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1.
J Immunol ; 207(6): 1652-1661, 2021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34426543

RESUMO

The IκB kinase (IKK) complex plays a vital role in regulating the NF-κB activation. Aberrant NF-κB activation is involved in various inflammatory diseases. Thus, targeting IKK activation is an ideal therapeutic strategy to cure and prevent inflammatory diseases related to NF-κB activation. In a previous study, we demonstrated that IKK-interacting protein (IKIP) inhibits the phosphorylation of IKKα/ß and the activation of NF-κB through disruption of the formation of IKK complex. In this study, we identified a 15-aa peptide derived from mouse IKIP (46-60 aa of IKIP), which specifically suppressed IKK activation and NF-κB targeted gene expression via disrupting the association of IKKß and NEMO. Importantly, administration of the peptide reduced LPS-induced acute inflammation and attenuated Zymosan-induced acute arthritis in mice. These findings suggest that this IKIP peptide may be a promising therapeutic reagent in the prevention and treatment of inflammatory diseases.


Assuntos
Quinase I-kappa B/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , NF-kappa B/metabolismo , Peptídeos/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Animais , Artrite Experimental/induzido quimicamente , Artrite Experimental/tratamento farmacológico , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Lipopolissacarídeos/efeitos adversos , Camundongos , Camundongos Knockout , Ligação Proteica , Transdução de Sinais/genética , Zimosan/efeitos adversos
2.
Molecules ; 26(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34443321

RESUMO

Berberine (BBR), a plant alkaloid, is known for its therapeutic properties of anticancer, cardioprotective, antidiabetic, hypolipidemic, neuroprotective, and hepatoprotective activities. The present study was to determine the molecular mechanism of BBR's pharmacological activity in human monocytic (THP-1) cells induced by arachidonic acid (AA) or lipopolysaccharide (LPS). The effect of BBR on AA/LPS activated proinflammatory markers including TNF-α, MCP-1, IL-8 and COX-2 was measured by ELISA or quantitative real-time PCR. Furthermore, the effect of BBR on LPS-induced NF-κB translocation was determined by immunoblotting and confocal microscopy. AA/ LPS-induced TNF-α, MCP-1, IL-6, IL-8, and COX-2 markers were markedly attenuated by BBR treatment in THP-1 cells by inhibiting NF-κB translocation into the nucleus. Molecular modeling studies suggested the direct interaction of BBR to IKKα at its ligand binding site, which led to the inhibition of the LPS-induced NF-κB translocation to the nucleus. Thus, the present study demonstrated the anti-inflammatory potential of BBR via NF-κB in activated monocytes, whose interplay is key in health and in the pathophysiology of atherosclerotic development in blood vessel walls. The present study findings suggest that BBR has the potential for treating various chronic inflammatory disorders.


Assuntos
Anti-Inflamatórios/farmacologia , Berberina/farmacologia , Quinase I-kappa B/metabolismo , Lipopolissacarídeos/farmacologia , NF-kappa B/metabolismo , Linhagem Celular , Quimiocina CCL2/metabolismo , Ciclo-Oxigenase 2/metabolismo , Humanos , Interleucina-8/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo
3.
J Virol ; 95(19): e0092321, 2021 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-34260286

RESUMO

Peroxiredoxin 1 (PRDX1) is a cellular antioxidant enzyme that is crucial for diverse fundamental biological processes, such as autophagy, inflammation, and carcinogenesis. However, molecular mechanisms underpinning its diverse roles are not well understood. Here, we report that PRDX1 positively regulates interferon (IFN) induction and that pseudorabies virus (PRV) targets PRDX1 to evade IFN induction. PRV UL13 encodes a serine/threonine kinase important for PRV infection, although its biological function remains obscure. We identified PRDX1 as a UL13-interacting protein. Virological and biochemical assays demonstrate that PRDX1 promotes IFN induction by interacting with TANK-binding kinase 1 (TBK1) and IκB kinase ε (IKKε). Conversely, UL13 accelerates PRDX1 degradation via the ubiquitin-proteosome pathway in a kinase-dependent manner. In doing so, PRV inhibits IFN induction during productive infection, which requires PRDX1 expression. This study uncovers an essential role of PRDX1 in the innate immune response and reveals a new viral immune evasion strategy to counteract cellular defenses. IMPORTANCE PRV interacts with numerous cellular proteins during productive infection. Here, we demonstrated the interaction of viral protein UL13 with the antioxidant enzyme PRDX1, which functions in multiple signal transduction pathways. We found that PRDX1 participates in the type I IFN pathway by interacting with TBK1 and IKKε, thereby negatively regulating PRV propagation. However, UL13 ubiquitinates PRDX1, which routes PRDX1 into proteasomes for degradation and effectively reduces its expression. These results illuminate the fundamental role that PRDX1 plays in the IFN pathway, and they identify a potential target for the control of PRV infection.


