Ulinastatin Alleviates Rhabdomyolysis-Induced Acute Kidney Injury by Suppressing Inflammation and Apoptosis via Inhibiting TLR4/NF-κB Signaling Pathway

Inflammation - Acute kidney injury (AKI) is an important complication of rhabdomyolysis (RM), but there is lack of effective treatments. Ulinastatin (UTI) is a broad-spectrum serine protease...     

PD-L1 Regulates Inflammation in LPS-Induced Lung Epithelial Cells and Vascular Endothelial Cells by Interacting with the HIF-1α Signaling Pathway

Inflammation - Sepsis-induced lung injury was the most common cause of death in patients. This study aimed to investigate whether PD-L1 regulates the inflammation in LPS-induced lung epithelial...     

Thyroid Hormone Promotes Neuronal Differentiation of Embryonic Neural Stem Cells by Inhibiting STAT3 Signaling Through TRα1

A deficiency of maternal thyroid hormones (THs) during pregnancy may have severe impacts on fetal brain development. However, the cellular targets of THs and their underlying mechanisms are still unclear. In this study, we found that maternal hypothyroidism during pregnancy in mice inhibited neurogenesis in the embryonic telencephalon and caused learning and memory impairment in the offspring. To explore the underlying mechanisms, we treated cultured mouse embryonic neural stem cells (eNSCs) with a physiological level of 3, 5, 3'-triiodo-L-thyronine (T3). We found that T3 promoted the neuronal differentiation of eNSCs, while inhibiting astrocytic differentiation. In addition, the proliferation and maintenance of eNSCs were inhibited by T3. Furthermore, the TH receptor alpha 1 (TRα1) was detected in the eNSCs both in vivo and in vitro. Silencing TRα1 protein expression with specific siRNA eliminated the effects of T3 on eNSCs. We also found that T3 decreased STAT3 phosphorylation and STAT3-DNA binding activity through TRα1. The over expression of STAT3 attenuated the promotive effects of T3 on neuronal differentiation of eNSCs. Taken together, these results suggest that T3 promotes the neuronal differentiation of eNSCs by inhibiting STAT3 signaling activity through TRα1 and contributes to early neurogenesis in the embryonic telencephalon. Our studies reveal the physiological effects of TH in regulating eNSCs differentiation and suggest that eNSCs are one of the major cellular targets in the central nervous system by which TH influences early brain development. These findings also provide new insights into the mechanisms of neurological deficits caused by TH deficiency during embryogenesis.     

Etanercept Impairs Maturation of Human Monocyte-Derived Dendritic Cells by Inhibiting the Autocrine TNFα-mediated Signaling

The success of anti-tumor necrosis factor alpha (TNFα) therapies has led to increased interest as to the mechanisms and consequences of TNFα blockade. The aim of the study was to examine the effects of TNFα blockade by etanercept on lipopolysaccharide (LPS) or peptidoglycan (PG)-induced maturation of human monocyte-derived dendritic cells (MDDCs). MDDCs grown from peripheral blood of healthy donors were stimulated by LPS or PG with/without the presence of etanercept. Concentrations of TNFα in cell supernatants were assessed by ELISA, while the cells were stained with monoclonal antibodies to CD83, CD80, CD86, CD11c, CD40, HLA-DR, and annexin-V and acquired using a flow cytometer. Etanercept significantly decreased the stimulated cell surface expression of HLA-DR, CD80, CD86, CD40 and CD83 on MDDCs in all examined samples. Etanercept in the same dose, but denatured to loss of specificity for TNFα, failed to change any of the aforementioned markers. In the presence of etanercept, concentrations of TNFα in cell supernatants were decreased by 53% on average, with a range of 25%–87%. Etanercept impaired the stimulated maturation of MDDCs by neutralizing the induced TNFα, produced by the same MDDCs after antigenic stimulation. The reported data confirms that TNFα blockade may have a direct effect on DCs, with a wide spectrum of potential secondary effects downstream. The data also suggests the presence of TNFα-mediated autocrine signaling, serving to accelerate or catalyze the maturation process of MDDCs.     

