Nuclear factor-kappaB Inhibitor Research Articles

Interleukin-33 increases the sensitivity of multiple myeloma cells to the proteasome inhibitor bortezomib through reactive oxygen species-mediated inhibition of nuclear factor kappa-B signal and stemness properties.

The proteasome inhibitor bortezomib (BTZ) is the first-line therapy for multiple myeloma (MM). BTZ resistance largely limits its clinical application in MM. Interleukin-33 (IL-33) exerts antitumor effects through various mechanisms, including enhancing antitumor immunity and promoting the apoptosis of cancer cells. Here, the synergistic anti-MM effect of IL-33 and BTZ was verified, and the underlying mechanisms were elucidated. Bioinformatic analysis indicated that IL-33 expression levels were downregulated in MM, and that BTZ-treated MM patients with high IL-33 levels had better prognosis than those with low IL-33 levels. Moreover, the patients with high IL-33 levels had a better treatment response to BTZ. Further immune analysis suggested that IL-33 can enhance the anti-MM immunity. IL-33 and BTZ synergistically inhibited proliferation and induced apoptosis of MM cells, which was mediated by the excessive accumulation of cellular reactive oxygen species (ROS). Furthermore, increased ROS hindered the nuclear translocation of NF-κB-p65, thereby decreasing the transcription of target stemness-related genes (SOX2, MYC, and OCT3/4). These effects induced by the combination therapy could be reversed by eliminating ROS by N-acetylcysteine. In conclusion, our results indicated that IL-33 enhanced the sensitivity of MM to BTZ through ROS-mediated inhibition of nuclear factor kappa-B (NF-κB) signal and stemness properties.

Salvimulticanol from Salvia multicaulis suppresses LPS-induced inflammation in RAW264.7 macrophages: in vitro and in silico studies.

Sustained inflammatory responses can badly affect several vital organs and lead to chronic inflammation-related disorders, such as atherosclerosis, pneumonia, rheumatoid arthritis, obesity, diabetes, Alzheimer's disease, and cancers. Salvia multicaulis is one of the widely distributed plants that contains several biologically active phytochemicals and diterpenoids with anti-inflammatory effects. Therefore, finding alternative and safer natural plant-extracted compounds with good curative anti-inflammatory efficiencies is an urgent need for the clinical treatment of inflammation-related diseases. In the current study, S. multicaulis Vahl was used to extract and isolate two compounds identified as salvimulticanol and candesalvone B methyl ester to examine their effects against inflammation in murine macrophage RAW264.7 cells that were induced by lipopolysaccharide (LPS). Accordingly, after culturing RAW264.7 cells and induction of inflammation by LPS (100ng/ml), cells were exposed to different concentrations (9, 18, 37.5, 75, and 150µM) of each compound. Then, Griess assay for detection of nitric oxide (NO) levels and western blotting for the determination of inducible nitric oxide synthase (iNOS) expression were performed. Molecular docking and molecular dynamics (MD) simulation studies were employed to investigate the anti-inflammatory mechanism. Our obtained results validated that the level of NO was significantly decreased in the macrophage cell suspensions as a response to salvimulticanol treatment in a dose-dependent manner (IC50: 25.1 ± 1.2µM) as compared to the methyl ester of candesalvone B which exerted a weaker inhibition (IC50: 69.2 ± 3.0µM). This decline in NO percentage was comparable with a down-regulation of iNOS expression by western blotting. Salvimulticanol strongly interacted with both the Toll-like receptor 4 (TLR4)/myeloid differentiation factor 2 (MD-2) complex and the inhibitor of nuclear factor kappa-B (NF-κB) kinase subunit beta (IKKβ) to disrupt their inflammatory activation due to the significant hydrogen bonds and effective interactions with amino acid residues present in the target proteins' active sites. S.multicaulis is a rich natural source of the aromatic abietane diterpenoid, salvimulticanol, which exerted a strong anti-inflammatory effect through targeting iNOS and diminishing NO production in LPS-induced RAW264.7 cells in a mechanism that is dependent on the inhibition of TLR4-MD-2 and IKKβ as activators of the classical NF-κB-mediated inflammatory pathway.

Inhibition of NF-κB Activation Improves Non-Nitric Oxide-Mediated Cutaneous Microvascular Function in Reproductive-Aged Healthy Women