Assuntos
Herpesvirus Suídeo 1/fisiologia , Quinase I-kappa B/metabolismo , Imunidade Inata , Peroxirredoxinas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Virais/metabolismo , Animais , Linhagem Celular , Células HEK293 , Herpesvirus Suídeo 1/imunologia , Humanos , Evasão da Resposta Imune , Interferon Tipo I/biossíntese , Mutação , Complexo de Endopeptidases do Proteassoma/metabolismo , Transdução de Sinais , Ubiquitinação , Proteínas Virais/genética , Replicação Viral
4.
Int J Mol Sci ; 22(14)2021 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-34299264

RESUMO

Inflammation has a fundamental impact on the pathophysiology of osteoarthritis (OA), a common form of degenerative arthritis. It has previously been established that curcumin, a component of turmeric (Curcuma longa), has anti-inflammatory properties. This research evaluates the potentials of curcumin on the pathophysiology of OA in vitro. To explore the anti-inflammatory efficacy of curcumin in an inflamed joint, an osteoarthritic environment (OA-EN) model consisting of fibroblasts, T-lymphocytes, 3D-chondrocytes is constructed and co-incubated with TNF-α, antisense oligonucleotides targeting NF-kB (ASO-NF-kB), or an IkB-kinase (IKK) inhibitor (BMS-345541). Our results show that OA-EN, similar to TNF-α, suppresses chondrocyte viability, which is accompanied by a significant decrease in cartilage-specific proteins (collagen II, CSPG, Sox9) and an increase in NF-kB-driven gene proteins participating in inflammation, apoptosis, and breakdown (NF-kB, MMP-9, Cox-2, Caspase-3). Conversely, similar to knockdown of NF-kB at the mRNA level or at the IKK level, curcumin suppresses NF-kB activation, NF-kB-promotes gene proteins derived from the OA-EN, and stimulates collagen II, CSPG, and Sox9 expression. Furthermore, co-immunoprecipitation assay shows that curcumin reduces OA-EN-mediated inflammation and chondrocyte apoptosis, with concomitant chondroprotective effects, due to modulation of Sox-9/NF-kB signaling axis. Finally, curcumin selectively hinders the interaction of p-NF-kB-p65 directly with DNA-this association is disrupted through DTT. These results suggest that curcumin suppresses inflammation in OA-EN via modulating NF-kB-Sox9 coupling and is essential for maintaining homeostasis in OA by balancing chondrocyte survival and inflammatory responses. This may contribute to the alternative treatment of OA with respect to the efficacy of curcumin.


Assuntos
Curcumina/farmacologia , Osteoartrite/tratamento farmacológico , Osteoartrite/metabolismo , Apoptose/efeitos dos fármacos , Cartilagem/metabolismo , Células Cultivadas , Condrócitos/metabolismo , Curcuma/metabolismo , Curcumina/metabolismo , Ciclo-Oxigenase 2/metabolismo , Humanos , Quinase I-kappa B/antagonistas & inibidores , Quinase I-kappa B/metabolismo , Imidazóis/farmacologia , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Interleucina-1beta/metabolismo , NF-kappa B/metabolismo , Osteoartrite/fisiopatologia , Cultura Primária de Células , Quinoxalinas/farmacologia , Fatores de Transcrição SOX9/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo
5.
Mol Med Rep ; 23(5)2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34240224

RESUMO

Atopic dermatitis (AD) is a chronic inflammatory skin disease that seriously affects quality of life. Quinine is a bitter taste receptor agonist that exhibits antimalarial effects. The aim of the present study was to examine the therapeutic effects of quinine in AD­like mice. AD was induced with 2,4­dinitrochlorobenzene, and the mice were treated with 10 mg/kg quinine for 1, 4 and 7 days. A total of 60 BALB/c mice were divided into the following groups: Healthy, AD­like, AD­like + quinine and healthy + quinine, with 1, 4 and 7 days groups for each treatment. Blood was extracted from all mice and ELISA was performed to detect immunoglobulin E (IgE) levels. H&E­stained tissue sections were prepared from skin lesions on the backs of the mice and pathological changes were observed. Cytokines were detected via ELISA, and the filaggrin (FLG) and kallikrein­7 (KLK7) proteins were detected via western blotting and immunohistochemistry. IKKα and NF­κB mRNA were analyzed via reverse transcription­quantitative PCR. Quinine ameliorated skin damage in the AD­like mice, reduced IgE expression in the blood, inhibited expression of IKKα and NF­κB, reduced cytokine secretion, reduced KLK7 expression, reduced scratching frequency, increased FLG expression and repaired the skin barrier. These results suggested that quinine exhibited therapeutic effects in AD­like mice.


