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1.
INTRODUCTION: Intracellular signaling/synthetic pathways are being increasingly extensively characterized. However, while these pathways can be displayed in static diagrams, in reality they exist with a degree of dynamic complexity that is responsible for heterogeneous cellular behavior. Multiple parallel pathways exist and interact concurrently, limiting the ability to integrate the various identified mechanisms into a cohesive whole. Computational methods have been suggested as a means of concatenating this knowledge to aid in the understanding of overall system dynamics. Since the eventual goal of biomedical research is the identification and development of therapeutic modalities, computational representation must have sufficient detail to facilitate this 'engineering' process. Adding to the challenge, this type of representation must occur in a perpetual state of incomplete knowledge. We present a modeling approach to address this challenge that is both detailed and qualitative. This approach is termed 'dynamic knowledge representation,' and is intended to be an integrated component of the iterative cycle of scientific discovery. METHODS: BioNetGen (BNG), a software platform for modeling intracellular signaling pathways, was used to model the toll-like receptor 4 (TLR-4) signal transduction cascade. The informational basis of the model was a series of reference papers on modulation of (TLR-4) signaling, and some specific primary research papers to aid in the characterization of specific mechanistic steps in the pathway. This model was detailed with respect to the components of the pathway represented, but qualitative with respect to the specific reaction coefficients utilized to execute the reactions. Responsiveness to simulated lipopolysaccharide (LPS) administration was measured by tumor necrosis factor (TNF) production. Simulation runs included evaluation of initial dose-dependent response to LPS administration at 10, 100, 1000 and 10,000, and a subsequent examination of preconditioning behavior with increasing LPS at 10, 100, 1000 and 10,000 and a secondary dose of LPS at 10,000 administered at approximately 27h of simulated time. Simulations of 'knockout' versions of the model allowed further examination of the interactions within the signaling cascade. RESULTS: The model demonstrated a dose-dependent TNF response curve to increasing stimulus by LPS. Preconditioning simulations demonstrated a similar dose-dependency of preconditioning doses leading to attenuation of response to subsequent LPS challenge - a 'tolerance' dynamic. These responses match dynamics reported in the literature. Furthermore, the simulated 'knockout' results suggested the existence and need for dual negative feedback control mechanisms, represented by the zinc ring-finger protein A20 and inhibitor kappa B proteins (IkappaB), in order for both effective attenuation of the initial stimulus signal and subsequent preconditioned 'tolerant' behavior. CONCLUSIONS: We present an example of detailed, qualitative dynamic knowledge representation using the TLR-4 signaling pathway, its control mechanisms and overall behavior with respect to preconditioning. The intent of this approach is to demonstrate a method of translating the extensive mechanistic knowledge being generated at the basic science level into an executable framework that can provide a means of 'conceptual model verification.' This allows for both the 'checking' of the dynamic consequences of a mechanistic hypothesis and the creation of a modular component of an overall model directed at the engineering goal of biomedical research. It is hoped that this paper will increase the use of knowledge representation and communication in this fashion, and facilitate the concatenation and integration of community-wide knowledge. 相似文献
2.
Background
Adjuvants serve as catalysts of the innate immune response by initiating a localized site of inflammation that is mitigated by the interactions between antigens and toll like receptor (TLR) proteins. Currently, the majority of vaccines are formulated with aluminum based adjuvants, which are associated with various side effects. In an effort to develop a new class of adjuvants, agonists of TLR proteins, such as bacterial products, would be natural candidates. Lipopolysaccharide (LPS), a major structural component of gram negative bacteria cell walls, induces the systemic inflammation observed in septic shock by interacting with TLR-4. The use of synthetic peptides of LPS or TLR-4 agonists, which mimic the interaction between TLR-4 and LPS, can potentially regulate cellular signal transduction pathways such that a localized inflammatory response is achieved similar to that generated by adjuvants.Methodology/Principal Findings
We report the identification and activity of several peptides isolated using phage display combinatorial peptide technology, which functionally mimicked LPS. The activity of the LPS-TLR-4 interaction was assessed by NF-κB nuclear translocation analyses in HEK-BLUE™-4 cells, a cell culture model that expresses only TLR-4, and the murine macrophage cell line, RAW264.7. Furthermore, the LPS peptide mimics were capable of inducing inflammatory cytokine secretion from RAW264.7 cells. Lastly, ELISA analysis of serum from vaccinated BALB/c mice revealed that the LPS peptide mimics act as a functional adjuvant.Conclusions/Significance
Our data demonstrate the identification of synthetic peptides that mimic LPS by interacting with TLR-4. This LPS mimotope-TLR-4 interaction will allow for the development and use of these peptides as a new class of adjuvants, namely TLR-4 agonists. 相似文献3.
