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1.
Nucleic Acids Res ; 52(9): 4969-4984, 2024 May 22.
Artículo en Inglés | MEDLINE | ID: mdl-38452206

RESUMEN

Proteasome-mediated degradation of chromatin-bound NF-κB is critical in terminating the transcription of pro-inflammatory genes and can be triggered by Set9-mediated lysine methylation of the RelA subunit. However, the E3 ligase targeting methylated RelA remains unknown. Here, we find that two structurally similar substrate-recognizing components of Cullin-RING E3 ligases, WSB1 and WSB2, can recognize chromatin-bound methylated RelA for polyubiquitination and proteasomal degradation. We showed that WSB1/2 negatively regulated a subset of NF-κB target genes via associating with chromatin where they targeted methylated RelA for ubiquitination, facilitating the termination of NF-κB-dependent transcription. WSB1/2 specifically interacted with methylated lysines (K) 314 and 315 of RelA via their N-terminal WD-40 repeat (WDR) domains, thereby promoting ubiquitination of RelA. Computational modeling further revealed that a conserved aspartic acid (D) at position 158 within the WDR domain of WSB2 coordinates K314/K315 of RelA, with a higher affinity when either of the lysines is methylated. Mutation of D158 abolished WSB2's ability to bind to and promote ubiquitination of methylated RelA. Together, our study identifies a novel function and the underlying mechanism for WSB1/2 in degrading chromatin-bound methylated RelA and preventing sustained NF-κB activation, providing potential new targets for therapeutic intervention of NF-κB-mediated inflammatory diseases.


Asunto(s)
Cromatina , Complejo de la Endopetidasa Proteasomal , Factor de Transcripción ReIA , Ubiquitinación , Humanos , Cromatina/metabolismo , Células HEK293 , Lisina/metabolismo , Metilación , FN-kappa B/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Unión Proteica , Proteolisis , Factor de Transcripción ReIA/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genética
2.
J Med Virol ; 95(7): e28967, 2023 07.
Artículo en Inglés | MEDLINE | ID: mdl-37496184

RESUMEN

As an intracellular parasite, the virus usurps cellular machinery and modulates cellular metabolism pathways to replicate itself in cells. Lipid droplets (LDs) are universally conserved energy storage organelles that not only play vital roles in maintaining lipid homeostasis but are also involved in viral replication. Increasing evidence has demonstrated that viruses take advantage of cellular lipid metabolism by targeting the biogenesis, hydrolysis, and lipophagy of LD during viral infection. In this review, we summarize the current knowledge about the modulation of cellular LD by different viruses, with a special emphasis on the Hepatitis C virus, Dengue virus, and SARS-CoV-2.


Asunto(s)
COVID-19 , Gotas Lipídicas , Humanos , Gotas Lipídicas/metabolismo , COVID-19/metabolismo , SARS-CoV-2 , Metabolismo de los Lípidos , Hepacivirus
3.
J Med Virol ; 95(3): e28630, 2023 03.
Artículo en Inglés | MEDLINE | ID: mdl-36861654

RESUMEN

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection leads to the accumulation of lipid droplets (LD), the central hubs of the lipid metabolism, in vitro or in type II pneumocytes and monocytes from coronavirus disease 19 (COVID-19) patients and blockage of LD formation by specific inhibitors impedes SARS-CoV-2 replication. Here, we showed that ORF3a is necessary and sufficient to trigger LD accumulation during SARS-CoV-2 infection, leading to efficient virus replication. Although highly mutated during evolution, ORF3a-mediated LD modulation is conserved in most SARS-CoV-2 variants except the Beta strain and is a major difference between SARS-CoV and SARS-CoV-2 that depends on the genetic variations on the amino acid position 171, 193, and 219 of ORF3a. Importantly, T223I substitution in recent Omicron strains (BA.2-BF.8) impairs ORF3a-Vps39 association and LD accumulation, leading to less efficient replication and potentially contributing to lower pathogenesis of the Omicron strains. Our work characterized how SARS-CoV-2 modulates cellular lipid homeostasis to benefit its replication during virus evolution, making ORF3a-LD axis a promising drug target for the treatment of COVID-19.


Asunto(s)
COVID-19 , SARS-CoV-2 , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo , Humanos , Gotas Lipídicas , SARS-CoV-2/genética , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/genética
4.
Immunopharmacol Immunotoxicol ; 45(2): 213-223, 2023 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-36218392

