Syringin from Tinospora crispa downregulates pro-inflammatory mediator production through MyD88-dependent pathways in lipopolysaccharide (LPS)-induced U937 macrophages.

BACKGROUND: Syringin, a phenylpropanoid glycoside, has exhibited numerous biological properties including inhibitory activities against various immune and inflammatory disorders. In this study, syringin isolated from Tinospora crispa was evaluated for its ability to down-regulate activated nuclear factor-kappa B (NF-κB), phosphoinositide-3-kinase-Akt (PI3K-Akt) and mitogen-activated protein kinases (MAPKs) signal transducing networks in U937 macrophages activated by lipopolysaccharide. METHODS: The attenuating effects of syringin on the productions of prostaglandin E2 (PGE2), cyclooxygenase-2 (COX-2), interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α), and the expressions of signaling molecules of the signaling pathways were investigated by using ELISA, Western blot, and qRT-PCR. RESULTS: Syringin downregulated the NF-κB, MAPKs, and PI3K-Akt signal networks by significantly reducing PGE2 production in the macrophages via suppression of COX-2 gene and protein expression levels. It also reduced TNF-α and IL-1ß secretion and their mRNA expression, suppressed phosphorylation of NF-κB (p65), IKKα/ß, and IκBα, and restored ability of IκBα to degrade. Syringin dose-dependently attenuated Akt, p38 MAPKs, JNK, and ERK phosphorylation. Also, the expression of corresponding upstream signaling molecules toll-like receptor 4 (TLR4) and myeloid differentiation primary response gene 88 (MyD88) were down-regulated in response to syringin treatment. CONCLUSION: The suppressive effect of syringin on the inflammatory signaling molecules in MyD88-dependent pathways suggested it's potential as a drug candidate for development into an agent for treatment of various immune-mediated inflammatory disorders.

Mitochondrial GRIM19 Loss Induces Liver Fibrosis through NLRP3/IL33 Activation via Reactive Oxygen Species/NF-кB Signaling.

Background and Aims: Hepatic fibrosis (HF) is a critical step in the progression of hepatocellular carcinoma (HCC). Gene associated with retinoid-IFN-induced mortality 19 (GRIM19), an essential component of mitochondrial respiratory chain complex I, is frequently attenuated in various human cancers, including HCC. Here, we aimed to investigate the potential relationship and underlying mechanism between GRIM19 loss and HF pathogenesis. Methods: GRIM19 expression was evaluated in normal liver tissues, hepatitis, hepatic cirrhosis, and HCC using human liver disease spectrum tissue microarrays. We studied hepatocyte-specific GRIM19 knockout mice and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 (Cas9) lentivirus-mediated GRIM19 gene-editing in murine hepatocyte AML12 cells in vitro and in vivo. We performed flow cytometry, immunofluorescence, immunohistochemistry, western blotting, and pharmacological intervention to uncover the potential mechanisms underlying GRIM19 loss-induced HF. Results: Mitochondrial GRIM19 was progressively downregulated in chronic liver disease tissues, including hepatitis, cirrhosis, and HCC tissues. Hepatocyte-specific GRIM19 heterozygous deletion induced spontaneous hepatitis and subsequent liver fibrogenesis in mice. In addition, GRIM19 loss caused chronic liver injury through reactive oxygen species (ROS)-mediated oxidative stress, resulting in aberrant NF-кB activation via an IKK/IкB partner in hepatocytes. Furthermore, GRIM19 loss activated NLRP3-mediated IL33 signaling via the ROS/NF-кB pathway in hepatocytes. Intraperitoneal administration of the NLRP3 inhibitor MCC950 dramatically alleviated GRIM19 loss-driven HF in vivo. Conclusions: The mitochondrial GRIM19 loss facilitates liver fibrosis through NLRP3/IL33 activation via ROS/NF-кB signaling, providing potential therapeutic approaches for earlier HF prevention.

Copper Promotes LPS-Induced Inflammation via the NF-кB Pathway in Bovine Macrophages.

Inflammation is a complex physiological process that enables the clearance of pathogens and repairing damaged tissues. Elevated serum copper concentration has been reported in cases of inflammation, but the role of copper in inflammatory responses remains unclear. This study used bovine macrophages to establish lipopolysaccharide (LPS)-induced inflammation model. There were five groups in the study: a group treated with LPS (100 ng/ml), a group treated with either copper chelator (tetrathiomolybdate, TTM) (20 µmol) or CuSO4 (25 µmol or 50 µmol) after LPS stimulation, and a control group. Copper concentrations increased in macrophages after the LPS treatment. TTM decreased mRNA expression of pro-inflammatory factors (IL-1ß, TNF-α, IL-6, iNOS, and COX-2), whereas copper supplement increased them. Compared to the control group, TLP4 and MyD88 protein levels were increased in the TTM and copper groups. However, TTM treatment decreased p-p65 and increased IкB-α while the copper supplement showed reversed results. In addition, the phagocytosis and migration of bovine macrophages decreased in the TTM treatment group while increased in the copper treatment groups. Results mentioned above indicated that copper could promote the LPS-induced inflammatory response in bovine macrophages, promote pro-inflammatory factors by activating the NF-кB pathway, and increase phagocytosis capacity and migration. Our study provides a possible targeted therapy for bovine inflammation.