Shishido T Nozaki N Takahashi H Arimoto T Niizeki T Koyama Y Abe J Takeishi Y Kubota I 《Biochemical and biophysical research communications》2006,345(4):1446-1453
Background
It is now evident that inflammation after vascular injury has significant impact on the restenosis after revascularization procedures such as angioplasty, stenting, and bypass grafting. However, the mechanisms that regulate inflammation and repair after vascular injury are incompletely understood. Here, we report that vascular injury-mediated cytokine expression, reactive oxygen species (ROS) production, as well as subsequent neointimal formation requires Toll-like receptor-2 (TLR-2) mediated signaling pathway in vivo.Methods and results
Vascular injury was induced by cuff-placement around the femoral artery in non-transgenic littermates (NLC) and TLR-2 knockout (TLR-2KO) mice. After cuff-placement in NLC mice, expression of TLR-2 was significantly increased in both smooth muscle medial layer and adventitia. Interestingly, we found that inflammatory genes expression such as tumor necrosis factor-α, interleukin-1β (IL-1β), IL-6, and monocyte chemoattractant protein-1 were markedly decreased in TLR-2KO mice compared with NLC mice. In addition, ROS production after vascular injury was attenuated in TLR-2KO mice compared with NLC mice. Since we observed the significant role of endogenous TLR-2 activation in regulating inflammatory responses and ROS production after vascular injury, we determined whether inhibition of endogenous TLR-2 activation can inhibit neointimal proliferation after vascular injury. Neointimal hyperplasia was markedly suppressed in TLR-2KO mice compared with WT mice at both 2 and 4 weeks after vascular injury.Conclusions
These findings suggested that endogenous TLR-2 activation might play a central role in the regulation of vascular inflammation as well as subsequent neointimal formation in injured vessels. 相似文献4.
The complex biology of Gram-negative bacterial lipopolysaccharide (LPS) is central to the acute inflammatory response in sepsis and related diseases. Repeated treatment with LPS can lead to desensitization or enhancement of subsequent responses both in vitro and in vivo (a phenomenon known as preconditioning). Previous computational studies have demonstrated a role for anti-inflammatory influences in this process (J. Day, J. Rubin, Y. Vodovotz, C.C. Chow, A. Reynolds, G. Clermont, A reduced mathematical model of the acute inflammatory response: II. Capturing scenarios of repeated endotoxin administration. J. Theor. Biol. 242 (2006) 237). Since LPS signals via Toll-like receptor 4 (TLR4), we created a simple mathematical model in order to address the role of this receptor in both the normal and preconditioned response to LPS. We created a non-linear system of ordinary differential equations, consisting of free LPS, free TLR4, bound complex LPS-TLR4, and an intracellular signaling cascade (lumped into a single variable). We simulate the effects of preconditioning by small and large repeated doses of LPS on the system, varying the timing of the doses as well as the rate of expression of TLR4. Our simulations suggest that a simplified model of LPS/TLR4 signaling can account for complex preconditioning phenomena without invoking a specific signaling inhibition mechanism, but rather based on the dynamics of the signaling response itself, as well as the timing and magnitude of the LPS stimuli. 相似文献
5.
