LPS-induced IL-6 Research Articles

GSK3179106 ameliorates lipopolysaccharide-induced inflammation and acute lung injury by targeting P38 MAPK

Acute lung injury (ALI) is a serious acute respiratory disease that can cause alveolar-capillary barrier disruption and pulmonary edema, respiratory failure and multiple organ dysfunction syndrome. However, there is no effective drugs in clinic until now. GSK3179106 has been reported can alleviate intestinal stress syndrome, but the protective effect of GSK3179106 on ALI has not been elucidated. The present study will evaluate the pharmacological activity of GSK3179106 on lipopolysaccharide (LPS)-induced inflammation and lung injury and clarify its underlying mechanism. We found that GSK3179106 significantly attenuated LPS-induced lung injury in vivo, accompanied by inhibited infiltration of inflammatory cells and reduced expression of inflammatory cytokines. Meanwhile, GSK3179106 dose-dependently reduced the LPS-induced IL-6 expression both in protein and gene levels in macrophages. Mechanistically, GSK3179106 could inhibited the phosphorylation of P38 MAPK induced by LPS. Importantly, results showed that there is a direct combination between GSK3179106 and P38 MAPK. Together, our findings not only clarified the anti-inflammatory activity of GSK3179106 but also discovered its new clinical indications. Therefore, compound GSK3179106 may be a potential candidate for the treatment of acute inflammatory diseases.

Human gut commensal Alistipes timonensis modulates the host lipidome and delivers anti-inflammatory outer membrane vesicles to suppress colitis in an Il10 -deficient mouse model.

Correlative studies have linked human gut microbes to specific health conditions. Alistipes is one such microbial genus negatively linked to inflammatory bowel disease (IBD). However, the protective role of Alistipes in IBD has not been studied and the underlying molecular mechanisms also remain unknown. In this study, colonization of Il10 -deficient mice with Alistipes timonensis DSM 27924 delays the development of colitis. Colonization with Alistipes does not significantly alter the gut microbiome composition during colitis development, but instead shifts the host plasma lipidome, increasing phosphatidic acids while decreasing triglycerides. Outer membrane vesicles (OMVs) derived from Alistipes are also detected in the plasma of colonized mice, which carry metabolites with immunomodulatory potential into the host circulatory system. We further demonstrate that fractions of A. timonensis OMVs suppress LPS-induced Il6 , Il1b , and Tnfa expression in vitro in murine macrophages. We detect immunomodulatory sulfonolipids (SoLs) in the active fraction, which are also increased in the blood of A. timonensis -colonized mice; and we identify other putative bioactive lipids in the A. timonensis OMVs. Thus, A. timonensis OMVs represent a potential mechanism for Alistipes -mediated delay of colitis progression in Il10 -deficient mice through the delivery of immunomodulatory lipids, including SoLs, and modulation of the host plasma lipidome.

Edodes Cultured Extract Regulates Immune Stress During Puberty and Modulates MicroRNAs Involved in Mammary Gland Development and Breast Cancer Suppression.

Immune stressors, such as lipopolysaccharides (LPS), profoundly affect microbiota balance, leading to gut dysbiosis. This imbalance disrupts the metabolic phenotype and structural integrity of the gut, increasing intestinal permeability. During puberty, a critical surge in estrogen levels is crucial for mammary gland development. However, inflammation originating from the gut in this period may interfere with this development, potentially heightening breast cancer risk later. The long-term effects of pubertal inflammation on mammary development and breast cancer risk are underexplored. Such episodes can dysregulate cytokine levels and microRNA expression, altering mammary cell gene expression, and predisposing them to tumorigenesis. This study hypothesizes that prebiotics, specifically Lentinula edodes Cultured Extract (AHCC), can counteract LPS's adverse effects. Using BALB/c mice, an acute LPS dose was administered at puberty, and breast cancer predisposition was assessed at 13 weeks. Cytokine and tumor-related microRNA levels, tumor development, and cancer stem cells were explored through immunoassays and qRT-PCR. Results show that LPS induces lasting effects on cytokine and microRNA expression in mammary glands and tumors. AHCC modulates cytokine expression, including IL-1β, IL-17A/F, and IL-23, and mitigates LPS-induced IL-6 in mammary glands. It also regulates microRNA expression linked to tumor progression and suppression, particularly counteracting the upregulation of oncogenic miR-21, miR-92, and miR-155. Although AHCC slightly alters some tumor-suppressive microRNAs, these changes are modest, highlighting a complex regulatory role that warrants further study. These findings underscore the potential of dietary interventions like AHCC to mitigate pubertal LPS-induced inflammation on mammary gland development and tumor formation, suggesting a preventive strategy against breast cancer.

