Tumor Necrosis Factor-alpha Secretion Research Articles

723 High glucose environment induces macrophage M1 polarization that contributes to impaired wounding via reducing keratinocyte migration

Diabetes is an important global health issue due to its increasing prevalence and association with serious co-morbidities including impaired wound healing. A hallmark of impaired diabetic healing is suboptimal re-epithelialization, an event that is regulated by infiltrating macrophages. The present study aimed to explore the effects of high glucose environment on macrophage polarization and explore the resulting effect on keratinocyte function using relevant cell and animal models. Our results showed that high glucose environment polarized cultured human macrophages toward classical M1 phenotype as demonstrated by significant upregulation of CCR7 expression. Additionally, condition media from the M1 macrophages significantly decreased human keratinocyte migration via reducing the metalloproteinase 1 (MMP1) expression in keratinocytes by more than 50%. Highly increased tumor necrosis factor-alpha (TNF-a) secretion from the M1 macrophages was responsible for inhibiting MMP1 expression in keratinocytes. These in vitro results were validated in vivo using a diabetic animal model. More specifically, the wound healing was delayed in diabetic rats with prolonged infiltration of M1 macrophages, elevated TNF-a expression in the wound area, and reduced MMP1 expression in the perilesional skin. In summary, our results demonstrated that high glucose environment induced macrophage M1 polarization, an event that leads to TNF-a mediated MMP1 suppression in keratinocytes and results in impaired keratinocyte migration. Restoring the macrophage phenotype and neutralizing elevated TNF-a in the diabetic wound environment are potential strategies to promote better diabetic wound healing in the near future.

Salidroside suppresses solar ultraviolet-induced skin inflammation by targeting cyclooxygenase-2.

Solar ultraviolet (SUV) irradiation causes skin disorders such as inflammation, photoaging, and carcinogenesis. Cyclooxygenase-2 (COX-2) plays a key role in SUV-induced skin inflammation, and targeting COX-2 may be a strategy to prevent skin disorders. In this study, we found that the expression of COX-2, phosphorylation of p38 or JNKs were increased in human solar dermatitis tissues and SUV-irradiated human skin keratinocyte HaCaT cells and mouse epidermal JB6 Cl41 cells. Knocking down COX-2 inhibited the production of prostaglandin E2 (PGE2), the phosphorylation of p38 or JNKs in SUV-irradiated cells, which indicated that COX-2 is not only the key enzyme for PGs synthesis, but also an upstream regulator of p38 or JNKs after SUV irradiation. The virtual ligand screening assay was used to search for natural drugs in the Chinese Medicine Database, and indicated that salidroside might be a COX-2 inhibitor. Molecule modeling indicated that salidroside can directly bind with COX-2, which was proved by in vitro pull-down binding assay. Ex vivo studies showed that salidroside has no toxicity to cells, and inhibits the production of PGE2, phosphorylation of p38 or JNKs, and secretion of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) caused by SUV irradiation. In vivo studies demonstrated that salidroside attenuates the skin inflammation induced by SUV. In brief, our data provided the evidences for the protective role of salidroside against SUV-induced inflammation by targeting COX-2, and salidroside might be a promising drug for the treatment of SUV-induced skin inflammation.

Role of testosterone in regulating induction of TNF-α in rat spleen via ERK signaling pathway

Spleen is a pivotal organ for regulating immune homeostasis. It has been shown that testosterone diminishes secretion of various inflammatory molecules under multiple conditions. However, the mechanisms of action of endogenous testosterone affecting immune responses in the spleen remain unknown. The aim of the present study was to evaluate the immune functions of the spleen in response to testosterone withdrawal after orchidectomy, and the impact of splenocytes on the bacterial endotoxin lipopolysaccharide (LPS)-induced secretion of inflammatory molecules. Male rats were divided into 3 groups, i.e. intact, orchidectomized (Orch) and orchidectomized plus replacement of testosterone propionate (TP) (Orch+TP). The Orch and Orch+TP rats underwent bilateral orchidectomy one week before TP replacement (2mg/kg body weight) or sesame oil in intact rats as controls for seven days. Orch resulted in a significant increase of spleen weight and basal secretion of nitric oxide (NO) from splenocytes. Additionally, LPS up-regulated cell proliferation and the secretion of tumor necrosis factor-alpha (TNF-α) in splenocytes of Orch rats. Orch further up-regulated phosphorylation of extracellular signal-regulated kinases. Interestingly, the plasma corticosterone concentration in the Orch group was higher than that in the intact and Orch+TP groups. Deficiency of testosterone-elevated TNF-α and NO secretion in response to LPS were confirmed in the rat splenocytes. Testosterone also significantly attenuated LPS-elicited release of TNF-α and NO in a dose-dependent manner. However, testosterone did not suppress splenic blastogenesis at doses in the 10−10–10−7M concentration range. In this context, testosterone might have a protective role against inflammatory responses in the spleen. The present study provides evidence to indicate that testosterone might modulate the immune system.