Diallyl Disulfide Suppresses the Inflammation and Apoptosis Resistance Induced by DCA Through ROS and the NF-κB Signaling Pathway in Human Barrett’s Epithelial Cells

Barrett’s esophagus (BE) is generally accepted as the only precursor to esophageal adenocarcinoma (EAC). Deoxycholic acid (DCA)-induced inflammation and apoptotic resistance play an important role in the carcinogenesis and progression from BE to EAC. Diallyl disulfide (DADS) is a garlic-derived natural organosulfur compound. This study investigated whether DADS has chemopreventive effects against BE and the potentially related signaling pathway. BAR-T cells were treated with DCA in the presence or absence of DADS. An MTT assay was used to detect the viability of the cells. The apoptosis rate of the cells was measured by light microscopy and flow cytometry. ROS levels were determined by fluorescence microscopy and flow cytometry. Real-time PCR and ELISA were used to detect mRNA and protein levels, respectively. The levels of target proteins were also determined by western blot analysis. DADS did not inhibit cell viability in a certain concentration range. DADS, similar to the NF-κB inhibitor PDTC, inhibited the DCA-induced ROS production, inflammatory factors, IκBα phosphorylation, and expression of p50 in the nucleus in a dose-dependent manner. DADS also increased the cell apoptosis rate through down-regulating the level of Bcl-2. DADS has low cytotoxicity in BAR-T cells. It has an anti-inflammatory effect in BAR-T cells through inhibiting ROS and the NF-κB signaling pathway. Further, it abolishes the apoptotic resistance induced by DCA in an NF-κB/Bcl-2 dependent manner. DADS may be a good candidate for BE and EAC chemical prevention and therapy.     

TNF-α Induces Transient Resistance to Fas-Induced Apoptosis in Eosinophilic Acute Myeloid Leukemia Cells

Tumor necrosis factor α （TNF-α） has been recognized as an activator of nuclear factor kB （NF-kB）, a factor implicated in the protection of many cell types from apoptosis. We and others have presented evidence to suggest that Fas-induced apoptosis may be an important aspect of the resolution of inflammation, and that delayed resolution of inflammation may be directly associated with NF-kB-dependent resistance to Fas. Because TNF-α activates NF-kB in many cell types including inflammatory cells such as eosinophils, we examined effects of TNF-α signaling on the Fas-mediated killing of an eosinophilic cell line AML14. While agonist anti-Fas （CHII） treatment induced apoptosis in AML14 cells, no significant cell death occurred in response to TNF-α alone. Electrophoretic mobility shift assay （EMSA） revealed that TNF-α induced NF-kB transactivation in AMLI4 cells in a time- and dose-dependent fashion, and subsequent supershift assays indicated that the translocated NF-kB was the heterodimer p65 （RelA）/p50. Pre-treatment of cells with TNF-α dramatically decreased the CHll-induced cell death in a transient fashion, accompanied by suppression of activation of caspase-8 and caspase-3 activation. Inhibition of NF-kB transactivation by inhibitors, BAY 11-7085 and parthenolide, reversed the suppression of Fas-mediated apoptosis by TNF-α. Furthermore, TNF-α up-regulated X-linked inhibitor of apoptosis protein （XIAP） transiently and XIAP levels were correlated with the temporal pattern of TNF-α protection against Fas-mediated apoptosis. This finding suggested that TNF-α may contribute to the prolonged survival of inflammatory cells by suppression of Fas-mediated apoptosis, the process involved with NF-kB transactivation, anti-apoptotic XIAP up-regulation and caspase suppression. Cellular ＆ Molecular Immunology. 2007;4（1）：43-52.     