Dexamethasone, Remdesivir and Azithromycin modulate ACE2 and IL-6 in Lung Epithelial Cells.

The optimal use of steroids in COVID-19 patients remains challenging. Current S3-guidelines "Recommendations for patients with COVID-19" recommend dexamethasone (DEX) for patients requiring respiratory support, remdesivir (RD) in the early disease phase and azythromycin (AZ) is no longer recommended. We investigated effects of DEX, RD and AZ in a lipopolysaccharide induced inflammation in lung cells in vitro and analyzed publicly available datasets with a focus on the Angiotensin-converting enzyme 2 (ACE2) to better understand drugs' mechanisms of action. human bronchial (Calu) and alveolar (A549) lung epithelial cells were treated with DEX, AZ or RDV in the presence of lipopolysaccharides (LPS). Gene expression (GE) of ACE2, IL-6 and the IL-6 protein release were measured. Publicly available GE data from lung tissues of COVID-19 patients and from lung cells treated with DEX were analyzed for the GE of ACE2. DEX increased and RDV and AZ reduced the GE of ACE2 in LPS-stimulated bronchial and alveolar epithelial cells. Only DEX significantly reduced LPS-induced IL-6 releases in alveolar cells substantially. The database analyses showed an, albeit not always significant, increase in ACE2 for lung tissue or cell lines treated with DEX. Lung tissue from patients after COVID-19 infection as well as bronchial cell cultures after COVID-19 infection showed lower GEs of ACE2. DEX can increase ACE2 expression in vitro and thereby the portal of entry of SARS-CoV-2 into lung cells during an LPS induced inflammation. Simultaneously the inflammatory marker IL-6 is reduced. Comparative database analyses indicate that these processes can also take place in vivo.

Desaminotyrosine is a redox-active microbial metabolite that bolsters macrophage antimicrobial functions while attenuating IL-6 production.

Intestinal microbiota contributes to host defense against pathogens while avoiding the induction of inflammation in homeostatic conditions, but the mechanism is not fully understood. To investigate the potential role of the bacterial metabolite desaminotyrosine (DAT) in regulating host defense and inflammation, we pretreated mouse bone marrow-derived macrophages (BMDMs) with DAT for 12 hours and then challenged with bacterial lipopolysaccharide (LPS). We found that DAT priming-enhanced type I interferon response while selectively inhibiting proinflammatory interleukin (IL)-6 production after exposure to LPS. This is related to the fact that DAT is a natural antioxidant determined by radical scavenging assay in a cell-free system. DAT-primed cells had increased levels of the reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) upon LPS stimulation. Countering the increased NADPH by supplementing extra oxidized NADP+ to cells reversed DAT's effect on LPS-induced Il-6 and interferon-stimulated gene expressions. DAT-primed cells also were more resistant to oxidative stress-induced generation of reactive oxygen species and cell death. DAT promoted the production of antimicrobial effector nitric oxide in a cellular redox-dependent manner, leading to enhanced macrophage antimicrobial activity during Salmonella enterica infection. Our data suggest that DAT acts as a host-microbiota crosstalk signal in shaping host immune defense and inflammatory response.

Galectin-3 inhibition alleviated LPS-induced periodontal inflammation in gingival fibroblasts and experimental periodontitis mice.

Clinical studies have confirmed that galectin-3 (Gal-3) levels are significantly elevated in periodontitis patients. The present study aimed to explore the effects of Gal-3 inhibition on periodontal inflammation in vitro and in vivo. Human gingival fibroblasts (HGFs) with or without Gal-3 knockdown were stimulated by lipopolysaccharide (LPS), and a ligation-induced mouse periodontitis model treated with a Gal-3 inhibitor was established. Hematoxylin-eosin (H&E) and immunohistochemistry (IHC) staining were used to evaluate Gal-3 levels in gingival tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used to detect Gal-3, interleukin (IL)-6, IL-8, and C-C motif ligand 2 (CCL2) expression. Immunofluorescence and western blotting were used to detect NF-κB and ERK signaling pathway activation. Micro-computed tomography was used to analyse the degree of bone loss. Gal-3 was significantly up-regulated in inflamed gingival tissues and LPS-induced HGFs. Gal-3 knockdown markedly decreased LPS-induced IL-6, IL-8, and CCL2 expression and blocked NF-κB and ERK signaling pathway activation in HGFs. In the mouse periodontitis model, Gal-3 inhibition significantly alleviated IL-1β and IL-6 infiltration in gingival tissue and mitigated periodontal bone loss. Gal-3 inhibition notably alleviated periodontal inflammation partly through blocking NF-κB and ERK signaling pathway activation.