Faure E Equils O Sieling PA Thomas L Zhang FX Kirschning CJ Polentarutti N Muzio M Arditi M 《The Journal of biological chemistry》2000,275(15):11058-11063
A missense mutation in the cytoplasmic domain of the Toll-like receptor-4 (TLR-4) has been identified as the defect responsible for lipopolysaccharide (LPS) hyporesponsiveness in C3H/HeJ mice. TLR-4 and TLR-2 have recently been implicated in LPS signaling in studies where these receptors were overexpressed in LPS non-responsive 293 human embryonic kidney cells. However, the signaling role of TLR-4 or TLR-2 in human cells with natural LPS response remains largely undefined. Here we show that human dermal microvessel endothelial cells (HMEC) and human umbilical vein endothelial cells express predominantly TLR-4 but very weak TLR-2 and respond vigorously to LPS but not to Mycobacterium tuberculosis 19-kDa lipoprotein. Transient transfection of non-signaling mutant forms of TLR-4 and anti-TLR-4 monoclonal antibody inhibited LPS-induced NF-kappaB activation in HMEC, while a monoclonal antibody against TLR-2 was ineffective. In contrast to LPS responsiveness, the ability of HMEC to respond to 19-kDa lipoprotein correlated with the expression of TLR-2. Transfection of TLR-2 into HMEC conferred responsiveness to 19-kDa lipoprotein. These data indicate that TLR-4 is the LPS signaling receptor in HMEC and that human endothelial cells (EC) express predominantly TLR-4 and weak TLR-2, which may explain why they do not respond to 19-kDa lipoprotein. The differential expression of TLRs on human EC may have important implications in the participation of vascular EC in innate immune defense mechanisms against various infectious pathogens, which may use different TLRs to signal. 相似文献
6.
7.
Jianhua Rao Jianjie Qin Xiaofeng Qian Ling Lu Ping Wang Zhengshan Wu Yuan Zhai Feng Zhang Guoqiang Li Xuehao Wang 《PloS one》2013,8(6)
Background
Low-dose lipopolysaccharide (LPS) preconditioning-induced liver protection has been demonstrated during ischemia-reperfusion injury (IRI) in several organs but has not been sufficiently elucidated underlying causal mechanism. This study investigated the role of low-dose LPS preconditioning on ATF4-CHOP pathway as well as the effects of the pathway on tissue injury and inflammation in a mouse model of liver partial-warm IRI.Methods
LPS (100 µg/kg/d) was injected intraperitoneally two days before ischemia. Hepatic injury was evaluated based on serum alanine aminotransferase levels, histopathology, and caspase-3 activity. The ATF4-CHOP pathway and its related apoptotic molecules were investigated after reperfusion. The role of LPS preconditioning on apoptosis and ATF4-CHOP pathway was examined in vitro. Moreover, the effects of the ATF4-CHOP pathway on apoptosis, Caspase-12, and Caspase-3 were determined with ATF4 small interfering RNA (siRNA). Inflammatory cytokine expression was also checked after reperfusion. Inflammatory cytokines and related signaling pathways were analyzed in vitro in macrophages treated by LPS preconditioning or ATF4 siRNA.Results
LPS preconditioning significantly attenuated liver injury after IRI. As demonstrated by in vitro experiments, LPS preconditioning significantly reduced the upregulation of the ATF4-CHOP pathway and inhibited Caspase-12 and Caspase-3 activation after IRI. Later experiments showed that ATF4 knockdown significantly suppressed CHOP, cleaved caspase-12 and caspase-3 expression, as well as inhibited hepatocellular apoptosis. In addition, in mice pretreated with LPS, TNF-α and IL-6 were inhibited after reperfusion, whereas IL-10 was upregulated. Similarly, low-dose LPS significantly inhibited TNF-α, IL-6, ATF4-CHOP pathway, NF-κB pathway, and ERK1/2 in high-dose LPS-stimulated macrophages, whereas IL-10 and cytokine signaling (SOCS)-3 suppressor were induced. Importantly, ATF4 siRNA is consistent with results of LPS preconditioning in macrophages.Conclusions
This work is the first time to provide evidence for LPS preconditioning protects hepatocytes from IRI through inhibiting ATF4-CHOP pathway, which may be critical to reducing related apoptosis molecules and modulating innate inflammation. 相似文献8.
