IκB Kinase Research Articles

Potential effect of kaempferol against various malignancies: recent advances and perspectives

ABSTRACT Kaempferol is a flavonoid present in many eatable plants. Researchers discovered a link between consuming foods high in kaempferol and lowering the threat for acquiring many diseases, including cardiovascular disease, diabetes, obesity and cancer. Kaempferol can inhibit AHR transcription, modulate ERK signalling pathway and NF-κB pathways, block MAPK and AP1 signalling pathways and perform more anticancer roles. Kaempferol also has the ability to act against diabetes via suppressed phosphorylation of (IKK) IkB kinase, (IRS-1) insulin receptor substrate-1 and (IKK) IkB kinase through the hepatic IKK/NF-B signalling pathways and significantly enhanced insulin secretion and synthesis. Kaempferol protects cardiomyocytes from anoxia/reoxygenation (A/R)-induced damage by lowering LDH release, improving cell survival, reducing A/R-induced ROS formation, and release of cytochrome c. This knowledge may aid in understanding health advantages of medicinal plants that contain kaempferol and may lead to the development of the flavonoid as a potential agents for disease aversion and therapy.

A multiomic investigation of lung adenocarcinoma molecular subtypes.

Lung adenocarcinoma-an aggressive and life-threatening malignancy-is a type of non-small-cell lung cancer. Despite medical advancements, the prognosis of lung adenocarcinoma remains unfavorable, likely because of its heterogeneous nature. Furthermore, few subtype-specific treatments are available for lung adenocarcinoma. This study was conducted to explore the molecular subtypes of lung adenocarcinoma. We performed a joint analysis of transcriptome and proteome data from East Asian patients with lung adenocarcinoma (nonsmokers, 86.5%). Four novel subtypes were identified based on distinct molecular characteristics: subtypes I, II, III, and IV. In patients with subtype I lung adenocarcinoma, eukaryotic translation initiation factor 4 gamma 1 activates cell proliferation; inhibiting this factor suppresses tumor growth, and reducing its level induces autophagy. Subtype II is characterized by Kristen rat sarcoma viral oncogene homolog-activating oncogenesis; the onset age of this subtype is the lowest among all subtypes. Subtype III manifests as an advanced disease at diagnosis; it is characterized by a core serum response-related oncogenic signature, which indicates poor overall survival in Western patients with lung cancer. Subtype IV is more common in men than in women; it has astroglial characteristics. A Connectivity Map analysis revealed that the oncogenic expression patterns corresponding to subtypes I, II, III, and IV can be reversed by the inhibitors of Inhibitor of κB (IκB) kinase (eg, withaferin A), mammalian target of rapamycin (eg, everolimus), Src proto-oncogene (Src) (eg, saracatinib), and Transforming Growth Factor (TGF)-β/Smad (eg, LY-364947), respectively. This study introduced an innovative multiomics data analysis pipeline. Using this approach, we successfully identified four molecular subtypes of lung adenocarcinoma and their candidate therapeutic agents. The newly identified subtypes can be combined with the current biomarkers to generate a comprehensive roadmap for treatment decision-making.

Molecular cloning and functional characterization of pigeon IKKε

Inhibitor of nuclear factor kappa-B kinase ε (IKKε), a member of the non-canonical IκB kinase family, plays a critical role in connecting various signaling pathways associated with the initiation of type I interferon (IFN) production. Although the importance of IKKε in innate immunity has been well established in mammals and fish, its characterization and function in pigeons have remained largely unexplored. In this study, we successfully cloned pigeon IKKε (piIKKε) from pigeon embryo fibroblasts (PEFs) for the first time. This gene encodes 722 amino acids and shares high amino acid similarity with its duck and goose counterparts. piIKKε showed a diffuse cytoplasmic distribution and broad expression in all tissues examined. Overexpression of piIKKε in PEFs significantly activated the IFN-β promoter, with both the kinase and CC domains of piIKKε playing key roles in initiating IFN-β expression. Knockdown of piIKKε using small interfering RNA significantly reduced the levels of IFN-β induced by NDV, AIV, poly (I:C), or SeV. Furthermore, the presence of piIKKε resulted in a remarkable reduction in the replication of both avian influenza virus (AIV) H9N2 and Newcastle disease virus (NDV) in PEFs. Our results demonstrate that piIKKε plays a critical role in mediating antiviral innate immunity in pigeons.