IL-1 and TNF mediates IL-6 signaling at the maternal-fetal interface during intrauterine inflammation.

IL6 signaling plays an important role in triggering labor and IL6 is an established biomarker of intrauterine infection/inflammation (IUI) driven preterm labor (PTL). The biology of IL6 during IUI at the maternal-fetal interface was investigated in samples from human subjects and non-human primates (NHP). Pregnant women with histologic chorioamnionitis diagnosed by placenta histology were recruited (n=28 term, n=43 for preterm pregnancies from 26-36 completed weeks of gestation). IUI was induced in Rhesus macaque by intraamniotic injection of lipopolysachharide (LPS, n=23). IL1 signaling was blocked using Anakinra (human IL-1 receptor antagonist, n=13), and Tumor necrosis factor (TNF) signaling was blocked by anti TNF-antibody (Adalimumab n=14). The blockers were given before LPS. All animals including controls (intraamniotic injection of saline n=27), were delivered 16h after LPS/saline exposure at about 80% gestation. IUI induced a robust expression of IL6 mRNAs in the fetal membranes (chorion-amnion-decidua tissue) both in humans (term and preterm) and NHP. The major sources of IL6 mRNA expression were the amnion mesenchymal cells (AMC) and decidua stroma cells. Additionally, during IUI in the NHP, ADAM17 (a protease that cleaves membrane bound IL6 receptor (IL6R) to release a soluble form) and IL6R mRNA increased in the fetal membranes, and the ratio of IL6 and soluble forms of IL6R, gp130 increased in the amniotic fluid signifying upregulation of IL6 trans-signaling. Both IL1 and TNF blockade suppressed LPS-induced IL6 mRNAs in the AMC and variably decreased elements of IL6 trans-signaling. These data suggest that IL1 and TNF blockers may be useful anti-inflammatory agents via suppression of IL6 signaling at the maternal-fetal interface.

Ursodeoxycholic Acid Modulates the Interaction of miR-21 and Farnesoid X Receptor and NF-κB Signaling.

(1) Background: This study investigates the effects of Ursodeoxycholic acid (UDCA) on NF-κB signaling, farnesoid X receptor (FXR) singling, and microRNA-21 in HepG2 cells. (2) Methods: HepG2 cells were treated with lipopolysaccharide (LPS) to simulate hepatic inflammation. The investigation focused on the expression of NF-κB activation, which was analyzed using Western blot, confocal microscopy, and Electrophoretic Mobility-shift Assays (EMSA). Additionally, NF-κB and farnesoid X receptor (FXR) singling expressions of micro-RNA-21, COX-2, TNF-α, IL-6, cyp7A1, and shp were assessed by RT-PCR. (3) Results: UDCA effectively downregulated LPS-induced expressions of NF-κB/65, p65 phosphorylation, and also downregulated FXR activity by Western blot. Confocal microscopy and EMSA results confirmed UDCA's role in modulating NF-κB signaling. UDCA reduced the expressions of LPS-induced COX-2, TNF-α, and IL-6, which were related to NF-κB signaling. UDCA downregulated LPS-induced cyp7A1 gene expression and upregulated shp gene expression, demonstrating selective gene regulation via FXR. UDCA also significantly decreased micro-RNA 21 levels. (4) Conclusions: This study demonstrates UDCA's potent anti-inflammatory effects on NF-κB and FXR signaling pathways, and thus its potential to modulate hepatic inflammation and carcinogenesis through interactions with NF-κB and FXR. The decrease in micro-RNA 21 expression further underscores its therapeutic potential.