Chronic HIV-1 Tat action induces HLA-DR downregulation in B cells: A mechanism for lymphoma immune escape in people living with HIV.

Despite the success of combination antiretroviral therapy, people living with human immunodeficiency virus (HIV) still have an increased risk of Epstein-Barr virus (EBV)-associated B cell malignancies. In the HIV setting, B cell physiology is altered by coexistence with HIV-infected cells and the chronic action of secreted viral proteins, for example, HIV-1 Tat that, once released, efficiently penetrates noninfected cells. We modeled the chronic action of HIV-1 Tat on B cells by ectopically expressing Tat or TatC22G mutant in two lymphoblastoid B cell lines. The RNA-sequencing analysis revealed that Tat deregulated the expression of hundreds of genes in B cells, including the downregulation of a subset of major histocompatibility complex (MHC) class II-related genes. Tat-induced downregulation of HLA-DRB1 and HLA-DRB5 genes led to a decrease in HLA-DR surface expression; this effect was reproduced by coculturing B cells with Tat-expressing T cells. Chronic Tat presence decreased the NF-ᴋB pathway activity in B cells; this downregulated NF-ᴋB-dependent transcriptional targets, including MHC class II genes. Notably, HLA-DRB1 and surface HLA-DR expression was also decreased in B cells from people with HIV. Tat-induced HLA-DR downregulation in B cells impaired EBV-specific CD4+ T cell response, which contributed to the escape from immune surveillance and could eventually promote B cell lymphomagenesis in people with HIV.

Helicobacter pylori induces a novel form of innate immune memory via accumulation of NF-кB proteins.

Helicobacter pylori is a widespread Gram-negative pathogen involved in a variety of gastrointestinal diseases, including gastritis, ulceration, mucosa-associated lymphoid tissue (MALT) lymphoma and gastric cancer. Immune responses aimed at eradication of H. pylori often prove futile, and paradoxically play a crucial role in the degeneration of epithelial integrity and disease progression. We have previously shown that H. pylori infection of primary human monocytes increases their potential to respond to subsequent bacterial stimuli - a process that may be involved in the generation of exaggerated, yet ineffective immune responses directed against the pathogen. In this study, we show that H. pylori-induced monocyte priming is not a common feature of Gram-negative bacteria, as Acinetobacter lwoffii induces tolerance to subsequent Escherichia coli lipopolysaccharide (LPS) challenge. Although the increased reactivity of H. pylori-infected monocytes seems to be specific to H. pylori, it appears to be independent of its virulence factors Cag pathogenicity island (CagPAI), cytotoxin associated gene A (CagA), vacuolating toxin A (VacA) and γ-glutamyl transferase (γ-GT). Utilizing whole-cell proteomics complemented with biochemical signaling studies, we show that H. pylori infection of monocytes induces a unique proteomic signature compared to other pro-inflammatory priming stimuli, namely LPS and the pathobiont A. lwoffii. Contrary to these tolerance-inducing stimuli, H. pylori priming leads to accumulation of NF-кB proteins, including p65/RelA, and thus to the acquisition of a monocyte phenotype more responsive to subsequent LPS challenge. The plasticity of pro-inflammatory responses based on abundance and availability of intracellular signaling molecules may be a heretofore underappreciated form of regulating innate immune memory as well as a novel facet of the pathobiology induced by H. pylori.

Korean Chestnut Honey Suppresses HSV-1 Infection by Regulating the ROS-NLRP3 Inflammasome Pathway.