The Secreted Protein Rv1860 of Mycobacterium tuberculosis Stimulates Human Polyfunctional CD8+ T Cells.

We previously reported that Rv1860 protein from Mycobacterium tuberculosis stimulated CD4(+)and CD8(+)T cells secreting gamma interferon (IFN-γ) in healthy purified protein derivative (PPD)-positive individuals and protected guinea pigs immunized with a DNA vaccine and a recombinant poxvirus expressing Rv1860 from a challenge with virulent M. tuberculosis We now show Rv1860-specific polyfunctional T (PFT) cell responses in the blood of healthy latently M. tuberculosis-infected individuals dominated by CD8(+) T cells, using a panel of 32 overlapping peptides spanning the length of Rv1860. Multiple subsets of CD8(+) PFT cells were significantly more numerous in healthy latently infected volunteers (HV) than in tuberculosis (TB) patients (PAT). The responses of peripheral blood mononuclear cells (PBMC) from PAT to the peptides of Rv1860 were dominated by tumor necrosis factor alpha (TNF-α) and interleukin-10 (IL-10) secretions, the former coming predominantly from non-T cell sources. Notably, the pattern of the T cell response to Rv1860 was distinctly different from those of the widely studied M. tuberculosis antigens ESAT-6, CFP-10, Ag85A, and Ag85B, which elicited CD4(+) T cell-dominated responses as previously reported in other cohorts. We further identified a peptide spanning amino acids 21 to 39 of the Rv1860 protein with the potential to distinguish latent TB infection from disease due to its ability to stimulate differential cytokine signatures in HV and PAT. We suggest that a TB vaccine carrying these and other CD8(+) T-cell-stimulating antigens has the potential to prevent progression of latent M. tuberculosis infection to TB disease.

Polysaccharides from raw and cooked chayote modulate macrophage function

Chayote (Sechium edule) pulp is consumed raw or cooked and is source of polysaccharides with potential immunomodulatory properties in macrophages, which are cells that play important roles in homeostasis and innate immune responses to pathogens; however, these immunomodulatory effects of chayote polysaccharides remain unknown. Therefore, polysaccharides from raw (SeR) and cooked (SeC) chayote and from the hot water extract obtained after heating (SeH) were analyzed for their composition and effects on macrophages. Chayote has a high- (340kDa) and a low- (46kDa) molecular weight (MW) polysaccharide fraction containing high amounts of galactose, arabinose and galacturonic acid. After cooking, SeC was enriched in high-MW galactose-rich fractions, whereas SeH was enriched in low-MW arabinans and homogalaturonans-rich fractions. The polysaccharides induced tumor necrosis factor alpha (TNF-α) and nitric oxide (NO) secretion in macrophages; however, only SeR and SeH reduced TNF-α, NO and interleukin-6 secretion in activated macrophages. Furthermore, SeR and SeCW, but not SeC, affected phagocytic activity. In conclusion, bioactive polysaccharides in chayote modulate macrophage functions, and minor changes in composition resulting from the solubilization of a small proportion of low-MW arabinans and homogalacturonans during cooking accounted for the different effects of raw and cooked chayote.

Effects of selected bioactive food compounds on human white adipocyte function.