Xiaowugui decoction alleviates experimental rheumatoid arthritis by suppressing Rab5a-mediated TLR4 internalization in macrophages

BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by exacerbated synovial inflammation and joint destruction. Recent studies suggest toll-like receptor 4 (TLR4) internalization facilitate inflammatory response of macrophage. The role of TLR4 internalization in the pathogenesis of RA is unknown. PurposeTo investigate the role and mechanism of TLR4 internalization in macrophage inflammatory response of RA and explore whether TLR4 internalization mediates the anti-arthritic effect of Xiaowugui (XWG) decoction, a patented herbal formula used in China. MethodsThe co-expression of TLR4 and the internalization marker, early endosome antigen 1 (EEA1), in the synovial samples of RA patients and joint tissue of collagen-induced arthritis (CIA) mice, were evaluated using immunofluorescence. The effect of Rab5a-mediated early internalization of TLR4 on the activation induced by lipopolysaccharide (LPS) in RAW264.7 cells was investigated using small interfering RNAs that act against Rab5a. CIA was induced in Rab5a−/− mice to evaluate the role of Rab5a in vivo. The disease progression and expression of Rab5a and TLR4 in the joint tissue were evaluated in CIA mice treated with XWG. Inflammatory factors production, TLR4 internalization, and activation of downstream signaling pathways were examined in RAW264.7 cells treated with XWG in vitro. ResultsThe co-expression and co-localization of TLR4 and EEA1 were elevated in the synovial samples of RA patients and joint tissue of CIA mice. Pharmaceutical inhibition of TLR4 internalization reduced macrophages inflammatory responses induced by LPS. The co-expression and co-localization of Rab5a and TLR4 were significantly increased in macrophages treated with LPS. Silencing Rab5a reduced LPS-induced TLR4 internalization, inflammatory factors production, and phosphorylation of Jun N-terminal kinases (JNK) and p65. Genetic deletion of Rab5a inhibited TLR4 internalization and the development of arthritis in vivo. The co-expression of TLR4 and Rab5a was also elevated in the synovial samples of RA patients. XWG treatment of mice with CIA alleviated arthritis and reduced the co-expression of Rab5a and TLR4 in the joint tissue. XWG treatment of macrophage inhibited LPS-induced IL-6 and TNF-α production, co-expression of Rab5a and TLR4, and phosphorylation of JNK and p65. ConclusionsOur findings highlight the pathogenic role of TLR4 internalization in patients with RA and identify a novel Rab5a-dependent internalization pathway that promotes macrophage inflammatory response. XWG treatment demonstrated outstanding therapeutic effects in experimental arthritis, and targeting the Rab5a-mediated internalization of TLR4 may be the main underlying mechanism.

Antimicrobial Peptide Reduces Cytotoxicity and Inflammation in Canine Epidermal Keratinocyte Progenitor Cells Induced by Pseudomonas aeruginosa Infection.

The direct effects and antimicrobial activity of synthetic antimicrobial peptides (AMPs) obtained from dogs, including cBD, cBD103, and cCath, against P. aeruginosa wild-type strain PAO1 and canine keratinocytes were analyzed. Antibacterial effects on planktonic bacteria were assessed by determining the minimum bactericidal concentrations (MBCs) of AMPs and by a time-kill assay. Antibiofilm effects were assessed using the microtiter plate assay. We also evaluated the effects of AMPs on cell cytotoxicity and host immune response induced by stimulating canine epidermal keratinocyte progenitor (CPEK) cells with PAO1 and its LPS. cBD, cBD103, and cCath all exhibited dose-dependent antimicrobial and antibiofilm effects. In particular, 25 μg/mL cBD103 showed rapid bactericidal activity within 60 min and inhibited biofilm formation. In addition, pretreatment with cBD103 (25 µg/mL) and cCath (50 µg/mL) 1 h before stimulation significantly reduced the cytotoxicity of the CPEK cells by PAO1 and LPS-induced IL-6 and TNF-a expressions. cBD had little effect on the response to PAO1 and LPS in the cells. These results indicate the therapeutic potential of AMPs in P. aeruginosa skin infections. However, further studies on the mechanism of action of AMPs in keratinocytes and clinical trials are needed.