PURPOSE: The human cutaneous circulation is used as an in vivo bioassay to examine mechanisms of systemic vascular function and dysfunction in health and disease. In cohorts of patients with accelerated cardiovascular disease risk, systemic interventions to investigate inflammatory mechanisms [e.g., oral salsalate; nuclear factor kappa-B (NF-κB) inhibitor] have been used to elucidate mechanisms underlying microvascular dysfunction. In healthy control subjects, inhibition of inflammatory mechanisms modestly reduces microvascular function. We hypothesize that inhibition of NF-κB activation may impair nitric oxide (NO)-mediated cutaneous microvascular function in reproductive-aged healthy women. METHODS: In a randomized, single-blind, placebo-controlled design, oral salsalate (1500mg/b.i.d./5 days) was used to assess the impact of systemic inflammation on cutaneous microvascular function. Two intradermal microdialysis fibers were placed into the ventral forearm skin of eleven healthy women [Mean (SEM): 34 (2) yrs]. Laser-Doppler flowmetry was used to measure cutaneous blood flow while increasing concentrations of acetylcholine (10−10 to 10−1 M; endothelium-dependent vasodilator) were perfused with a control (lactated Ringer’s) or a NO synthase inhibitor (15 mM NG-nitro-L-arginine methyl ester). Following the dose-response protocol, maximal cutaneous vasodilation was induced by increasing local skin temperature to 43°C and perfusing 28 mM sodium nitroprusside. Cutaneous vascular conductance (CVC) was calculated as the quotient of red blood cell flux over mean arterial pressure and normalized as a percentage of the site-specific maximum (%CVCmax). The NO-dependent contribution was calculated as the difference between the area under the dose-response curve (AUC) of the control and the NO synthase-inhibited sites. RESULTS: There was an overall main effect of salsalate treatment ( p < 0.001). Both the control and NO synthase-inhibited sites shifted upwards following salsalate treatment (Placebo - Control: 45.78 (8.81)% vs NO synthase inhibited: 33.62 (8.10)%; Salsalate - Control: 56.30 (8.48)% vs NO synthase inhibited: 41.57 (7.85)%; both p < 0.05). There was no change in the NO-dependent contribution between treatments (Placebo: 164.30 (35.40) AU vs Salsalate: 180.30 (51.17) AU; p=0.7028).CONCLUSION: Inhibition of NF-κB activation improved cutaneous microvascular function in reproductive-aged healthy women. This improvement was mediated via non-nitric oxide-dependent mechanisms. Supported by NIH Grant R01 HL161000 (LMA) and Diversity Supplement R01 HL161000-02S2 (VGC). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Equisetum arvense standardized dried extract hinders age-related osteosarcopenia

Age-associated osteosarcopenia is an unresolved syndrome characterized by the concomitant loss of bone (osteopenia) and skeletal muscle (sarcopenia) tissues increasing falls, immobility, morbidity, and mortality. Unbalanced resorption of bone in the remodeling process and excessive protein breakdown, especially fast type II myosin heavy chain (MyHC-II) isoform and myofiber metabolic shift, are the leading causes of bone and muscle deterioration in the elderly, respectively. Equisetum arvense (EQ) is a plant traditionally recommended for many pathological conditions due to its anti-inflammatory properties. Thus, considering that a chronic low-grade inflammatory state predisposes to both osteoporosis and sarcopenia, we tested a standardized hydroalcoholic extract of EQ in in vitro models of muscle atrophy [C2C12 myotubes treated with proinflammatory cytokines (TNFα/IFNγ), excess glucocorticoids (dexamethasone), or the osteokine, receptor activator of nuclear factor kappa-B ligand (RANKL)] and osteoclastogenesis (RAW 264.7 cells treated with RANKL). We found that EQ counteracted myotube atrophy, blunting the activity of several pathways depending on the applied stimulus, and reduced osteoclast formation and activity. By in silico target fishing, IKKB-dependent nuclear factor kappa-B (NF-κB) inhibition emerges as a potential common mechanism underlying EQ′s anti-atrophic effects. Consumption of EQ (500 mg/kg/day) by pre-geriatric C57BL/6 mice for 3 months translated into: i) maintenance of muscle mass and performance; ii) restrained myofiber oxidative shift; iii) slowed down age-related modifications in osteoporotic bone, significantly preserving trabecular connectivity density; iv) reduced muscle- and spleen-related inflammation. EQ can preserve muscle functionality and bone remodeling during aging, potentially valuable as a natural treatment for osteosarcopenia.

Insights into the mechanism of action of pterostilbene against influenza A virus-induced acute lung injury

