Induction of phospholipase A2 gene expression in human hepatoma cells by mediators of the acute phase response.

Serum levels of phospholipase A2 (PLA2) activity have been shown to be elevated in cases of septic shock and rheumatoid arthritis. The cellular origin of serum PLA2, however, is not known. In this report, we demonstrate that human group II PLA2 expression and secretion are induced in hepatoma cells (HepG2) following treatment with interleukin-6 (IL-6), tumor necrosis factor (TNF), and interleukin-1 (IL-1). Of the three cytokines, IL-6 is the most potent. Significant synergy is observed between IL-6 and IL-1 and between IL-6 and TNF, but not between IL-1 and TNF. PLA2 induction does not occur in human YT cells, which are known to have receptors for both IL-1 and IL-6, indicating that the regulatory mechanism involved is cell type-specific. The results of RNA blot analysis indicate that the PLA2 gene is regulated in HepG2 cells at the pretranslational level. Induction of PLA2 synthesis in HepG2 cells in response to these cytokines resembles the induction of the acute phase plasma proteins which are synthesized in cultured hepatocytes and hepatoma cells following exposure to the same cytokines and in liver in response to inflammation and infection. In addition, a putative IL-6-responsive element, which is homologous to a similar element found in several acute phase genes, is present in the 5'-promoter-proximal region of the PLA2 gene. These results suggest that serum PLA2 is synthesized in and secreted from liver cells in response to inflammatory stimuli, mediated primarily by IL-6, and therefore should be classified as an acute phase protein.     

The role of interleukin-6 in development.

Interleukin-6 is a member of a class of hormone-like molecules termed cytokines. The actions of IL-6 are highly pleiotropic. In adults IL-6 functions as a major mediator of inflammatory responses as well as inducing the synthesis of acute phase proteins by the liver following infection or injury. Based on in vitro and in vivo studies IL-6 also has important functions in regulating the development of multiple lineages of hemopoietic cells. It may also be an inflammatory mediator in the central nervous system. Although IL-6 has been found in early mouse embryos, its function has not yet been determined. Its expression by placental trophoblasts and maternal decidua suggests that it has some role in fetal-maternal interactions. Finally, the response of fetal hemopoietic progenitor cells to IL-6 suggests that IL-6 may have a broader action on the expansion and maturation of fetal precursors. New approaches such as those involving the disruption of the IL-6 gene in mice will be needed for a more complete understanding of IL-6's role in embryonic development.     

Changes of hepatic lactoferrin gene expression in two mouse models of the acute phase reaction

Lactoferrin (Ltf), an iron binding glycoprotein, is a pleiotropic molecule whose serum concentration increases under acute phase conditions. The physiological roles of this protein have been well elucidated, but the source and serum regulation of Ltf gene expression have not been investigated in detail as part of the acute phase reaction (APR). In the current work, the changes in hepatic Ltf-gene-expression during turpentine oil- (TO-) or LPS-induced APR were investigated. Ltf was upregulated at both the mRNA and protein levels in the liver of TO- and LPS-treated wild type (WT) mice. The pattern of induction however was different in both animal models indicating distinctive signalling patterns resulting in an acute phase reaction. Cytokines are the core regulators of APR. Among the major cytokines, IL-6 is an important signalling molecule, which also regulates iron homeostasis in response to an inflammatory situation. In this study, the administration of IL-6 induced Ltf gene expression in the liver of WT mice, in murine hepatocytes and in hepa 1-6 cells. Ltf-gene-expression was upregulated also in the liver of TO- and LPS-treated IL-6 knockout (KO) mice. The increase in serum Ltf after LPS injection was greater than after TO-injection both in WT and IL-6-KO mice. To evaluate the contribution of other acute phase cytokines in the regulation of Ltf-gene-expression in the liver, both in vitro and in vivo studies with IL-1β, TNF-α, or IFN-γ were performed. The results demonstrate that TNF-α and IFN-γ also upregulated Ltf-gene-expression, while IL-1β has no role in the regulation of Ltf-gene-expression.     

Contribution of vascular cell-derived cytokines to innate and inflammatory pathways in atherogenesis

