda Grave/virologia , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Sulfonamidas/farmacologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/virologia
3.
Mol Med Rep ; 22(1): 219-226, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32468047

RESUMO

Bladder outlet obstruction (BOO), which is primarily caused by benign prostatic hyperplasia, is a common chronic disease. However, previous studies have most commonly investigated BOO using the acute obstruction model. In the present study, a chronic obstruction model was established to investigate the different pathological alterations in the bladder between acute and chronic obstruction. Compared with chronic obstruction, acute obstruction led to increased expression of proliferating cell nuclear antigen and interleukin­1ß, which are markers of proliferation and inflammation, respectively. Furthermore, increased fibrosis in the bladder at week 2 was observed. Low pressure promoted mice bladder smooth muscle cell (MBSMC) proliferation, and pressure overload inhibited cell proliferation and increased the proportion of dead MBSMCs. Further investigation using serum/glucocorticoid regulated kinase 1 (SGK1) small interfering RNAs indicated that low pressure may promote MBSMC proliferation by upregulating SGK1 and nuclear factor of activated T­cell expression levels. Therefore, the present study suggested that acute obstruction led to faster decompensation of bladder function and chronic bladder obstruction displayed an enhanced ability to progress to BOO.


Assuntos
Proteínas Imediatamente Precoces/metabolismo , Interleucina-1beta/metabolismo , Fatores de Transcrição NFATC/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Piroptose , Obstrução do Colo da Bexiga Urinária/metabolismo , Doença Aguda , Animais , Morte Celular , Proliferação de Células/genética , Doença Crônica , Colágeno/metabolismo , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Miócitos de Músculo Liso/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo , Piroptose/genética , RNA Interferente Pequeno , Transdução de Sinais/genética , Bexiga Urinária/citologia , Bexiga Urinária/patologia , Obstrução do Colo da Bexiga Urinária/genética , Obstrução do Colo da Bexiga Urinária/patologia , Urodinâmica/genética , Urodinâmica/fisiologia
4.
Nat Commun ; 11(1): 2212, 2020 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-32371889

RESUMO

Gasdermin-D (GSDMD) in inflammasome-activated macrophages is cleaved by caspase-1 to generate N-GSDMD fragments. N-GSDMD then oligomerizes in the plasma membrane (PM) to form pores that increase membrane permeability, leading to pyroptosis and IL-1ß release. In contrast, we report that although N-GSDMD is required for IL-1ß secretion in NLRP3-activated human and murine neutrophils, N-GSDMD does not localize to the PM or increase PM permeability or pyroptosis. Instead, biochemical and microscopy studies reveal that N-GSDMD in neutrophils predominantly associates with azurophilic granules and LC3+ autophagosomes. N-GSDMD trafficking to azurophilic granules causes leakage of neutrophil elastase into the cytosol, resulting in secondary cleavage of GSDMD to an alternatively cleaved N-GSDMD product. Genetic analyses using ATG7-deficient cells indicate that neutrophils secrete IL-1ß via an autophagy-dependent mechanism. These findings reveal fundamental differences in GSDMD trafficking between neutrophils and macrophages that underlie neutrophil-specific functions during inflammasome activation.


Assuntos
Membrana Celular/metabolismo , Interleucina-1beta/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neutrófilos/metabolismo , Organelas/metabolismo , Proteínas de Ligação a Fosfato/metabolismo , Animais , Autofagossomos/metabolismo , Autofagia/genética , Caspase 1/metabolismo , Permeabilidade da Membrana Celular/genética , Humanos , Inflamassomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Elastase de Leucócito/genética , Elastase de Leucócito/metabolismo , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas de Ligação a Fosfato/genética , Transporte Proteico , Piroptose/genética
5.
Oxid Med Cell Longev ; 2020: 8351342, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32190178

RESUMO

Endothelial cell damage caused by oxidative stress is widely considered to be a triggering event in atherosclerosis (AS). However, the specific effect elicited by autophagy in endothelial cells undergoing oxidative stress remains controversial, especially during end-stage autophagy. The inhibition of end-stage autophagy has been reported to increase cell pyroptosis and contribute to endothelial damage. Several studies have shown that microRNA-103 is involved in end-stage autophagy; however, its specific mechanism of action is not yet characterized. In this study, we addressed the regulatory role of miR-103 in autophagy during oxidative stress of endothelial cells. Hydrogen peroxide (H2O2) treatment was used as an in vitro model of oxidative stress. MTS and ROS levels were measured to evaluate cell activity. qRT-PCR was used to detect the expression of miR-103. Autophagy was examined using western blot, immunofluorescence staining, and electron microscopy, while western blot analysis detected pyroptosis-related proteins. Results show that miR-103 expression decreased under oxidative stress. Further, miR-103 repressed transcription of Bcl-2/adenovirus E1B 19 kDa interacting protein (BNIP3). The oxidative stress caused by H2O2 caused cell damage from 2 hours (P < 0.05) and increased the level of intracellular reactive oxygen species (P < 0.05); at the same time, the damage could be further aggravated by the stimulation of bafA1 (P < 0.05). Under the stimulation of H2O2, the expression of miR-103 decreased (P < 0.05). However, high expression of miR-103 could reduce the accumulation of LC3II and P62 (P < 0.05) by inhibiting the downstream target gene Bcl-2/adenovirus E1B 19 kDa interacting protein (BNIP3), thus reducing the occurrence of cell pyroptosis (P < 0.05). This process could be blocked by end-stage autophagy inhibitor bafA1 (P < 0.05), which further indicated that miR-103 affected cell injury by autophagy. On the contrary, the low expression of miR-103 promoted the accumulation of autophagy protein and increased the occurrence of pyroptosis (P < 0.05). In conclusion, inhibition of miR-103 restrained end-stage of autophagy by regulating BNIP3, thus changing the occurrence of cell pyroptosis.