Thrombospondin-1 Production Regulates the Inflammatory Cytokine Secretion in THP-1 Cells Through NF-κB Signaling Pathway

Thrombospondin-1 (TSP-1) is upregulated in several inflammatory diseases. Recent data have shown that macrophages from TSP-1-deficient mice have a reduced inflammatory phenotype, suggesting that TSP-1 plays a part in macrophage activation. DNA microarray approach revealed that Porphyromonas gingivalis lipopolysaccharide (P. gingivalis LPS) may induce the enhanced TSP-1 expression in human monocytes, suggesting a role of TSP-1-mediated pathogenesis in periodontitis. Until recently, the function of TSP-1 has been a matter of debate. In this study, we explored the role of TSP-1 in inflammatory cytokine secretions and its putative mechanism in pathogenesis of periodontitis. We demonstrated that TSP-1 expression was significantly upregulated in gingival tissues with periodontitis and in P. gingivalis LPS-stimulated THP-1 cells. Deficiency of TSP-1 by transfecting siRNAs decreased IL-6, IL-1β, and TNF-α secretions in THP-1 cells, whereas overexpression of TSP-1 resulted in an upregulation of IL-6, IL-1β, and TNF-α productions. Additional experiments showed that Pyrrolidine dithiocarbamate (PDTC) inhibited IL-6, IL-1β, and TNF-α expression induced by overexpression of TSP-1, accompanying with downregulation of phosphorylated p65 and IκBα protein levels in response to P. gingivalis LPS. These results indicated that TSP-1 played a significant role in P. gingivalis LPS-initiated inflammatory cytokines (IL-6, IL-1β, and TNF-α) secretions of THP-1 cells, and the NF-κB signaling is involved in its induction of expression. Thus, TSP-1 effectively elevated P. gingivalis LPS-induced inflammation mediated by the NF-κB pathway and may be critical for pathology of periodontitis.     

Ophiopogonin D Inhibiting Epithelial NF-κB Signaling Pathway Protects Against Experimental Colitis in Mice

Inflammation - The sustained activation of the nuclear factor κB (NF-κB) signaling pathway has been observed in human inflammatory bowel disease (IBD). Ophiopogonin D (OP-D) is a small...     

Luteolin Prevents LPS-Induced TNF-α Expression in Cardiac Myocytes Through Inhibiting NF-κB Signaling Pathway

Luteolin, a plant flavonoid, has been shown to suppress inflammatory responses; however, the mechanism of luteolin on cardiac myocyte inflammation is still unknown. Because tumor necrosis factor-α (TNF-α), an inflammatory cytokine, is elevated in the failing heart and exerts multiple potentially harmful effects on cardiac myocytes, we therefore sought to examine the effects of luteolin on the expression of TNF-α in neonatal rat cardiac myocytes. In the present study, enzyme-linked immunosorbent assay (ELISA), real-time PCR, immunoblot, immunochemistry staining, and electrophoretic mobility shift assays (EMSA) were performed. ELISA assay showed that luteolin decreased lipopolysaccharide (LPS)-induced production of TNF-α in the medium. Real-time PCR assay confirmed that luteolin also inhibited LPS-induced increase in TNF-α mRNA in myocytes. Furthermore, immunoblot and immunochemistry staining assays represented that luteolin blocked LPS-induced IκB-β degradation and NF-κB p65 subunit nuclear translocation. In addition, EMSA demonstrated that luteolin reduced LPS-induced NF-κB DNA binding activity. Luteolin protects against LPS-induced TNF-α expression via inhibition of the NF-κB signaling pathway, suggesting that luteolin may be a potential therapeutic agent for the treatment of inflammation-related myocardial diseases.     