RESUMEN

BACKGROUND: Secoeudesma sesquiterpenes lactone A (SESLA) is a sesquiterpene derived from Inula japonica Thunb. and is known to possess many pharmacological properties, e.g. anti-tumor and anti-inflammatory activities. However, the immunomodulatory role of SESLA in gram-positive (G+) bacterial infection is not clear. MATERIALS AND METHODS: To set up a G+ bacterial infection model in vitro, we carried out a bacterial mimic (PGN or Pam3CSK4) or Methicillin-resistant Staphylococcus aureus (MRSA) stimulated experiment using macrophages or dendritic cells (DCs). ELISA and qPCR were performed to measure the expression of inflammatory cytokines. Flow cytometry was used to detect the expression of MHC II and co-stimulatory molecules on the surface of DCs. The network pharmacology was used to identify the molecular mechanism and potential targets of SESLA that are predicted to be involved in the MRSA-elicited inflammation. Western blot and dual luciferase reporter assay were adopted to certify possible molecular mechanism of SESLA. RESULTS: This study demonstrated that SESLA treatment significantly reduced the levels of inflammatory cytokines stimulated by PGN, Pam3CSK4 or even MRSA in vitro, and it also reduced PGN-induced expression of MHC II and co-stimulatory molecules on the surface of DCs. Mechanistically, the inhibition of IκBα phosphorylation and the suppression of T cells activation could account for its anti-inflammatory activity. CONCLUSION: The present study validated the notable anti-inflammatory activity of SESLA and discovered its previously uncharacterized immunoregulatory role and the underlying mechanism in G+ bacterial infections. Overall, SESLA has a potential to be an antibiotic adjuvant for the treatment of G+ bacterial infections.


Asunto(s)
Staphylococcus aureus Resistente a Meticilina , Infecciones Estafilocócicas , Humanos , Staphylococcus aureus Resistente a Meticilina/metabolismo , Macrófagos/metabolismo , Citocinas/metabolismo , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Células Dendríticas/metabolismo , Infecciones Estafilocócicas/tratamiento farmacológico , Infecciones Estafilocócicas/metabolismo , Infecciones Estafilocócicas/microbiología
5.
Zhongguo Zhong Yao Za Zhi ; 47(17): 4698-4706, 2022 Sep.
Artículo en Zh | MEDLINE | ID: mdl-36164877

RESUMEN

This study aimed to explore the protective effect of Reduning Injection(RDN) on mice infected by influenza virus A/PR/8(PR8) and its immune regulatory roles during viral infection. In in vivo experiments, female C57 BL/6 mice were randomly divided into phosphate buffered saline(PBS) group, PR8-infected group, oseltamivir treatment group(OSV) and RDN treatment group. After 2 h of PR8 infection, mice in the oseltamivir group were gavaged with oseltamivir 30 mg·kg~(-1), and those in the RDN treatment group were injected intraperitoneally with RDN 1.5 mL·kg~(-1)once per day for seven consecutive days. The body weight of mice in each group was recorded at the same time every morning for 16 consecutive days. The line chart of body weight change was created to analyze the protective effect of RDN on flu-infected mice. The relative mRNA expression of different cytokines(IL-6, TNF-α, MCP-1, IL-1ß, MIP-2, IP-10 and IL-10) in lung samples of flu-infected mice was detected by PCR. Flow cytometry was utilized to analyze the composition of immune cells of mouse BALF samples on day 5 after infection. Mouse macrophage cell line RAW264.7 was planted and treated by different concentrations of RDN(150, 300, 600 µg·mL~(-1)) for 24 h or 48 h, and cell proliferation was detected by CCK-8 assay. RAW264.7 cells and mouse primary peritoneal macrophages were stimulated with synthetic single stranded RNA(R837), which elicited the inflammatory response by mimicking the infection of single-stranded RNA viruses. The expression of cytokines and chemokines in the supernatants of above culture system was detected by ELISA and qPCR. On days 4, 5, 6, 7 and 15 after infection, the body weight loss of mice in the RDN treatment group was alleviated compared with that of PR8-infected mice(P<0.05). RDN treatment obviously reduced lung index and the production of IL-6, TNF-α, MCP-1 and MIP-2 in lung tissues of flu-infected mice(P<0.05). The proportions of macrophages, neutrophils and T cells in mouse BALF samples were analyzed by flow cytometry, and compared with PR8-infected mice, RDN decreased the proportion of macrophages in BALF of flu-infected mice(P<0.05), and the proportion of T cells was recovered dramatically(P<0.001). In CCK-8 assay, the concentrations of RDN(150, 300, 600 µg·mL~(-1)) failed to cause cytotoxicity to RAW264.7 cells. In addition, RDN lowered the expression of inflammatory cytokines such as IL-6, TNF-α,MCP-1, IL-1ß, RANTES, and IP-10 and even anti-inflammatory cytokine IL-10 in R837-induced macrophages. RDN reduced the infiltration of inflammatory macrophages and the production of excessive inflammatory cytokines, alleviated the body weight loss of flu-infected mice. What's more, RDN restored the depletion of T cells, which might prevent secondary infection and deteriorative progression of the disease. Taken together, RDN may inhibit cytokine production and therefore down-regulate cytokine storm during the infection of influenza virus.