Assuntos
Dermatite Atópica/tratamento farmacológico , Quinina/farmacologia , Quinina/uso terapêutico , Animais , Citocinas/metabolismo , Dermatite Atópica/induzido quimicamente , Dermatite Atópica/metabolismo , Dermatite Atópica/patologia , Dinitroclorobenzeno/toxicidade , Modelos Animais de Doenças , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Imunoglobulina E/sangue , Calicreínas/genética , Calicreínas/metabolismo , Masculino , Camundongos Endogâmicos BALB C , Inibidor de NF-kappaB alfa/genética , Inibidor de NF-kappaB alfa/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Pele/efeitos dos fármacos , Pele/patologia
6.
Phytother Res ; 35(8): 4547-4554, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34132431

RESUMO

In the current study, the pivotal roles of serum and glucocorticoid-induced protein kinase (SGK1) and NF-kB related signalings known as prognostic biomarkers in cervical cancers were explored in the antitumor effect of a ginseng saponin metabolite compound K (CK) in HeLa and SiHa cervical cancer cells. CK exerted significant cytotoxicity, induced sub-G1 accumulation, and attenuated the expression of proPoly (ADP-ribose) polymerase (pro-PARP) and Pro-cysteine aspartyl-specific protease (pro-caspase3) in HeLa cells more than in SiHa cells. CK inhibited phosphorylation of SGK1 and its upstream genes, phosphoinositide 3-kinases (PI3K), and phosphoinositide-dependent kinase-1 (PDK1) in HeLa cells. In addition, CK suppressed the phosphorylation of SGK1, NF-κB, and inhibitor of kappa B (IκB) and also NF-κB target genes such as X-linked inhibitor of apoptosis protein and B-cell lymphoma 2 (Bcl-2) in HeLa cells. Notably, Immunoprecipitation revealed that SGK1 binds to PI3K or PDK1 and also CK disturbed the binding between SGK1 and PI3K or PDK1 in HeLa cells. Furthermore, PI3K inhibitor LY294002 decreased expression of PI3K, p-PDK1, p-SGK1, and pro-caspase3 and SGK1 inhibitor GSK650394 also reduced expression of NF-κB and pro-caspase3 just like CK in HeLa cells. Overall, these findings suggest that CK induces apoptosis via suppression of PI3K/PDK1/SGK1 and NF-κB signaling axis.


Assuntos
Ginsenosídeos/farmacologia , Proteínas Imediatamente Precoces/metabolismo , Fosfatidilinositol 3-Quinases , Proteínas Serina-Treonina Quinases/metabolismo , Células HeLa , Humanos , Quinase I-kappa B/metabolismo , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Quinases , Transdução de Sinais
7.
Commun Biol ; 4(1): 663, 2021 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-34079066

RESUMO

The reciprocal interactions between pathogens and hosts are complicated and profound. A comprehensive understanding of these interactions is essential for developing effective therapies against infectious diseases. Interferon responses induced upon virus infection are critical for establishing host antiviral innate immunity. Here, we provide a molecular mechanism wherein isoform switching of the host IKKε gene, an interferon-associated molecule, leads to alterations in IFN production during EV71 infection. We found that IKKε isoform 2 (IKKε v2) is upregulated while IKKε v1 is downregulated in EV71 infection. IKKε v2 interacts with IRF7 and promotes IRF7 activation through phosphorylation and translocation of IRF7 in the presence of ubiquitin, by which the expression of IFNß and ISGs is elicited and virus propagation is attenuated. We also identified that IKKε v2 is activated via K63-linked ubiquitination. Our results suggest that host cells induce IKKε isoform switching and result in IFN production against EV71 infection. This finding highlights a gene regulatory mechanism in pathogen-host interactions and provides a potential strategy for establishing host first-line defense against pathogens.


Assuntos
Enterovirus Humano A/imunologia , Enterovirus Humano A/patogenicidade , Quinase I-kappa B/genética , Quinase I-kappa B/imunologia , Processamento Alternativo , Linhagem Celular , Genes de Troca , Células HEK293 , Interações entre Hospedeiro e Microrganismos/genética , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Quinase I-kappa B/metabolismo , Imunidade Inata/genética , Fator Regulador 7 de Interferon/metabolismo , Interferon beta/biossíntese , Isoenzimas/genética , Isoenzimas/imunologia , Fosforilação , Ubiquitina/metabolismo
8.
Front Immunol ; 12: 662989, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34084167

RESUMO

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the causative pathogen of current COVID-19 pandemic, and insufficient production of type I interferon (IFN-I) is associated with the severe forms of the disease. Membrane (M) protein of SARS-CoV-2 has been reported to suppress host IFN-I production, but the underlying mechanism is not completely understood. In this study, SARS-CoV-2 M protein was confirmed to suppress the expression of IFNß and interferon-stimulated genes induced by RIG-I, MDA5, IKKϵ, and TBK1, and to inhibit IRF3 phosphorylation and dimerization caused by TBK1. SARS-CoV-2 M could interact with MDA5, TRAF3, IKKϵ, and TBK1, and induce TBK1 degradation via K48-linked ubiquitination. The reduced TBK1 further impaired the formation of TRAF3-TANK-TBK1-IKKε complex that leads to inhibition of IFN-I production. Our study revealed a novel mechanism of SARS-CoV-2 M for negative regulation of IFN-I production, which would provide deeper insight into the innate immunosuppression and pathogenicity of SARS-CoV-2.