DLG1 promotes the antiviral innate immune response by inhibiting p62-mediated autophagic degradation of IKKε.

The type-I interferon (IFN-I) signaling pathway is the first line of antiviral innate immunity. It must be precisely regulated against virus-induced damage. The tightly regulated mechanisms of action of host genes in the antiviral innate immune signaling pathway are still worth studying. Here, we report a novel role of DLG1 in positively regulating the IκB kinase epsilon (IKKε)-mediated IFN-I signaling response against negative-stranded RNA virus replication, whereas the RNA virus inhibits the expression of DLG1 for immune escape. Importantly, the E3 ligase March2 interacts with and promotes K27-linked polyubiquitination of IKKε, and p62 is a cargo receptor that recognizes ubiquitinated IKKε for eventual autophagic degradation. Together, the current findings elucidate the role of DLG1 in the antiviral IFN-I signaling pathway and viral infection repression.

FAT10 is phosphorylated by IKKβ to inhibit the antiviral type-I interferon response.

IFN-I secretion provides a rapid host defense against infection with RNA viruses. Within the host cell, viral RNA triggers the activation of the RIG-I signaling pathway, leading to the production of IFN-I. Because an exaggerated IFN-I response causes severe tissue damage, RIG-I signaling is tightly regulated. One of the factors that control the IFN-I response is the ubiquitin-like modifier FAT10, which is induced by TNF and IFNγ and targets covalently FAT10-linked proteins for proteasomal degradation. However, the mechanism of how FAT10 modulates IFN-I secretion remains to be fully elucidated. Here, we provide strong evidence that FAT10 is phosphorylated by IκB kinase β (IKKβ) upon TNF stimulation and during influenza A virus infection on several serine and threonine residues. FAT10 phosphorylation increases the binding of FAT10 to the TRAF3-deubiquitylase OTUB1 and its FAT10-mediated activation. Consequently, FAT10 phosphorylation results in a low ubiquitylation state of TRAF3, which is unable to maintain interferon regulatory factor 3 phosphorylation and downstream induction of IFN-I. Taken together, we reveal a mechanism of how phosphorylation of FAT10 limits the production of tissue-destructive IFN-I in inflammation.

Methyl 2-[3-(4-hydroxyphenyl)prop-2-enoylamino]-3-phenylpropanoate Is a Potent Cell-Permeable Anti-Cytokine Compound To Inhibit Inflammatory Cytokines in Monocyte/Macrophage-Like Cells.

Cytokines are signaling molecules involved in inflammation process. Interleukin (IL)-6 is one of pivotal inflammatory cytokines associated with many human diseases. Therefore, there are on-going efforts to find a therapeutic to inhibit IL-6 and other cytokines. Methyl 2-[3-(4-hydroxyphenyl)prop-2-enoylamino]-3-phenylpropanoate (MHPAP) is a phenolic amide ester, transported better than its non-ester form (NEF) in monocyte/macrophage-like cells. However, there is no information about the effects of their cell permeability on cytokines. Therefore, the effects of MHPAP and NEF on cytokines were investigated in lipopolysaccharide (LPS)-stimulated THP-1 and human peripheral blood mononuclear cells (PBMCs). In the THP-1 cells, MHPAP significantly inhibited IL-6, IL-1beta, IL-8, and tumor necrosis factor (TNF)-alpha (P < 0.05), but NEF showed no effects. MHPAP also inhibited nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 phosphorylation in the THP-1 cells (P < 0.05), without significant effects on c-FOS, ATF-2, and JUN phosphorylations. Because NF-κB p65 is phosphorylated by IκB kinase (IKK), in silico analysis was performed on IKK. MHPAP was found to bind to IKK better than an IKK inhibitor ((E)-2-fluoro-4'-methoxystilbene). Furthermore, MHPAP inhibited the luminescence increased in the LPS-stimulated NF-κB-Luc2 THP-1 cells. As anticipated, MHPAP was also found to inhibit IL-6, IL-1beta, IL-8, and TNF-alpha significantly in LPS-stimulated PBMCs (P < 0.05). Especially, MHPAP inhibited IL-6 and IL-1beta with an IC50 of 0.85 and 0.87 µM, better than IL-8 (1.58 µM) and TNF-alpha (1.22 µM) in the cells. Altogether, the data suggest that cell permeability may have a significant impact on MHPAP's ability to inhibit cytokines and MHPAP may be used as a potent cell-permeable compound to inhibit inflammatory cytokines in monocyte/macrophage-like cells. SIGNIFICANCE STATEMENT: Potential effects of MHPAP and NEF on inflammatory cytokines (IL-6, IL-8, IL-1beta, and TNF-alpha) were investigated in LPS-stimulated THP-1 and PBMCs. Cell transport had a great impact on cytokine inhibition in the cells. MHPAP was also found to inhibit NF-κB pathway, which was supported by in silico and NF-κB reporter (Luc)-THP-1 data. Also, in LPS-stimulated PBMCs, MHPAP significantly inhibited IL-6, IL-1beta, IL-8, and TNF-alpha, suggesting that MHPAP may be a potent cell-permeable compound to inhibit inflammatory cytokines in monocyte/macrophage-like cells.