BackgroundSevere respiratory system illness caused by influenza A virus infection is associated with excessive inflammation and abnormal apoptosis in alveolar epithelial cells (AEC). However, there are limited therapeutic options for influenza-associated lung inflammation and apoptosis. Pterostilbene (PTE, trans-3,5-dimethoxy-4-hydroxystilbene) is a dimethylated analog of resveratrol that has been reported to limit influenza A virus infection by promoting antiviral innate immunity, but has not been studied for its protective effects on virus-associated inflammation and injury in AEC. PurposeOur study aimed to investigate the protective effects and underlying mechanisms of PTE in modulating inflammation and apoptosis in AEC, as well as its effects on macrophage polarization during influenza virus infection. Study design and methodsA murine model of influenza A virus-mediated acute lung injury was established by intranasal inoculation with 5LD50 of mouse-adapted H1N1 viruses. Hematoxylin and eosin staining, immunofluorescence, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, western blotting, Luminex and flow cytometry were performed. ResultsPTE effectively mitigated lung histopathological changes and injury induced by H1N1 viruses in vivo. These beneficial effects of PTE were attributed to the suppression of inflammation and apoptosis in AEC, as well as the modulation of M1 macrophage polarization. Mechanistic investigations revealed that PTE activated the phosphorylated AMP-activated protein kinase alpha (P-AMPKα)/sirtui1 (Sirt1)/PPARγ coactivator 1-alpha (PGC1α) signal axis, leading to the inhibition of nuclear factor kappa-B (NF-κB) and p38 mitogen-activated protein kinase (MAPK) signaling induced by H1N1 viruses, thereby attenuating inflammation and apoptosis in AEC. PTE also forced activation of the P-AMPKα/Sirt1/PGC1α signal axis in RAW264.7 cells, counteracting the activation of phosphorylated signal transducer and activator of transcription 1 (P-STAT1) induced by H1N1 viruses and the augment of P-STAT1 activation in RAW264.7 cells with interferon-gamma (IFN-γ) pretreatment before viral infection, thereby reducing H1N1 virus-mediated M1 macrophage polarization as well as the enhancement of macrophages into M1 phenotypes elicited by IFN-γ pretreatment. Additionally, the promotion of the transition of macrophages towards the M2 phenotype by PTE was also related to activation of the P-AMPKα/Sirt1/PGC1α signal axis. Moreover, co-culturing non-infected AEC with H1N1 virus-infected RAW264.7 cells in the presence of PTE inhibited apoptosis and tight junction disruption, which was attributed to the suppression of pro-inflammatory mediators and pro-apoptotic factors in an AMPKα-dependent manner. ConclusionIn conclusion, our findings suggest that PTE may serve as a promising novel therapeutic option for treating influenza-associated lung injury. Its ability to suppress inflammation and apoptosis in AEC, modulate macrophage polarization, and preserve alveolar epithelial cell integrity highlights its potential as a therapeutic agent in influenza diseases.

Transcriptome analysis reveals the oxidative damage and immune-suppression of leech (Whitmania pigra Whitman) intestine induced by high-temperature stress

Temperature is a key component in regulating the survival and growth of aquatic animals. However, the mechanism of high-temperature stress in Whitmania pigra Whitman has received little attention. Therefore, the aim of this study was to investigate the possible mechanisms of high-temperature stress in W. pigra using transcriptome analysis and detection of antioxidant markers. Samples were collected at 1 d, 3 d, 5 d, and 7 d under stress in two experimental groups: the control group at 27 °C and the high-temperature stress group at 35 °C. High-temperature stress caused oxidative damage in the leech intestine, as indicated by decreased antioxidant enzyme activities (superoxide dismutase and total antioxidant capacity) and increased lipid hydroperoxide malondialdehyde. Transcriptome analysis revealed 425 DEGs in leech intestine during high-temperature stress, with 275 up-regulated and 150 down-regulated genes. According to KEGG enrichment analysis, 425 DEGs were involved in the immune system and induced abnormal expression of disease-related genes. Furthermore, when leeches were exposed to high-temperature stress, B-cell lymphoma-2, heat shock protein 70, heat shock protein 90, and nuclear factor erythroid 2-related factor 2 mRNAs were dramatically upregulated, whereas activating transcription factor, inhibitor of Nuclear factor kappa-B, sequestosome 1, and toll-like receptor 4 mRNAs were significantly downregulated. In conclusion, 35 °C high-temperature stress impaired leech intestinal function by decreasing antioxidant capacity, causing oxidative damage, and suppressing the activation of immune capacity. This study contributes to future research on the detailed mechanisms of high-temperature stress in leeches.

Lower vitamin D levels are associated with the pathogenesis of inflammatory bowel diseases.

Vitamin D plays a role in regulating immune homeostasis, inflammation and has an impact on the pathogenesis of inflammatory bowel diseases (IBD). IBD has a multifactorial pathogenesis primarily associated with immune dysregulation, dysbiosis, structurally altered intestinal mucosa, and genetic factors. The immunomodulatory function of this vitamin is linked to its control over innate and adaptive immunity, facilitated through its nuclear vitamin D receptor, leading to the inhibition of nuclear factor kappa-B. This study aimed to investigate serum vitamin D levels in patients with IBD compared to healthy individuals and to evaluate the relationship between vitamin D and inflammatory markers. Cross-sectional study. The study included 106 participants divided into 2 groups: patients with IBD (92), and healthy controls (14). The diagnosis of IBD was based on clinical, laboratory, fecal, endoscopic, and histological findings, following the European guidelines for diagnosis and follow-up ECCO-ESGAR guidelines for diagnostic assessment of IBD from 2019. Serum vitamin D levels were measured along with laboratory tests, imaging, and endoscopic examinations. IBD activity was evaluated using the Montreal classification and clinical and endoscopic indices. Data analysis involved calculating the mean, minimum, and maximum values, standard deviation, and Pearson coefficient. The level of statistical significance for this study was set at P < .05. The study found a prevalence of vitamin D deficiency in 32.6% of patients with IBD, while 66.3% had insufficiency, as compared with healthy individuals. The mean levels of vitamin D in UC and CD were 16 ± 8.6 ng/mL, whereas in the control healthy group, they were 26 ± 9.73 ng/mL. A statistically significant reverse correlation was observed between lower vitamin D levels and higher levels of the inflammatory markers. The study concluded that IBD patients exhibit lower levels of vitamin D, which is associated with inflammation and may contribute to the pathogenesis of the disease.

HM-chromanone alleviates hyperglycemia and inflammation in mice with endotoxin-induced insulin resistance.