Assuntos
Interferon Tipo I/biossíntese , Proteínas Serina-Treonina Quinases/metabolismo , SARS-CoV-2/imunologia , Ubiquitina/metabolismo , Proteínas da Matriz Viral/imunologia , Proteína DEAD-box 58/metabolismo , Células HEK293 , Humanos , Quinase I-kappa B/metabolismo , Fator Regulador 3 de Interferon/metabolismo , Helicase IFIH1 Induzida por Interferon/metabolismo , Proteólise , Receptores Imunológicos/metabolismo , Transdução de Sinais , Fator 3 Associado a Receptor de TNF/metabolismo
9.
Int J Mol Sci ; 22(11)2021 May 26.
Artigo em Inglês | MEDLINE | ID: mdl-34073390

RESUMO

Activation of nuclear factor-kappa B (NF-κB) in microglia plays a decisive role in the progress of neuropathic pain, and the inhibitor of kappa B (IκB) is a protein that blocks the activation of NF-κB and is degraded by the inhibitor of NF-κB kinase subunit beta (IKBKB). The role of IKBKB is to break down IκB, which blocks the activity of NF-kB. Therefore, it prevents the activity of NK-kB. This study investigated whether neuropathic pain can be reduced in spinal nerve ligation (SNL) rats by reducing the activity of microglia by delivering IKBKB small interfering RNA (siRNA)-encapsulated poly (lactic-co-glycolic acid) (PLGA) nanoparticles. PLGA nanoparticles, as a carrier for the delivery of IKBKB genes silencer, were used because they have shown potential to enhance microglial targeting. SNL rats were injected with IKBKB siRNA-encapsulated PLGA nanoparticles intrathecally for behavioral tests on pain response. IKBKB siRNA was delivered for suppressing the expression of IKBKB. In rats injected with IKBKB siRNA-encapsulated PLGA nanoparticles, allodynia caused by mechanical stimulation was reduced, and the secretion of pro-inflammatory mediators due to NF-κB was reduced. Delivering IKBKB siRNA through PLGA nanoparticles can effectively control the inflammatory response and is worth studying as a treatment for neuropathic pain.


Assuntos
Portadores de Fármacos/farmacologia , Quinase I-kappa B/antagonistas & inibidores , Nanopartículas/uso terapêutico , Neuralgia/tratamento farmacológico , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacologia , RNA Interferente Pequeno/farmacologia , Animais , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Masculino , Microglia/patologia , Neuralgia/genética , Neuralgia/metabolismo , Neuralgia/patologia , RNA Interferente Pequeno/genética , Ratos , Ratos Sprague-Dawley
10.
Lab Invest ; 101(9): 1238-1253, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34059758

RESUMO

Spinal cord injury (SCI) is one common neurological condition which involves primary injury and secondary injury. Neuron inflammation and apoptosis after SCI is the most important pathological process of this disease. Here, we tried to explore the influence and mechanism of miRNAs on the neuron inflammatory response and apoptosis after SCI. First, by re-analysis of Gene Expression Omnibus dataset (accession GSE19890), miR-182 was selected for further study because of its suppressive effects on the inflammatory response in the various types of injuries. Functional experiments demonstrated that miR-182 overexpression promoted functional recovery, reduced histopathological changes, and alleviated spinal cord edema in mice. It was also observed that miR-182 overexpression reduced apoptosis and attenuated the inflammatory response in spinal cord tissue, as evidenced by the reduction of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1ß, and the induction of IL-10. Using a lipopolysaccharide (LPS)-induced SCI model in BV-2 cells, we found that miR-182 was downregulated in the BV-2 cells following LPS stimulation, and upregulation of miR-182 improved LPS-induced cell damage, as reflected by the inhibition of apoptosis and the inflammatory response. IκB kinase ß (IKKß), an upstream target of the NF-κB pathway, was directly targeted by miR-182 and miR-182 suppressed its translation. Further experiments revealed that overexpression of IKKß reversed the anti-apoptosis and anti-inflammatory effects of miR-182 in LPS stimulated BV-2 cells. Finally, we found that miR-182 overexpression blocked the activation of the NF-κB signaling pathway in vitro and in vivo, as demonstrated by the downregulation of phosphorylated (p­) IκB-α and nuclear p-p65. Taken together, these data indicate that miR-182 improved SCI-induced secondary injury through inhibiting apoptosis and the inflammatory response by blocking the IKKß/NF-κB pathway. Our findings suggest that upregulation of miR-182 may be a novel therapeutic target for SCI.