Assuntos
Autofagia/genética , Vasos Coronários/patologia , Células Endoteliais/patologia , Peróxido de Hidrogênio/toxicidade , Proteínas de Membrana/metabolismo , MicroRNAs/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Proteínas Proto-Oncogênicas/metabolismo , Piroptose/genética , Autofagia/efeitos dos fármacos , Citoproteção/efeitos dos fármacos , Citoproteção/genética , Regulação para Baixo/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Humanos , MicroRNAs/genética , Estresse Oxidativo/genética , Piroptose/efeitos dos fármacos
6.
Curr Opin Cell Biol ; 63: 186-193, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32163825

RESUMO

Necroptosis and pyroptosis are inflammatory forms of regulated necrotic cell death as opposed to apoptosis that is generally considered immunologically silent. Recent studies revealed unexpected links in the pathways regulating and executing cell death in response to activation of signaling cascades inducing apoptosis, necroptosis, and pyroptosis. Emerging evidence suggests that receptor interacting protein kinase 1 and caspase-8 control the cross-talk between apoptosis, necroptosis, and pyroptosis and determine the type of cell death induced in response to activation of cell death signaling.


Assuntos
Apoptose/genética , Caspase 8/fisiologia , Necroptose/genética , Piroptose/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/fisiologia , Animais , Caspase 8/genética , Caspase 8/metabolismo , Humanos , Necrose/genética , Necrose/patologia , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Transdução de Sinais/genética
7.
Biol Pharm Bull ; 43(3): 558-564, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32115515

RESUMO

Diabetic kidney disease (DKD) is the leading cause of chronic kidney disease. Current therapies for DKD are insufficient. Therefore, there is an urgent need for identifying new therapies. An increasing number of micro RNAs (miRNAs) and long noncoding RNAs (lncRNAs) have been demonstrated to modulate the progression of diabetic kidney disease. Nevertheless, until now, there have been few reports evaluating the relevance of circular RNAs (circRNAs) in DKD. circRNAs have been reported to regulate the occurrence and development of multiple diseases. In this study, we intended to explore the circRNA expression profiles and determine the role of circRNA in DKD. We identified a series of dysregulated circRNAs in glucose-stressed HK-2 cells using circRNA microarray analysis. Among the candidate circRNAs, we found that circACTR2 was upregulated and may be involved in inflammation and pyroptosis. Knockdown of circACTR2 significantly decreased pyroptosis, interleukin (IL)-1ß release and collagen IV and fibronectin production, indicating the effective regulation by circACTR2 of cell death and inflammation. Overall, our study identified a new circRNA, circACTR2, that regulates high glucose-induced pyroptosis, inflammation and fibrosis in proximal tubular cells. The present study preliminarily explores the role of circRNAs in pyroptosis of tubular cells, and provides novel insight into the pathogenesis of DKD and new therapeutic strategies.


Assuntos
Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Glucose/efeitos adversos , Piroptose/genética , Piroptose/fisiologia , RNA Circular/deficiência , RNA Circular/genética , Proteína 2 Relacionada a Actina , Linhagem Celular , Colágeno/metabolismo , Células Epiteliais , Fibronectinas/metabolismo , Fibrose/genética , Fibrose/metabolismo , Expressão Gênica , Humanos , Interleucina-1beta/metabolismo , Rim/metabolismo
8.
Cell Death Dis ; 11(1): 76, 2020 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-32001670