Inflammation is a central element of atherogenesis. Innate pathways contribute to vascular inflammation. However, the initial molecular process(es) starting atherogenesis remain elusive. The various risk factors, represented by particular compounds (activators), may cause altered cellular functions in the endothelium (e.g. vascular endothelial cell activation or -dysfunction), in invading cells (e.g. inflammatory mediator production) or in local vessel wall cells (e.g. inflammatory mediators, migration), thereby triggering the innate inflammatory process. The cellular components of innate immunology include granulocytes, natural killer cells and monocytes. Among the molecular innate constituents are innate molecules, such as the toll-like receptors or innate cytokines. Interleukin-1 (IL-1) and IL-6 are among the innate cytokines. Cytokines are potent activators of a great number of cellular functions relevant to maintain or commove homeostasis of the vessel wall. Within the vessel wall, vascular smooth muscle cells (SMCs) can significantly contribute to the cytokine-dependent inflammatory network by: (i) production of cytokines, (ii) response to cytokines and (iii) cytokine-mediated interaction with invading leucocytes. The cytokines IL-1 and IL-6 are involved in SMC-leucocyte interaction. The IL-6 effects are proposed to be mediated by trans-signalling. Dysregulated cellular functions resulting from dysregulated cytokine production may be the cause of cell accumulation, subsequent low-density lipoprotein accumulation and deposition of extracellular matrix (ECM). The deposition of ECM, increased accumulation of leucocytes and altered levels of inflammatory mediators may constitute an 'innate-immunovascular-memory' resulting in an ever-growing response to anew invasion. Thus, SMC-fostered inflammation, promoted by invading innate cells, may be a potent component for development and acceleration of atherosclerosis.     

Immunomodulatory effects of inactivated parapoxvirus ovis (ORF virus) on human peripheral immune cells: induction of cytokine secretion in monocytes and Th1-like cells

Inactivated parapoxvirus ovis (Orf virus; PPVO) recently displayed strong immunostimulating and modulating capacities in several animal models for acute and chronic virus infections through the induction of gamma interferon (IFN-gamma) as a key mediator of antiviral activity. The data presented in this work demonstrate that inactivated PPVO has strong effects on cytokine secretion by human immune cells, including the upregulation of inflammatory and Th1-related cytokines (IFN-gamma, tumor necrosis factor alpha [TNF-alpha], interleukin 6 [IL-6], IL-8, IL-12, and IL-18) as well as anti-inflammatory and Th2-related cytokines (IL-4, IL-10, and IL-1 receptor antagonist [IL-1ra]). Studies on the mechanism of action revealed virus particles to be the effective components of the preparation. The virus particles activate monocytes or other antigen-presenting cells (APC), e.g., plasmacytoid dendritic cells, through signaling over CD14 and a Toll-like receptor and the intracellular presence of certain PPVO-specific components. The activation of monocytes or APC is followed by the release of early proinflammatory cytokines (TNF-alpha, IL-6, and IL-8) as well as the Th1-related cytokines IL-12 and IL-18. Both IL-18 and IL-12 are involved in PPVO-mediated IFN-gamma release by T cells and/or NK cells. The proinflammatory response is accompanied by the induction of anti-inflammatory and Th2-related cytokines (IL-4, IL-10, and IL-1ra), which exert a limiting efffect on the inflammatory response induced by PPVO. We conclude that the induction of a natural immune response with physiologically significant amounts of different cytokines and with antiviral potential might provide advantages over existing antiviral immunotherapies.     

Expression of the phospholipid scramblase (PLSCR) gene family during the acute phase response

Phospholipid scramblase 1 (PLSCR1) is a member of PLSCR gene family that has been implicated in multiple cellular processes including movement of phospholipids, gene regulation, immuno-activation, and cell proliferation/apoptosis. In the present study, we identified PLSCR1 as a positive intracellular acute phase protein that is upregulated by LPS in liver, heart, and adipose tissue, but not skeletal muscle. LPS administration resulted in a marked increase in PLSCR1 mRNA and protein levels in the liver. This stimulation occurred rapidly (within 2 h), and was very sensitive to LPS (half-maximal response at 0.1 microg/mouse). Moreover, two other APR-inducers, zymosan and turpentine, also produced significant increases in PLSCR1 mRNA and protein levels, indicating that PLSCR1 was stimulated in a number of models of the APR. To determine signaling pathways by which LPS stimulated PLSCR1, we examined the effect of proinflammatory cytokines in vitro and in vivo. TNFalpha, IL-1beta, and IL-6 all stimulated PLSCR1 in cultured Hep B3 hepatocytes, whereas only TNFalpha stimulated PLSCR1 in cultured 3T3-L1 adipocytes, suggesting cell type-specific effects of cytokines. Furthermore, the LPS-stimulated increase in liver PLSCR1 mRNA was greatly attenuated by 80% in TNFalpha and IL-1beta receptor null mice as compared to wild-type controls. In contrast, PLSCR1 levels in adipose tissue were induced to a similar extent in TNFalpha and IL-1beta receptor null mice and controls. These results indicate that maximal stimulation of PLSCR1 by LPS in liver required TNFalpha and/or IL-1beta, whereas the stimulation of PLSCR1 in adipose tissue is not dependent on TNFalpha and/or IL-1beta. These data provide evidence that PLSCR1 is a positive intracellular acute phase protein with a tissue-specific mechanism for up-regulation.     