A conserved core region of the scaffold NEMO is essential for signal-induced conformational change and liquid-liquid phase separation

Scaffold proteins help mediate interactions between protein partners, often to optimize intracellular signaling. Herein, we use comparative, biochemical, biophysical, molecular, and cellular approaches to investigate how the scaffold protein NEMO contributes to signaling in the NF-κB pathway. Comparison of NEMO and the related protein optineurin from a variety of evolutionarily distant organisms revealed that a central region of NEMO, called the Intervening Domain (IVD), is conserved between NEMO and optineurin. Previous studies have shown that this central core region of the IVD is required for cytokine-induced activation of IκB kinase (IKK). We show that the analogous region of optineurin can functionally replace the core region of the NEMO IVD. We also show that an intact IVD is required for the formation of disulfide-bonded dimers of NEMO. Moreover, inactivating mutations in this core region abrogate the ability of NEMO to form ubiquitin-induced liquid-liquid phase separation droplets invitro and signal-induced puncta invivo. Thermal and chemical denaturation studies of truncated NEMO variants indicate that the IVD, while not intrinsically destabilizing, can reduce the stability of surrounding regions of NEMO due to the conflicting structural demands imparted on this region by flanking upstream and downstream domains. This conformational strain in the IVD mediates allosteric communication between the N-and C-terminal regions of NEMO. Overall, these results support a model in which the IVD of NEMO participates in signal-induced activation of the IKK/NF-κB pathway by acting as a mediator of conformational changes in NEMO.

Molecular insights into the anti-inflammatory activity of fermented pineapple juice using multimodal computational studies.

Pineapple has been recognized for its potential to enhance health and well-being. This study aimed to gain molecular insights into the anti-inflammatory properties of fermented pineapple juice using multimodal computational studies. In this study, pineapple juice was fermented using Lactobacillus paracasei, and the solution underwent liquid chromatography-mass spectrometry analysis. Network pharmacology was applied to investigate compound interactions and targets. In silico methods assessed compound bioactivities. Protein-protein interactions, network topology, and enrichment analysis identified key compounds. Molecular docking explored compound-receptor interactions in inflammation regulation. Molecular dynamics simulations were conducted to confirm the stability of interactions between the identified crucial compounds and their respective receptors. The study revealed several compounds including short-chain fatty acids, peptides, dihydroxyeicosatrienoic acids, and glycerides that exhibited promising anti-inflammatory properties. Leucyl-leucyl-norleucine and Leu-Leu-Tyr exhibited robust and stable interactions with mitogen-activated protein kinase 14 and IκB kinase β, respectively, indicating their potential as promising therapeutic agents for inflammation modulation. This proposition is grounded in the pivotal involvement of these two proteins in inflammatory signaling pathways. These findings provide valuable insights into the anti-inflammatory potential of these compounds, serving as a foundation for further experimental validation and exploration. Future studies can build upon these results to advance the development of these compounds as effective anti-inflammatory agents.

SERPINB1 promotes Senecavirus A replication by degrading IKBKE and regulating the IFN pathway via autophagy.