RESUMO

Only a few types of inflammasomes have been described in central nervous system cells. Among these, the absent in melanoma 2 (AIM2) inflammasome is primarily found in neurons, is highly specific and can be activated only by double-stranded DNA. Although it has been demonstrated that the AIM2 inflammasome is activated by poly(deoxyadenylic-deoxythymidylic) acid sodium salt and leads to pyroptotic neuronal cell death, the role of AIM2 inflammasome-mediated pyroptosis in early brain injury (EBI) after subarachnoid haemorrhage (SAH) has rarely been studied. Thus, we designed this study to explore the mechanism of gasdermin D(GSDMD)-induced pyroptosis mediated by the AIM2 inflammasome in EBI after SAH. The level of AIM2 from the cerebrospinal fluid (CSF) of patients with SAH was detected. The pathway of AIM2 inflammasome-mediated pyroptosis, the AIM2/Caspase-1/GSDMD pathway, was explored after experimental SAH in vivo and in primary cortical neurons stimulated by oxyhaemoglobin (oxyHb) in vitro. Then, we evaluated GSDMD-induced pyroptosis mediated by the AIM2 inflammasome in AIM2 and caspase-1- deficient mice and primary cortical neurons generated through lentivirus (LV) knockdown. Compared with that of the control samples, the AIM2 level in the CSF of the patients with SAH was significantly increased. Pyroptosis-associated proteins mediated by the AIM2 inflammasome were significantly increased in vivo and in vitro following experimentally induced SAH. After AIM2 and caspase-1 were knocked down by an LV, GSDMD-induced pyroptosis mediated by the AIM2 inflammasome was alleviated in EBI after SAH. Intriguingly, when caspase-1 was knocked down, apoptosis was significantly suppressed via impeding the activation of caspase-3. GSDMD-induced pyroptosis mediated by the AIM2 inflammasome may be involved in EBI following SAH. The inhibition of AIM2 inflammasome activation caused by knocking down AIM2 and caspase-1 alleviates GSDMD-induced pyroptosis in EBI after SAH.


Assuntos
Lesões Encefálicas/metabolismo , Proteínas de Ligação a DNA/líquido cefalorraquidiano , Inflamassomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neurônios/metabolismo , Proteínas de Ligação a Fosfato/metabolismo , Piroptose/genética , Hemorragia Subaracnóidea/metabolismo , Animais , Lesões Encefálicas/genética , Caspase 1/genética , Caspase 1/metabolismo , Caspase 3/metabolismo , Células Cultivadas , Proteínas de Ligação a DNA/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Varredura , Neurônios/ultraestrutura , Proteínas de Ligação a Fosfato/genética , Piroptose/fisiologia , Hemorragia Subaracnóidea/genética
9.
Cell Death Dis ; 11(1): 78, 2020 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-32001671

RESUMO

Preeclampsia (PE) remains a leading cause of maternal and neonatal morbidity and mortality. Numerous studies have shown that women with PE develop autoantibody, termed angiotensin II type 1 receptor autoantibody (AT1-AA), and key features of the disease result from it. Emerging evidence has indicated that inflammatory cell necrosis, such as pyroptosis, could lead to autoantigen exposure and stimulate autoantibody production. Caspase-1, the central enzyme of inflammasome and key target of pyroptosis, may play roles in AT1R exposure and AT1-AA production. Exploring endogenous regulator that could inhibit AT1-AA production by targeting pyroptosis will be essential for treating PE. Lipoxin A4 (LXA4), endogenous dual anti-inflammatory and proresolving lipid mediator, may inhibit AT1-AA production via modulating caspase-1. Thus, we explore whether caspase-1 is essential for AT1-AA production and LXA4 inhibits AT1-AA via modulating caspase-1. PE patients and mice developed AT1-AA associated with caspase-1 activation. Caspase-1 deletion leaded to AT1-AA decrease in PE mice. Consistent with these findings, we confirmed caspase-1 activation, trophoblast pyroptosis and AT1R exposure in PE mice and trophoblast model, while caspase-1 deficiency showed decreased trophoblast pyroptosis and AT1R exposure in vitro and in vivo. Interestingly, LXA4 could suppress AT1-AA production via regulating caspase-1 as well as enhancing phagocytosis of dead trophoblasts by macrophages. These results suggest that caspase-1 promotes AT1-AA production via inducing trophoblast pyroptosis and AT1R exposure, while LXA4 suppresses AT1-AA production via modulating caspase-1, supporting caspase-1 serving as a therapeutic target for attenuating AT1-AA and LXA4 protecting patients from AT1-AA and PE.