Herpes simplex virus 1 (HSV-1) is double-stranded DNA virus that belongs to the Orthoherpesviridae family. It causes serious neurological diseases of the central nervous system, such as encephalitis. The current U.S. Food and Drug Administration (FDA)-approved drugs for preventing HSV-1 infection include acyclovir (ACV) and valacyclovir; however, their long-term use causes severe side effects and often results in the emergence of drug-resistant strains. Therefore, it is important to discover new antiviral agents that are safe and effective against HSV-1 infection. Korean chestnut honey (KCH) has various pharmacological activities, such as antioxidant, antibacterial, and anti-inflammation effects; however, antiviral effects against HSV-1 have not yet been reported. Therefore, we determined the antiviral activity and mechanism of action of KCH after HSV-1 infection on the cellular level. KCH inhibited the HSV-1 infection of host cells through binding and virucidal steps. KCH decreased the production of reactive oxygen species (ROS) and calcium (Ca2+) following HSV-1 infection and suppressed the production of inflammatory cytokines by inhibiting nuclear factor kappa-light-chain-enhancer of activated B cells (NF-кB) activity. Furthermore, we found that KCH inhibited the expression of the nod-like receptor protein 3 (NLRP3) inflammasome during HSV-1 infection. Taken together, the antiviral effects of KCH occur through multiple targets, including the inhibition of viral replication and the ROS-mediated NLRP3 inflammasome pathway. Our findings suggest that KCH has potential for the treatment of HSV-1 infection and related diseases.

Investigating the mechanism of action of Yanghe Pingchuan Granule in the treatment of bronchial asthma based on bioinformatics and experimental validation.

Background: Yanghe Pingchuan Granule (YPG) is a patented Chinese medicine developed independently by the Anhui Provincial Hospital of Traditional Chinese Medicine. For many years, it has been used for the treatment of asthma with remarkable clinical effects. However, the composition of YPG is complex, and its potential active ingredients and mechanism of action for the treatment of asthma are unknown. Materials and methods: In this study, we investigated the potential mechanism of action of YPG in the treatment of asthma through a combination of bioinformatics and in vivo experimental validation. We searched for active compounds in YPG and asthma targets from multiple databases and obtained common targets. Subsequently, a protein-protein interaction (PPI) network for compound disease was constructed using the protein interaction database for Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Finally, hematoxylin and eosin (H&E) staining, Masson staining, enzyme-linked immunosorbent assay (ELISA) analysis, immunofluorescence (IF) experiments, and Western blot (WB) experiments were performed to verify the possible mechanism of action of YPG for asthma treatment. Results: We obtained 72 active ingredients and 318 drug target genes that overlap with asthma. Serine/threonine-protein kinase (AKT1), tumor protein p53 (TP53), tumor necrosis factor (TNF), interleukin (IL)-6, IL-1ß, vascular endothelial growth factor-A (VEGFA), prostaglandin-endoperoxide synthase 2 (PTGS2), caspase-3 (CASP3), mitogen-activated protein kinase 3 (MAPK3) and epidermal growth factor receptor (EGFR) were the most relevant genes in the PPI network. KEGG analysis showed a high number of genes enriched for the nuclear factor kappa-B (NF-κB) signaling pathway. Animal experiments confirmed that YPG reduced inflammatory cell infiltration and down-regulated the expression of ovalbumin-induced inflammatory factors. Furthermore, YPG treatment decreased the protein expression of NFĸB1, nuclear factor kappa B kinase subunit beta (IKBKB), vascular endothelial growth factor (VEGF), and vascular endothelial growth factor receptor 2 (VEGFR2) in lung tissue. Conclusion: YPG has a positive effect on asthma by interfering with multiple targets. Furthermore, YPG may significantly inhibit the follicle-induced inflammatory response through the NF-ĸB signaling pathway.

Xianglian Zhixie Tablet Antagonizes Dextran Sulfate Sodium-Induced Ulcerative Colitis by Attenuating Systemic Inflammation and Modulating Gut Microbiota.

Purpose: Xianglian Zhixie Tablet (XLZXT), a classical traditional Chinese medicine formulation, is commonly used to treat Ulcerative Colitis (UC) in China. However, the therapeutic mechanisms of XLZXT for UC have yet to be fully understood. This study aimed to investigate the curative benefits of XLZXT and its associated mechanisms for healing UC in mice. Methods: In the present study, the 1% dextran sulfate sodium (DSS) solution was used to establish the UC model in C57BL/6N mice. To investigate the therapeutic effects of XLZXT on DSS-induced UC mice, several parameters were measured, including DAI score, colon length, spleen index, pathological changes in colon tissue, and levels of inflammatory factors in plasma and colon tissue. By investigating the gut microbiota, assessing the levels of intestinal mucosal protein expression, and looking at the proteins involved in the TLR4/MyD88/NF-B p65 signaling pathway, the mechanisms of XLZXT impact on UC were investigated. Mouse feces were examined for patterns of gut microbiota expression using high-throughput sequencing of 16S rRNA. Results: XLZXT effectively alleviated UC symptoms and colon pathological damage in DSS-induced UC mice. It improved body weight loss, stool consistency, and hematochezia, while also repairing colon damage. Moreover, it down-regulated pro-inflammatory cytokines (such as TNF-α, IL-1ß, and IL-6), and up-regulated anti-inflammatory cytokines (such as IL-10). XLZXT also increased the expression of MUC-2, Occludin and ZO-1, while decreasing the expression of NF-κB, MyD88 and TLR4. Additionally, it regulated gut microbiota disorder by increasing the abundance of beneficial bacteria and reducing the adhesion of intestinal harmful bacteria. Conclusion: XLZXT demonstrated therapeutic effects on DSS-induced UC mice. The mechanisms may be associated with repairing the intestinal mucosal barrier, regulating the TLR4/MyD88/NF-κB p65 signaling pathway, and restoring the balance of gut microbiota.