Assuntos
Apoptose/efeitos dos fármacos , Inflamação/metabolismo , MicroRNAs/farmacologia , Traumatismos da Medula Espinal/metabolismo , Animais , Modelos Animais de Doenças , Feminino , Quinase I-kappa B/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Transdução de Sinais/efeitos dos fármacos
11.
Eur J Med Chem ; 222: 113579, 2021 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-34098465

RESUMO

Genetic models validated Inhibitor of nuclear factor (NF) kappa B kinase beta (IKKß) as a therapeutic target for KRAS mutation associated pancreatic cancer. Phosphorylation of the activation loop serine residues (S177, S181) in IKKß is a key event that drives tumor necrosis factor (TNF) α induced NF-κB mediated gene expression. Here we conducted structure activity relationship (SAR) study to improve potency and oral bioavailability of a quinoxaline analog 13-197 that was previously reported as a NFκB inhibitor for pancreatic cancer therapy. The SAR led to the identification of a novel quinoxaline urea analog 84 that reduced the levels of p-IKKß in dose- and time-dependent studies. When compared to 13-197, analog 84 was ∼2.5-fold more potent in TNFα-induced NFκB inhibition and ∼4-fold more potent in inhibiting pancreatic cancer cell growth. Analog 84 exhibited ∼4.3-fold greater exposure (AUC0-∞) resulting in ∼5.7-fold increase in oral bioavailability (%F) when compared to 13-197. Importantly, oral administration of 84 by itself and in combination of gemcitabine reduced p-IKKß levels and inhibited pancreatic tumor growth in a xenograft model.


Assuntos
Antineoplásicos/farmacologia , Quinase I-kappa B/antagonistas & inibidores , Neoplasias Pancreáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Quinoxalinas/farmacologia , Ureia/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Quinase I-kappa B/metabolismo , Camundongos , Estrutura Molecular , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Quinoxalinas/síntese química , Quinoxalinas/química , Relação Estrutura-Atividade , Ureia/análogos & derivados , Ureia/química
12.
Chem Commun (Camb) ; 57(38): 4678-4681, 2021 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-33977973

RESUMO

Inhibitor of nuclear factor kappa-B kinase subunit beta (IKKß) is a key regulator of the cannonical NF-κB pathway. IKKß has been validated as a drug target for pathological conditions, which include chronic inflammatory diseases and cancer. Pharmacological studies revealed that chronic administration of ATP-competitive IKKß inhibitors resulted in unexpected toxicity. We previously reported the discovery of 13-197 as a non-toxic IKKß inhibitor that reduced tumor growth. Here, we show that 13-197 inhibits IKKß in a ATP non-competitive manner and an allosteric pocket at the interface of the kinase and ubiquitin like domains was identified as the potential binding site.


Assuntos
Trifosfato de Adenosina/metabolismo , Quinase I-kappa B/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Sítios de Ligação/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Quinase I-kappa B/metabolismo , Estrutura Molecular , Inibidores de Proteínas Quinases/química , Bibliotecas de Moléculas Pequenas/química
13.
PLoS Comput Biol ; 17(5): e1009040, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34043616

RESUMO

Dorsal-ventral patterning of the Drosophila embryo depends on the NFκB superfamily transcription factor Dorsal (Dl). Toll receptor activation signals for degradation of the IκB inhibitor Cactus (Cact), leading to a ventral-to-dorsal nuclear Dl gradient. Cact is critical for Dl nuclear import, as it binds to and prevents Dl from entering the nuclei. Quantitative analysis of cact mutants revealed an additional Cact function to promote Dl nuclear translocation in ventral regions of the embryo. To investigate this dual Cact role, we developed a predictive model based on a reaction-diffusion regulatory network. This network distinguishes non-uniform Toll-dependent Dl nuclear import and Cact degradation, from the Toll-independent processes of Cact degradation and reversible nuclear-cytoplasmic Dl flow. In addition, it incorporates translational control of Cact levels by Dl. Our model successfully reproduces wild-type data and emulates the Dl nuclear gradient in mutant dl and cact allelic combinations. Our results indicate that the dual role of Cact depends on the dynamics of Dl-Cact trimers along the dorsal-ventral axis: In the absence of Toll activation, free Dl-Cact trimers retain Dl in the cytoplasm, limiting the flow of Dl into the nucleus; in ventral-lateral regions, Dl-Cact trimers are recruited by Toll activation into predominant signaling complexes and promote Dl nuclear translocation. Simulations suggest that the balance between Toll-dependent and Toll-independent processes are key to this dynamics and reproduce the full assortment of Cact effects. Considering the high evolutionary conservation of these pathways, our analysis should contribute to understanding NFκB/c-Rel activation in other contexts such as in the vertebrate immune system and disease.