Assuntos
Autoanticorpos/metabolismo , Caspase 1/metabolismo , Lipoxinas/farmacologia , Pré-Eclâmpsia/imunologia , Piroptose/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/imunologia , Trofoblastos/efeitos dos fármacos , Antagonistas de Receptores de Angiotensina , Animais , Autoanticorpos/genética , Autoanticorpos/imunologia , Caspase 1/sangue , Caspase 1/deficiência , Caspase 1/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Células Cultivadas , Citocinas/metabolismo , Feminino , Humanos , Lipopolissacarídeos/farmacologia , Lipoxinas/sangue , Lipoxinas/deficiência , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pré-Eclâmpsia/sangue , Pré-Eclâmpsia/metabolismo , Gravidez , Piroptose/genética , Piroptose/imunologia , RNA Interferente Pequeno , Receptor Tipo 1 de Angiotensina/sangue , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/metabolismo , Baço/crescimento & desenvolvimento , Baço/imunologia , Baço/patologia , Trofoblastos/metabolismo
10.
Sci Rep ; 10(1): 2263, 2020 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-32041990

RESUMO

Activation of the inflammasome is involved in the progression of retinal degenerative diseases, in particular, in the pathogenesis of Age-Related Macular Degeneration (AMD), with NLRP3 activation the focus of many investigations. In this study, we used genetic and pharmacological approaches to explore the role of the inflammasome in a mouse model of retinal degeneration. We identify that Casp1/11-/- mice have better-preserved retinal function, reduced inflammation and increased photoreceptor survivability. While Nlrp3-/- mice display some level of preservation of retinal function compared to controls, pharmacological inhibition of NLRP3 did not protect against photoreceptor cell death. Further, Aim2-/-, Nlrc4-/-, Asc-/-, and Casp11-/- mice show no substantial retinal protection. We propose that CASP-1-associated photoreceptor cell death occurs largely independently of NLRP3 and other established inflammasome sensor proteins, or that inhibition of a single sensor is not sufficient to repress the inflammatory cascade. Therapeutic targeting of CASP-1 may offer a more promising avenue to delay the progression of retinal degenerations.


Assuntos
Caspase 1/metabolismo , Inflamassomos/imunologia , Degeneração Macular/imunologia , Células Fotorreceptoras/patologia , Piroptose/imunologia , Animais , Caspase 1/genética , Caspases Iniciadoras/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Células Cultivadas , Modelos Animais de Doenças , Progressão da Doença , Feminino , Compostos Heterocíclicos de 4 ou mais Anéis/administração & dosagem , Humanos , Inflamassomos/antagonistas & inibidores , Inflamassomos/metabolismo , Injeções Intravítreas , Luz/efeitos adversos , Degeneração Macular/tratamento farmacológico , Degeneração Macular/patologia , Masculino , Camundongos , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Estresse Oxidativo/imunologia , Estresse Oxidativo/efeitos da radiação , Células Fotorreceptoras/imunologia , Piroptose/efeitos dos fármacos , Piroptose/genética , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/imunologia , Epitélio Pigmentado da Retina/patologia , Sulfonas/administração & dosagem
11.
Cell ; 180(5): 941-955.e20, 2020 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-32109412

RESUMO

The pyroptosis execution protein GSDMD is cleaved by inflammasome-activated caspase-1 and LPS-activated caspase-11/4/5. The cleavage unmasks the pore-forming domain from GSDMD-C-terminal domain. How the caspases recognize GSDMD and its connection with caspase activation are unknown. Here, we show site-specific caspase-4/11 autoprocessing, generating a p10 product, is required and sufficient for cleaving GSDMD and inducing pyroptosis. The p10-form autoprocessed caspase-4/11 binds the GSDMD-C domain with a high affinity. Structural comparison of autoprocessed and unprocessed capase-11 identifies a ß sheet induced by the autoprocessing. In caspase-4/11-GSDMD-C complex crystal structures, the ß sheet organizes a hydrophobic GSDMD-binding interface that is only possible for p10-form caspase-4/11. The binding promotes dimerization-mediated caspase activation, rendering a cleavage independently of the cleavage-site tetrapeptide sequence. Crystal structure of caspase-1-GSDMD-C complex shows a similar GSDMD-recognition mode. Our study reveals an unprecedented substrate-targeting mechanism for caspases. The hydrophobic interface suggests an additional space for developing inhibitors specific for pyroptotic caspases.


Assuntos
Inflamassomos/ultraestrutura , Complexos Multiproteicos/ultraestrutura , Proteínas de Ligação a Fosfato/ultraestrutura , Piroptose/genética , Animais , Caspase 1/química , Caspase 1/genética , Caspase 1/ultraestrutura , Caspases Iniciadoras/química , Caspases Iniciadoras/genética , Cristalografia por Raios X , Células HEK293 , Células HeLa , Humanos , Interações Hidrofóbicas e Hidrofílicas , Inflamassomos/genética , Peptídeos e Proteínas de Sinalização Intracelular/química , Peptídeos e Proteínas de Sinalização Intracelular/genética , Complexos Multiproteicos/química , Complexos Multiproteicos/genética , Proteínas de Ligação a Fosfato/química , Proteínas de Ligação a Fosfato/genética , Conformação Proteica em Folha beta/genética , Domínios Proteicos/genética , Processamento de Proteína Pós-Traducional/genética , Proteólise
12.
Mol Med Rep ; 21(1): 360-370, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31939622