Asunto(s)
Interleucina-10 , Oseltamivir , Animales , Antiinflamatorios/farmacología , Peso Corporal , Quimiocina CCL5/farmacología , Quimiocina CXCL10/farmacología , Síndrome de Liberación de Citoquinas , Citocinas/genética , Medicamentos Herbarios Chinos , Femenino , Imiquimod/farmacología , Interleucina-6 , Pulmón , Ratones , Ratones Endogámicos C57BL , Oseltamivir/farmacología , Fosfatos/farmacología , ARN , ARN Mensajero , Factor de Necrosis Tumoral alfa/genética , Pérdida de Peso
6.
Mediators Inflamm ; 2021: 1805147, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34790063

RESUMEN

Mycobacterium tuberculosis (Mtb) remains a significant threat to global health as it induces granuloma and systemic inflammatory responses during active tuberculosis. Mtb can induce macrophage pyroptosis, leading to the release of IL-1ß and tissue damage, promoting its spread. Here, we established an in vitro Mtb-infected macrophage model to seek an effective antipyroptosis agent. Baicalin, isolated from Radix Scutellariae, was found to reduce pyroptosis in Mtb-infected macrophages. Baicalin could inhibit activation of the PERK/eIF2α pathway and thus downregulates TXNIP expression and subsequently reduces activation of the NLRP3 inflammasome, resulting in reduced pyroptosis in Mtb-infected macrophages. In conclusion, baicalin reduced pyroptosis by inhibiting the PERK/TXNIP/NLRP3 axis and might thus be a new adjuvant host-directed therapy (HDT) drug.


Asunto(s)
Proteínas Portadoras/antagonistas & inhibidores , Flavonoides/farmacología , Mycobacterium tuberculosis/patogenicidad , Proteína con Dominio Pirina 3 de la Familia NLR/antagonistas & inhibidores , Piroptosis/efectos de los fármacos , Tiorredoxinas/antagonistas & inhibidores , eIF-2 Quinasa/antagonistas & inhibidores , Animales , Proteínas Portadoras/fisiología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Macrófagos/microbiología , Macrófagos/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína con Dominio Pirina 3 de la Familia NLR/fisiología , Células RAW 264.7 , Tiorredoxinas/fisiología , eIF-2 Quinasa/fisiología
7.
Nat Immunol ; 9(5): 542-50, 2008 May.
Artículo en Inglés | MEDLINE | ID: mdl-18391954

RESUMEN

Unbalanced production of proinflammatory cytokines and type I interferons in immune responses may lead to immunopathology; thus, the mechanisms that ensure the beneficial production of proinflammatory cytokines and type I interferons are of particular importance. Here we demonstrate that the phosphatase SHP-1 negatively regulated Toll-like receptor-mediated production of proinflammatory cytokines by inhibiting activation of the transcription factor NF-kappaB and mitogen-activated protein kinase. Simultaneously, SHP-1 increased the production of type I interferon mediated by Toll-like receptors and the helicase RIG-I by directly binding to and inhibiting activation of the kinase IRAK1. Our data demonstrate that SHP-1 contributes to immune homeostasis by balancing the production of proinflammatory cytokines and type I interferons in the innate immune response.


Asunto(s)
Interferón Tipo I/biosíntesis , Quinasas Asociadas a Receptores de Interleucina-1/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 6/metabolismo , Receptores Toll-Like/metabolismo , Animales , Dominio Catalítico/fisiología , Citocinas/biosíntesis , Homeostasis/inmunología , Inmunidad Innata , Factor 1 Regulador del Interferón/metabolismo , Quinasas Asociadas a Receptores de Interleucina-1/antagonistas & inhibidores , Quinasas Asociadas a Receptores de Interleucina-1/química , Macrófagos Peritoneales , Proteínas de la Membrana/inmunología , Ratones , Ratones Endogámicos C57BL , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , FN-kappa B/antagonistas & inhibidores , FN-kappa B/metabolismo , Proteínas del Tejido Nervioso/inmunología , Unión Proteica , Proteína Tirosina Fosfatasa no Receptora Tipo 6/inmunología , Receptores de Superficie Celular , Transducción de Señal , Receptores Toll-Like/inmunología
8.
J Autoimmun ; 113: 102491, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32565049

RESUMEN

Cell specific and cytokine targeted therapeutics have underperformed in systemic lupus erythematosus (SLE). Mesenchymal stem cells (MSCs) have emerged as a novel therapy to address the dysregulation in autoimmune diseases but also have limitations. Human gingiva derived MSCs (GMSCs) are superior in regulating immune responses. Here, we demonstrate that the adoptive transfer of GMSCs homes to and maintains in the kidney and has a robust therapeutic effect in a spontaneous lupus nephritis model. Specifically, GMSCs limits the development of autoantibodies as well as proteinuria, decreases the frequency of plasma cells and lupus nephritis histopathological scores by directly suppressing B cells activation, proliferation and differentiation. The blockage of CD39-CD73 pathway dramatically abrogates the suppressive capacities of GMSCs in vitro and in vivo and highlights the significance of this signaling pathway in SLE. Collectively, manipulation of GMSCs provides a promising strategy for the treatment of patients with SLE and other autoimmune diseases.