Barsness KA Arcaroli J Harken AH Abraham E Banerjee A Reznikov L McIntyre RC 《American journal of physiology. Regulatory, integrative and comparative physiology》2004,287(3):R592-R599
Toll-like receptor 4 (TLR-4), initially identified as an LPS receptor, is critical to the signaling of a variety of danger signals, including heat shock protein 60, fibrinogen, and fibronectin. Recent data also suggest that TLR-4 plays a role in determining survival in both endotoxemia and hemorrhagic shock. We hypothesized that a functional TLR-4 would be required for hemorrhage and endotoxin-induced acute lung injury. Hemorrhage- and endotoxin-induced lung TNF-alpha mRNA and protein production, neutrophil accumulation, and protein permeability were dependent on a functional TLR-4. Hemorrhage-induced nuclear factor (NF)-kappaB activation was independent of functional TLR-4, whereas endotoxin-induced activation of NF-kappaB requires a functional TLR-4 for full response. Therefore, we conclude that 1) hemorrhage-induced acute lung injury is TLR-4 dependent and 2) hemorrhage has a different and distinct TLR-4-dependent intracellular activation mechanism compared with endotoxemia. 相似文献
9.
Siri L. Feruglio Marius Tr?seid Jan Kristian Dam?s Dag Kvale Anne Ma Dyrhol-Riise 《PloS one》2013,8(7)
Background
Biomarkers to differentiate between active tuberculosis (TB) and latent TB infection (LTBI) and to monitor treatment responses are requested to complement TB diagnostics and control, particularly in patients with multi-drug resistant TB. We have studied soluble markers of the Toll-like-receptor 4 (TLR-4) pathway in various stages of TB disease and during anti-TB treatment.Methods
Plasma samples from patients with culture confirmed drug-sensitive TB (n = 19) were collected before and after 2, 8 and 24 weeks of efficient anti-TB treatment and in a LTBI group (n = 6). Soluble (s) CD14 and myeloid differentiation-2 (MD-2) were analyzed by the Enzyme-linked immunosorbent assay (ELISA). Lipopolysaccharide (LPS) was analyzed by the Limulus Amebocyte Lysate colorimetric assay. Nonparametric statistics were applied.Results
Plasma levels of sCD14 (p<0.001), MD-2 (p = 0.036) and LPS (p = 0.069) were elevated at baseline in patients with untreated active TB compared to the LTBI group. MD-2 concentrations decreased after 2 weeks of treatment (p = 0.011), while LPS levels decreased after 8 weeks (p = 0.005). In contrast, sCD14 levels increased after 2 weeks (p = 0.047) with a subsequent modest decrease throughout the treatment period. There was no significant difference in concentrations of any of these markers between patients with pulmonary and extrapulmonary TB or between patients with or without symptoms.Conclusion
Our data suggest that plasma levels of LPS, MD-2 and sCD14 can discriminate between active TB and LTBI. A decline in LPS and MD-2 concentrations was associated with response to anti-TB treatment. The clinical potential of these soluble TLR-4 pathway proteins needs to be further explored. 相似文献10.