Senecavirus A (SVA) is an emerging picornavirus associated with vesicular disease, which wide spreads around the world. It has evolved multiple strategies to evade host immune surveillance. The mechanism and pathogenesis of the virus infection remain unclear. In this study, we show that SERPINB1, a member of the SERPINB family, promotes SVA replication, and regulates both innate immunity and the autophagy pathway. SERPINB1 catalyzes K48-linked polyubiquitination of IκB kinase epsilon (IKBKE) and degrades IKBKE through the proteasome pathway. Inhibition of IKBKE expression by SERPINB1 induces autophagy to decrease type I interferon signaling, and ultimately promotes SVA proliferation. These results provide importantly the theoretical basis of SVA replication and pathogenesis. SERPINB1 could be a potential therapeutic target for the control of viral infection.

The effects of berteroin on inflammatory mediators and antioxidant enzymes expression in human periodontal ligament cells.

Berteroin is a bioactive substance classified as an isothiocyanate found in cruciferous vegetables such as cabbage, arugula, and salad leaves. In this study, we aimed to determine whether berteroin exerts anti-inflammatory effects on human periodontal ligament cells (HPDLCs), a resident cells of periodontal tissue. Berteroin suppressed interleukin (IL)-1β or tumor necrosis factor (TNF)-α-induced chemokines (C-C motif chemokine ligand (CCL)2, CCL20, C-X-C motif chemokine ligand (CXCL)10, IL-8, and IL-6) production and intercellular adhesion molecule (ICAM)-1 expression in HPDLCs. In addition, berteroin inhibited phosphorylation of IκB kinase (IKK)- α/ β, nuclear factor (NF)- κB p65, and IκB- α and degradation of IκB- α in the NF-κB pathway induced by IL-1 β or TNF- α stimulation. Moreover, berteroin could inhibit signal transducer and activator of transcription (STAT)3 phosphorylation in TNF- α -stimulated HPDLC. Furthermore, berteroin increased the expression of the antioxidant enzymes, heme oxygenase (HO)-1 and NAD(P)H quinone dehydrogenase (NQO)1, in IL-1 β or TNF- α -stimulated HPDLCs. These results suggest that berteroin may decrease the production of inflammatory mediators in HPDLCs by suppressing the NF-κB pathway, and may also decrease the local reactive oxygen species (ROS) production in periodontal lesions by increasing the production of antioxidant enzymes.

Methiothepin downregulates SNAP-23 and inhibits degranulation of rat basophilic leukemia cells and mouse bone marrow-derived mast cells.

In the present study, we found that methiothepin (a nonselective 5-hydroxytryptamine [5-HT] receptor antagonist) inhibited antigen-induced degranulation in rat basophilic leukemia cells and mouse bone marrow-derived mast cells. Although antigen stimulation induces release of histamine and serotonin (5-HT) by exocytosis and mast cells express several types of 5-HT receptor, the detailed role of these receptors remains unclear. Here, pretreatment of cells with methiothepin attenuated increased intracellular Ca2+ concentration, phosphorylated critical upstream signaling components (Src family tyrosine kinases, Syk, and PLCγ1), and suppressed TNF-α secretion via inhibition of Akt (a Ser/Thr kinase activated by PI3K)and ERK phosphorylation. Furthermore, it inhibited PMA/ionomycin-induced degranulation; this finding suggested that methiothepin affected downstream signaling. IκB kinase β phosphorylates synaptosomal associated protein 23, which regulates the fusion events of the secretory granule/plasma membrane after mast cell activation, resulting in degranulation. We showed that methiothepin blocked PMA/ionomycin-induced phosphorylation of synaptosomal associated protein 23 by inhibiting its interaction with IκB kinase β. Together with the results of selective 5-HT antagonists, it is suggested that methiothepin inhibits mast cell degranulation by downregulating upstream signaling pathways and exocytotic fusion machinery through mainly 5-HT1A receptor. Our findings provide that 5-HT antagonists may be used to relieve allergic reactions.

IκB kinase-α coordinates BRD4 and JAK/STAT signaling to subvert DNA damage-based anticancer therapy.