Asunto(s)
Encía/citología , Nefritis Lúpica/terapia , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/inmunología , 5'-Nucleotidasa/antagonistas & inhibidores , 5'-Nucleotidasa/metabolismo , Animales , Antígenos CD/metabolismo , Apirasa/antagonistas & inhibidores , Apirasa/metabolismo , Linfocitos B/inmunología , Linfocitos B/metabolismo , Diferenciación Celular/inmunología , Proliferación Celular , Células Cultivadas , Técnicas de Cocultivo , Modelos Animales de Enfermedad , Femenino , Proteínas Ligadas a GPI/antagonistas & inhibidores , Proteínas Ligadas a GPI/metabolismo , Humanos , Nefritis Lúpica/inmunología , Activación de Linfocitos , Ratones , Células Plasmáticas/inmunología , Células Plasmáticas/metabolismo , Cultivo Primario de Células , RNA-Seq , Transducción de Señal/efectos de los fármacos , Transducción de Señal/inmunología , Análisis de la Célula Individual
9.
Mediators Inflamm ; 2017: 2432904, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-28642632

RESUMEN

Mycobacterium tuberculosis (Mtb) remains a significant menace to global health as it induces granulomatous lung lesions and systemic inflammatory responses during active tuberculosis (TB). Micheliolide (MCL), a sesquiterpene lactone, was recently reported to have a function of relieving LPS-induced inflammatory response, but the regulative role of MCL on the immunopathology of TB still remains unknown. In this experiment, we examined the inhibitory effect of MCL on Mtb-induced inflammatory response in mouse macrophage-like cell line Raw264.7 by downregulating the activation of nuclear factor kappa B (NF-κB) and NLRP3 inflammasome. Evidences showed that MCL decreased the secretion of Mtb-induced inflammatory cytokines (IL-1ß and TNF-α) in a dose-dependent manner. Meanwhile, MCL dramatically suppressed Mtb-induced activation of iNOS and COX2 as well as subsequent production of NO. Furthermore, MCL inhibited Mtb-induced phosphorylation of Akt (Ser 473) in Raw264.7. According to our results, MCL plays an important role in modulating Mtb-induced inflammatory response through PI3K/Akt/NF-κB pathway and subsequently downregulating the activation of NLRP3 inflammasome. Therefore, MCL may represent as a potential drug candidate in the adjuvant treatment of TB by regulating host immune response.


Asunto(s)
Antiinflamatorios/uso terapéutico , Inflamasomas/efectos de los fármacos , Inflamasomas/metabolismo , Mycobacterium tuberculosis/inmunología , FN-kappa B/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Sesquiterpenos de Guayano/uso terapéutico , Animales , Interleucina-1beta/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Fosforilación/efectos de los fármacos , Células RAW 264.7 , Transducción de Señal/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo
10.
BMC Complement Altern Med ; 14: 400, 2014 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-25315519

RESUMEN

BACKGROUND: The Mycobacterium tuberculosis (Mtb) proteasome has been established as a viable target for the development of anti-tuberculosis agents. In this study, the inhibitory activities of 100 plant-derived natural products on the Mtb proteasome were analyzed to identify novel potential inhibitors. METHODS: The fluorescent substrate Suc-Leu-Leu-Val-Tyr-AMC can be hydrolyzed by the proteasome to release free AMC, the fluorescence of which is proportional to the proteasome activity. The inhibitory activities of 100 natural products (each at a final concentration of 200 µM) were detected by this method using MG132 as a positive control. RESULTS: Twelve of these natural products (10 of which were flavonoids) inhibited the activity of the Mtb proteasome by more than 65%. Comparison of the structural differences between the flavonoids with good inhibitory activity and those without inhibitory activity revealed that the hydroxyl at the flavonoid C ring C-3 or the hydroxyl/methoxyl at the flavonoid A ring C-6 were critical for the inhibition of proteasomal activity. CONCLUSIONS: These data indicate that flavonoids represent a basis for rational structural design in the process of novel anti-tuberculosis drug discovery.


Asunto(s)
Proteínas Bacterianas/antagonistas & inhibidores , Mycobacterium tuberculosis/efectos de los fármacos , Extractos Vegetales/química , Extractos Vegetales/farmacología , Plantas/química , Complejo de la Endopetidasa Proteasomal/efectos de los fármacos , Inhibidores de Proteasoma/química , Inhibidores de Proteasoma/farmacología , Proteínas Bacterianas/metabolismo , Cumarinas/metabolismo , Flavonoides/química , Flavonoides/farmacología , Leupeptinas/química , Mycobacterium tuberculosis/enzimología , Oligopéptidos/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo
11.
J Leukoc Biol ; 2024 Mar 22.
Artículo en Inglés | MEDLINE | ID: mdl-38518381