RESUMO

The gasdermin (GSDM) superfamily has been demonstrated to consist of several important molecules that modulate multifunctional signal processes, such as cell pyroptosis. In this research, the roles of the GSDM superfamily on the occurrence and prognosis of lung adenocarcinoma (LUAD) were evaluated using integrative bioinformatic analyses and in vitro methods. Here, data from several bioinformatic platforms revealed that GSDMC is significantly upregulated in LUAD tissues and cell lines. Real­time fluorescence quantitative PCR (qPCR) demonstrated that GSDMC was obviously upregulated in radio­resistant LUAD cells, compared with their parental cells. Moreover, upregulated GSDMC expression was confirmed to be an independent indicator of poor first progression (FP) and overall survival (OS) in LUAD patients. DNA methylation analysis showed an evidently negative correlation between GSDMC expression and methylation status of one CpG site (cg05316065) in its DNA sequence. Patients with high methylation values had significantly higher Karnofsky performance scores (KPSs) and prolonged OS rates. Together, we confirmed that overexpression of GSDMC acts as a promising predictive factor for the poor prognosis of LUAD patients.


Assuntos
Adenocarcinoma de Pulmão/metabolismo , Biomarcadores Tumorais/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Neoplasias Pulmonares/metabolismo , Células A549 , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Adenocarcinoma de Pulmão/patologia , Biomarcadores Tumorais/genética , Biologia Computacional , Metilação de DNA , Proteínas de Ligação a DNA/genética , Bases de Dados Genéticas , Ontologia Genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Prognóstico , Piroptose/genética , Regulação para Cima
13.
Am J Physiol Cell Physiol ; 318(2): C346-C359, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31693400

RESUMO

Diabetic corneal endothelial keratopathy is an intractable ocular complication characterized by corneal edema and endothelial decompensation, which seriously threaten vision. It has been suggested that diabetes is associated with pyroptosis, a type of programmed cell death via the activation of inflammation. Long noncoding RNA KCNQ1OT1 is commonly associated with various pathophysiological mechanisms of diabetic complications, including diabetic cardiomyopathy and diabetic retinopathy. However, whether KCNQ1OT1 is capable of regulating pyroptosis and participates in the pathogenesis of diabetic corneal endothelial keratopathy remains unknown. The aim of this study was to investigate the mechanisms of KCNQ1OT1 in diabetic corneal endothelial keratopathy. Here, we reveal that KCNQ1OT1 and pyroptosis can be triggered in diabetic human and rat corneal endothelium, along with the high glucose-treated corneal endothelial cells. However, miR-214 expression was substantially decreased in vivo and in experiments with cultured cells. LDH assay was also used to verify the existence of pyroptosis in high glucose-treated cells. Bioinformatics prediction and luciferase assays showed that KCNQ1OT1 may function as a competing endogenous RNA binding miR-214 to regulate the expression of caspase-1. To further analyze the KCNQ1OT1-mediated mechanism, miR-214 mimic and inhibitor were introduced into the high glucose-treated corneal endothelial cells. The results showed that upregulation of miR-214 attenuated pyroptosis; conversely, knockdown of miR-214 promoted it. In addition, KCNQ1OT1 knockdown by a small interfering RNA decreased pyroptosis factors expressions but enhanced miR-214 expression in corneal endothelial cells. To understand the signaling mechanisms underlying the prepyroptotic properties of KCNQ1OT1, si-KCNQ1OT1 was cotransfected with or without miR-214 inhibitor. The results showed that pyroptosis was repressed after silencing KCNQ1OT1 but was reversed by cotransfection with miR-214 inhibitor, suggesting that KCNQ1OT1 mediated pyroptosis induced by high glucose via targeting miR-214. Therefore, the KCNQ1OT1/miR-214/caspase-1 signaling pathway represents a new mechanism of diabetic corneal endothelial keratopathy progression, and KCNQ1OT1 could potentially be a novel therapeutic target.


Assuntos
Complicações do Diabetes/genética , Cardiomiopatias Diabéticas/genética , Células Endoteliais/metabolismo , Epitélio Posterior/metabolismo , Piroptose/genética , Animais , Estudos de Casos e Controles , Caspase 1/genética , Linhagem Celular , Complicações do Diabetes/metabolismo , Diabetes Mellitus/genética , Diabetes Mellitus/metabolismo , Cardiomiopatias Diabéticas/metabolismo , Feminino , Humanos , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , RNA Longo não Codificante/genética , RNA Interferente Pequeno/genética , Ratos , Ratos Sprague-Dawley , Regulação para Cima/genética
14.
Life Sci ; 240: 117138, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31809715

RESUMO

Pyroptosis is a form of cell death mediated by gasdermin D (GSDMD); it is characterised by NLRP3 inflammasome activation, caspase activation, cell membrane pore formation, and the release of interleukin-1ß and interleukin-18. NLRP3 inflammasome activation plays a central role in pyroptosis. Recent research has suggested that NLRP3 inflammasome activation may be involved in the occurrence and development of diabetes mellitus and its associated complications. This finding provided the impetus for us to clarify the significance of pyroptosis in diabetes. In this review, we summarise the current understanding of the molecular mechanisms involved in pyroptosis, as well as recent advances in the role of NLRP3 inflammasome activation and pyroptosis in the development of diabetes and diabetic complications.