Baicalin suppresses macrophage JNK-mediated adipose tissue inflammation to mitigate insulin resistance in obesity

Ethnopharmacological relevanceRadix scutellariae (the root of Scutellaria baicalensis Georgi) is a traditional Chinese medicine (TCM) used to treat a wide range of inflammation-related diseases, such as obesity, diabetes, diabetic kidney disease, and COVID-19-associated inflammatory states in the lung and kidney. Baicalin is the major anti-inflammatory component of Radix scutellariae and has shown the potential to inhibit inflammation in metabolic disorders. In this study, we explored the ability and underlying mechanisms of baicalin to modulate the macrophage to mitigate insulin resistance in obesity. Materials and methodsObese mice were administered baicalin (50 mg/kg/day) intraperitoneally for 3 weeks. RAW264.7 and BMDM cells were stimulated with LPS and treated with baicalin for 24 h, while 3T3-L1 and primary white adipocytes were treated with the supernatants from baicalin-treated RAW264.7 cells for 24 h. ResultsThe results showed that baicalin significantly improved glucose and insulin tolerance as well as decreased fat and adipose tissue macrophage levels in obese mice. Besides, baicalin significantly reduced serum and adipose tissue IL-1β, TNF-α and IL-6 levels in obese mice, as well as suppressed LPS-induced IL-1β, TNF-α and IL-6 expression and release in macrophages. Furthermore, treatment with the supernatant from baicalin-treated RAW264.7 cells increased the levels of PGC-1α, SIRT1, p-IRS-1 and p-AKT in adipocytes. Moreover, baicalin treatment dramatically downregulated macrophage p-p38, p-JNK, and Ac-p65Lys310 levels while increasing SIRT1 both in vivo and in vitro. Importantly, JNK inhibitor SP600125 blocked most of the effects of baicalin on SIRT1, Ac-p65Lys310 and pro-inflammatory factors in macrophages. ConclusionTherefore, these results demonstrated for the first time that baicalin exerts its anti-inflammatory effects in obese adipose tissue macrophages mainly through suppressing JNK/SIRT1/p65 signaling. These findings amplified the mechanisms of baicalin and its potential to attenuate insulin resistance.

TRPV1 Regulates Proinflammatory Properties of M1 Macrophages in Periodontitis Via NRF2.

Periodontitis, characterized by progressive alveolar bone destruction, leads to the loss of attachment and stability of the affected teeth. Macrophages, especially the proinflammatory M1 subtype, are key in periodontitis pathogenesis, driving the disease's inflammatory and destructive processes. Despite existing insight into their involvement, comprehensive understanding of the underlying molecular mechanisms remains limited. TRPV1 is a non-selective cation channel protein and is known to regulate cellular function and homeostasis in macrophages. Our research objective was to investigate the impact of TRPV1 on the proinflammatory attributes of M1 macrophages in periodontal tissues, exploring potential mechanistic pathways. A mouse model of periodontitis was established using Porphyromonas gingivalis inoculation and ligature application around the maxillary second molar. Immunohistological analysis showed a significant reduction in macrophage TRPV1 expression in periodontitis-induced mice. Treatment with capsaicin, a TRPV1 agonist, was observed to effectively elevate TRPV1 expression in these macrophages. Furthermore, micro-computed tomography analysis revealed a marked decrease in alveolar bone resorption in the capsaicin -treated group, compared with vehicle and healthy control groups. Our in vitro findings show that capsaicin treatment successfully attenuated LPS-induced TNF-α and IL-6 production in macrophages, mediated through NRF2 activation, consequently reducing intracellular ROS levels. These findings suggest that TRPV1 agonists, through modulating M1 macrophage activity and up-regulating TRPV1, could be a novel therapeutic approach in periodontal disease management.

Obesity-associated inflammatory macrophage polarization is inhibited by capsaicin and phytolignans.

Obesity is often accompanied by increased adipose tissue inflammation, a process that is partially driven by adipose tissue-resident macrophages. In this study, we explored the potential for plant-derived dietary compounds to exert anti-inflammatory effects in macrophages that alleviate obesity-associated adipocyte dysfunction. Capsaicin (CAP), schisandrin A (SA), enterodiol (END), and enterolactone (ENL) treatment polarized J774 macrophages to an "M2" or anti-inflammatory phenotype and inhibited responses to stimulation with lipopolysaccharide (LPS). Furthermore, these compounds blocked inflammasome activation when administered just before ATP-induced NLRP3 activation, as evidenced by the abrogation of IL-1β release in mouse macrophages and human peripheral blood monocytes. The addition of CAP, SA, or ENL during the differentiation of bone marrow-derived macrophages was also sufficient to inhibit LPS-induced IL-6 and TNFα production. Finally, CAP, END, and ENL treatment during differentiation of 3T3-L1 adipocytes induced an adiponectin-high phenotype accompanied by increases in thermogenic gene expression, and conditioned media from these adipocytes inhibited LPS-induced production of IL-1β, IL-6, and TNFα from J774 macrophages. These polarizing effects were partially mediated by the elevated adiponectin and decreased syndecan-4 in the adipocyte-conditioned media. These results implicate the contribution of plant-derived dietary components to the modulation of macrophages and adipocytes in obesity.NEW & NOTEWORTHY The utility of food-based products to prevent or alleviate chronic conditions such as obesity and its associated comorbidities is an attractive approach. Capsaicin, schisandrin A, enterodiol, and enterolactone, phytochemicals present in traditional medicinal food, decreased proinflammatory cytokine production from macrophages that, in turn, reduced obesity-associated adipocyte dysfunction. These results implicate the contribution of plant-derived dietary components to the modulation of macrophages and adipocytes in obesity.