RESUMEN

Influenza virus infection is a worldwide challenge that causes heavy burdens on public health. The mortality rate of severe influenza patients is often associated with hyperactive immunological abnormalities characterized by hypercytokinemia. Due to the continuous mutations and the occurrence of drug-resistant influenza virus strains, the development of host-directed immunoregulatory drugs is urgently required. Platycodon grandiflorum is among the top 10 herbs of traditional Chinese medicine used to treat pulmonary diseases. As one of the major terpenoid saponins extracted from Platycodon grandiflorum, Platycodin D (PD) has been reported to play several roles, including anti-inflammation, analgesia, anti-cancer, hepatoprotection, and immunoregulation. However, the therapeutic roles of PD to treat influenza virus infection remains unknown. Here, we show that PD can protect the body weight loss in severely infected influenza mice, alleviate lung damage, and thus improve the survival rate. More specifically, PD protects flu mice via decreasing the immune cell infiltration into lungs and downregulating the overactivated inflammatory response. Western blot and immunofluorescence assays exhibited that PD could inhibit the activation of TAK1/IKK/NF-κB and MAPK pathways. Besides that, CETSA, SPR and immunoprecipitation assays indicated that PD binds with TRAF6 to decrease its K63 ubiquitination after R837 stimulation. Additionally, siRNA interference experiments exhibited that PD could inhibit the secretion of IL-1ß and TNF-α in TRAF6-dependent manner. Altogether, our results suggested that PD is a promising drug candidate for treating influenza. Our study also offered a scientific explanation for the commonly used Platycodon grandiflorum in many anti-epidemic classic formulas. Due to its host-directed regulatory role, PD may serve as an adjuvant therapeutic drug in conjunction with other antiviral drugs to treat the flu.

12.
Heliyon ; 10(1): e23225, 2024 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-38170002

RESUMEN

Bacterial infection remains a big concern in the patients of ICU, which is the main cause of life-threatening organ dysfunction, or even sepsis. The poor control of bacterial infection caused by antibiotic resistance, etc. or the overwhelming immune response are the most important patho genic factors in intensive care unit (ICU) patients. As main pathogens, antibiotic-resistant bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), impose serious challenges during sepsis and require alternative therapeutic options. Irisflorentin (IFL) is one of the major bioactive compounds isolated from the roots of Belamcanda chinensis (Shegan). In this study, IFL could suppress inflammatory response induced by MRSA or a synthetic mimic of bacterial lipoprotein (Pam3CSK4). IFL treatment enhanced the ability of macrophages to phagocytose bacteria likely through up-regulating the expression of phagocytic receptors SR-A1 and FcγR2a. Furthermore, IFL inhibited Pam3CSK4-induced production of pro-inflammatory cytokines, including IL-6 and TNF-α in Raw 264.7 cells, mouse primary macrophages or dendritic cells. IFL treatment also inhibited heat-killed MRSA-induced secretion of IL-6 and TNF-α in mouse bone marrow-derived macrophages. Moreover, IFL attenuated M1 polarization of macrophages as indicated by the down-regulated expression of its polarization markers CD86 and iNOS. Mechanistically, IFL markedly decreased the Pam3CSK4-induced activation of ERK, JNK or p38 MAPK pathways in macrophages. Taken together, IFL may serve as a promising compound for the therapy of bacterial infection, particularly those caused by antibiotic-resistant bacteria, such as MRSA.

13.
Cancers (Basel) ; 16(14)2024 Jul 13.
Artículo en Inglés | MEDLINE | ID: mdl-39061173

RESUMEN

BACKGROUNDS: Abnormal metabolism is the hallmark of hepatocellular carcinoma. Targeting energy metabolism has become the major focus of cancer therapy. The natural product, sanguinarine, displays remarkable anti-tumor properties by disturbing energy homeostasis; however, the underlying mechanism has not yet been elucidated. METHODS: The anticancer activity of sanguinarine was determined using CCK-8 and colony formation assay. Morphological changes of induced cell death were observed under electron microscopy. Necroptosis and apoptosis related markers were detected using western blotting. PKM2 was identified as the target by transcriptome sequencing. Molecular docking assay was used to evaluate the binding affinity of sanguinarine to the PKM2 molecule. Furthermore, Alb-CreERT2; PKM2loxp/loxp; Rosa26RFP mice was used to construct the model of HCC-through the intervention of sanguinarine in vitro and in vivo-to accurately explore the regulation effect of sanguinarine on cancer energy metabolism. RESULTS: Sanguinarine inhibited tumor proliferation, metastasis and induced two modes of cell death. Molecular docking of sanguinarine with PKM2 showed appreciable binding affinity. PKM2 kinase activity and aerobic glycolysis rate declined, and mitochondrial oxidative phosphorylation was inhibited by sanguinarine application; these changes result in energy deficits and lead to necroptosis. Additionally, sanguinarine treatment prevents the translocation of PKM2 into the nucleus and suppresses the interaction of PKM2 with ß-catenin; the transcriptional activity of PKM2/ß-catenin signaling and its downstream genes were decreased. CONCLUSIONS: Sanguinarine showed remarkable anti-HCC activity via regulating energy metabolism by PKM2/ß-catenin signaling. On the basis of these investigations, we propose that sanguinarine might be considered as a promising compound for discovery of anti-HCC drugs.