Assuntos
Complicações do Diabetes/tratamento farmacológico , Complicações do Diabetes/genética , Diabetes Mellitus/tratamento farmacológico , Diabetes Mellitus/genética , Inflamassomos/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Piroptose/genética , Animais , Complicações do Diabetes/patologia , Diabetes Mellitus/patologia , Humanos , Inflamassomos/efeitos dos fármacos , Piroptose/efeitos dos fármacos
15.
Biochem Pharmacol ; 176: 113779, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-31881190

RESUMO

Previous studies from this lab and others have demonstrated that nitric oxide (NO) in a concentration dependent manner, modulated neutrophil and leukemic cell survival. Subsequent studies delineated importance of iNOS in neutrophil differentiation and leukemic cell death. On the contrary, role of nNOS in survival of these cells remains least understood. Present study was therefore undertaken to assess and compare the role of iNOS and nNOS in the survival of NOS overexpressing myelocytic K562 cells. Cells with almost similar iNOS and nNOS activities displayed comparable cell cycle perturbation, Annexin V positivity, mitochondrial dysfunction, augmented DCF fluorescence, and also attenuated expression of antioxidants. Moreover, induction in cell death was also accompanied by the activation of pJNK/p38MAPK/Erk1/2 and reduction in PI3K/Akt/mTOR signaling. Treatment of NOS isoform overexpressing K562 cells with NAC, a potent free radical scavenger prevented cell death and also the modulations in the signaling proteins. In addition, enhanced expression of CASP1 and CASP4 genes, along with increased Caspase-1 cleavage and increased IL-1ß release were significantly more in K562iNOS cells, which indicate priming of these cells for pyroptotic cell death. On the other hand, K562nNOS cells, displayed much enhanced CASP3 gene expression, Caspase-3 cleavage and Caspase-3 activity. Results obtained indicate that similar level of iNOS or nNOS activation in K562 cells, preferred pyroptotic and apoptotic cell death respectively.


Assuntos
Apoptose/fisiologia , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Piroptose/fisiologia , Acetilcisteína/farmacologia , Apoptose/genética , Caspase 1/genética , Caspase 1/metabolismo , Caspase 3/genética , Caspase 3/metabolismo , Ciclo Celular/genética , Ciclo Celular/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Depuradores de Radicais Livres/farmacologia , Humanos , Células K562 , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo I/genética , Óxido Nítrico Sintase Tipo II/genética , Piroptose/genética , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética
16.
Cell Death Dis ; 10(12): 927, 2019 12 05.
Artigo em Inglês | MEDLINE | ID: mdl-31804457

RESUMO

Systemic manifestation of preeclampsia (PE) is associated with circulating factors, including inflammatory cytokines and damage-associated molecular patterns (DAMPs), or alarmins. However, it is unclear whether the placenta directly contributes to the increased levels of these inflammatory triggers. Here, we demonstrate that pyroptosis, a unique inflammatory cell death pathway, occurs in the placenta predominantly from early onset PE, as evidenced by elevated levels of active caspase-1 and its substrate or cleaved products, gasdermin D (GSDMD), IL-1ß, and IL-18. Using cellular models mimicking pathophysiological conditions (e.g., autophagy deficiency, hypoxia, and endoplasmic reticulum (ER) stress), we observed that pyroptosis could be induced in autophagy-deficient human trophoblasts treated with sera from PE patients as well as in primary human trophoblasts exposed to hypoxia. Exposure to hypoxia elicits excessive unfolded protein response (UPR) and ER stress and activation of the NOD-like receptor pyrin-containing 3 (NLRP3) inflammasome in primary human trophoblasts. Thioredoxin-interacting protein (TXNIP), a marker for hyperactivated UPR and a crucial signaling molecule linked to NLRP3 inflammasome activation, is significantly increased in hypoxia-treated trophoblasts. No evidence was observed for necroptosis-associated events. Importantly, these molecular events in hypoxia-treated human trophoblasts are significantly observed in placental tissue from women with early onset PE. Taken together, we propose that placental pyroptosis is a key event that induces the release of factors into maternal circulation that possibly contribute to severe sterile inflammation and early onset PE pathology.