BackgroundPrevious studies suggest that intake of specific bioactive compounds may have beneficial clinical effects on adipose tissue partly due to their anti-inflammatory and insulin-sensitizing properties. With the overall aim to contribute to better understanding of the mechanisms of selected bioactive nutrients on fat metabolism, we investigated their role on human white adipocyte function.MethodsThe influence of the omega-3-fatty acid docosahexaenoic acid (DHA), the anthocyanin (AC) cyanidin-3-glucoside and its metabolite protocatechuic acid, and the beta-glucan metabolite propionic acid (PI) on adipokine secretion, fatty acid metabolism (lipolysis/lipogenesis) and adipocyte differentiation (lipid accumulation) was studied in human fat cells differentiated in vitro. To investigate possible synergistic, additive or antagonistic effects, DHA was also combined with AC or PI.ResultsEach compound, alone or together with DHA, suppressed basal adipocyte lipolysis compared to control treated cells. DHA alone attenuated the secretion of pro-inflammatory adipokines such as chemerin, interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1/CCL2), whereas AC suppressed only the latter two. Treatment with PI decreased IL-6, tumour necrosis factor alpha (TNFα) and adiponectin secretion. A combination of DHA and AC decreased TNFα secretion and increased insulin-stimulated lipogenesis. No effect was found on adipocyte differentiation. At the selected concentrations, none of the compounds was found to be cytotoxic.ConclusionThe studied bioactive food compounds or their metabolites have beneficial effects in human primary fat cells measured as decreased basal lipolytic activity and secretion of inflammatory markers. A minor effect was also observed on insulin-stimulated glucose uptake albeit only with the combination of DHA and AC. Taken together, our results may link the reported health benefits of the selected bioactives on metabolic disorders such as insulin resistance, hypertension and dyslipidemia to effects on white adipocytes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12986-016-0064-3) contains supplementary material, which is available to authorized users.

Diet-induced obesity reduces the production of influenza vaccine-induced antibodies via impaired macrophage function.

Obesity is a metabolic disease characterized by low-level chronic inflammation. Obese individuals are susceptible to infection by viruses, and vaccination against these pathogens is less effective than in nonobese individuals. Here, we sought to explore the immunological environment in a mouse model of obesity induced by a high-fat diet (HFD). HFD treatment increased the body weight and epididymal fat mass. The proportion of activated B cells, T cells, and macrophages was similar between mice in the HFD group and the regular-fat diet (RFD) group. The Th1 cell subpopulation in the HFD group was increased, whereas the proportion of Treg cells was reduced compared with the RFD group. Moreover, T-cell proliferation and cytokine production did not differ between the groups when cells were stimulated with anti-CD3 and anti-CD28 antibodies in vitro. In macrophages, phagocytic activity was higher in mice fed an HFD than in those fed an RFD, but expression levels of CD86 and MHC class II antigens were similar. When macrophages were cultured in vitro, the proportion of CD86-expressing macrophages was lower in those isolated from mice in the HFD group than in those isolated from the RFD group. Furthermore, lipopolysaccharide-induced interleukin 6 (IL-6) and tumor necrosis factor alpha secretions were significantly reduced in macrophages isolated from the HFD group. In addition, influenza vaccine-induced antibodies in the HFD group diminished more rapidly than in the RFD group. These results suggest that poor functionality of macrophages during obesity might contribute to a reduction in vaccine efficacy.

Serrulatane Diterpenoid from Eremophila neglecta Exhibits Bacterial Biofilm Dispersion and Inhibits Release of Pro-inflammatory Cytokines from Activated Macrophages.

The purpose of this study was to assess the biofilm-removing efficacy and inflammatory activity of a serrulatane diterpenoid, 8-hydroxyserrulat-14-en-19-oic acid (1), isolated from the Australian medicinal plant Eremophila neglecta. Biofilm breakup activity of compound 1 on established Staphylococcus epidermidis and Staphylococcus aureus biofilms was compared to the antiseptic chlorhexidine and antibiotic levofloxacin. In a time-course study, 1 was deposited onto polypropylene mesh to mimic a wound dressing and tested for biofilm removal. The ex-vivo cytotoxicity and effect on lipopolysaccharide-induced pro-inflammatory cytokine release were studied in mouse primary bone-marrow-derived macrophage (BMDM) cells. Compound 1 was effective in dispersing 12 h pre-established biofilms with a 7 log10 reduction of viable bacterial cell counts, but was less active against 24 h biofilms (approximately 2 log10 reduction). Compound-loaded mesh showed dosage-dependent biofilm-removing capability. In addition, compound 1 displayed a significant inhibitory effect on tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) secretion from BMDM cells, but interleukin-1 beta (IL-1β) secretion was not significant. The compound was not cytotoxic to BMDM cells at concentrations effective in removing biofilm and lowering cytokine release. These findings highlight the potential of this serrulatane diterpenoid to be further developed for applications in wound management.