Assuntos
Núcleo Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Quinase I-kappa B/metabolismo , Modelos Biológicos , NF-kappa B/metabolismo , Fosfoproteínas/metabolismo , Animais , Transporte Proteico , Transdução de Sinais
14.
Biochem Biophys Res Commun ; 560: 165-171, 2021 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-33992959

RESUMO

Alkaliptosis is a recently discovered form of regulated cell death driven by intracellular alkalization. However, the immune characteristics and mechanisms of alkaliptosis are still poorly understood. Here, we show that HMGB1, a multifunctional alarm protein that drives innate immunity, is necessary for inflammation caused by alkaliptotic damage. During alkaliptosis, HMGB1 translocation and release from the nucleus to the cytoplasm to the extracellular space requires nuclear DNA damage signals, whereas the FANCD2-dependent (but not ATM-mediated) DNA repair pathway inhibits this process. Once released by alkaliptotic cancer cells, extracellular HMGB1 binds to the AGER receptor in macrophages and then activates the STING1 pathway to produce pro-inflammatory cytokines (e.g., TNF and IL6). Consequently, the pharmacological or genetic inhibition of the HMGB1-AGER-STING1 pathway limits cytokine production during alkaliptosis. These findings provide new insight into the sterile inflammatory response to cell death.


Assuntos
Proteína HMGB1/metabolismo , Proteínas de Membrana/metabolismo , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Morte Celular Regulada , Animais , Linhagem Celular , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/metabolismo , Humanos , Quinase I-kappa B/metabolismo , Mediadores da Inflamação/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos Knockout , Transdução de Sinais
15.
Genes (Basel) ; 12(5)2021 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-33946816

RESUMO

Preclinical studies conducted to date suggest that depression could be elicited by the elevated expression of proinflammatory molecules: these play a key role in the mediation of neurochemical, neuroendocrine and behavioral changes. Thus, this study investigates the effect of chronic mild stress (CMS) and administration of venlafaxine (SSRI) on the expression and methylation status of new target inflammatory genes: TGFA, TGFB, IRF1, PTGS2 and IKBKB, in peripheral blood mononuclear cells (PMBCs) and in selected brain structures of rats. Adult male Wistar rats were subjected to the CMS and further divided into matched subgroups to receive vehicle or venlafaxine. TaqMan gene expression assay and methylation-sensitive high-resolution melting (MS-HRM) were used to evaluate the expression of the genes and the methylation status of their promoters, respectively. Our results indicate that both CMS and chronic treatment with venlafaxine were associated with changes in expression of the studied genes and their promoter methylation status in PMBCs and the brain. Moreover, the effect of antidepressant administration clearly differed between brain structures. Summarizing, our results confirm at least a partial association between TGFA, TGFB, IRF1, PTGS2 and IKBKB and depressive disorders.


Assuntos
Encéfalo/metabolismo , Metilação de DNA , Leucócitos Mononucleares/metabolismo , Inibidores da Recaptação de Serotonina e Norepinefrina/farmacologia , Estresse Psicológico/genética , Transcriptoma , Cloridrato de Venlafaxina/farmacologia , Animais , Encéfalo/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Fator Regulador 1 de Interferon/genética , Fator Regulador 1 de Interferon/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Masculino , Ratos , Ratos Wistar , Inibidores da Recaptação de Serotonina e Norepinefrina/uso terapêutico , Estresse Psicológico/tratamento farmacológico , Estresse Psicológico/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Cloridrato de Venlafaxina/uso terapêutico
16.
J Biochem Mol Toxicol ; 35(8): e22799, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33949057

RESUMO

The anti-inflammatory activity of cirsilineol in in vivo condition was assessed by measuring the relative organ weight, lung dry/wet weight ratio, protein concentration, and infiltration of inflammatory cells in bronchoalveolar lavage fluid. We estimated the myeloperoxidase activity and levels of cytokines, chemokines, and inflammatory markers to analyze the efficacy of cirsilineol against lipopolysaccharide (LPS)-induced lung inflammation. Furthermore, we quantified the gene expression of NFkB/IKK signaling molecules in cirsilineol-treated and untreated acute lung injury mice to confirm the anti-inflammatory property of cirsilineol. The lung histology was assessed with hematoxylin and eosin staining. Apart from in vivo experiments, in vitro tests with LPS-stimulated RAW 264.7 macrophages were also performed. Cell viability assay was performed in the presence and absence of LPS in RAW 264.7 macrophages to determine the cytotoxic effect of cirsilineol against macrophages. Reverse-transcription polymerase chain reaction (RT-PCR) analysis was done to analyze the gene expression of inflammatory markers in LPS-treated RAW 264.7 macrophages to prove that cirsilineol effectively inhibits inflammation in vitro. The results of our study prove that cirsilineol effectively inhibits inflammation in both in vivo and in vitro conditions. RT-PCR analysis results of NFkB/IKK signaling molecules clearly illustrate that cirsilineol inhibited the expression of NFkB/IKK signaling protein and thereby prevented inflammation in in vivo condition, and it is further confirmed with the results of inflammatory protein expression in vitro model. The lung histopathological studies authentically confirm that cirsilineol potentially prevented the mice from LPS-induced lung inflammation.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Flavonas/farmacologia , Quinase I-kappa B/metabolismo , Lipopolissacarídeos/toxicidade , NF-kappa B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptor 4 Toll-Like/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/metabolismo , Animais , Modelos Animais de Doenças , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Masculino , Camundongos , Células RAW 264.7
17.
Development ; 148(8)2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33913480