Regina Sordi Fausto Chiazza Nimesh S. A. Patel Rachel A. Doyle Massimo Collino Christoph Thiemermann 《PloS one》2015,10(4)
Methods
Male rats were ‘pretreated’ with phosphate-buffered saline (PBS; i.p.) or LPS (1 mg/kg; i.p.) 24 h prior to HS. Mean arterial pressure (MAP) was maintained at 30 ± 2 mmHg for 90 min or until 25% of the shed blood had to be re-injected to sustain MAP. This was followed by resuscitation with the remaining shed blood. Four hours after resuscitation, parameters of organ dysfunction and systemic inflammation were assessed.Results
HS resulted in renal dysfunction, and liver and muscular injury. At a first glance, LPS preconditioning attenuated organ dysfunction. However, we discovered that HS-rats that had been preconditioned with LPS (a) were not able to sustain a MAP at 30 mmHg for more than 50 min and (b) the volume of blood withdrawn in these animals was significantly less than in the PBS-control group. This effect was associated with an enhanced formation of the nitric oxide (NO) derived from inducible NO synthase (iNOS). Thus, a further control group in which all animals were resuscitated after 50 min of hemorrhage was performed. Then, LPS preconditioning aggravated both circulatory failure and organ dysfunction. Most notably, HS-rats pretreated with LPS exhibited a dramatic increase in NF-κB activation and pro-inflammatory cytokines.Conclusion
In conclusion, LPS preconditioning predisposed animals to an earlier vascular decompensation, which may be mediated by an excess of NO production secondary to induction of iNOS and activation of NF-κB. Moreover, LPS preconditioning increased the formation of pro-inflammatory cytokines, which is likely to have contributed to the observed aggravation of organ injury/dysfunction caused by HS. 相似文献11.
Nemoto S Vallejo JG Knuefermann P Misra A Defreitas G Carabello BA Mann DL 《American journal of physiology. Heart and circulatory physiology》2002,282(6):H2316-H2323
The precise molecular mechanisms responsible for sepsis-induced myocardial dysfunction remain undefined. Toll-like receptor-4 (TLR-4) engages lipopolysaccharide (LPS) and activates signaling pathways leading to the expression of proinflammatory cytokines implicated in myocardial dysfunction. We determined whether TLR-4 was necessary for LPS-induced myocardial dysfunction in vivo. The effects of LPS on left ventricular (LV) function were studied in mice with defective TLR-4 signaling (C3H/HeJ, TLR-4 deficient) and wild-type mice (C3HeB/FeJ). Mice (n = 5/group) were injected with LPS or diluent, and LV function was examined by using two-dimensional echocardiography and conductance catheters. LPS significantly decreased all indexes of LV function in wild-type mice when compared with controls; LV function was not depressed in the LPS-treated TLR-4-deficient mice relative to controls. LPS increased myocardial nitric oxide synthase-2 expression and cGMP only in wild-type mice. This study suggests that TLR-4 mediates the LV dysfunction that occurs in LPS-induced shock. Therefore, TLR-4 might be a therapeutic target for attenuating the effects of LPS on the heart. 相似文献
12.
Regulator of G-protein signaling 4 (RGS4), an intracellular modulator of G-protein coupled receptor (GPCR)-mediated signaling, is regulated by multiple processes including palmitoylation and proteasome degradation. We found that co-expression of DHHC acyltransferases (DHHC3 or DHHC7), but not their acyltransferase-inactive mutants, increased expression levels of RGS4 but not its Cys2 to Ser mutant (RGS4C2S). DHHC3 interacts with and palmitoylates RGS4 but not RGS4C2S in vivo. Palmitoylation prolongs the half-life of RGS4 by over 8-fold and palmitoylated RGS4 blocked α1A-adrenergic receptor-stimulated intracellular Ca2+ mobilization. Together, our findings revealed that DHHC proteins could regulate GPCR-mediated signaling by increasing RGS4 stability.
Structured summary
MINT-8049215: Rgs4 (uniprotkb:P49799) physically interacts (MI:0915) with DHHC3 (uniprotkb:Q8R173) by anti-tag coimmunoprecipitation (MI:0007) 相似文献13.