Epidermal growth factor and transforming growth factor-beta differently modulate the acute phase response elicited by interleukin-6 in cultured liver cells from man, rat and mouse.

1. Complex effects of principal inflammatory cytokines (IL-6, IL-1, TNF, IFN-gamma) on acute phase protein synthesis and other metabolic processes in cultured liver cells are briefly reviewed. 2. Molecular properties and biological functions of transforming growth factor-beta and epidermal growth factor are compared. 3. The effects of these factors with respect to both amino acid uptake and acute phase protein synthesis are described in detail. The results are found to be different for rat or mouse hepatocytes and human hepatoma cells.     

IL-6 in autoimmune disease and chronic inflammatory proliferative disease

Interleukin 6 (IL-6), which was originally identified as a B-cell differentiation factor, is now known to be a multifunctional cytokine that regulates the immune response, hematopoiesis, the acute phase response, and inflammation. Deregulation of IL-6 production is implicated in the pathology of several disease processes. The expression of constitutively high levels of IL-6 in transgenic mice results in fatal plasmacytosis, which has been implicated in human multiple myeloma. Increased IL-6 levels are also observed in several diseases, including rheumatoid arthritis (RA), systemic-onset juvenile chronic arthritis (JCA), osteoporosis, and psoriasis. IL-6 is critically involved in experimentally induced autoimmune disease, such as antigen-induced arthritis (AIA), and experimental allergic encephalomyelitis. All these clinical data and animal models suggest that IL-6 plays critical roles in the pathogenesis of autoimmune diseases. Here we review the evidence for the involvement of IL-6 in the pathophysiology of autoimmune diseases and chronic inflammatory proliferative diseases (CIPD) and discuss the possible molecular mechanisms of its involvement.     

Are lipid mediators implicated in the production of pro- and anti-inflammatory cytokines during cardiopulmonary bypass graft with extracorporeal circulation?

In this study the authors assessed the sequential release of lipid mediators (TXB2, PGE2, 6-keto-PGF1alpha, LTB4, LTC4, PAF), pro-inflammatory cytokines (IL-6, IL-8, TNF-alpha) and anti-inflammatory cytokines (IL-4, IL-10) in 17 patients undergoing coronary artery bypass graft (CABG) with extracorporeal circulation (ECC). Time course of appearance of inflammatory mediators revealed the early and transient increase in lipid mediator plasma concentrations (6-keto-PGF1alpha, LTB4, LTC4, PAF) whereas cytokines (IL-6, IL-8, IL-10) were involved only in late pre- and post-operative periods. No variation of TXB2, PGE2, IL-4 and TNF-alpha levels were found. No correlation was documented between the levels of lipid mediators and pro- or anti-inflammatory cytokines suggesting that lipidic compounds are not implicated in the genesis of cytokines which appear much later involved. Despite the common use of high doses of aprotinin (a non-specific enzyme inhibitor) in hope to abrogate the inflammatory response to cardiopulmonary bypass procedure, this study reports the persistent release of several inflammatory compounds that might be involved in the post-CABG multiple organ failure syndromes.     

Phorbol ester modulates interleukin 6- and interleukin 1-regulated expression of acute phase plasma proteins in hepatoma cells

Interleukin 6 (IL 6) and interleukin 1 (IL-1) regulate the expression of acute phase plasma proteins in rat and human hepatoma cells. Phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), partially mimics the stimulatory effect of IL-6 but reduces that effect of IL-1. TPA and IL-6 act synergistically. These regulatory properties of TPA are also manifested in HepG2 cells transiently transfected with an indicator gene construct carrying the IL-1/IL-6 regulatory enhancer element of the rat alpha 1-acid glycoprotein gene. IL-6 and IL-1 act independently of TPA-inducible kinase C, and of changes in intracellular Ca2+ concentrations. However, prolonged pretreatment of HepG2 cells with TPA results in a drastically reduced cytokine response that is proportional to the loss of cell surface binding activity for the cytokine. These data suggest that hormones activating protein kinase C probably play a contributing role in stimulating the expression of acute phase plasma protein genes but they may be crucial in controlling the responsiveness of liver cells to inflammatory cytokines during subsequent stages of the hepatic acute phase reaction.     