The gut lactic acid bacteria metabolite, 10-oxo-cis-6,trans-11-octadecadienoic acid, suppresses inflammatory bowel disease in mice by modulating the NRF2 pathway and GPCR-signaling.

Various gut bacteria, including Lactobacillus plantarum, possess several enzymes that produce hydroxy fatty acids (FAs), oxo FAs, conjugated FAs, and partially saturated FAs from polyunsaturated FAs as secondary metabolites. Among these derivatives, we identified 10-oxo-cis-6,trans-11-octadecadienoic acid (γKetoC), a γ-linolenic acid (GLA)-derived enon FA, as the most effective immunomodulator, which inhibited the antigen-induced immunoactivation and LPS-induced production of inflammatory cytokines. The treatment with γKetoC significantly suppressed proliferation of CD4+ T cells, LPS-induced activation of bone marrow-derived dendritic cells (BMDCs), and LPS-induced IL-6 release from peritoneal cells, splenocytes, and CD11c+ cells isolated from the spleen. γKetoC also inhibited the release of inflammatory cytokines from BMDCs stimulated with poly-I:C, R-848, or CpG. Further in vitro experiments using an agonist of GPR40/120 suggested the involvement of these GPCRs in the effects of γKetoC on DCs. We also found that γKetoC stimulated the NRF2 pathway in DCs, and the suppressive effects of γKetoC and agonist of GPR40/120 on the release of IL-6 and IL-12 were reduced in Nrf2-/- BMDCs. We evaluated the role of NRF2 in the anti-inflammatory effects of γKetoC in a dextran sodium sulfate-induced colitis model. The oral administration of γKetoC significantly reduced body weight loss, improved stool scores, and attenuated atrophy of the colon, in wild-type C57BL/6 and Nrf2+/- mice with colitis. In contrast, the pathology of colitis was deteriorated in Nrf2-/- mice even with the administration of γKetoC. Collectively, the present results demonstrated the involvement of the NRF2 pathway and GPCRs in γKetoC-mediated anti-inflammatory responses.

Essential phospholipids impact cytokine secretion and alter lipid-metabolizing enzymes in human hepatocyte cell lines

BackgroundEssential phospholipids (EPL) are hepatoprotective.MethodsThe effects on interleukin (IL)-6 and -8 secretion and on certain lipid-metabolizing enzymes of non-cytotoxic concentrations of EPL (0.1 and 0.25 mg/ml), polyenylphosphatidylcholine (PPC), and phosphatidylinositol (PtdIns) (both at 0.1 and 1 mg/ml), compared with untreated controls, were assessed in human hepatocyte cell lines (HepG2, HepaRG, and steatotic HepaRG).ResultsLipopolysaccharide (LPS)-induced IL-6 secretion was significantly decreased in HepaRG cells by most phospholipids, and significantly increased in steatotic HepaRG cells with at least one concentration of EPL and PtdIns. LPS-induced IL-8 secretion was significantly increased in HepaRG and steatotic HepaRG cells with all phospholipids. All phospholipids significantly decreased amounts of fatty acid synthase in steatotic HepaRG cells and the amounts of acyl-CoA oxidase in HepaRG cells. Amounts of lecithin cholesterol acyltransferase were significantly decreased in HepG2 and HepaRG cells by most phospholipids, and significantly increased with 0.1 mg/ml PPC (HepaRG cells) and 1 mg/ml PtdIns (steatotic HepaRG cells). Glucose-6-phosphate dehydrogenase activity was unaffected by any phospholipid in any cell line.ConclusionsEPL, PPC, and PtdIns impacted the secretion of pro-inflammatory cytokines and affected amounts of several key lipid-metabolizing enzymes in human hepatocyte cell lines. Such changes may help liver function improvement, and provide further insights into the EPL’s mechanism of action.