C-Reactive Protein Induces TNF-α Secretion by p38 MAPK–TLR4 Signal Pathway in Rat Vascular Smooth Muscle Cells

Atherosclerosis is a chronic inflammatory disease. C-reactive protein (CRP) not only is an inflammatory marker but also regulates the expressions of other inflammatory cytokines associated with the pathogenesis of atherosclerosis. Toll-like receptor 4 (TLR4) also contributes to atherogenesis via transducting inflammatory signals. Herein, our studies focused on characterizing the effect of CRP on tumor necrosis factor α (TNF-α) production and TLR4-related molecular mechanisms in rat vascular smooth muscle cells (VSMCs). The results showed that CRP stimulated VSMCs to secrete TNF-α and enhanced TLR4 expression in a time-concentration-dependent manner. TLR4 knockdown significantly inhibited CRP-induced TNF-α generation, and p38 mitogen-activated protein kinase (MAPK) blocker SB203580 depressed TLR4 expression and TNF-α production initiated by CRP in VSMCs. The data demonstrate that CRP triggers an inflammatory response in rat VSMCs by inducing TNF-α secretion, which is mediated by p38 MAPK–TLR4 signaling pathway.     

Active Uptake of Dendritic Cell-Derived Exovesicles by Epithelial Cells Induces the Release of Inflammatory Mediators through a TNF-α-Mediated Pathway

Dendritic cells (DCs) can release hundreds of membrane vesicles, called exovesicles, which are able to activate resting DCs and distribute antigen. Here, we examined the role of mature DC-derived exovesicles in innate and adaptive immunity, in particular their capacity to activate epithelial cells. Our analysis of exovesicle contents showed that exovesicles contain major histocompatibility complex-II, CD40, and CD83 molecules in addition to tumor necrosis factor (TNF) receptors, TNFRI and TNFRII, and are important carriers of TNF-α. These exovesicles are rapidly internalized by epithelial cells, inducing the release of cytokines and chemokines, but do not transfer an alloantigen-presenting capacity to epithelial cells. Part of this activation appears to involve the TNF-α-mediated pathway, highlighting the key role of DC-derived exovesicles, not only in adaptive immunity, but also in innate immunity by triggering innate immune responses and activating neighboring epithelial cells to release cytokines and chemokines, thereby amplifying the magnitude of the innate immune response.Dendritic cells (DCs) are antigen-presenting cells with a unique ability to induce primary immune responses. They are present in trace amounts in most tissues, but they are particularly abundant and act as sentinels in organs with an environmental interface, such as the skin, the respiratory system, and the gastrointestinal tract. Due to their location, immature dendritic cells are profoundly influenced by the environment and transmit danger signals to cells of the adaptive immune system. The presence of pathogens activates immature dendritic cells and triggers their maturation, resulting in enhanced expression of costimulatory molecules such as CD86 and CD80, and of maturation markers such as CD83. Once activated, DCs migrate to lymph nodes where antigen presentation leads to the maturation and proliferation of specific T-cell clones, which in turn migrate to the injured tissue.¹Depending on their location, DCs are able to release a specific array of cytokines to amplify the innate response. In addition, we would like to suggest that the innate and adaptive immune response may also be amplified through the release of tiny DC-derived microparticles. At least two types of vesicles released from DCs into the extracellular medium have been described. The first type are membrane vesicles, or exovesicles, which are between 0.1 and 1 μm in diameter; they are produced by membrane surface shedding, and released through a process similar to viral budding.^2,3,4 The second type of vesicle is defined as an exosome, ie, microvesicle of endocytic origin, cup-shaped, and ∼0.05 μm in diameter; exosomes are released through exocytosis of multivesicular bodies.^4,5Initially, the secretion of these tiny microparticles was described as a process designed to regulate membrane functions and eliminate unnecessary membrane proteins.⁵ However, exosomes have raised immunological interest because they originate from compartments of the endocytic pathway, which are sites of peptide loading on major histocompatibility complex (MHC) class II molecules. Indeed, both exovesicles and exosomes have been shown to be highly immunogenic, expressing on their surface not only MHC II molecules, but also costimulatory molecules such as CD86,^5,6,7 and specific proteins lacking secretory signals sequence, such as interleukin (IL)-1β.^8,9,10 Recently, we were able to quantify, on a per cell basis, the release of exovesicles from activated DCs; these exovesicles represent the most relevant microparticles released by DCs. Using double vital staining, we demonstrated that exovesicles released from activated DCs can fuse with the membrane of resting DCs, thereby allowing them to present alloantigens to T-lymphocytes.²In the present study, we analyzed the composition and the fate of exovesicles regarding the epithelium. We were able to show that exovesicles from lipopolysaccharide (LPS)-activated DCs are important carriers of tumor necrosis factor (TNF)-α. Using double vital staining, we demonstrated that they are internalized by epithelial cells (ECs), and that this process induces the release of inflammatory mediators such as IL-8, Monocyte chemotactic protein-1 (MCP-1), Macrophage inflammatory protein 1β (MIP-1β), Regulated on Activation, Normal T Cell Expressed and Secreted (RANTES), and TNF-α. Furthermore, we demonstrate that the TNF-α cascade is one of the pathways involved in the activation of these cytokines. In contrast to the well-characterized transfer of alloantigens of exovesicles to heterologous resting DCs, exovesicles in the co-culture with ECs do not transfer an antigen presenting capacity to ECs. Our results demonstrate the potential role of exovesicles not only in adaptive immunity, as a relevant source of antigens fusing with the cytoplasmic membrane of resting DCs, allowing them to present antigens, but also in innate immunity by triggering ECs to release cytokines-chemokines, thereby amplifying the magnitude of the innate immune response.     