Cytokine-induced gene expression of a neutrophil chemotactic factor/IL-8 in human hepatocytes

The liver participates in inflammation via the elaboration of acute phase proteins from hepatocytes in response to IL-1, TNF-alpha, and IL-6/INF-beta 2/hepatocyte-stimulating factor. In addition, some inflammatory states of the liver are characterized by leukocyte infiltrates. Here we demonstrate that human hepatocyte lines are capable of expressing mRNA and biologic activity for a neutrophil chemotactic factor (NCF)/IL-8 in response to the inflammatory mediators IL-1 alpha, IL-1 beta, and TNF. Two human hepatoma cell lines (SK-Hep and Hep-G2) displayed a time- and dose-dependent increase in steady state levels of NCF/IL-8 mRNA and secretion of chemotactic activity in response to TNF and IL-1. Neutralizing antibody to NCF/IL-8 inhibited hepatocyte-derived chemotactic activity by 88%. In contrast to IL-1 and TNF, hepatocytes did not respond to LPS or IL-6 within the time and dose parameters used above. Although the expression of NCF/IL-8 mRNA (1.8 kb) was first detectable between 1 and 2 h poststimulation, significant chemotactic bioactivity was not observed until about 4 h. Heat-inactivated (100 degrees C, 30 min) cytokine failed to induced NCF/IL-8 mRNA synthesis, and cotreatment of cells with cytokine and cycloheximide super-induced NCF/IL-8 mRNA while inhibiting production of bioactivity. Thus, NCF/IL-8 expression is a primary induction phenomenon. Our data demonstrate the stimulus specific induction of NCF/IL-8 in hepatocytes and suggest that cytokine cell-to-cell communication circuits may be important in neutrophil-mediated inflammatory processes in the liver.     

CCAAT enhancer-binding protein alpha is required for interleukin-6 receptor alpha signaling in newborn hepatocytes

The acute phase response is an evolutionarily conserved response of the liver to inflammatory stimuli, which aids the body in host defense and homeostasis. We have previously reported that CCAAT enhancer-binding protein alpha (C/EBPalpha) is required for the induction of acute phase protein (APP) genes in newborn mice in response to lipopolysaccharide. In this paper, we describe a mechanism by which C/EBPalpha knock-out mice are unable to induce APP gene expression in response to inflammatory stimuli. We demonstrate that the lack of acute phase response in C/EBPalpha knock-out mice is because of a hepatocyte autonomous defect. C/EBPalpha knock-out hepatocytes do not activate STAT3 in response to recombinant interleukin (IL)-6, indicating a defect in the IL-6 pathway. C/EBPalpha knock-out hepatocytes also do not show activation of other IL-6 receptor (IL-6R)-mediated Janus kinase substrates, gp130, SHP-2, and Tyk2. Further examination of the IL-6 pathway demonstrated that C/EBPalpha knock-out hepatocytes have decreased IL-6Ralpha protein levels caused, in part, by reduced protein stability. However, other components of the IL-6 pathway are intact, as demonstrated by rescue of STAT3 activation and APP gene induction with recombinant-soluble IL-6R linked to IL-6 cytokine (Hyper-IL-6) or with another gp130 signaling cytokine, Oncostatin M. In conclusion, C/EBPalpha is required for the proper regulation of IL-6Ralpha protein in hepatocytes resulting in a lack of acute phase protein gene induction in newborn C/EBPalpha null mice in response to lipopolysaccharide or cytokines.     

Effects of cytokine combinations on acute phase protein production in two human hepatoma cell lines.

We evaluated the effects of binary combinations of four cytokines on production of the positive acute phase proteins alpha-1 antichymotrypsin, haptoglobin and fibrinogen, and the negative acute phase proteins albumin and alpha-fetoprotein (AFP) in two human hepatoma cell lines. The effects of the cytokine combinations on the five proteins varied; each protein exhibited a unique and specific pattern of response to the cytokine combinations. In Hep G2 cells, antichymotrypsin was induced by all four cytokines, IL-6, IL-1, TNF-alpha, and transforming growth factor beta 1 alone, and their effects in binary combinations could be attributed to additive or minimally synergistic interactions. Fibrinogen was induced only by IL-6 and this induction was inhibited by IL-1 alpha, TNF-alpha or transforming growth factor beta 1. Haptoglobin was also induced only by IL-6, but TNF-alpha was the only cytokine that inhibited this induction at all concentrations of IL-6. Each of the four cytokines alone down regulated production of AFP and albumin. However, binary combinations of the four cytokines were simply additive, for the most part, in inhibiting AFP production, whereas the inhibitory effects of combinations of cytokines on albumin production differed significantly from simple additive effects. These observations, taken together with studies of effects of cytokine combinations on other acute phase proteins, indicate that the various acute phase proteins respond differently to different combinations of cytokines and that the potential exists for highly specific regulation of synthesis of individual plasma proteins by cytokine interactions. These findings imply that the acute phase response in vivo represents the integrated sum of multiple, separately regulated changes in gene expression.     

白细胞介素-6在酒精性肝病中的作用

酒精性肝病(alcoholic liver disease,ALD)是由于长期过量饮酒导致肝的内部组织发生炎症损伤的慢性肝病.乙醇及其衍生物在代谢过程中直接或间接诱导引起的肝炎症反应可能是ALD发病的重要机制.然而,该过程内在的细胞分子机制尚不明确.最新研究发现,白细胞介素-6(interleukin-6,IL-6)对...