This study was designed to investigate whether (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone alleviates inflammation and hyperglycemia in mice with endotoxin-induced insulin resistance. (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone (10, 30, and 50mg/kg bodyweight) was orally pre-administered to C57BL/6J mice. An hour later, lipopolysaccharides (20mg/kg bodyweight) was administered intraperitoneally to induce endotoxins. Blood samples were collected from the tail vein of the mice every 0, 30, and 90min. The results indicated that (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone effectively regulated blood glucose levels in mice with endotoxin-induced insulin resistance. Furthermore, (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone significantly reduced the phosphorylation of mammalian target of rapamycin, ribosomal protein S6 kinase 1, and protein kinase C θ. Additionally, (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone suppressed the phosphorylation of c-Jun-NH2-terminal kinase and IkB kinase β, thereby decreasing the phosphorylation of inhibitor of nuclear factor kappa-B α and activating the nuclear factor-κB and activator protein-1 in the liver. Therefore, the expression of tumor necrosis factor-α, interleukin-6, and interleukin-1β was significantly reduced by suppressing the nuclear factor-κB and activator protein 1 activity. Suppression of mammalian target of rapamycin, S6 kinase 1, protein kinase C θ, c-Jun-NH2-terminal kinase, and IkB kinase β also ameliorated insulin resistance by reducing the phosphorylation of insulin receptor substrate-1 serine 307, thereby decreasing hyperglycemia. These findings suggest that (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone can alleviate hyperglycemia and inflammation in mice with endotoxin-induced insulin resistance.

Atractylodes Processing Products Protect against Gastric Ulcers in Rats by Influencing the NF-κB-MMP-9/TIMP-1 Regulatory Mechanism and Intestinal Flora.

Atractylodes macrocephala Koidz. (AM) is a Chinese herbal medicine that is widely used for treating gastrointestinal diseases. However, little research has focused on it as a single medicine for treating gastric ulcers. Honey-bran stir-frying is a characteristic method of concocting AM, so we speculated that AM is more effective after this preparation process. Analysis by ultra-high-performance liquid chromatography-hybrid quadrupole-Orbitrap high-resolution mass spectrometry revealed changes in the chemical composition of raw Atractylodes (SG), bran-fried Atractylodes (FG), and honey-bran-fried Atractylodes (MFG). MFG was superior to SG and FG in improving the pathological structure of gastric tissue in rats with acute gastric ulcers, reducing inflammatory cell infiltration in gastric tissue, and significantly reducing malondialdehyde while increasing superoxide dismutase and glutathione peroxidase, and reducing the damage caused by free radical accumulation in the gastric mucosa. In addition, MFG reduced the expression of matrix metalloproteinase-9 (MMP-9), an inhibitor of metalloproteinase-1 (TIMP-1) and nuclear factor kappa-B (NF-κB)proteins, inhibited inflammatory response, and regulated the degradation and rebalancing of the extracellular matrix. Fecal microbiota analysis also revealed that MFG normalized the intestinal flora to some extent. Our study shows that AM had a protective effect on rats with alcohol-induced acute gastric ulcers before and after processing, and AM-processed products were more effective than raw ones. Compared with MF, MFG had a higher rate of ulcer inhibition and a stronger anti-inflammatory effect, and its mechanism of action was related to the NF-κB-MMP-9/TIMP-1 signaling pathway.

Tumor necrosis factor-α upregulates polymeric immunoglobulin receptor expression by NF-κB signaling pathways in grass carp (Ctenopharyngodon idellus) liver cells

The polymeric immunoglobulin receptor (pIgR) is essential for controlling polymeric immunoglobulin to defend species from invading pathogens. However, the modulation pathway of pIgR expression in teleosts remains unclear. In this paper, to define that the cytokine TNF-α impacted the expression of pIgR, the recombinant proteins of TNF-α of grass carp were first prepared after approving that natural pIgR was expressed in liver cells of grass carp (Ctenopharyngodon idellus) (L8824). L8824 cells were incubated with variable amounts of recombinant TNF-α at various times, the results revealed that pIgR expressions showed a significant dose-dependent elevation at the gene and proteins, and a similar alteration trend was detected for the pIgR protein (secretory component: SC) secreted by L8824 cells into the culture supernatant. Moreover, nuclear factor kappa-B (NF-κB) inhibitors PDTC was used to study whether TNF-α regulated pIgR expressions through the NF-κB signaling pathways. L8824 cells were treated with TNF-α, inhibitor PDTC, and TNF-α + PDTC mixtures, respectively, and the levels of pIgR genes and pIgR protein in cells and SC in the culture supernatant decreased in cells treated with PDTC contrasted to the control, and subjected to reduced expression of PDTC + TNF-α reduced expression contrasted to that treated just with TNF-α, demonstrating that suppression of NF-κB obstructed the ability of TNF-α to elevate pIgR gene and pIgR protein in cells and SC in the culture supernatant. These outcomes indicated that TNF-α raised pIgR gene expression, pIgR protein, and SC creation, and this pIgR expression induced by TNF-α was modulated by complicated pathways that included NF-κB signaling mechanism, confirming TNF-α as a pIgR expression modulator and enhancing a deeper insight of the regulatory pathway for pIgR expression in teleosts.