Targeting transitioning lung monocytes/macrophages as treatment strategies in lung disease related to environmental exposures

BackgroundEnvironmental/occupational exposures cause significant lung diseases. Agricultural organic dust extracts (ODE) and bacterial component lipopolysaccharide (LPS) induce recruited, transitioning murine lung monocytes/macrophages, yet their cellular role remains unclear.MethodsCCR2 RFP+ mice were intratracheally instilled with high concentration ODE (25%), LPS (10 μg), or gram-positive peptidoglycan (PGN, 100 μg) for monocyte/macrophage cell-trafficking studies. CCR2 knockout (KO) mice and administration of intravenous clodronate liposomes strategies were employed to reduce circulating monocytes available for lung recruitment following LPS exposure. Lung tissues and bronchoalveolar lavage fluid (BALF) were collected. Pro-inflammatory and/or pro-fibrotic cytokines, chemokines, and lung extracellular matrix mediators were quantitated by ELISA. Infiltrating lung cells including monocyte/macrophage subpopulations, neutrophils, and lymphocytes were characterized by flow cytometry. Lung histopathology, collagen content, vimentin, and post-translational protein citrullination and malondialdehyde acetaldehyde (MAA) modification were quantitated. Parametric statistical tests (one-way ANOVA, Tukey’smultiple comparison) and nonparametric statistical (Kruskal–Wallis, Dunn’s multiple comparison) tests were used following Shapiro–Wilk testing for normality.ResultsIntratracheal instillation of ODE, LPS, or PGN robustly induced the recruitment of inflammatory CCR2+ CD11cintCD11bhi monocytes/macrophages and both CCR2+ and CCR2− CD11c−CD11bhi monocytes at 48 h. There were also increases in CCR2+ CD4+ and CD8+ T cells and NK cells. Despite reductions in LPS-induced lung infiltrating CD11cintCD11bhi cells (54% reduction), CCR2 knockout (KO) mice were not protected against LPS-induced inflammatory and pro-fibrotic consequences. Instead, compensatory increases in lung neutrophils and CCL2 and CCL7 release occurred. In contrast, the depletion of circulating monocytes through the administration of intravenous clodronate (vs. vehicle) liposomes 24 h prior to LPS exposure reduced LPS-induced infiltrating CD11cintCD11bhi monocyte-macrophage subpopulation by 59% without compensatory changes in other cell populations. Clodronate liposome pre-treatment significantly reduced LPS-induced IL-6 (66% reduction), matrix metalloproteinases (MMP)-3 (36%), MMP-8 (57%), tissue inhibitor of metalloproteinases (61%), fibronectin (38%), collagen content (22%), and vimentin (40%). LPS-induced lung protein citrullination and MAA modification, post-translational modifications implicated in lung disease, were reduced (39% and 48%) with clodronate vs. vehicle liposome.ConclusionHighly concentrated environmental/occupational exposures induced the recruitment of CCR2+ and CCR2− transitioning monocyte-macrophage and monocyte subpopulations and targeting peripheral monocytes may reduce the adverse lung consequences resulting from exposures to LPS-enriched inhalants.

Four new diterpenoids from the aerial parts of Leucas zeylanica (L.) R. Br.

Four new undescribed halimane- and labdane-type diterpenoids, named zeylleucapenoids E–H (1–4), along with four known analogues (5–8), were isolated from the aerial parts of Leucas zeylanica (L.) R. Br. Their structures were determined by comprehensive spectroscopic analysis and computational calculations. Compounds 1 and 2 are the highly modified halimane diterpenoids featuring a 6/6/6-fused tricyclic system with an unusual six-membered 6,11-ether ring. Compound 8 exhibits nontoxic effects for zebrafish embryo, while it displays efficient reduction against NO production in a dose-dependent manner and strongly suppresses the secretion of LPS-induced TNF-α and IL-6 cytokines in RAW264.7 macrophages. In addition, marked reductions of iNOS and COX-2 expression were observed. Molecular docking analysis indicated that 8 has high affinities with the target amino acid residues on protein-binding sites, which may be a possible mechanism contributing to the anti-inflammatory potential of this molecule.