Activation of the IκB kinase (IKK) complex has recurrently been linked to colorectal cancer (CRC) initiation and progression. However, identification of downstream effectors other than NF-κB has remained elusive. Here, analysis of IKK-dependent substrates in CRC cells after UV treatment revealed that phosphorylation of BRD4 by IKK-α is required for its chromatin-binding at target genes upon DNA damage. Moreover, IKK-α induces the NF-κB-dependent transcription of the cytokine LIF, leading to STAT3 activation, association with BRD4 and recruitment to specific target genes. IKK-α abrogation results in defective BRD4 and STAT3 functions and consequently irreparable DNA damage and apoptotic cell death upon different stimuli. Simultaneous inhibition of BRAF-dependent IKK-α activity, BRD4, and the JAK/STAT pathway enhanced the therapeutic potential of 5-fluorouracil combined with irinotecan in CRC cells and is curative in a chemotherapy-resistant xenograft model. Finally, coordinated expression of LIF and IKK-α is a poor prognosis marker for CRC patients. Our data uncover a functional link between IKK-α, BRD4, and JAK/STAT signaling with clinical relevance.

Classical swine fever virus NS5A protein antagonizes innate immune response by inhibiting the NF-κB signaling

The NS5A non-structural protein of classical swine fever virus (CSFV) is a multifunctional protein involved in viral genomic replication, protein translation, assembly of infectious virus particles, and regulation of cellular signaling pathways. Previous report showed that NS5A inhibited nuclear factor kappa B (NF-κB) signaling induced by poly(I:C); however, the mechanism involved has not been elucidated. Here, we reported that NS5A directly interacted with NF-κB essential modulator (NEMO), a regulatory subunit of the IκB kinase (IKK) complex, to inhibit the NF-κB signaling pathway. Further investigations showed that the zinc finger domain of NEMO and the aa 126–250 segment of NS5A are essential for the interaction between NEMO and NS5A. Mechanistic analysis revealed that NS5A mediated the proteasomal degradation of NEMO. Ubiquitination assay showed that NS5A induced the K27-linked but not the K48-linked polyubiquitination of NEMO for proteasomal degradation. In addition, NS5A blocked the K63-linked polyubiquitination of NEMO, thus inhibiting IKK phosphorylation, IκBα degradation, and NF-κB activation. These findings revealed a novel mechanism by which CSFV inhibits host innate immunity, which might guide the drug design against CSFV in the future.

Panax notoginseng saponins alleviate diabetic retinopathy by inhibiting retinal inflammation: Association with the NF-κB signaling pathway

Ethnopharmacological relevanceDiabetic retinopathy (DR) is a neurovascular disease that causes blindness in adults and is the most serious and common complication of diabetes mellitus. Retinal inflammation is an early stage of DR, and it is believed to play a crucial role in the development of DR. Panax notoginseng saponins (PNS) are the major active constituent in the main root of P. notoginseng, and they exhibit various biological activities, including anti-inflammatory, antioxidant, neuroprotective, and immunomodulatory functions. However, the protective effects and underlying mechanisms of PNS against DR remain unclear. Aim of the studyThis study aimed to investigate the alleviation effects of PNS on DR and the mechanisms involved. Furthermore, it intended to explore the major components that exert efficacy in vivo. Materials and methodsStreptozotocin (STZ) was administered intraperitoneally to Sprague Dawley rats, and PNS was administered orally for 1 month after 2 months of STZ injection. The morphological structure of the retina and retinal acellular capillaries were assessed via hematoxylin and eosin (H&E) staining assay. The disruption of the blood–retinal barrier (BRB) was detected through Evans blue dye leakage assay, and retinal leukocyte adhesion was achieved via fluorescein isothiocyanate-coupled concanavalin A lectin labeling assay. Immunofluorescence staining and Western blot assays were conducted to detect the expression of tight junction proteins, adhesion molecules, and the ionized calcium-binding adapter molecule-1 (Iba-1) in the retina. Enzyme-linked immunosorbent assay was performed to detect the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β in serum. In addition, the protein expression levels of nuclear factor (NF)-κB p65, phosphorylated IκB kinase (p-IKK), phosphorylated NF-κB inhibitor (p-IκB), and phosphorylated NF-κB p65 (p-p65) were measured using Western blot assay. The ocular tissue distribution of PNS in normal and diabetic rats was determined through ultra-performance liquid chromatography–tandem mass spectrometry. The in vitro anti-inflammatory effects of PNS, notoginsenoside (NGR1), ginsenoside Rg1, Re, Rb1, and Rd (GRg1, GRe, GRb1, and GRd) were evaluated on human Müller (MIO-M1) cells. ResultsPNS increased the reduction in retinal inner nuclear layer thickness, reduced the increase in retinal acellular capillaries, and attenuated elevated BRB disruption by upregulating the decrease in protein expression of claudin-1 and occludin. Furthermore, PNS significantly abrogated microglial cell activation and reversed the increase in leukocyte adhesion by downregulating the increase in the protein expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1. Moreover, PNS reduced the elevated levels of TNF-α, IL-6, and IL-1β in serum and inhibited the increased protein expression of p-IKK, p-IκB, and p-p65, and the nuclear translocation of p65. The tissue distribution results revealed that NGR1, GRg1, GRe, GRb1, and GRd were detected in the ocular tissue, while GRg1 and GRb1 were found at the highest levels compared with the other components. The cellular results showed that PNS, NGR1, GRg1, GRe, GRb1, and GRd suppressed the development of cellular inflammatory responses by inhibiting the activation of the NF-κB signaling pathway in MIO-M1 cells and that their anti-inflammatory effects were comparable. ConclusionPNS suppressed retinal inflammation by inhibiting the activation of the NF-κB signaling pathway, alleviating DR. GRg1 and GRb1 may be the primary components that exert anti-inflammatory effects in vivo.