Anti-inflammatory potential of selective small compounds by targeting TNF-α & NF-kB signaling: a comprehensive molecular docking and simulation study

Tumor necrosis factor alpha (TNF-α) is the major cause of inflammation in autoimmune diseases like rheumatoid arthritis (RA). It’s mechanisms of signal transduction through nuclear factor kappa B (NF-kB) pathway via small molecules such as metabolite crosstalk are still elusive. In this study, we have targeted TNF-α and NF-kB through metabolites of RA, to inhibit TNF-α activity and deter NF-kB signaling pathways, thereby mitigating the disease severity of RA. TNF-α and NF-kB structure was obtained from PDB database and metabolites of RA were selected from literature survey. In-silico studies were carried out by molecular docking using AutoDock Vina software and further, known TNF-α and NF-kB inhibitors were compared and revealed metabolite’s capacity to targets the respective proteins. Most suitable metabolite was then validated by MD simulation to verify its efficiency against TNF-α. Total 56 known differential metabolites of RA were docked with TNF-α and NF-kB compared to their corresponding inhibitor compounds. Four metabolites such as Chenodeoxycholic acid, 2-Hydroxyestrone, 2-Hydroxyestradiol (2-OHE2), and 16-Hydroxyestradiol were identified as a common TNF-α inhibitor’s having binding energies ranging from −8.3 to −8.6 kcal/mol, followed by docking with NF-kB. Further, 2-OHE2 was selected because of having binding energy −8.5 kcal/mol, found to inhibit inflammation and the effectiveness was validated by root mean square fluctuation, radius of gyration and molecular mechanics with generalized born and surface area solvation against TNF-α. Thus 2-OHE2, an estrogen metabolite was identified as the potential inhibitor, attenuated inflammatory activation and can be utilized as a therapeutic target to disseminate severity of RA.

Exploration of bioactive compounds from Mangifera indica (Mango) as probable inhibitors of thymidylate synthase and nuclear factor kappa-B (NF-Κb) in colorectal cancer management

Abstract This research is aimed at investigating the anti-colorectal cancer activities of phytochemicals from Mangifera indica (Mango) via the inhibition of thymidylate synthase (TS) and Nuclear Factor kappa B (NF–κB) using computational chemistry tools. Ligands (141 phytochemicals previously isolated from mangoes) and reference drugs (Raltitrexed and Emetine), the drug inhibitors of TS and NF–κB, respectively) were subjected to screening via ADMET profiling, drug-likeness analysis, oral bioavailability, PASS profile, and molecular interactions. Ligands that passed the previously mentioned screening were docked in duplicate against the target receptors (TS and NF–κB) using PyRx software. The mean values were calculated to obtain suitable docking scores. The analysis showed that TS was strongly inhibited by Friedelan-3beta-Ol with its lower binding energy of −9.0 kcal/mol more than Raltitrexed with a binding energy of −8.7 kcal/mol. NF–κB was also inhibited by Friedelan-3beta-Ol and Friedelin with binding energies of −8.0 and −8.1 kcal/mol, respectively, more than Emetine with a binding energy of −6.4 kcal/mol. These two phytochemicals performed much better than the standard drugs, thus selected as the best hits compounds because of their ADMET profile, drug-likeness properties, bioactivity, oral bioavailability, PASS prediction, binding affinities, and their interactions with the amino acids in the active sites of the receptors. Therefore, further studies are necessary for the validation of these claims toward the development of new effective and safer anti-colorectal cancer drugs.

Activation of ROS-PERK-TFEB by filbertone ameliorates neurodegenerative diseases via enhancing the autophagy-lysosomal pathway

The molecular mechanisms underlying the pathogenesis of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease (PD), and Huntington's disease remain enigmatic, resulting in an unmet need for therapeutics development. Here, we suggest that filbertone, a key flavor compound found in the fruits of hazel trees of the genus Corylus, can ameliorate PD via lowering the abundance of aggregated α-synuclein. We previously reported that inhibition of hypothalamic inflammation by filbertone is mediated by suppression of nuclear factor kappa-B. Here, we report that filbertone activates PERK through mitochondrial reactive oxygen species production, resulting in the increased nuclear translocation of transcription factor-EB in SH-SY5Y human neuroblastoma cells. TFEB activation by filbertone promotes the autophagy-lysosomal pathway, which in turn alleviates the accumulation of α-synuclein. We also demonstrate that filbertone prevented the loss of dopaminergic neurons in the substantia nigra and striatum of mice on high-fat diet. Filbertone treatment also reduced high-fat diet-induced α-synuclein accumulation through upregulation of the autophagy-lysosomal pathway. In addition, filbertone improved behavioral abnormalities (i.e., latency time to fall and decrease of running distance) in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced PD murine model. In conclusion, filbertone may show promise as a potential therapeutic for neurodegenerative disease.