Increased CD271 expression by the NF-kB pathway promotes melanoma cell survival and drives acquired resistance to BRAF inhibitor vemurafenib.

Specific BRAFV600E inhibitors (BRAFi) are highly effective in the treatment of melanoma. However, acquired drug resistances invariably develop after the initial response. Therefore, the identification of new mechanisms of acquired resistance gives important clues towards the development of therapies that could elicit long lasting responses. Here we report that CD271 confers resistance to BRAFi in melanoma cells. The expression of CD271 is increased by BRAFi through a stimulation of tumor necrosis factor-alpha (TNFα) secretion that leads to NF-κB signaling pathway activation. CD271 is upregulated in a subset of BRAFi-resistant melanoma cells. The inhibition of TNFα/NF-κB pathway and CD271 silencing restore the BRAFi sensitivity of resistant melanoma cells. Finally, increase of CD271 expression is validated in BRAFi-resistant xenografts tumors and also in tumors from the patients who relapsed under BRAFi. In summary, these results reveal a novel TNFα/NF-κB/CD271 axis whose activation contributes to the acquisition of resistance to BRAFi and therefore may represent a novel therapeutic target to improve the efficacy of therapy in melanoma.

Saturated fatty acids enhance TLR4 immune pathways in human trophoblasts.

What are the effects of fatty acids on placental inflammatory cytokine with respect to toll-like receptor-4/nuclear factor-kappa B (TLR4/NF-kB)? Exogenous fatty acids induce a pro-inflammatory cytokine response in human placental cells in vitro via activation of TLR4 signaling pathways. The placenta is exposed to changes in circulating maternal fatty acid concentrations throughout pregnancy. Fatty acids are master regulators of innate immune pathways through recruitment of toll-like receptors and activation of cytokine synthesis. Trophoblast cells isolated from 14 normal term human placentas were incubated with long chain fatty acids (FA) of different carbon length and degree of saturation. The expression and secretion of interleukin-6 (IL-6), IL-8 and tumor necrosis factor-alpha (TNF-α) were measured by reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. Antibodies against TLR4 ligand binding domain, downstream signaling and anti-p65 NFkB-inhibitor were used to characterize the pathways of FA action. General approach used primary human term trophoblast cell culture. Methods and end-points used real-time quantitative PCR, cytokine measurements, immunohistochemistry, western blots. The long chain saturated fatty acids, stearic and palmitic (PA), stimulated the synthesis as well as the release of TNF-α, IL-6 and IL-8 by trophoblast cells (2- to 6-fold, P < 0.001). In contrast, the unsaturated (palmitoleic, oleic, linoleic) acids did not modify cytokine expression significantly. Palmitate-induced inflammatory effects were mediated via TLR4 activation, NF-kB phosphorylation and nuclear translocation. TNF-α protein level was close to the limit of detection in the culture medium even when cells were cultured with PA. These mechanisms open the way to a better understanding of how changes in maternal lipid homeostasis may regulate placental inflammatory status. X.Y. was recipient of fellowship award from West China Second University Hospital, Sichuan University (NIH HD 22965-19). The authors have nothing else to disclose. None.