Chang Chen Fan Zhang Zongze Zhang Mian Peng Yanlin Wang Yingying Chen 《Molecular biology reports》2013,40(2):1167-1172
Resuscitated hemorrhagic shock is believed to promote the development of acute lung injury (ALI) by priming the immune system for an exaggerated inflammatory response to a second trivial stimulus. This work explored effects of TLR4 on hemorrhage-induced ALI and “second-hit” responses, and further explore the mechanisms involved in “second-hit” responses. Expression of HO-1, IL-10, lung W/D and MPO markedly increased at nearly all time-points examined in HSR/LPS group as compared with sham/LPS group in WT mice. In HSR/LPS mice, the induced amount of IL-10 and the expressions of HO-1 of WT mice were significantly higher compared with TLR-4d/d. This study provides in vivo evidence that pulmonary infections after LPS instillation contribute to local tissue release of pro-inflammatory mediators after HSR systemic. Activation of TLR4 might induce HO-1 expression and HO-1 modulates proinflammatory responses that are triggered via TLR4 signaling. 相似文献
14.
Claudio Lera Orsatti Eliana Aguiar Petri Nahas Jorge Nahas-Neto Fabio Lera Orsatti Vanessa Innocenti Giorgi Steven S. Witkin 《PloS one》2014,9(10)
Objective
To evaluate the gene expression of Toll-Like (TLR-2 and TLR-4) receptors and cytokine profile in postmenopausal women with or without metabolic syndrome (MetS).Methods
In this cross-sectional study, 311 Brazilian women (age≥45 years and amenorrhea≥12 months) were included. Women showing three or more of the following diagnostic criteria were diagnosed as positive for MetS: waist circumference>88 cm, triglycerides≥150 mg/dL, HDL cholesterol<50 mg/dL, blood pressure≥130/85 mmHg, and fasting glucose≥100 mg/dL. The expression of TLR-2 and TLR-4 in peripheral blood was evaluated by RNA extraction and subsequent real time PCR analysis. The cytokine profile, tumor necrosis factor alpha (TNF-α) and interleukins 1β, 6, and 10, were measured by ELISA.Results
The expression of TLR-2 RNA was demonstrated in 32.5% and TLR-4 in 20.6% of the subjects. There was no association between the expression of TLR-2 and TLR-4 and the presence or absence of MetS (P>0.05). A greater production of IL-6 was associated with TLR-2 and TLR-4 expressions and greater production of TNF-α was associated only with TLR-2 expression (P>0.05). Only the lower quartile of IL-10 was associated with the presence of the MetS (P>0.05).Conclusions
TLR-2 and TLR-4 expressions were associated with increased pro-inflammatory cytokines, IL-6 and TNF-α, with no association with biomarkers of MetS. The low concentrations of IL-10 may suggest an anti-inflammatory modulation in postmenopausal women with MetS. 相似文献15.
Mara MassimiAlberta Tomassini Fabio SciubbaAnatoli P. Sobolev Laura Conti DevirgiliisAlfredo Miccheli 《Biochimica et Biophysica Acta (BBA)/General Subjects》2012,1820(1):1-8
Background
Resveratrol, a polyphenol found in plant products, has been shown to regulate many cellular processes and to display multiple protective and therapeutic effects. Several in vitro and in vivo studies have demonstrated the influence of resveratrol on multiple intracellular targets that may regulate metabolic homeostasis.Methods
We analysed the metabolic modifications induced by resveratrol treatment in a human hepatoblastoma line, HepG2 cells, using a 1H-NMR spectroscopy-based metabolomics approach that allows the simultaneous screening of multiple metabolic pathways.Results
Results demonstrated that cells cultured in the presence or absence of resveratrol displayed different metabolic profiles: the treatment induced a decreased utilisation of glucose and amino acids for purposes of energy production and synthesis associated to a decreased release of lactate in the culture medium and an increase in succinate utilisation. At the same time, resveratrol treatment slowed the cell cycle in the S phase without inducing apoptosis, and increased Sirt1 expression, also affecting its intracellular localisation.Conclusions
Our results show that the metabolomic analysis of the exometabolome of resveratrol-treated HepG2 cells indicates a metabolic switch from glucose and amino acid utilisation to fat utilisation for the production of energy, and seem in agreement with an effect mediated via AMPK- and Sirt1-activation.General significance
NMR-based metabolomics has been applied in a hepatocyte cell culture model in relation to resveratrol treatment; such an approach could be transferred to evaluate the effects of nutritional compounds with health impact. 相似文献16.