Exosomes derived from bone marrow mesenchymal stem cells regulate pyroptosis via the miR-143-3p/myeloid differentiation factor 88 axis to ameliorate intestinal ischemia-reperfusion injury.

Intestinal ischemia-reperfusion (I/R) injury is a condition in which tissue injury is aggravated after ischemia due to recovery of blood supply. Bone marrow mesenchymal stem cell-derived exosome (BMSC-exo) showed a protective effect on I/R injury. This study aimed to investigate the possible mechanisms by which BMSC-exos ameliorate intestinal I/R injury. We isolated mouse BMSC-exos by super-centrifugation and found that they effectively increased cell viability in a cell model, alleviated intestinal barrier injury in a mouse model, and downregulated the expression of inflammatory cytokines and pyroptosis-related proteins, suggesting that BMSC-exos may alleviate intestinal I/R injury in vitro and in vivo by regulating pyroptosis. We identified miR-143-3p as a differentially expressed miRNA by microarray sequencing. Bioinformatic analysis predicted a binding site between miR-143-3p and myeloid differentiation factor 88 (MyD88); a dual-luciferase reporter assay confirmed that miR-143-3p could directly regulate the expression of MyD88. Our findings suggest that miR-143-3p regulates pyroptosis by regulating NOD-like receptor thermal protein domain associated protein 3 (NLRP3) through the toll-like receptor (TLR)-4/MyD88/nuclear factor kappa-B (NF-кB) pathway. This study describes a potential strategy for the treatment of intestinal I/R injury using BMSC-exos that act by regulating pyroptosis through the miR-143-3p mediated TLR4/MyD88/NF-кB pathway.

BMSC-exos ameliorate intestinal ischemia/reperfusion (I/R) injurymiR-143-3p levels were reduced in I/R injury and increased with BMSC-exo treatmentmiR-143-3p directly targeted and downregulated the expression of MyD88BMSC-exos regulate pyroptosis in intestinal I/R injury via the miR-143-3p-MyD88 axis.

Biodistribution and intestinal inflammatory response following voluntary oral intake of silver nanoparticles by C57BL/6J mice.

Silver nanoparticles (AgNP) are among the most widely commercialized nanomaterials globally, with applications in medicine and the food industry. Consequently, the increased use of AgNP in the food industry has led to an unavoidable rise  in human exposure to these nanoparticles. Their widespread use raises concerns about potential hazards to human health, specifically their intestinal pro-inflammatory effects. Thus, the main objective of this study was to evaluate the biological effects of two subacute doses of 5 nm polyvinylpyrrolidone (PVP)-AgNP in C57BL/6J mice. One mg/kg body weight or 10 mg/kg bw was provided once a day for 14 days, using a new technology (HaPILLness) that allows voluntary, stress-free, and accurate oral dosing. It was observed that after oral ingestion, while AgNP is biodistributed throughout the entire organism, most of the ingested dose is excreted in the feces. The passage and accumulation of AgNP throughout the intestine instigated a prominent inflammatory response, marked by significant histological, vascular, and cellular transformations. This response was driven by the activation of the nuclear factor-кB (NF-кB) inflammatory pathway, ultimately leading to the generation of multiple cytokines and chemokines.

Babao Dan alleviates gut immune and microbiota disorders while impacting the TLR4/MyD88/NF-кB pathway to attenuate 5-Fluorouracil-induced intestinal injury.