Assuntos
Proteínas de Transporte/genética , Inflamação/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Pré-Eclâmpsia/genética , Adulto , Hipóxia Celular/genética , Estresse do Retículo Endoplasmático/genética , Feminino , Humanos , Inflamação/metabolismo , Inflamação/patologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Ligação a Fosfato/genética , Placenta/metabolismo , Placenta/patologia , Pré-Eclâmpsia/metabolismo , Pré-Eclâmpsia/patologia , Gravidez , Piroptose/genética , Transdução de Sinais , Trofoblastos/metabolismo , Trofoblastos/patologia , Resposta a Proteínas não Dobradas/genética
17.
Oxid Med Cell Longev ; 2019: 3753485, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31814872

RESUMO

Background: The neuronal injury and cognitive dysfunction after liver transplantation have severe effects on the prognosis and life quality of patients. Accumulating evidence suggests that both exosomes and pyroptosis could participate in hepatic ischemia-reperfusion injury (HIRI) and play key roles in neuronal death. However, the link between exosomes and neuronal pyroptosis in HIRI awaits further investigation. Methods: After establishing the HIRI rat models, we primarily studied the role of pyroptosis in hippocampal and cortical neuron injury through detecting NOD-like receptor protein 3 (NLRP3), pro-caspase-1, cleaved-caspase-1, apoptosis-associated speck-like protein containing CARD (ASC), gasdermin D (GSDMD), interleukin-1beta (IL-1ß), and interleukin-18 (IL-18) expressions with western blotting, immunohistochemical staining, and enzyme-linked immunosorbent assay (ELISA). Then, we intravenously injected normal male rats with exosomes isolated from the sera of HIRI-challenged rats and pretreated rats with MCC950, a specific inhibitor of NLRP3, and carried out the same assay. We also detected the levels of reactive oxygen species (ROS), superoxide dismutase (SOD), and malondialdehyde (MDA) in the hippocampal and cortical tissues. Results: The results indicated that NLRP3 inflammasome and caspase-1-dependent pyroptosis were activated in the hippocampus and cortex of HIRI rats. Furthermore, serum-derived exosomes from HIRI-challenged rats not only had the ability to cross the blood-brain barrier (BBB) but also had the similar effects on neuronal pyroptosis. Moreover, ROS and MDA productions were induced in the HIRI and exosome-challenged groups. In addition, to some degree, MCC950 could alleviate HIRI-mediated hippocampal and cortical neuronal pyroptosis. Conclusion: This study experimentally demonstrated that circulating exosomes play a critical role in HIRI-mediated hippocampal and cortical injury through regulating neuronal pyroptosis.


Assuntos
Barreira Hematoencefálica/metabolismo , Citocinas/metabolismo , Exossomos/metabolismo , Hipocampo/fisiopatologia , Fígado/fisiopatologia , Neurônios/metabolismo , Piroptose/genética , Traumatismo por Reperfusão/metabolismo , Animais , Modelos Animais de Doenças , Humanos , Masculino , Ratos , Ratos Sprague-Dawley
18.
Mediators Inflamm ; 2019: 2917128, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31871426

RESUMO

Background: Atherosclerosis (AS) is recognized as a chronic inflammatory disease. It is caused by the interaction between inflammatory cells such as macrophages, dendritic cells, and lipoproteins. Evidence has revealed that macrophage pyroptosis in lesion contributes to the formation of the necrotic core and thinning of the fibrous cap, which plays crucial roles in the onset of acute coronary syndrome (ACS). IFN regulatory factor 1 (IRF-1) is a pleiotropic transcription factor involved in various immune processes and cell death. We propose that IRF-1 may be implicated in macrophage pyroptosis in the pathogenesis of AS and ACS. Methods: Patients with stable angina, unstable angina, acute myocardial infarction, and clinical presentation of chest pain were enrolled. The expression of IRF-1 in human PBMC-derived macrophages was analyzed. Then, overexpression and inhibition of IRF-1 was performed in macrophages from patients with ACS to explore the possible role and mechanism of IRF-1 involvement in macrophage pyroptosis. Results: The expression of IRF-1 in macrophages was upregulated in ACS patients. The overexpression or inhibition of IRF-1 effectively modulated caspase-1 activation, as well as macrophage lysis, expression of gasdermin D-N (GSDMD-N), production of IL-1ß and IL-18, and activation of NLRP3-ASC inflammasome, which were all inhibited by caspase-1 inhibitor. Further experiments revealed that pyroptosis and the downstream inflammatory response in AS induced by IRF-1 is a process that is dependent on reactive oxygen species (ROS) generation. Conclusion: Our observations suggest that IRF-1 potently activates ox-LDL-induced macrophage pyroptosis and may play an important role in AS and ACS.