Identification of potential inhibitors of thymidylate synthase (TS) (PDB ID: 6QXH) and nuclear factor kappa-B (NF–κB) (PDB ID: 1A3Q) from Capsicum annuum (bell pepper) towards the development of new therapeutic drugs against colorectal cancer (CRC)

Abstract Colorectal cancer is the third most deadly cancer globally. Drug resistance and attendant side effects make the available standard anti-colorectal cancer drugs against target receptors inefficient. Phytochemicals from medicinal plants are safer, cheaper, effective, and heal diseases from the cellular level. This study is aimed at identifying potential inhibitors of thymidylate synthase (TS) and nuclear factor kappa-B (NF–κB) target receptors from Capsicum annuum towards the development of new therapeutic drugs against colorectal cancer via in silico approach. One hundred and fifty (150) ligands previously reported from Capsicum annuum were downloaded from the PubChem database and were subjected to chemo-informatics analyses such as ADMET, drug-likeness, oral bioavailability, bioactivity, and PASS prediction to ascertain their therapeutic and safety profile before docking. The ligands that passed the analyses were docked against TS and NF–κB in duplicate using a creditable docking tool (PyRx). Raltitrexed and emetine were used as the standard drug inhibitors for TS and NF–κB, respectively. The results obtained from this study showed that feruloyl-beta-D-glucose (8.45 kcal/mol), 5-O-caffeoylquinic acid (−8.40 kcal/mol), 5-O-caffeoylquinic acid methyl ester (−7.89 kcal/mol), feruloyl hexoside (−7.40 kcal/mol), O-glucopyranoside (−7.55 kcal/mol), and quercetin (−7.00 kcal/mol) shared the same binding pocket with TS while feruloyl-beta-D-glucose (−7.00 kcal/mol), chlorogenic acid (−6.90 kcal/mol), 5-O-caffeoylquinic acid (−6.90 kcal/mol) and feruloyl hexoside (−6.50 kcal/mol) shared the same pocket with NF–κB. These compounds were selected as best hits due to their excellent inhibitory efficiency and chemoinformatic profiles. Thus, the compounds may function as prospective lead compounds for developing a new anti-colorectal cancer drug.

Molecular Targets and Mechanisms of 6,7-Dihydroxy-2,4-dimethoxyphenanthrene from Chinese Yam Modulating NF-κB/COX-2 Signaling Pathway: The Application of Molecular Docking and Gene Silencing

Chinese yam (Dioscorea opposita) tuber has a significant effect of invigorating the intestine and improving the symptoms of long-term diarrhea according to the records of the Chinese Pharmacopoeia. Phenanthrene polyphenols from Chinese yam, with higher inhibition of cyclooxygenase-2 (COX-2) than anti-inflammatory drugs, are an important material basis in alleviating ulcerative colitis via nuclear factor kappa-B (NF-κB)/COX-2 pathway, based on our previous research. The present study further explored the target and molecular mechanisms of phenanthrenes' modulation of the NF-κB/COX-2 signaling pathway by means of molecular docking and gene silencing. Firstly, interleukin-8 (IL-8) and tumor necrosis factor-α (TNF-α) expression of 6-hydroxy-2,4,7-trimethoxyphenanthrene (PC2)/6,7-dihydroxy-2,4-dimethoxyphe-nanthrene (PC4) were compared on TNF-α induced human colon adenocarcinoma (Caco-2) cells. Secondly, molecular docking and dynamics simulation were implemented for PC2/PC4 and COX-2. Finally, COX-2 silencing was performed on TNF-α induced Caco-2 cells to confirm the target of PC4 on NF-κB/COX-2 pathway. Lower expression of IL-8 and TNF-α in PC4 treated Caco-2 cells indicated that PC4 had stronger anti-inflammatory activity than PC2. The binding of PC4 and COX-2 was stronger due to the hydrogen bond between hydroxyl group and Tyr385. No significant differences were found in phosphorylation nuclear factor kappa-B inhibitor alpha (pIkBα), phosphorylation NF-κB (pNF-κB) and phosphorylation extracellular signal-regulated kinase 1/2 (pERK1/2) expression between control and PC4 group after silencing, while these protein expressions significantly decreased in PC4 group without silencing, which confirmed that COX-2 was the important target for PC4 in alleviating ulcerative colitis. These findings indicate that PC4 was supposed to have inhibited NF-κB pathway mediated inflammation via suppression of positive feedback targeting COX-2.

ADAR1 Non-Editing Function in Macrophage Activation and Abdominal Aortic Aneurysm.