Adjuvant chemotherapy based on 5-fluorouracil (5-FU), such as FOLFOX, is suggested as a treatment for gastrointestinal cancer. Yet, intestinal damage continues to be a prevalent side effect for which there are no practical prevention measures. We investigated whether Babao Dan (BBD), a Traditional Chinese Medicine, protects against intestinal damage induced by 5-FU by controlling immune response and gut microbiota. 5-FU was injected intraperitoneally to establish the mice model, then 250 mg/kg BBD was gavaged for five days straight. 5-FU led to marked weight loss, diarrhea, fecal blood, and histopathologic intestinal damage. Administration of BBD reduced these symptoms, inhibited proinflammatory cytokine (IL-6, IL-1ß, IFN-γ, TNF-α) secretion, and upregulated the ratio of CD3(+) T cells and the CD4(+)/CD8(+) ratio. According to 16S rRNA sequencing, BBD dramatically repaired the disruption of the gut microbiota caused in a time-dependent way, and increased the Firmicutes/Bacteroidetes (F/B) ratio. Transcriptomic results showed that the mechanism is mainly concentrated on the NF-κB pathway, and we found that BBD reduced the concentration of LPS in the fecal suspension and serum, and inhibited TLR4/MyD88/NF-κB pathway activation. Furthermore, at the genus level on the fifth day, BBD upregulated the abundance of unidentified_Corynebacteriaceae, Aerococcus, Blautia, Jeotgalicoccus, Odoribacter, Roseburia, Rikenella, Intestinimonas, unidentified_Lachnospiraceae, Enterorhabdus, Ruminiclostridium, and downregulated the abundance of Bacteroides, Parabacteroides, Parasutterella, Erysipelatoclostridium, which were highly correlated with intestinal injury or the TLR4/MyD88/NF-κB pathway. In conclusion, we established a network involving 5-FU, BBD, the immune response, gut microbiota, and key pathways to explain the pharmacology of oral BBD in preventing 5-FU-induced intestinal injury.

Exosomal lncRNA GAS5 promotes M1 macrophage polarization in allergic rhinitis via restraining mTORC1/ULK1/ATG13-mediated autophagy and subsequently activating NF-кB signaling.

Macrophages are involved in the pathogenesis of allergic rhinitis (AR), but how these macrophages are polarized to M1 or M2 type is undetermined. Long non-coding RNA growth arrest specific transcript 5 (GAS5) is upregulated in exosomes isolated from nasal mucus of AR patients (AR-EXO) and aggravates nasal symptoms in AR mice. In the present study, we are aimed to elucidate the potential role of GAS5 in macrophage polarization during AR pathogenesis. An AR mice model was constructed. The potential function of GAS5 was evaluated by western blot, RNA immunoprecipitation (RIP), biotinylated RNA pull-down assay, co-immunoprecipitation (co-IP) assay, flow cytometry, enzyme-linked immunosorbent assay (ELISA) assay, and immunohistochemistry (IHC) staining. We found that GAS5 is upregulated in ovalbumin-treated human nasal epithelial cells RPMI 2650 (OVA-EXO) and nasal mucus of AR mice. OVA-EXO treatment or forced GAS5 expression promoted M1 macrophage polarization of peripheral blood monocytes (PB monocytes) and THP-1 macrophages in vitro. GAS5 overexpression aggravated the allergic nasal symptoms induced by OVA in AR mice and facilitated M1 macrophage polarization and allergic inflammation, while knockdown of GAS5 exhibited opposite effects in vivo. GAS5 activated NF-кB signaling via suppressing autophagy-dependent degradation of IKKα/ß in macrophages. Furthermore, GAS5 acted as a scaffold to strengthen the interaction between mTORC1 and ULK1, thus impaired ULK1/ATG13-mediated autophagy via increasing mTORC1 activity. Finally, restored autophagy by ATG13 overexpression suppressed the effect of GAS5 on M1 macrophage polarization. In conclusion, these results suggested that exosomal transfer of GAS5 promoted M1 macrophage polarization via restraining mTORC1/ULK1/ATG13-mediated autophagy and subsequently activating NF-кB signaling in allergic rhinitis.

Gentisic acid ameliorates type 2 diabetes induced by Nicotinamide-Streptozotocin in male mice by attenuating pancreatic oxidative stress and inflammation through modulation of Nrf2 and NF-кB pathways.

AIMS: There is a close link between oxidative stress, inflammation, and type 2 diabetes mellitus (T2DM). Gentisic acid (GA) is a di-phenolic compound and an active metabolite of aspirin that possesses antioxidant and anti-inflammatory properties, but its potential anti-diabetic effects have not been evaluated so far. Therefore, this study aimed to evaluate GA's potential antidiabetic effects through the Nuclear Factor Erythroid 2-Related Factor (Nrf2) and Nuclear Factor Kappa Beta (NF-кB) signaling pathways. MATERIAL AND METHODS: In this study, T2DM induced by a single intraperitoneal injection of STZ (65 mg/kg B.W) after 15 min nicotinamide (120 mg/kg B.W) injection. After seven days of injections, fasting blood glucose (FBS) was measured. Seven days after FBS monitoring treatments started. Grouping and treatments were as follows: 1) Normal control group; NC, 2) Diabetic control group; DC, 3) Metformin group; MT (150 mg/kg B.W, daily), 4) Test group; GA (100 mg/kg B.W, daily). Treatments continued for 14 consecutive days. KEY FINDINGS: Diabetic mice treatment with GA significantly decreased FBS, improved plasma lipid profiles and pancreatic antioxidant status. GA modulated Nrf2 pathway by upregulation of Nrf2 protein, NAD(P)H: quinone oxidoreductase 1 (Nqo1), and p21, and downregulation of miR-200a, Kelch-like ECH-associated protein 1 (Keap1), and nicotinamide adenine dinucleotide phosphate oxidase-2 (NOX2). Also, GA attenuated inflammation by upregulation of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and interleukin-10 (IL-10) and downregulation of miR-125b, NF-кB, tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1ß). SIGNIFICANCE: GA attenuates T2DM, possibly by improving antioxidant status through the Nrf2 pathway and attenuation of inflammation.