The Crossroad of Inflammation and Diabetes: Role of Toll-Like Receptor

A different facet of type 2 diabetes was recognized in the early 80s with the demonstration of the “state of coagulation activation” and “endothelial dysfunction / activation” in not only type 2 diabetes but also in obesity. The discovery of tumor necrosis factor (TNF) alpha and its physiological role including many aspects of coagulation activation and endothelial dysfunction was later found to have a correlation with adiposity and type 2 diabetes. Further knowledge in the metabolic function of TNF and concomitant discovery of elevated cytokines released from adipose tissue suggested the integral involvement of inflammation in the pathogenesis of type 2 diabetes and obesity. The increased influx of monocytes in adipose tissue, an inflammatory state, elevates TNF alpha secretion leading to the induction of insulin resistance through the release of free fatty acids from visceral adipose tissue; furthermore, TNF alpha directly causes insulin receptor signaling dysfunction by shifting the phosphorylation residues. The fundamental question of why excess adiposity stimulates inflammatory response, thereby insulin resistance, lingered for a decade. The demonstration of free fatty acids (lauric, palmitic and stearic acid) engaging Toll Like Receptor 4 to stimulate innate immunity has some similarity to bacterial lipopolysacharides; providing the link of visceral adiposity to inflammatory syndrome. This article provides a historical review of the progression of knowledge in unmasking the inflammatory aspect in diabetes and visceral obesity, which were diseases often associated with hyperglycemia. In fact, it is the inflammatory aspect of the disease that governs the basic pathophysiology with complex interplays at the cross road of inflammation and metabolism.

Perfluorooctane sulfonate mediates microglial activation and secretion of TNF-α through Ca²⁺-dependent PKC-NF-кB signaling.

Perfluorooctane sulfonate (PFOS), a ubiquitous pollutant widely found in the environment and biota, can cause numerous adverse effects on human health. In recent years, PFOS's toxic effects on the central nervous system (CNS) have been shown. However, we still have a lot to study in the underlying molecular mechanism of PFOS's neurotoxicity. Microglia, the innate immune cells of CNS, are critically implicated in various neurological diseases caused by pro-inflammatory mediators. In our research, we found that HAPI microglia secreted tumor necrosis factor-alpha (TNF-α) after PFOS exposure in time-dependent and dose-dependent way. We also discovered that intracellular concentration of free Ca(2+) ([Ca(2+)]i) significantly increased after PFOS treatments. It was noteworthy here the secretion of TNF-α mediated by PFOS was blocked by Ca(2+) inhibitor and protein kinase C (PKC) inhibitor. Besides these, we had learned as well that PFOS brought about the up-regulation of phosphorylated nuclear factor kappa B (NF-кB) p65 expression and accelerated degradation of NF-κB inhibitor alpha (IкBα), however, these effects could be attenuated or blocked by Ca(2+) inhibitor and PKC inhibitor. Finally, through treating SH-SY5Y cells with PFOS-treated microglial conditioned medium, we demonstrated that TNF-α mediated neuronal apoptosis. To sum up, our research had shown, for the first time, that the distinct TNF-α secretion brought by PFOS in HAPI microglia, was achieved through the Ca(2+)-dependent PKC-NF-кB signaling, subsequently participating in neuronal loss.

Anti-inflammatory effects of isoacteoside from Abeliophyllum distichum

Isoacteoside, a dihydroxypheynylethyl glycoside, is a major bioactive component of Abeliophyllum distichum (White Forsythia) which is a deciduous shrub native to the south and central areas of Korea. The present study is designed to evaluate the anti-inflammatory activities and underlying mechanisms of isoacteoside in human mast cell line, HMC-1 cells. We isolated isoacteoside from A. distichum. The anti-inflammatory effect of isoacteoside was investigated in HMC-1 cells by studying the following markers: phorbol 12-myristate 13-acetate and calcium ionophore A23187 (PMACI)-induced interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor alpha (TNF-α) secretion and mRNA expression by ELISA and RT-PCR, respectively. In addition, mechanism related to anti-inflammatory was investigated by Western blotting. Isoacteoside significantly suppressed the production and mRNA expression of proinflammatory cytokines including IL-1β, IL-6, IL-8 and TNF-α in PMACI-stimulated HMC-1 cells without cytotoxicity. It was found that anti-inflammatory effects of isoacteoside are mediated by action on caspase-1, mitogen-activated protein kinases (c-Jun N-terminal kinase, p38, extracellular signal-regulated protein kinase) and nuclear factor-kappa B pathways. Taken together, the present findings provide new insights that isoacteoside may be a promising anti-inflammatory agent for inflammatory disorders.