Gossypol improves myocardial dysfunction caused by sepsis by regulating histone acetylation.

Gossypol is a polyphenol from the cotton plant with anti-inflammatory and anti-oxidation activities and can also function as a histone deacetylase (HDAC) inhibitor. Sepsis is an inflammatory disease with high mortality. Inflammation, oxidative stress, and epigenetic factors are involved in sepsis and its complications. The biological activities of gossypol strongly suggest the potential effects of gossypol on sepsis. In the present study, the beneficial effects of gossypol on sepsis were evaluated. We established a cecal ligation and puncture (CLP) mouse model of sepsis and treated CLP mice with gossypol. The survival rate, serum level of myocardial injury markers, and myocardial level of oxidation markers were measured. We also administered gossypol to lipopolysaccharide (LPS)-treated primary cardiomyocytes. The production of pro-inflammatory cytokines, activation of protein kinase B (AKT) and IκB kinase (IKK), acetylation of histone, and expression of HDACs were measured. Gossypol prevented the death of CLP mice and ameliorated myocardial damage in CLP mice. Moreover, gossypol decreased oxidative factors, while promoting antioxidant production in CLP mice. Gossypol prevented LPS and cytosine-phosphate-guanosine-induced expression of pro-inflammatory cytokines, suppressed LPS-induced activation of AKT and IKK, inhibited histone acetylation, and decreased the expression of HDACs. In conclusion, gossypol ameliorates myocardial dysfunction in mice with sepsis.

Damaged mitochondria recruit the effector NEMO to activate NF-κB signaling

Failure to clear damaged mitochondria via mitophagy disrupts physiological function and may initiate damage signaling via inflammatory cascades, although how these pathways intersect remains unclear. We discovered that nuclear factor kappa B (NF-κB) essential regulator NF-κB effector molecule (NEMO) is recruited to damaged mitochondria in a Parkin-dependent manner in a time course similar to recruitment of the structurally related mitophagy adaptor, optineurin (OPTN). Upon recruitment, NEMO partitions into phase-separated condensates distinct from OPTN but colocalizing with p62/SQSTM1. NEMO recruitment, in turn, recruits the active catalytic inhibitor of kappa B kinase (IKK) component phospho-IKKβ, initiating NF-κB signaling and the upregulation of inflammatory cytokines. Consistent with a potential neuroinflammatory role, NEMO is recruited to mitochondria in primary astrocytes upon oxidative stress. These findings suggest that damaged, ubiquitinated mitochondria serve as an intracellular platform to initiate innate immune signaling, promoting the formation of activated IKK complexes sufficient to activate NF-κB signaling. We propose that mitophagy and NF-κB signaling are initiated as parallel pathways in response to mitochondrial stress.