Panax ginseng abuse exhibits a pro-inflammatory effect by activating the NF-κB pathway.

P. ginseng (Panax ginseng C. A. Meyer) is a well-known traditional medicine that has been used for thousands of years to treat diseases. However, "ginseng abuse syndrome" (GAS) often occurs due to an inappropriate use such as high-dose or long-term usage of ginseng; information about what causes GAS and how GAS occurs is still lacking. In this study, the critical components that potentially caused GAS were screened through a step-by-step separation strategy, the pro-inflammatory effects of different extracts on messenger RNA (mRNA) or protein expression levels were evaluated in RAW 264.7 macrophages through quantitative real-time polymerase chain reaction (qRT-PCR) or Western blot, respectively. It was found that high-molecular water-soluble substances (HWSS) significantly increased the expression of cytokines (cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and interleukin 6 (IL-6)) and cyclooxygenase 2 (COX-2) protein; gel filtration chromatography fraction 1 (GFC-F1) further purified from HWSS showed prominent pro-inflammatory effects by increasing the transcription of cytokines (COX-2, iNOS, tumor necrosis factor alpha (TNF-α), and interleukin 1ß (IL-1ß)) as well as the expression of COX-2 and iNOS protein. Moreover, GFC-F1 activated nuclear factor-kappa B (NF-кB) (p65 and inhibitor of nuclear factor-kappa B alpha (IκB-α)) and the p38/MAPK (mitogen-activated protein kinase) signaling pathways. On the other hand, the inhibitor of the NF-κB pathway (pyrrolidine dithiocarbamate (PDTC)) reduced GFC-F1-induced nitric oxide (NO) production, while the inhibitors of the MAPK pathways did not. Taken together, GFC-F1 is the potential composition that caused GAS through the production of inflammatory cytokines by activating the NF-кB pathway.

Isochlorogenic acid A alleviates dextran sulfate sodium-induced ulcerative colitis in mice through STAT3/NF-кB pathway.

Isochlorogenic acid A (ICGA-A) is a dicaffeoylquinic acid widely found in various medicinal plants or vegetables, such as Lonicerae japonicae Flos and chicory, and multiple properties of ICGA-A have been reported. However, the therapeutic effect of ICGA-A on colitis is not clear, and thus were investigated in our present study, as well as the underlying mechanisms. Here we found that ICGA-A alleviated clinical symptoms of dextran sodium sulfate (DSS) induced colitis model mice, including disease activity index (DAI) and histological damage. In addition, DSS-induced inflammation was significantly attenuated in mice given ICGA-A supplementation. ICGA-A reduced the fraction of neutrophils in peripheral blood and the infiltration of neutrophils and macrophages in colon tissue, and reduced pro-inflammatory cytokine production and tight junctions in mouse models. Furthermore, ICGA-A down-regulated expression of STAT3 and up-regulated the protein level of IκBα. Our in vitro studies confirmed that ICGA-A inhibited the mRNA expression of pro-inflammatory cytokines. ICGA-A blocked the phosphorylation of STAT3, p65, and IκBα, suppressed the expression STAT3 and p65. In addition, the present study also demonstrated that ICGA-A had no obvious toxicity on normal cells and organs. Taken together, we conclude that ICGA-A mitigates experimental ulcerative colitis (UC) at least in part by inhibiting the STAT3/NF-кB signaling pathways. Hence, ICGA-A may be a promising and effective drug for treating UC.

Characterization of effects of chitooligosaccharide monomer addition on immunomodulatory activity in macrophages.