MiR-124 disinhibits neurite outgrowth in an inflammatory environment

Lesions of the central nervous system elicit inflammatory responses that counteract the regeneration of neurites. Microglia and infiltrating macrophages that were activated by trauma have been identified as cellular sources of inhibitory factors. We examine cultured macrophage (RAW264.7) and neuronal (PC12) cell lines to ascertain the potential modulators of the inflammatory impact on neurons. By exposing quiescent macrophages to lipopolysaccharide (LPS) and interferon γ (IFN-γ), cells can be transformed into an activated M1 phenotype. Neurite extension was induced in PC12 cells by culturing them in the presence of nerve growth factor. Neurite outgrowth was quantified by analyzing immunofluorescence and phase contrast microscopy images. Activated macrophages significantly reduced neurite extension. Macrophage activation by LPS/IFN-γ induced a 1000-fold increase in tumor necrosis factor alpha (TNF-α) secretion, as quantified by enzyme-linked immunosorbent assays (ELISA). Recombinant TNF-α inhibited neurite formation at concentrations as low as 0.016ng/ml. In contrast, the masking of TNF-α with specific functional antibodies abrogated neurite growth inhibition by activated macrophages. Taken together, these results indicated that TNF-α is a key component of inhibitory macrophage action. The transfection of PC12 neurons with microRNA-124 (miR-124) counteracted the inhibition of neurites mediated by both recombinant TNF-α and macrophages. miR-124 did not stimulate neurite formation per se, nor was cell viability affected. These data suggest that miR-124 might be a valuable tool for desensitizing neurons to a repulsive inflammatory environment.

Mouse aorta-derived mesenchymal progenitor cells contribute to and enhance the immune response of macrophage cells under inflammatory conditions.

IntroductionMesenchymal progenitor cells interact with immune cells and modulate inflammatory responses. The cellular characteristics required for this modulation are under fervent investigation. Upon interaction with macrophage cells, they can contribute to or suppress an inflammatory response. Current studies have focused on mesenchymal progenitors derived from bone marrow, adipose, and placenta. However, the arterial wall contains many mesenchymal progenitor cells, which during vascular disease progression have the potential to interact with macrophage cells. To examine the consequence of vascular-tissue progenitor cell-macrophage cell interactions in an inflammatory environment, we used a recently established mesenchymal progenitor cell line derived from the mouse aorta.MethodsMouse bone marrow-derived macrophage (MΦ) cells and mouse aorta-derived mesenchymal progenitor (mAo) cells were cultured alone or co-cultured directly and indirectly. Cells were treated with oxidized low-density lipoprotein (ox-LDL) or exposed to the inflammatory mediators lipopolysaccharide (LPS) and interferon-gamma (IFNγ) or both. A Toll-like receptor-4 (TLR4)-deficient macrophage cell line was used to determine the role of the mAo cells. To monitor inflammation, nitric oxide (NO), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNFα) secretions were measured.ResultsMesenchymal progenitor cells isolated from aorta and cloned by high proliferative capacity (mAo) can differentiate into multiple mesenchymal lineages and are positive for several commonly used mouse mesenchymal stem cell markers (that is, CD29, CD44, CD105, CD106, and Sca-1) but are negative for CD73 and ecto-5′-nucleotidase. In co-culture with MΦ cells, they increase MΦ oxidized-LDL uptake by 52.2%. In an inflammatory environment, they synergistically and additively contribute to local production of both NO and IL-6. After exposure to ox-LDL, the inflammatory response of MΦ cells to LPS and LPS/IFNγ is muted. However, when lipid-laden MΦ cells are co-cultured with mAo cell progenitors, the muted response is recovered and the contribution by the mAo cell progenitor is dependent upon cell contact.ConclusionsThe resident mesenchymal progenitor cell is a potential contributor to vascular inflammation when in contact with inflamed and lipid-laden MΦ cells. This interaction represents an additional target in vascular disease treatment. The potential for resident cells to contribute to the local immune response should be considered when designing therapeutics targeting inflammatory vascular disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0071-8) contains supplementary material, which is available to authorized users.