Methyl lucidone inhibits airway inflammatory response by reducing TAK1 activity in human bronchial epithelial NCI–H292 cells

BackgroundMethyl lucidone (ML), a methyl derivative of lucidone, has anti-inflammatory properties. However, the molecular mechanisms that reduce the inflammatory effect of ML in human lung epithelial cells remain unkown. This study aimed to elucidate the molecular mechanisms underlying the anti-inflammatory effect of ML. MethodsFour compounds (ML, methyl linderone, kanakugiol, and linderone) from Lindera erythrocarpa Makino were evaluated for their ability to reduce MUC5AC secretion levels in phorbol-12-myristate-13-acetate (PMA)-stimulated NCI–H292 cells using ELISA. The expression and secretion levels of inflammatory response-related proteins were analyzed using quantitative reverse transcription-PCR, ELISA, and western blotting. To determine whether ML directly regulates TGF-β-activated kinase 1 (TAK1), we performed an in vitro kinase assay. ResultsML treatment effectively reduced the levels of inflammatory cytokines, including interleukin-1β and TNF-α, increased by stimulation. Furthermore, ML downregulated the pathway cascade of both IκB kinase (IKK)/NF-κB and p38 mitogen-activated protein (MAP) kinase/CREB by inhibiting the upstream kinase TAK1. An in vitro kinase analysis confirmed that ML treatment significantly reduced the kinase activity of TAK1. ConclusionML pretreatment repressed the PMA-stimulated inflammation reaction by reducing the TAK1-mediated IKK/NF-κB and p38 MAP kinase/CREB signaling. These findings suggest that ML may improve respiratory health and can be used as a dietary supplement or functional food to prevent inflammatory lung diseases.

Mechanism of Zhenwu Decoction in improving renal inflammatory injury in mice with DN of spleen-kidney Yang deficiency syndrome by regulating ROCK/IKK/NF-κB pathway

To investigate the intervention effect and mechanism of Zhenwu Decoction on diabetic nephropathy(DN) mice of spleen-kidney Yang deficiency syndrome based on the Rho-associated coiled-coil kinase(ROCK)/IκB kinase(IKK)/nuclear factor-κB(NF-κB) pathway. Ninety-five 7-week-old db/db male mice and 25 7-week-old db/m male mice were fed adaptively for one week. The DN model of spleen-kidney Yang deficiency syndrome was induced by Dahuang Decoction combined with hydrocortisone by gavage, and then the model was evaluated. After modeling, they were randomly divided into a model group, high-dose, medium-dose, and low-dose Zhenwu Decoction groups(33.8, 16.9, and 8.45 g·kg~(-1)·d~(-1)), and an irbesartan group(25 mg·kg~(-1)·d~(-1)), with at least 15 animals in each group. The intervention lasted for eight weeks. After the intervention, body weight and food intake were measured. Serum crea-tinine(Scr), blood urea nitrogen(BUN), fasting blood glucose(FBG), urinary albumin(uALb), and urine creatinine(Ucr) were determined. The uALb/Ucr ratio(ACR) and 24 h urinary protein(UTP) were calculated. Renal pathological morphology was evaluated by HE staining and Masson staining. The levels of key molecular proteins in the ROCK/IKK/NF-κB pathway were detected by Western blot. Enzyme-linked immunosorbent assay(ELISA) was used to detect interleukin-1β(IL-1β), interleukin-6(IL-6), interleukin-8(IL-8), interleukin-10(IL-10), and tumor necrosis factor-α(TNF-α). Compared with the blank group, the model group showed increased content of BUN, uALb, and SCr, increased values of 24 h UTP and ACR, decreased content of Ucr(P&lt;0.05), enlarged glomeruli, thickened basement membrane, mesangial matrix proliferation, inflammatory cell infiltration, and collagen fiber deposition. The protein expression of ROCK1, ROCK2, IKK, NF-κB, phosphorylated IKK(p-IKK), phosphorylated NF-κB(p-NF-κB), and phosphorylated inhibitor of NF-κB(p-IκB) increased(P&lt;0.05), while the protein expression of inhibitor of NF-κB(IκB) decreased(P&lt;0.05). The levels of inflammatory factors IL-1β, IL-6, IL-8, and TNF-α increased(P&lt;0.05), while the level of IL-10 decreased(P&lt;0.05). Compared with the model group, the groups with drug treatment showed decreased levels of BUN, uALb, SCr, 24 h UTP, and ACR, increased level of Ucr(P&lt;0.05), and improved renal pathological status to varying degrees. The high-and medium-dose Zhenwu Decoction groups and the irbesartan group showed reduced protein expression of ROCK1, ROCK2, IKK, NF-κB, p-IKK, p-NF-κB, and p-IκB in the kidneys(P&lt;0.05), increased protein expression of IκB(P&lt;0.05), decreased levels of serum inflammatory factors IL-1β, IL-6, IL-8, and TNF-α(P&lt;0.05), and increased level of IL-10(P&lt;0.05). Zhenwu Decoction can significantly improve renal function and renal pathological damage in DN mice of spleen-kidney Yang deficiency syndrome, and its specific mechanism may be related to the inhibition of inflammatory response by down-regulating the expression of key molecules in the ROCK/IKK/NF-κB pathway in the kidney.