The present study aimed to investigate the effects of five chitooligosaccharide monomers of different molecular weights on immunomodulatory activity in macrophage-like RAW264.7 cells. The incubation of various chitooligosaccharide monomers enhanced phagocytosis and pinocytosis activity toward Staphylococcus aureus and Escherichia coli in RAW264.7 cells. The incorporation of chitooligosaccharide monomers significantly boosted the generation of reactive oxygen species and reactive nitrogen species, as well as the release of inflammatory cytokines. To further explore the mechanism of inflammation regulated by chitooligosaccharide, the activation inhibitors of NF-кB (CAPE) and TLR-4 (TAK-242) were utilized, the determination data demonstrated that chitobiose suppressed the expression of inflammatory cytokines and NF-кB p65. In addition, the investigation results revealed that the presence of the mannose receptor inhibitor (mannan) suppressed chitohexaose-induced phagocytic activity and inflammatory cytokines. These results suggested that the five distinct chitooligosaccharide monomers had inconsistent effects, the chitobiose and chitohexaose exhibiting the best biological activity in activating RAW264.7 cells, promoting cell proliferation, and increasing non-specific immunity.

Human Blood Extracellular Vesicles Activate Transcription of NF-kB-Dependent Genes in A549 Lung Adenocarcinoma Cells.

Extracellular vesicles (EVs) produced by various cell types are heterogeneous in size and composition. Changes in the RNA sets of EVs in biological fluids are considered the basis for the development of new approaches to minimally invasive diagnostics and the therapy of human diseases. In this study, EVs were obtained from the blood of healthy donors by centrifugation, followed by ultracentrifugation. It was shown that EVs consist of several populations including small exosome-like vesicles and larger microvesicle-like particles. The composition of EVs' RNAs was determined. A549 lung adenocarcinoma cells were incubated with EV and the NGS analysis of differentially expressed genes was performed. During the incubation of A549 cells with EVs, the levels of mRNA encoding components for the NF-kB signaling pathway increased, as well as the expression of genes controlled by the NF-kB transcription factor. Overall, our results suggest that components of EVs trigger the NF-kB signaling cascade in A549 cells, leading to the transcription of genes including cytokines, adhesion molecules, cell cycle regulators, and cell survival factors. Our data provide insight into the interaction between blood EVs and human cells and can be used for designing new tools for the diagnosis and treatment of human diseases.

Parental cadmium exposure during the spawning period reduces cadmium sensitivity through the antioxidant system in rare minnow (Gobiocypris rarus) larvae.

Cadmium (Cd) is a noxious heavy metal widely dispersed in aquatic systems. Parental Cd exposure of fish species at environmental concentrations has been shown to cause deformities and stunted growth in their offspring. However, the long-term effects and the mechanisms underlying parental Cd exposure in fish species on Cd sensitivity in their offspring remain unclear. To explore the impacts of parental Cd exposures on Cd sensitivity, rare minnow (Gobiocypris rarus) larvae whose parents were exposed to Cd at 0, 5 or 10 µg/L for 28 days were established. Results showed that parental Cd exposure in rare minnow increased the Cd content of its larvae. In terms of malformation rate, mortality rate and total length at 7 days of rare minnow larvae, parental Cd exposure at 5 or 10 µg/L reduced Cd sensitivity. Further mechanistic investigation demonstrated that parental Cd exposure significantly upregulated the expression of antioxidant gene regulated by nuclear factor erythroid 2-related factor 2 (Nrf2) and nuclear factor-kappa B (NF-кB) in rare minnow larvae. In addition, parental Cd exposure significantly elevated the level of reactive oxygen species (ROS) and malondialdehyde (MDA), but markedly decreased catalase (CAT), superoxide dismutase (SOD) and oxidized glutathione (GST) activity. The impact of parental Cd exposure to metallothionein (MT) content and the expression of MT mRNA, a detoxifying metallothionein, showed that parental Cd exposure of rare minnow induced oxidative stress in the larvae. Meanwhile, these results indicated that parental Cd exposure in rare minnow reduced the Cd sensitivity of the larvae via activating the Nrf2-mediated antioxidant system. This project helps us to further understand the toxicological mechanism of Cd in fish species and properly assess its potential ecological risk.

Hsa-microRNA-27b-3p inhibits hepatocellular carcinoma progression by inactivating transforming growth factor-activated kinase-binding protein 3/nuclear factor kappa B signalling.