Inhibitory Effect of Interleukin-1α-Induced Apoptosis by Polygala Tenuifolia in Hep G2 Cells

Abstract

A human hepatoma cell line, Hep G2 cells are reliable for the study of alcohol-induced hepatotoxicity. In this study, we investigated the effect of an aqueous extract of Polygala tenuifolia Willdenow (Polygalaceae) roots (PTAE) on ethanol (EtOH)-induced cytotoxicity in Hep G2 cells. PTAE (0.01–1 μg/ml) dose-dependently inhibited the EtOH-induced interluekin-1 α (IL-1α) secretion. PTAE (0.01–1 μg/ml) also inhibited the EtOH- and IL-1α-induced cytotoxicity. Furthermore, we found that PTAE inhibited the IL-1α-induced apoptosis of Hep G2 cells. These results suggest that PTAE may prevent the EtOH-induced cytotoxicity through inhibition of the apoptosis of Hep G2 cells.     

Linkage between PTK Signaling Pathway “Crosstalking” and Caspase-3／ CPP32-1ike Proteases Activation in Signaling Transduction of CD4^＋ T Lymphocytes Apoptosis Induced by Superantigen SEB

Exposure of naive murine CD4^+ T lymphocytes to superantigen such as staphylococcal enterotoxin B (SEB) induces a strong proliferative response. Prolonged exposure or subsequent restimulation of the responding T cell population with SEB leads to the apoptotic events of activation-induced cell death (AICD). The signaling mechanism responsible for the AICD is a target of intensive investigation. However, the precise downstream signahng pathways of SEB-induced AICD remains unclear. Our results here show that the sequential activation of caspase-1/ICE-hke and caspase-3/CPP32-hke cysteine proteases probably plays a role in the signaling transduction of SEB-induced AICD, but caspase-3/CPP32-hke proteases activation does not depend on caspase-1-like proteases activation. Herbimycin A, a specific inhibitor of protein tyresine kinases,inhibit caspase-3/CPP32-1ike cysteine proteases activation. However, it does not prevent DNA fragmentation of CD4^+ Tcells apoptosis induced by SEB. These results indicate that protein tyrosine kinases pathway is probably involved in the signaling transduction of CD4^+ T cells apoptosis induced by SEB and “crosstalks” with the pathway of caspase-3/CPP32-1ike proteases activation.