Effects of dietary eicosapentaenoic acid on the growth performance, fatty acid profile, immunity and heat tolerance of juvenile abalone Haliotis discus hannai Ino

A 93-day growth trial was performed to evaluate the influences of graded dietary EPA levels (0.03% (control group), 0.24%, 0.63%, 1.07%, 1.72% and 2.25%) on the growth performance, immunity and heat tolerance of juvenile abalone Haliotis discus hannai with a mean initial weight of 4.15 ± 0.18 g. After the feeding trial, the findings found that the highest weight gain rate (WGR) belonged to the groups fed with 0.63% ∼ 1.72% of dietary EPA (P < 0.05). The lowest feed conversion ratio was distinguished in the groups with 0.63% ∼ 1.07% of dietary EPA (P < 0.05). Also, EPA supplementation in abalone feed increased the content of crude lipid in the soft body and the activity of lipase in the intestine (P < 0.05). The total antioxidative capacity increased when dietary EPA levels rose from 0.03% to 1.07%, and then decreased (P < 0.05). Furthermore, compared to the control group, the 0.24% and 0.24% ∼ 0.63% of dietary EPA significantly down-regulated the mRNA levels of IκB kinase and kelch-like ECH-associated protein 1, respectively (P < 0.05). However, nuclear factor-κB and interleukin 1 receptor-associated kinase 4 genes expression in the digestive gland were significantly up-regulated by the use of 1.72% and 2.25% of EPA in diet (P < 0.05). The mRNA levels of tumor necrosis factor α and interleukin 16 were significantly up-regulated by the 2.25% of dietary EPA (P < 0.05). The 1.72% of dietary EPA significantly increased the falling rate of abalone after the heat stress compared with the control (P < 0.05). In conclusion, the optimal dietary EPA requirement of juvenile abalone was determined as 1.36% based on quadratic regression analysis against WGR. Moderate dietary EPA content (0.63% ∼ 1.07%) improved the immunity and heat resistance of abalone. Excessive dietary EPA content (1.72% ∼ 2.25%) decreased the growth, meanwhile, increasing the oxidative stress, inflammation and susceptibility of abalone to heat stress.

Protective effects of ginseng and ginsenosides in the development of osteoarthritis (Review).

Osteoarthritis (OA) is a chronic inflammatory joint disease. Traditional chinese medicine provides a resource for drug screening for OA treatment. Ginseng and the associated bioactive compound, ginsenosides, may reduce inflammation, which is considered a risk factor for the development of OA. Specifically, ginsenosides may exhibit anti-inflammatory and anti-oxidative stress activities, and inhibit extracellular matrix degradation by suppressing the NF-κB and MAPK signaling pathways. Notably, specific ginsenosides, such as compound K and Rk1, may physically interact with IκB kinase and inhibit the associated phosphorylation. Thus, ginsenosides exhibit potential as therapeutic candidates in the management of OA. However, the low water solubility limits the clinical applications of ginsenosides. Numerous effective strategies have been explored to improve bioavailability; however, further investigations are still required.