Assuntos
Síndrome Coronariana Aguda/metabolismo , Fator Regulador 1 de Interferon/metabolismo , Lipoproteínas LDL/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Síndrome Coronariana Aguda/genética , Idoso , Western Blotting , Morte Celular/genética , Morte Celular/fisiologia , Células Cultivadas , Feminino , Humanos , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Lipoproteínas LDL/genética , Masculino , Pessoa de Meia-Idade , Piroptose/genética , Piroptose/fisiologia
19.
Cell Death Dis ; 10(12): 957, 2019 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-31862870

RESUMO

Radiation-induced lung injury (RILI) is one of the most common and fatal complications of thoracic radiotherapy, whereas no effective interventions are available. Andrographolide, an active component extracted from Andrographis paniculate, is prescribed as a treatment for upper respiratory tract infection. Here we report the potential radioprotective effect and mechanism of Andrographolide on RILI. C57BL/6 mice were exposed to 18 Gy of whole thorax irradiation, followed by intraperitoneal injection of Andrographolide every other day for 4 weeks. Andrographolide significantly ameliorated radiation-induced lung tissue damage, inflammatory cell infiltration, and pro-inflammatory cytokine release in the early phase and progressive fibrosis in the late phase. Moreover, Andrographolide markedly hampered radiation-induced activation of the AIM2 inflammasome and pyroptosis in vivo. Furthermore, bone marrow-derived macrophages (BMDMs) were exposed to 8 Gy of X-ray radiation in vitro and Andrographolide significantly inhibited AIM2 inflammasome mediated-pyroptosis in BMDMs. Mechanistically, Andrographolide effectively prevented AIM2 from translocating into the nucleus to sense DNA damage induced by radiation or chemotherapeutic agents in BMDMs. Taken together, Andrographolide ameliorates RILI by suppressing AIM2 inflammasome mediated-pyroptosis in macrophage, identifying Andrographolide as a novel potential protective agent for RILI.


Assuntos
Anormalidades Induzidas por Radiação/tratamento farmacológico , Proteínas de Ligação a DNA/genética , Diterpenos/farmacologia , Lesão Pulmonar/tratamento farmacológico , Anormalidades Induzidas por Radiação/genética , Anormalidades Induzidas por Radiação/patologia , Animais , Modelos Animais de Doenças , Humanos , Inflamassomos/efeitos dos fármacos , Inflamassomos/genética , Inflamação/tratamento farmacológico , Inflamação/etiologia , Inflamação/genética , Inflamação/patologia , Pulmão/anormalidades , Pulmão/efeitos dos fármacos , Pulmão/efeitos da radiação , Lesão Pulmonar/etiologia , Lesão Pulmonar/patologia , Camundongos , Piroptose/efeitos dos fármacos , Piroptose/genética , Protetores contra Radiação/farmacologia
20.
Nature ; 575(7784): 683-687, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31748744

RESUMO

Caspase-8 is the initiator caspase of extrinsic apoptosis1,2 and inhibits necroptosis mediated by RIPK3 and MLKL. Accordingly, caspase-8 deficiency in mice causes embryonic lethality3, which can be rescued by deletion of either Ripk3 or Mlkl4-6. Here we show that the expression of enzymatically inactive CASP8(C362S) causes embryonic lethality in mice by inducing necroptosis and pyroptosis. Similar to Casp8-/- mice3,7, Casp8C362S/C362S mouse embryos died after endothelial cell necroptosis leading to cardiovascular defects. MLKL deficiency rescued the cardiovascular phenotype but unexpectedly caused perinatal lethality in Casp8C362S/C362S mice, indicating that CASP8(C362S) causes necroptosis-independent death at later stages of embryonic development. Specific loss of the catalytic activity of caspase-8 in intestinal epithelial cells induced intestinal inflammation similar to intestinal epithelial cell-specific Casp8 knockout mice8. Inhibition of necroptosis by additional deletion of Mlkl severely aggravated intestinal inflammation and caused premature lethality in Mlkl knockout mice with specific loss of caspase-8 catalytic activity in intestinal epithelial cells. Expression of CASP8(C362S) triggered the formation of ASC specks, activation of caspase-1 and secretion of IL-1ß. Both embryonic lethality and premature death were completely rescued in Casp8C362S/C362SMlkl-/-Asc-/- or Casp8C362S/C362SMlkl-/-Casp1-/- mice, indicating that the activation of the inflammasome promotes CASP8(C362S)-mediated tissue pathology when necroptosis is blocked. Therefore, caspase-8 represents the molecular switch that controls apoptosis, necroptosis and pyroptosis, and prevents tissue damage during embryonic development and adulthood.


Assuntos
Apoptose/genética , Caspase 8/genética , Caspase 8/metabolismo , Necroptose/genética , Piroptose/genética , Animais , Linhagem Celular , Células Cultivadas , Ativação Enzimática/genética , Perfilação da Expressão Gênica , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Inflamassomos/metabolismo , Mucosa Intestinal/citologia , Mucosa Intestinal/enzimologia , Queratinócitos/citologia , Queratinócitos/patologia , Camundongos , Mutação , Receptor TIE-2/genética , Receptor TIE-2/metabolismo
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