BACKGROUND: MicroRNAs (miRNAs) play crucial roles in the development of hepatocellular carcinoma (HCC). Hsa-microRNA-27b-3p (hsa-miR-27b) is involved in the formation and progression of various cancers, but its role and clinical value in HCC remain unclear. METHODS: The expression of hsa-miR-27b in HCC was examined by quantitative real-time PCR (qRT-PCR) and in situ hybridization (ISH) assays of clinical samples. Cell Counting Kit-8 assays (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays, Transwell assays, filamentous actin (F-actin) staining and western blot analyses were used to determine the effects of hsa-miR-27b on HCC cells in vitro. Subcutaneous xenograft and lung metastatic animal experiments were conducted to verify the role of hsa-miR-27b in HCC in vivo. In silico prediction, qRT-PCR, western blot, anti-Argonaute 2 (AGO2) RNA immunoprecipitation (RIP) and dual luciferase reporter assays were applied to identify the target genes of hsa-miR-27b. To detect the impacts of hsa-miR-27b on nuclear factor kappa B (NF-кB) signalling cascades mediated by transforming growth factor-activated kinase-binding protein 3 (TAB3), we performed qRT-PCR, western blot assays, immunofluorescence staining, immunohistochemistry (IHC) and dual-luciferase reporter assays. Recombinant oncolytic adenovirus (OncoAd) overexpressing hsa-miR-27b was constructed to detect their therapeutic value in HCC. RESULTS: The expression of hsa-miR-27b was lower in HCC than in adjacent non-tumourous tissues (ANTs), and the reduced expression of hsa-miR-27b was associated with worse outcomes in patients with HCC. Hsa-miR-27b significantly inhibited the proliferation, migration, invasion, subcutaneous tumour growth and lung metastasis of HCC cells. The suppression of hsa-miR-27b promoted the nuclear translocation of NF-κB by upregulating TAB3 expression. TAB3 was highly expressed in HCC compared with ANTs and was negatively correlated with the expression of hsa-miR-27b. The impaired cell proliferation, migration and invasion by hsa-miR-27b overexpression were recovered by ectopic expression of TAB3. Recombinant OncoAd with overexpression of hsa-miR-27b induced anti-tumour activity compared with that induced by negative control (NC) OncoAd in vivo and in vitro. CONCLUSIONS: By targeting TAB3, hsa-miR-27b acted as a tumour suppressor by inactivating the NF-кB pathway in HCC in vitro and in vivo, indicating its therapeutic value against HCC.

The effect of a chrysanthemum water extract in protecting the retina of mice from light damage.

BACKGROUND: Oxidative stress can induce age-related diseases. Age-related retinal diseases, such as age-related macular degeneration (AMD), are difficult to cure owing to their complicated mechanisms. Although anti-neovascular therapeutics are used to treat wet AMD, vision cannot always be completely restored, and disease progression cannot always be inhibited. Therefore, determining a method to prevent or slow retinal damage is important. This study aimed to investigate the protective effect of a chrysanthemum water extract rich in flavone on the oxidatively stressed retina of mice. METHODS: Light damage was induced to establish oxidative stress mouse models. For in vitro experiments, ARPE-19 cells were cultured and divided into four groups: control, light-damaged, and low- and high-dose chrysanthemum extract. No treatment was administered in the control group. The light-damaged and low- and high-dose chrysanthemum extract groups were exposed to a similar white light level. The chrysanthemum extract was added at a low dose of 0.4 mg/mL or a high dose of 1.0 mg/mL before cell exposure to 2500-lx white light. Reactive oxygen species (ROS) level and cellular viability were measured using MTT and immunofluorescence staining. For in vivo experiments, C57BL/6 J mice were divided into the same four groups. Low- (0.23 g/kg/day) and high-dose (0.38 g/kg/day) chrysanthemum extracts were continuously intragastrically administered for 8 weeks before mouse exposure to 10,000-lx white light. Retinal function was evaluated using electroretinography. In vivo optical coherence tomography and in vitro haematoxylin and eosin staining were performed to observe the pathological retinal changes in each group after light damage. Fluorescein fundus angiography of the arteriovenous vessel was performed, and the findings were analysed using the AngioTool software. TUNEL immunofluorescence staining was used to assess isolated retinal apoptosis. RESULTS: In vitro, increased ROS production and decreased ARPE-19 cell viability were found in the light-damaged group. Improved ARPE-19 cell viability and reduced ROS levels were observed in the chrysanthemum extract treatment groups. In vivo, dysfunctional retinas and abnormal retinal structures were found in the light-damaged group, as well as increased apoptosis in the retinal ganglion cells (RGCs) and inner and outer nuclear layers. The apoptosis rate in the same layers was lower in the chrysanthemum extract treatment groups than in the light-damaged group. The production of antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px), increased in the treatment groups. NF-κB in the nucleus and TNF-α were more highly expressed in the light-damaged group than in the low- and high-dose chrysanthemum extract groups. CONCLUSIONS: Light damage-induced retinal oxidative stress can lead to ROS accumulation in the retinal tissues. Herein, RGC and photoreceptor layer apoptosis was triggered, and NF-κB in the nucleus and TNF-α were highly expressed in the light-damaged group. Preventive chrysanthemum extract administration decreased ROS production by increasing SOD, CAT, and GSH-Px activities and reversing the negative changes, demonstrating a potential protective effect on the retina.