RESUMO

Multiple morphological abnormalities of the sperm flagella (MMAF) are a major cause of asthenoteratozoospermia. We have identified protease serine 50 (PRSS50) as having a crucial role in sperm development, because Prss50-null mice presented with impaired fertility and sperm tail abnormalities. PRSS50 could also be involved in centrosome function because these mice showed a threefold increase in acephalic sperm (head-tail junction defect), sperm with multiple heads (spermatid division defect) and sperm with multiple tails, including novel two conjoined sperm (complete or partial parts of several flagellum on the same plasma membrane). Our data support that, in the testis, as in tumorigenesis, PRSS50 activates NFκB target genes, such as the centromere protein leucine-rich repeats and WD repeat domain-containing protein 1 (LRWD1), which is required for heterochromatin maintenance. Prss50-null testes have increased IκκB, and reduced LRWD1 and histone expression. Low levels of de-repressed histone markers, such as H3K9me3, in the Prss50-null mouse testis may cause increases in post-meiosis proteins, such as AKAP4, affecting sperm formation. We provide important insights into the complex mechanisms of sperm development, the importance of testis proteases in fertility and a novel mechanism for MMAF.


Assuntos
Fertilidade , Serina Proteases/metabolismo , Cauda do Espermatozoide/enzimologia , Testículo/enzimologia , Animais , Astenozoospermia/enzimologia , Astenozoospermia/genética , Heterocromatina/enzimologia , Heterocromatina/genética , Histonas/biossíntese , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Masculino , Camundongos , Camundongos Mutantes , Proteínas dos Microtúbulos/genética , Proteínas dos Microtúbulos/metabolismo , Serina Proteases/deficiência , Cabeça do Espermatozoide/enzimologia
18.
J Clin Invest ; 131(10)2021 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-33822771

RESUMO

The protein kinases IKKε and TBK1 are activated in liver and fat in mouse models of obesity. We have previously demonstrated that treatment with the IKKε/TBK1 inhibitor amlexanox produces weight loss and relieves insulin resistance in obese animals and patients. While amlexanox treatment caused a transient reduction in food intake, long-term weight loss was attributable to increased energy expenditure via FGF21-dependent beiging of white adipose tissue (WAT). Amlexanox increased FGF21 synthesis and secretion in several tissues. Interestingly, although hepatic secretion determined circulating levels, it was dispensable for regulating energy expenditure. In contrast, adipocyte-secreted FGF21 may have acted as an autocrine factor that led to adipose tissue browning and weight loss in obese mice. Moreover, increased energy expenditure was an important determinant of improved insulin sensitivity by amlexanox. Conversely, the immediate reductions in fasting blood glucose observed with acute amlexanox treatment were mediated by the suppression of hepatic glucose production via activation of STAT3 by adipocyte-secreted IL-6. These findings demonstrate that amlexanox improved metabolic health via FGF21 action in adipocytes to increase energy expenditure via WAT beiging and that adipocyte-derived IL-6 has an endocrine role in decreasing gluconeogenesis via hepatic STAT3 activation, thereby producing a coordinated improvement in metabolic parameters.


Assuntos
Aminopiridinas/farmacologia , Glicemia/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Gluconeogênese/efeitos dos fármacos , Quinase I-kappa B/metabolismo , Fígado/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Glicemia/genética , Ingestão de Alimentos/efeitos dos fármacos , Ingestão de Alimentos/genética , Metabolismo Energético/efeitos dos fármacos , Metabolismo Energético/genética , Fatores de Crescimento de Fibroblastos/genética , Gluconeogênese/genética , Quinase I-kappa B/genética , Interleucina-6/genética , Interleucina-6/metabolismo , Camundongos , Camundongos Knockout , Proteínas Serina-Treonina Quinases/genética , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo
19.
J Immunol ; 206(9): 2184-2197, 2021 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-33858962