14.
Food Funct ; 15(10): 5287-5299, 2024 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-38639730

RESUMEN

Catechol-O-methyltransferase (COMT) plays a central role in the metabolic inactivation of endogenous neurotransmitters and xenobiotic drugs and hormones having catecholic structures. Its inhibitors are used in clinical practice to treat Parkinson's disease. In this study, a fluorescence-based visualization inhibitor screening method was developed to assess the inhibition activity on COMT both in vitro and in living cells. Following the screening of 94 natural products, Pu-erh tea extract exhibited the most potent inhibitory effect on COMT with an IC50 value of 0.34 µg mL-1. In vivo experiments revealed that Pu-erh tea extract substantially hindered COMT-mediated levodopa metabolism in rats, resulting in a significant increase in levodopa levels and a notable decrease in 3-O-methyldopa in plasma. Subsequently, the chemical components of Pu-erh tea were analyzed using UHPLC-Q-Exactive Orbitrap HRMS, identifying 24 major components. Among them, epigallocatechin gallate, gallocatechin gallate, epicatechin gallate, and catechin gallate exhibited potent inhibition of COMT activity with IC50 values from 93.7 nM to 125.8 nM and were the main bioactive constituents in Pu-erh tea responsible for its COMT inhibition effect. Inhibition kinetics analyses and docking simulations revealed that these compounds competitively inhibit COMT-mediated O-methylation at the catechol site. Overall, this study not only explained how Pu-erh tea catechins inhibit COMT, suggesting Pu-erh tea as a potential dietary intervention for Parkinson's disease, but also introduced a new strategy for discovering COMT inhibitors more effectively.


Asunto(s)
Catequina , Inhibidores de Catecol O-Metiltransferasa , Catecol O-Metiltransferasa , Levodopa , Extractos Vegetales , Ratas Sprague-Dawley , , Animales , Ratas , Inhibidores de Catecol O-Metiltransferasa/farmacología , Catecol O-Metiltransferasa/metabolismo , Catequina/análogos & derivados , Catequina/farmacología , Catequina/química , Levodopa/metabolismo , Té/química , Extractos Vegetales/farmacología , Extractos Vegetales/química , Masculino , Humanos , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/metabolismo , Camellia sinensis/química , Simulación del Acoplamiento Molecular
15.
Autoimmunity ; 57(1): 2387076, 2024 Jul 30.
Artículo en Inglés | MEDLINE | ID: mdl-39229919

RESUMEN

OBJECTIVE: This study aims to explore the effect of NONHSAT042241 on the function of rheumatoid arthritis -fibroblast-like synoviocyte (RA-FLS) and the underlying mechanisms. METHODS: RA-FLS was treated with NONHSAT042241 overexpression and NONHSAT042241 knockdown lentiviruses. Cell counting kit-8 (CCK-8) assay, colony formation assay, flow cytometry, Transwell assay, western-blot, ELISA, and qRT-PCR were used to measure the changes of cell proliferation, apoptosis, invasion, secretion of inflammatory cytokines and matrix metalloproteinases (MMPs). Fluorescent in situ hybridization (FISH) assay, RNA pull-down assay, mass spectrometry (MS) and RNA immunoprecipitation (RIP) were used to find the target proteins that bond to NONHSAT042241, and western-blot was used to detect the expression of related proteins of Wnt/ß-catenin signaling pathway. RESULTS: Overexpression of NONHSAT042241 inhibited the proliferation of RA-FLS (p < 0.05), invasion, secretion of pro-inflammatory factors (IL-1and IL-6) and MMPs (MMP-1 and MMP-3) (p < 0.05), and elevated the level of pro-apoptotic factors (Bax and cleaved caspase3), while NONHSAT042241 knockdown had the opposite effect. NONHSAT042241 can directly bind to hnRNP D, and down-regulated the expression of ß-catenin (p < 0.05), p-GSK-3ß (p < 0.05), Cyclin D1 (p < 0.05), PCNA (p < 0.05), and thus reduced the cell proliferation. CONCLUSION: NONHSAT042241 may inhibit FLS-mediated rheumatoid synovial proliferation, inflammation and aggression. The underlying mechanisms may be that NONHSAT042241 inhibits the activity of Wnt/ß-catenin signaling.