Monocyte hyporesponsiveness and Toll-like receptor expression profiles in coronary artery bypass grafting and its clinical implications for postoperative inflammatory response and pneumonia: An observational cohort study.

Cardiac surgery with cardiopulmonary bypass has a major impact on the congenital immune response, in which Toll-like receptors (TLRs) are the first line of defence. Decreased surface expression of TLRs and impaired monocyte responsiveness to TLR ligands occur following surgery. However, the clinical implications of this altered immune response are not clear. To study cardiac surgery induced changes in perioperative TLR expression and monocyte responsiveness, and the association with postoperative inflammatory complications. A prospective cohort study. Single university hospital, enrolment March to December 2007. Eighty-four out of 92 patients who underwent coronary artery bypass grafting (CABG) were followed up until hospital discharge. We assessed in-vivo TLR-2 and TLR-4 expression and ex-vivo monocyte responsiveness to TLR-2 and TLR-4 ligands, measured by Pam3Cys and lipopolysaccharide (LPS)-induced interleukin (IL)-6 and IL-8, and tumour necrosis factor alpha (TNF-α) secretion until 24 h after surgery. Patients were followed to identify adverse inflammatory and infectious outcomes. Monocyte TLR expression decreased during CABG but returned to baseline after 24 h [linear mixed effects (LME) over time P < 0.0001]. Monocyte responsiveness changed significantly over time, with marked postoperative hyporesponsiveness (LME P < 0.0001). Decreased monocyte responsiveness to LPS was associated with monocyte TLR-4 expression (LME for IL-6 P = 0.04, IL-8 P = 0.002, TNF-α P = 0.05). Intraoperative decrease of monocyte TLR-2 expression was associated with postoperative systemic inflammatory response syndrome (SIRS) and pneumonia [odds ratio (OR) 2.06, 95% confidence interval (95% CI) 1.14 to 3.72], but the perioperative decrease of monocyte TLR-4 expression was not (OR 1.10, 95% CI 0.80 to 1.52). CABG surgery induced a decrease in in-vivo monocyte TLR expression and also in ex-vivo monocyte responsiveness, with which monocyte TLR-4 expression and monocyte LPS responsiveness seemed to be associated. Decreased TLR-2 expression was associated with the occurrence of SIRS and pneumonia, suggesting a role in the cause of postoperative inflammatory conditions. Clinicaltrials.gov identifier: NCT00356746.

Suppression of the TNF-alpha level is mediated by Gan-Lu-Yin (traditional Chinese medicine) in human oral cancer cells through the NF-kappa B, AKT, and ERK-dependent pathways.

Oral cancer is one of the major causes of deaths in the male population of Taiwan. Gan-Lu-Yin (GLY) is used for an adjuvant treatment of Traditional Chinese Medicine in clinical patients. In this study, we investigated the molecular mechanisms in oral cancer cell lines after exposure to GLY. The cytometric bead-based array (CBA) method was used for the examining and analyzing of tumor necrosis factor-alpha (TNF-α) secretion level. TNF-α mRNA expression was determined by real-time PCR analysis. Nuclear factor kappa B (NF-κB) activity and other relative proteins were determined by NF-κB promoter assay, Western blotting, electrophoretic mobility shift assay (EMSA), and immuno-staining analyses. GLY decreased the secretion of TNF-α from the oral cancer CAL 27 cells. Furthermore, 2000 μg/mL of GLY significantly suppressed TNF-α mRNA expression of CAL 27 cells in a time-dependent manner. GLY reduced the levels of proteins, including nuclear NF-κB (p65 and p50), p-IKK (ser176), p-IκB, p-AKT, p-ERK, and nuclear Egr-1 in a time and dose-dependent manner. GLY also suppressed the NF-κB activity and translocation in CAL 27 cells. We suggest that GLY might promote the cure of oral cancer through decreasing the level of TNF-α cytokine, and these actions were mediated partially through the NF-κB, AKT, and ERK-dependent pathways in vitro. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1196-1205, 2016.