RESUMO

IFN-induced protein with tetratricopeptide repeats (IFITs), known as canonical IFN-stimulated genes (ISGs), play critical roles in regulating immune responses against pathogens and maintaining homeostasis. How the IFIT5 regulates innate immune responses is rarely reported and remains enigmatic. In this study, we discover that human IFIT5 (hIFIT5) functions as a negative regulator of the type I IFN (IFN) pathway in HEK293T cell lines. Our data illustrated that hIFIT5 inhibited the promotor activities of IFN-ß induced by IRF3 and its upstream factors but not by IRF3-5D (activated form of IRF3), suggesting that IRF3 might be a target of hIFIT5. Further investigations revealed that hIFIT5 downregulated the phosphorylation of IRF3 and IKKε and blocked the IRF3 nuclear translocation. Moreover, hIFIT5 impaired the IRF3-TBK1-IKKε complex, accompanied by IRF3 and IKKε degradation. In conclusion, these findings indicate that hIFIT5 is a negative modulator in the type I IFN signaling pathway, opening additional avenues for preventing hyperactivation and maintaining immunity homeostasis.


Assuntos
Quinase I-kappa B/imunologia , Fator Regulador 3 de Interferon/imunologia , Interferon Tipo I/imunologia , Proteínas de Neoplasias/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Animais , Linhagem Celular , Humanos , Quinase I-kappa B/metabolismo , Transdução de Sinais/imunologia
20.
Am J Physiol Heart Circ Physiol ; 320(6): H2324-H2338, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33929897

RESUMO

Ataxia-telangiectasia mutated (ATM) kinase deficiency exacerbates heart dysfunction late after myocardial infarction. Here, we hypothesized that ATM deficiency modulates Western-type diet (WD)-induced cardiac remodeling with an emphasis on functional and biochemical parameters of the heart. Weight gain was assessed in male wild-type (WT) and ATM heterozygous knockout (hKO) mice on weekly basis, whereas cardiac functional and biochemical parameters were measured 14 wk post-WD. hKO-WD mice exhibited rapid body weight gain at weeks 5, 6, 7, 8, and 10 versus WT-WD. WD decreased percent fractional shortening and ejection fraction, and increased end-systolic volumes and diameters to a similar extent in both genotypes. However, WD decreased stroke volume, cardiac output, peak velocity of early ventricular filling, and aortic ejection time and increased isovolumetric relaxation time (IVRT) and Tei index versus WT-NC (normal chow). Conversely, IVRT, isovolumetric contraction time, and Tei index were lower in hKO-WD versus hKO-NC and WT-WD. Myocyte apoptosis and hypertrophy were higher in hKO-WD versus WT-WD. WD increased fibrosis and expression of collagen-1α1, matrix metalloproteinase (MMP)-2, and MMP-9 in WT. WD enhanced AMPK activation, while decreasing mTOR activation in hKO. Akt and IKK-α/ß activation, and Bax, PARP-1, and Glut-4 expression were higher in WT-WD versus WT-NC, whereas NF-κB activation and Glut-4 expression were lower in hKO-WD versus hKO-NC. Circulating concentrations of IL-12(p70), eotaxin, IFN-γ, macrophage inflammatory protein (MIP)-1α, and MIP-1ß were higher in hKO-WD versus WT-WD. Thus, ATM deficiency accelerates weight gain, induces systolic dysfunction with increased preload, and associates with increased apoptosis, hypertrophy, and inflammation in response to WD.NEW & NOTEWORTHY Ataxia-telangiectasia mutated (ATM) kinase deficiency in humans associates with enhanced susceptibility to ischemic heart disease. Here, we provide evidence that ATM deficiency accelerates body weight gain and associates with increased cardiac preload, hypertrophy, and apoptosis in mice fed with Western-type diet (WD). Further investigations of the role of ATM deficiency in WD-induced alterations in function and biochemical parameters of the heart may provide clinically applicable information on treatment and/or nutritional counseling for patients with ATM deficiency.


Assuntos
Cardiomegalia/genética , Dieta Ocidental , Miocárdio/metabolismo , Remodelação Ventricular/genética , Ganho de Peso/genética , Adenilato Quinase/metabolismo , Animais , Apoptose/genética , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Débito Cardíaco/genética , Quimiocina CCL3/metabolismo , Quimiocina CCL4/metabolismo , Quimiocinas CC/metabolismo , Colágeno Tipo I/metabolismo , Fibrose/genética , Interação Gene-Ambiente , Transportador de Glucose Tipo 4/metabolismo , Heterozigoto , Quinase I-kappa B/metabolismo , Interferon gama/metabolismo , Interleucina-12/metabolismo , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Knockout , Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Poli(ADP-Ribose) Polimerase-1/metabolismo , Volume Sistólico/genética , Serina-Treonina Quinases TOR/metabolismo , Proteína X Associada a bcl-2/metabolismo
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