Asunto(s)
Artritis Reumatoide , Proliferación Celular , Inflamación , ARN Largo no Codificante , Sinoviocitos , Vía de Señalización Wnt , Humanos , Artritis Reumatoide/metabolismo , Artritis Reumatoide/patología , Artritis Reumatoide/genética , Sinoviocitos/metabolismo , Sinoviocitos/patología , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Inflamación/metabolismo , Membrana Sinovial/metabolismo , Membrana Sinovial/patología , Membrana Sinovial/inmunología , Apoptosis , beta Catenina/metabolismo , Células Cultivadas
16.
J Ethnopharmacol ; 321: 117553, 2024 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-38065349

RESUMEN

ETHNOPHARMACOLOGICAL RELEVANCE: Fei-Yan-Qing-Hua decoction (FYQHD), derived from the renowned formula Ma Xing Shi Gan tang documented in Zhang Zhong Jing's "Treatise on Exogenous Febrile Disease" during the Han Dynasty, has demonstrated notable efficacy in the clinical treatment of pneumonia resulting from bacterial infection. However, its molecular mechanisms underlying the therapeutic effects remains elusive. AIM OF THE STUDY: This study aimed to investigate the protective effects of FYQHD against lipopolysaccharide (LPS) and carbapenem-resistant Klebsiella pneumoniae (CRKP)-induced sepsis in mice and to elucidate its specific mechanism of action. MATERIALS AND METHODS: Sepsis models were established in mice through intraperitoneal injection of LPS or CRKP. FYQHD was administered via gavage at low and high doses. Serum cytokines, bacterial load, and pathological damage were assessed using enzyme-linked immunosorbent assay (ELISA), minimal inhibitory concentration (MIC) detection, and hematoxylin and eosin staining (H&E), respectively. In vitro, the immunoregulatory effects of FYQHD on macrophages were investigated through ELISA, MIC, quantitative real-time PCR (Q-PCR), immunofluorescence, Western blot, and a network pharmacological approach. RESULTS: The application of FYQHD in the treatment of LPS or CRKP-induced septic mouse models revealed significant outcomes. FYQHD increased the survival rate of mice exposed to a lethal dose of LPS to 33.3%, prevented hypothermia (with a rise of 3.58 °C), reduced pro-inflammatory variables (including TNF-α, IL-6, and MCP-1), and mitigated tissue damage in LPS or CRKP-induced septic mice. Additionally, FYQHD decreased bacterial load in CRKP-infected mice. In vitro, FYQHD suppressed the expression of inflammatory cytokines in macrophages activated by LPS or HK-CRKP. Mechanistically, FYQHD inhibited the PI3K/AKT/mTOR/4E-BP1 signaling pathway, thereby suppressing the translational level of inflammatory cytokines. Furthermore, it reduced the expression of HMGB1/RAGE, a positive feedback loop in the inflammatory response. Moreover, FYQHD was found to enhance the phagocytic activity of macrophages by upregulating the expression of phagocytic receptors such as CD169 and SR-A1. CONCLUSION: FYQHD provides protection against bacterial sepsis by concurrently inhibiting the inflammatory response and augmenting the phagocytic ability of immune cells.


Asunto(s)
Proteína HMGB1 , Sepsis , Ratones , Animales , Lipopolisacáridos/farmacología , Proteína HMGB1/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Transducción de Señal , Citocinas/metabolismo , Fagocitosis , Sepsis/tratamiento farmacológico
17.
Cell Mol Immunol ; 21(1): 80-90, 2024 01.
Artículo en Inglés | MEDLINE | ID: mdl-38082146

RESUMEN

Regulatory T (Treg) cells play an essential role in maintaining immune balance across various physiological and pathological conditions. However, the mechanisms underlying Treg homeostasis remain incompletely understood. Here, we report that RIPK1 is crucial for Treg cell survival and homeostasis. We generated mice with Treg cell-specific ablation of Ripk1 and found that these mice developed fatal systemic autoimmunity due to a dramatic reduction in the Treg cell compartment caused by excessive cell death. Unlike conventional T cells, Treg cells with Ripk1 deficiency were only partially rescued from cell death by blocking FADD-dependent apoptosis. However, simultaneous removal of both Fadd and Ripk3 completely restored the homeostasis of Ripk1-deficient Treg cells by blocking two cell death pathways. Thus, our study highlights the critical role of RIPK1 in regulating Treg cell homeostasis by controlling both apoptosis and necroptosis, thereby providing novel insights into the mechanisms of Treg cell homeostasis.


Asunto(s)
Apoptosis , Linfocitos T Reguladores , Animales , Ratones , Muerte Celular , Homeostasis
18.
Int Immunopharmacol ; 132: 111889, 2024 May 10.
Artículo en Inglés | MEDLINE | ID: mdl-38531202