Hepatotoxicants induce cytokine imbalance in response to innate immune system

In recent years, attention has been paid to innate immune systems as mechanisms to initiate or promote drug-induced liver injury (DILI). Kupffer cells are hepatic resident macrophages and might be involved in the pathogenesis of DILI by release of pro- and anti-inflammatory mediators such as cytokines, chemokines, reactive oxygen species, and/or nitric oxides. The purpose of this study was to investigate alterations in mediator levels induced by hepatotoxic compounds in isolated Kupffer cells and discuss the relation between balance of each cytokine or chemokine and potential of innate immune-mediated DILI. Primary cultured rat Kupffer cells were treated with hepatotoxic (acetaminophen, troglitazone, trovafloxacin) or non-hepatotoxic (pioglitazone, levofloxacin) compounds with or without lipopolysaccharide (LPS). After 24 hr treatment, cell supernatants were collected and various levels of mediators released by Kupffer cells were examined. Although hepatotoxicants had no effect on the LPS-induced tumor necrosis factor-alpha (TNF-α) secretion, they enhanced the release of pro-inflammatory cytokine interleukin-1 beta (IL-1β) and suppressed the anti-inflammatory cytokines interleukin-6 (IL-6) and interleukin-10 (IL-10) induced by LPS. These cytokine shifts were not associated with switching the phenotypes of M1 and M2 macrophages in Kupffer cells. In conclusion, the present study suggested that the levels of some specific cytokines are affected by DILI-related drugs with LPS stimulation, and imbalance between pro- and anti-inflammatory cytokines, induced by the up-regulation of IL-1β and the down-regulation of IL-6 or IL-10, plays a key role in innate immune-mediated DILI.

Oxymatrine inhibits lipopolysaccharide-induced inflammation by down-regulating Toll-like receptor 4/nuclear factor-kappa B in macrophages.

Oxymatrine (OMT) is the quinolizidine alkaloid extracted from the Chinese herb Sophora flavescens Ait. that has many pharmacological effects and is used for the treatment of some inflammatory diseases. In this study, RAW264.7 cells and THP-1 differentiated macrophages were pretreated with various concentrations of OMT at 2 h prior to treatment with lipopolysaccharide (LPS) (1.0 μg/mL) for different durations. We detected the anti-inflammatory effect of OMT in LPS-stimulated macrophages and investigated the molecular mechanism. We showed that OMT pretreatment significantly inhibited the LPS-induced secretion of nitric oxide (NO), interleukin-1 beta (IL-1β), and tumor necrosis factor-alpha (TNF-α) in supernatant, attenuated the mRNA levels of inducible nitric oxide synthase (iNOS), IL-1β, TNF-α, and Toll-like receptor 4 (TLR4), increased TLR4 and phosphorylation of inhibitor of kappa B-alpha (p-IBα) in cytosol, and decreased the nuclear level of nuclear factor-κB (NF-κB) p65 in macrophages. In conclusion, OMT exerts anti-inflammatory properties in LPS-stimulated macrophages by down-regulating the TLR4/NF-κB pathway.

Protective effects of pogostone from Pogostemonis Herba against ethanol-induced gastric ulcer in rats

We examined the protective effect of pogostone (PO), a chemical constituent isolated from Pogostemonis Herba, on the ethanol-induced gastric ulcer in rats. Administration of PO at doses of 10, 20 and 40mg/kg body weight prior to ethanol ingestion effectively protected the stomach from ulceration. The gastric lesions were significantly ameliorated by all doses of PO as compared to the vehicle group. Pre-treatment with PO prevented the oxidative damage and the decrease of prostaglandin E2 (PGE2) content. In addition, PO pretreatment markedly increased the mucosa levels of glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT), and decreased gastric malonaldehyde (MDA), relative to the vehicle group. In the mechanistic study, significant elevation of non-protein-sulfhydryl (NP-SH) was observed in the gastric mucosa pretreated by PO. Analysis of serum cytokines indicated that PO pretreatment obviously elevated the decrease of interleukin-10 (IL-10) level, while markedly mitigated the increment of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) secretions in ethanol-induced rats. Taken together, these results strongly indicate that PO could exert a gastro-protective effect against gastric ulceration, and the underlying mechanism might be associated with the stimulation of PGE2, improvement of antioxidant and anti-inflammatory status, as well as preservation of NP-SH.