Giuseppe M. Campo Angela AvenosoSalvatore Campo Angela D'AscolaGiancarlo Nastasi Alberto Calatroni 《Biochimie》2010
Hyaluronan (HA) action depends upon its molecular size. Low molecular weight HA elicits pro-inflammatory responses by modulating the toll-like receptor-4 (TLR-4) or by activating the nuclear factor kappa B (NF-kB). In contrast, high molecular weight HA manifests an anti-inflammatory effect via CD receptors and by inhibiting NF-kB activation. Lipopolysaccharide (LPS) –mediated activation of TLR-4 complex induces the myeloid differentiation primary-response protein (MyD88) and the tumor necrosis factor receptor-associated factor-6 (TRAF-6), and ends with the liberation of NF-kB/Rel family members. The aim of this study was to investigate the influence of HA at different MWs (low, medium, high) on TLR-4 modulation in LPS-induced inflammatory response in mouse chondrocyte cultures. 相似文献
17.
18.
Objective
To determine the signaling pathways and components involved in insulin-mediated regulation of Acyl-CoA: cholesterol acyltransferase1 (ACAT1).Methods
THP-1 cells were cultured in RPMI 1640 medium and were induced into macrophages in the presence of 160 nM phorbol 12-myristate 13-acetate (PMA). Before insulin was added in, macrophages were preincubated with the inhibitors of the insulin signaling pathway, including wortmannin, phosphatidylinositol 3-kinase (PI3 K) inhibitor; PD98059, extracellular signal-regulated kinase (ERK) inhibitor; SB203580, p38 mitogen-activated protein kinase (p38MAPK) inhibitor; SP600125, c-Jun N-terminal kinase (JNK) inhibitor and U73122, phospholipase C-γ (PLC-γ) inhibitor. ACAT1 mRNA and protein expression level in macrophages were determined by real-time quantitative polymerase chain reaction and western blotting, respectively.Results
Real-time quantitative polymerase chain reaction and western blotting demonstrated that PD98059, SB203580 or SP600125 down-regulated the expression of ACAT1 in a dose-dependent manner. However, no obvious alteration was found in wortmannin and U73122 groups.Conclusion
These results suggest that the ERK, p38MAPK and JNK signaling pathways may be involved in insulin-mediated regulation of ACAT1, but no PI3K and PLC-γ signaling pathways were involved in the present study. 相似文献19.
Background
Bupleurum polysaccharides (BPs), isolated from Bupleurum smithii var. parvifolium, possesses immunomodulatory activity, particularly on inflammation. Bacterial endotoxin lipopolysaccharide (LPS) triggers innate immune responses through Toll-like receptor 4 (TLR4) on host cell membrane. The present study was performed to evaluate whether the therapeutic efficacy of BPs on suppression of LPS’s pathogenecity could be associated with the modulating of TLR4 signaling pathway.Methodology/Principal Findings
LPS stimulated expression and activation of factors in the TLR4 signaling system, including TLR4, CD14, IRAK4, TRAF6, NF-κB, and JNK, determined using immunocytochemical and/or Western blot assays. BPs significantly inhibited these effects of LPS. LPS increased pro-inflammatory cytokines (TNF-α, IL-6, IL-1β, IL-12p40, and IFN-β) and NO production, evaluated using ELISA and Griess reaction assays, respectively. BPs antagonized these effects of LPS. Interestingly, BPs alone augmented secretion of some pro-inflammatory cytokines of non-LPS stimulated macrophages and enhanced phagocytic activity towards fluorescent E.coli bioparticles. In a rat model of acute lung injury (ALI) with pulmonary hemorrhage and inflammation, BPs ameliorated lung injuries and suppressed TLR4 expression.Significance
The therapeutic properties of BPs in alleviating inflammatory diseases could be attributed to its inhibitory effect on LPS-mediated TLR4 signaling. 相似文献20.
Trine M. Reine Tram Thu Vuong Trond G. Jenssen Svein O. Kolset 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014