RESUMEN

Host-directed therapy (HDT) is a new adjuvant strategy that interfere with host cell factors that are required by a pathogen for replication or persistence. In this study, we assessed the effect of dehydrozaluzanin C-derivative (DHZD), a modified compound from dehydrozaluzanin C (DHZC), as a potential HDT agent for severe infection. LPS-induced septic mouse model and Carbapenem resistant Klebsiella pneumoniae (CRKP) infection mouse model was used for testing in vivo. RAW264.7 cells, mouse primary macrophages, and DCs were used for in vitro experiments. Dexamethasone (DXM) was used as a positive control agent. DHZD ameliorated tissue damage (lung, kidney, and liver) and excessive inflammatory response induced by LPS or CRKP infection in mice. Also, DHZD improved the hypothermic symptoms of acute peritonitis induced by CRKP, inhibited heat-killed CRKP (HK-CRKP)-induced inflammatory response in macrophages, and upregulated the proportions of phagocytic cell types in lungs. In vitro data suggested that DHZD decreases LPS-stimulated expression of IL-6, TNF-α and MCP-1 via PI3K/Akt/p70S6K signaling pathway in macrophages. Interestingly, the combined treatment group of DXM and DHZD had a higher survival rate and lower level of IL-6 than those of the DXM-treated group; the combination of DHZD and DXM played a synergistic role in decreasing IL-6 secretion in sera. Moreover, the phagocytic receptor CD36 was increased by DHZD in macrophages, which was accompanied by increased bacterial phagocytosis in a clathrin- and actin-dependent manner. This data suggests that DHZD may be a potential drug candidate for treating bacterial infections.


Asunto(s)
Infecciones por Klebsiella , Klebsiella pneumoniae , Macrófagos , Fagocitosis , Sepsis , Animales , Ratones , Fagocitosis/efectos de los fármacos , Infecciones por Klebsiella/tratamiento farmacológico , Infecciones por Klebsiella/inmunología , Klebsiella pneumoniae/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Células RAW 264.7 , Sepsis/tratamiento farmacológico , Sepsis/inmunología , Masculino , Lipopolisacáridos , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Transducción de Señal/efectos de los fármacos , Citocinas/metabolismo , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico
19.
Heliyon ; 10(13): e34055, 2024 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-39071618

RESUMEN

Background: Rujin Jiedu decoction (RJJDD) is a classical prescription of Traditional Chinese Medicine that has long been applied to treat pneumonia caused by external infection, but whether and how it benefits influenza virus therapy remains largely unclear. The aim of this study was to investigate the anti-inflammatory effect of RJJDD on the mouse model of influenza and to explore its potential mechanism. Methods: The mice were mock-infected with PBS or infected with PR8 virus followed by treatment with RJJDD or antiviral oseltamivir. The weight loss and morbidity of mice were monitored daily. Network pharmacology is used to explore the potential pathways that RJJDD may modulate. qRT-PCR and ELISA were performed to assess the expression of inflammatory cytokines in the lung tissue and macrophages. The intestinal feces were collected for 16S rDNA sequencing to assess the changes in gut microbiota. Results: We demonstrate that RJJDD protects against IAV-induced pneumonia. Comprehensive network pharmacology analyses of the Mass Spec-identified components of RJJDD suggest that RJJDD may act through down-regulating key signaling pathways producing inflammatory cytokines, which was experimentally confirmed by cytokine expression analysis in IAV-infected mouse lung tissues and IAV single-strand RNA mimic R837-induced macrophages. Furthermore, gut microbiota analysis indicates that RJJDD prevented IAV-induced dysbiosis of host intestinal flora, thereby offering a mechanistic explanation for RJJDD's efficacy in influenza pneumonia. Conclusion: This study defines a previously uncharacterized role for RJJDD in protecting against influenza likely by maintaining homeostasis of gut microbiota, and provides a new therapeutic option for severe influenza.

20.
Front Pharmacol ; 15: 1309682, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38476329

RESUMEN

Introduction: Significant attention has been paid to myocardial damage mediated by the single-stranded RNA virus. Qingfei Paidu decoction (QFPDD) has been proved to protect the damage caused by the influenza virus A/PR/8/1934 (PR8), but its specific mechanism is unclear. Methods: Molecular biological methods, together with network pharmacology, were used to analyze the effects and underlying mechanism of QFPDD treatment on PR8-induced myocardial damage to obtain insights into the treatment of COVID-19-mediated myocardial damage. Results: Increased apoptosis and subcellular damage were observed in myocardial cells of mice infected by PR8. QFPDD treatment significantly inhibited the apoptosis and subcellular damage induced by the PR8 virus. The inflammatory factors IFN-ß, TNF-α, and IL-18 were statistically increased in the myocardia of the mice infected by PR8, and the increase in inflammatory factors was prevented by QFPDD treatment. Furthermore, the expression levels or phosphorylation of necroptosis-related proteins RIPK1, RIPK3, and MLKL were abnormally elevated in the group of infected mice, while QFPDD restored the levels or phosphorylation of these proteins. Our study demonstrated that HIF-1α is a key target of QFPDD in the treatment of influenza virus-mediated injury. The HIF-α level was significantly increased by PR8 infection. Both the knockdown of HIF-1α and treatment of the myocardial cell with QFPDD significantly reversed the increased inflammatory factors during infection. Overexpression of HIF-1α reversed the inhibition effects of QFPDD on cytokine expression. Meanwhile, seven compounds from QFPDD may target HIF-1α. Conclusion: QFPDD can ameliorate influenza virus-mediated myocardial damage by reducing the degree of cell necroptosis and apoptosis, inhibiting inflammatory response and the expression of HIF-1α. Thus, our results provide new insights into the treatment of respiratory virus-mediated myocardial damage.

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