Macrophage activation plays a critical role in abdominal aortic aneurysm (AAA) development. However, molecular mechanisms controlling macrophage activation and vascular inflammation in AAA remain largely unknown. The objective of the study was to identify novel mechanisms underlying adenosine deaminase acting on RNA (ADAR1) function in macrophage activation and AAA formation. Aortic transplantation was conducted to determine the importance of nonvascular ADAR1 in AAA development/dissection. Ang II (Angiotensin II) infusion of ApoE-/- mouse model combined with macrophage-specific knockout of ADAR1 was used to study ADAR1 macrophage-specific role in AAA formation/dissection. The relevance of macrophage ADAR1 to human AAA was examined using human aneurysm specimens. Moreover, a novel humanized AAA model was established to test the role of human macrophages in aneurysm formation in human arteries. Allograft transplantation of wild-type abdominal aortas to ADAR1+/- recipient mice significantly attenuated AAA formation, suggesting that nonvascular ADAR1 is essential for AAA development. ADAR1 deficiency in hematopoietic cells decreased the prevalence and severity of AAA while inhibited macrophage infiltration and aorta wall inflammation. ADAR1 deletion blocked the classic macrophage activation, diminished NF-κB (nuclear factor kappa B) signaling, and enhanced the expression of a number of anti-inflammatory microRNAs. Mechanistically, ADAR1 interacted with Drosha to promote its degradation, which attenuated Drosha-DGCR8 (DiGeorge syndrome critical region 8) interaction, and consequently inhibited pri- to pre-microRNA processing of microRNAs targeting IKKβ, resulting in an increased IKKβ (inhibitor of nuclear factor kappa-B) expression and enhanced NF-κB signaling. Significantly, ADAR1 was induced in macrophages and interacted with Drosha in human AAA lesions. Reconstitution of ADAR1-deficient, but not the wild type, human monocytes to immunodeficient mice blocked the aneurysm formation in transplanted human arteries. Macrophage ADAR1 promotes aneurysm formation in both mouse and human arteries through a novel mechanism, that is, Drosha protein degradation, which inhibits the processing of microRNAs targeting NF-kB signaling and thus elicits macrophage-mediated vascular inflammation in AAA.

HKDC1 upregulation promotes glycolysis and disease progression, and confers chemoresistance onto gastric cancer.

There is increasing evidence that hexokinase is involved in cell proliferation and migration. However, the function of the hexokinase domain containing protein-1 (HKDC1) in gastric cancer (GC) remains unclear. Immunohistochemistry analysis and big data mining were used to evaluate the correlation between HKDC1 expression and clinical features in GC. In addition, the biological function and molecular mechanism of HKDC1 in GC were studied by in vitro and in vivo assays. Our study indicated that HKDC1 expression was upregulated in GC tissues compared with adjacent nontumor tissues. High expression of HKDC1 was associated with worse prognosis. Functional experiments demonstrated that HKDC1 upregulation promoted glycolysis, cell proliferation, and tumorigenesis. In addition, HKDC1 could enhance GC invasion and metastasis by inducing epithelial-mesenchymal transition (EMT). Abrogation of HKDC1 could effectively attenuate its oncogenic and metastatic function. Moreover, HKDC1 promoted GC proliferation and migration in vivo. HKDC1 overexpression conferred chemoresistance to cisplatin, oxaliplatin, and 5-fluorouracil (5-Fu) onto GC cells. Furthermore, nuclear factor kappa-B (NF-κB) inhibitor PS-341 could attenuate tumorigenesis, metastasis, and drug resistance ability induced by HKDC1 overexpression in GC cells. Our results highlight a critical role of HKDC1 in promoting glycolysis, tumorigenesis, and EMT of GC cells via activating the NF-κB pathway. In addition, HKDC1-mediated drug resistance was associated with DNA damage repair, which further activated NF-κB signaling. HKDC1 upregulation may be used as a potential indicator for choosing an effective chemotherapy regimen for GC patients undergoing chemotherapy.

PLA2G12B Mediates Arachidonic Acid Metabolism through Activation of the NF-κB Pathway to Promote Membrane Nephropathy

Introduction: The disruption of podocyte structure and function are the main pathological mechanism of membranous nephropathy (MN). Phospholipases A2, Group XII B (PLA2G12B) was reported involved in the regulation of MN by interfering with arachidonic acid (AA) metabolism, but there is a lack of sufficient evidence. In this study, we investigated the role and molecular mechanism of PLA2G12B in MN. Methods: C57BL/6 mice were used to establish MN model to extract primary podocytes, then divided into control, model, si-phospholipases A2 receptor (PLA2R), PLA2G12B, PLA2G12B + si-PLA2R, PLA2G12B + nuclear factor kappa-B (NF-κB) inhibitor, PLA2G12B + NF-κB inhibitor + si-PLA2R groups. Hematoxylin-eosin staining and immunofluorescence were used to detect kidney histological arrangement, serum levels of cholesterol-related indices, and AA. Genes and proteins associated with metabolism and inflammatory factors were detected by quantitative real-time PCR and Western blot. Results: The results revealed that AA metabolites were activated in the MN model mice, and the expression of PLA2G12B and NF-κB pathway levels were elevated. Besides, cellular experiments demonstrated that prostaglandin I2 (PGI2), thromboxane A2 (TXA2), leukotriene B4 (LTB4), and NF-κB pathway were significantly increased in the PLA2G12B group. Also, PLA2G12B promotes apoptosis and suppresses cell activity in podocytes, and these effects could be antagonized by NF-κB inhibitors. Furthermore, with the inference of si-PLA2R, the NF-κB inhibitors’ effects were reversed. Conclusion: Promotional effects of PLA2G12B in primary MN are associated with the regulation of AA metabolism and NF-κB pathway.

Huoxiang Zhengqi alleviates azoxymethane/dextran sulfate sodium-induced colitis-associated cancer by regulating Nrf2/NF-κB/NLRP3 signaling.