Bruton's tyrosine kinase inhibition limits endotoxic shock by suppressing IL-6 production by marginal zone B cells in mice.

Sepsis is a systemic inflammatory response to a severe, life-threatening infection with organ dysfunction. Although there is no effective treatment for this fatal illness, a deeper understanding of the pathophysiological basis of sepsis and its underlying mechanisms could lead to the development of new treatment approaches. Here, we demonstrate that the selective Bruton's tyrosine kinase (Btk) inhibitor acalabrutinib augments survival rates in a lipopolysaccharide (LPS)-induced septic model. Our in vitro and in vivo findings both indicate that acalabrutinib reduces IL-6 production specifically in marginal zone B (MZ B) cells rather than in macrophages. Furthermore, Btk-deficient MZ B cells exhibited suppressed LPS-induced IL-6 production in vitro. Nuclear factor-kappa B (NF-κB) signaling, which is the downstream signaling cascade of Toll-like receptor 4 (TLR4), was also severely attenuated in Btk-deficient MZ B cells. These findings suggest that Btk blockade may prevent sepsis by inhibiting IL-6 production in MZ B cells. In addition, although Btk inhibition may adversely affect B cell maturation and humoral immunity, antibody responses were not impaired when acalabrutinib was administered for a short period after immunization with T-cell-independent (TI) and T-cell-dependent (TD) antigens. In contrast, long-term administration of acalabrutinib slightly impaired humoral immunity. Therefore, these findings suggest that Btk inhibitors may be a potential option for alleviating endotoxic shock without compromising humoral immunity and emphasize the importance of maintaining a delicate balance between immunomodulation and inflammation suppression.

Anti-inflammatory Polyketides from an Endophytic Fungus Chaetomium sp. UJN-EF006 of Vaccinium bracteatum.

Seven new polyketides including three chromone derivatives (1-3) and four linear ones incorporating a tetrahydrofuran ring (4-7), along with three known compounds (8-10), were obtained from the fermentation of an endophytic fungus (Chaetomium sp. UJN-EF006) isolated from the leaves of Vaccinium bracteatum. The structures of these fungal metabolites have been elucidated by spectroscopic means including MS, NMR and electronic circular dichroism. A preliminary anti-inflammatory screening with the lipopolysaccharide (LPS) induced RAW264.7 cell model revealed moderate NO production inhibitory activity for compounds 1 and 4. In addition, the expression of three LPS-induced inflammatory factors IL-6, iNOS and COX-2 was also blocked by 1 and 4.

Identification of a 10-mer peptide from the death domain of MyD88 which attenuates inflammation and insulin resistance and improves glucose metabolism.

Insulin resistance (IR) is the key pathophysiological cause of type 2 diabetes, and inflammation has been implicated in it. The death domain (DD) of the adaptor protein, MyD88 plays a crucial role in the transduction of TLR4-associated inflammatory signal. Herein, we have identified a 10-residue peptide (M10), from the DD of MyD88 which seems to be involved in Myddosome formation. We hypothesized that M10 could inhibit MyD88-dependent TLR4-signaling and might have effects on inflammation-associated IR. Intriguingly, 10-mer M10 showed oligomeric nature and reversible self-assembly property indicating the peptide's ability to recognize its own amino acid sequence. M10 inhibited LPS-induced nuclear translocation of NF-κB in L6 myotubes and also reduced LPS-induced IL-6 and TNF-α production in peritoneal macrophages of BALB/c mice. Remarkably, M10 inhibited IL-6 and TNF-α secretion in diabetic, db/db mice. Notably, M10 abrogated IR in insulin-resistant L6 myotubes, which was associated with an increase in glucose uptake and a decrease in Ser307-phosphorylation of IRS1, TNF-α-induced JNK activation and nuclear translocation of NF-κB in these cells. Alternate day dosing with M10 (10 and 20 mg/kg) for 30 days in db/db mice significantly lowered blood glucose and improved glucose intolerance after loading, 3.0 g/kg glucose orally. Furthermore, M10 increased insulin and adiponectin secretion in db/db mice. M10-induced glucose uptake in L6 myotubes involved the activation of PI3K/AKT/GLUT4 pathways. A scrambled M10-analog was mostly inactive. Overall, the results show the identification of a 10-mer peptide from the DD of MyD88 with anti-inflammatory and anti-diabetic properties, suggesting that targeting of TLR4-inflammatory pathway, could lead to the discovery of molecules against IR and diabetes.