Colitis-associated cancer (CAC) is a subtype of inflammatory bowel disease (IBD)-associated colorectal cancer. Huoxiang Zhengqi (HXZQ) is a classical Chinese herbal medicine and has been used to treat intestinal disorders, however, anti-CAC effects and underlying mechanisms of HXZQ have not been reported. An azoxymethane/dextran sulfate sodium-induced CAC mice model was used to investigate the anti-CAC effect of HXZQ. HXZQ significantly reduced colonic inflammation, suppressed the size and number of tumors, and reduced the levels of pro-inflammatory cytokines (interleukin [IL]-1α, IL-1β, IL-6, IL-17A, IL-21, IL-23, granulocyte macrophage-colony stimulating factor, and tumor necrosis factor-α) and oxidative stress markers (reactive oxygen species and malondialdehyde), and increased the levels of anti-inflammatory cytokines (IL-10 and IL-27) in CAC mice. Intestinal microbiota and serum metabolomics analyses indicated that HXZQ altered the gut microbial composition and the abundance of 29 serum metabolites in CAC mice. Additionally, HXZQ activated the nuclear factor-erythroid factor 2-related factor 2 (Nrf2) signaling pathway and increased the levels of antioxidants such as catalase (CAT), heme oxygenase-1 (HO-1), NAD(P)H quinone oxidoreductases-1 (NQO-1), and superoxide dismutase-1 (SOD-1). HXZQ inhibited the activation of the nuclear factor kappa-B (NF-κB) signaling pathway and decreased the expression of NLR family pyrin domain containing 3 (NLRP3) by inhibiting the phosphorylation of inhibitor of nuclear factor kappa-B (IκB), inhibitor of nuclear factor kappa-B kinase (IKK), and NF-κB. In conclusion, HXZQ alleviated CAC in mice by modulating the intestinal microbiota and metabolism, activating Nrf2-mediated antioxidant response, and inhibiting NF-κB-mediated NLRP3 inflammasome activation against inflammation. The present data provide a reference for the use of HXZQ as a therapeutic or combination agent for clinical CAC treatment.

Prolonged activation of NMDA receptors induces dedifferentiation of islet β cells in mice.

The β-cell dedifferentiation is one of the critical mechanisms in diabetic β-cell loss. Long-term activation of N-methyl-D-aspartate (NMDA) receptors plays an essential role in the development of diabetes, but the underlying mechanisms have not been fully elucidated. This study aims to investigate the effect of prolonged activation of NMDA receptors on islet β-cell dedifferentiation. Male C57BL/6 mice were randomly divided into a normal control group (control group) and an NMDA group. The mice in the NMDA group were intraperitoneally injected with NMDA (8 mg/kg body weight) and those in the control group were injected with the same volume of saline every day for 6 months. At the end of the 6 th month, glucose tolerance and enzyme linked immunosorbent assay (ELISA) were used to detect the function of islets, and pancreatic tissues were taken for immunofluorescence staining to detect the expressions of insulin, glucagon, and proliferating cell nuclear antigen (PCNA). Real-time PCR was used to detect the mRNA expression of pancreatic β cells, α cells, and islet progenitor cell markers.The primary islets were treated with NMDA to observe the effect of NMDA on the dedifferentiation of β cells. The nuclear factor kappa-B (NF-κB) inhibitor BAY 11-7082 was used at the cellular level via detecting insulin secretion and the expression of endocrine cell markers. Compared with the control group, the mice in the NMDA group had higher blood glucose levels at each time point after glucose injection, and the area under the glucose tolerance curve was significantly increased ( P <0.05). The serum insulin content and insulin stimulatory index of the mice in the NMDA group were significantly lower than those in the control group at 30 min after glucose injection (both P <0.05). The double immunofluorescence staining for insulin and glucagon showed that the number of insulin-positive β cells in the pancreatic tissues of mice was significantly decreased after intraperitoneal injection of NMDA in mice for 6 months, while the number of glucagon-positive α cells was significantly increased. Real-time PCR results showed that β-cell markers ( Insulin , Pdx1 , Neurod1 , and Mafa ) were significantly down-regulated in mouse pancreatic tissues after intraperitoneal injection of NMDA for 6 months, while pancreatic progenitor cell markers ( Neurog3 , Gata6 , Hnf4a , Notch1, and Hes1 ) were significantly down-regulated; α-cell markers ( Glucagon , Arx , Irx2 , Mafb , Pou6f2 , Fev , Kcnj3, and Sv2b ) were significantly up-regulated. NMDA treatment of mouse primary islets for 48 h cause significant down-regulation of β-cell marker gene expression ( P <0.05 or P <0.01), accompanied by significant up-regulation of pancreatic progenitor cell markers and α-cell markers ( P <0.05, P <0.01 or P <0.001). The NF-κB inhibitor BAY 11-7082 significantly blocked the down-regulation of β-cell marker expression (all P <0.05) and the up-regulation of α-cell and pancreatic progenitor cell marker after NMDA treatment of islets for 48 h ( P <0.05 or P <0.01). Prolonged activation of NMDA receptors induces islet β-cell dedifferentiation via regulating the NF-κB pathway.