FLIP Nature Reviews Immunology 6, 196-204, 2006

cFLIP regulation of lymphocyte activation and development

Ralph C. Budd1, Wen-Chen Yeh2 and Jürg Tschopp3 About the authors

Cellular caspase-8 (FLICE)-like inhibitory protein (cFLIP) was originally identified as an inhibitor of death-receptor signalling through competition with caspase-8 for recruitment to FAS-associated via death domain (FADD). More recently, it has been determined that both cFLIP and caspase-8 are required for the survival and proliferation of T cells following T-cell-receptor stimulation. This paradoxical finding launched new investigations of how these molecules might connect with signalling pathways that link to cell survival and growth following antigen-receptor activation. As discussed in this Review, insight gained from these studies indicates that cFLIP and caspase-8 form a heterodimer that ultimately links T-cell-receptor signalling to activation of nuclear factor-B through a complex that includes B-cell lymphoma 10 (BCL-10), mucosa-associated-lymphoid-tissue lymphoma-translocation gene 1 (MALT1) and receptor-interacting protein 1 (RIP1).

FLIPは炎症細胞がアポトーシスを起こさないためのkeyとなる蛋白です。主要臓器の炎症病態のFLIPの解析に取り掛かります。

Nature Medicine 11, 1161 - 1162 (2005)

TLRs play good cop, bad cop in the lung

Luke A J O'Neill

オニールもたいしたものだ。

Toll-like receptors act as mediators of injury or repair in the inflamed lung, and the balance depends on the integrity of a component of the extracellular matrix (pages 1173–1179).

現時点ではオニールと結託すると，僕のデータもグレードアップして，かなりすごい論文がいくつも出せそうに思われる。僕の秘策はまだまだあるのだ。だが時間がない。時間がないことに一番困っている。とにかく，瑣末なことを取り除き，仕事だけに専念させていただきたい毎日である。

Nature Medicine 11,1173 - 1179 (2005)

Regulation of lung injury and repair by Toll-like receptors and hyaluronan

僕の実験でも当たり前のことしかデータに出ていないが
Nature Medicineにでるということはすごい。
Alert cell starategyを早くまとめたいものです。

Mechanisms that regulate inflammation and repair after acute lung injury are incompletely understood. The extracellular matrix glycosaminoglycan hyaluronan is produced after tissue injury and impaired clearance results in unremitting inflammation. Here we report that hyaluronan degradation products require MyD88 and both Toll-like receptor (TLR)4 and TLR2 in vitro and in vivo to initiate inflammatory responses in acute lung injury. Hyaluronan fragments isolated from serum of individuals with acute lung injury stimulated macrophage chemokine production in a TLR4- and TLR2-dependent manner. Myd88-/- and Tlr4-/-Tlr2-/- mice showed impaired transepithelial migration of inflammatory cells but decreased survival and enhanced epithelial cell apoptosis after lung injury. Lung epithelial cell–specific overexpression of high-molecular-mass hyaluronan was protective against acute lung injury. Furthermore, epithelial cell–surface hyaluronan was protective against apoptosis, in part, through TLR-dependent basal activation of NF-B. Hyaluronan-TLR2 and hyaluronan-TLR4 interactions provide signals that initiate inflammatory responses, maintain epithelial cell integrity and promote recovery from acute lung injury.

J. Clin. Invest. 115:3378-3384, 2005

Platelets in inflammation and atherogenesis
Meinrad Gawaz, Harald Langer and Andreas E. May

Meinrad Gawaz, Medizinische Klinik III, Eberhard Karls Universität Tübingen, Otfried-Müller-Straße 10, D-72076 Tübingen, Germany. Phone: 49-7071-29-83688; Fax: 49-7071-29-5749; E-mail: meinrad.gawaz@med.uni-tuebingen.de.


Platelet-derived mediators stimulate inflammation
During the adhesion process, platelets become activated and release an arsenal of potent inflammatory and mitogenic substances into the local microenvironment, thereby altering chemotactic, adhesive, and proteolytic properties of ECs (24). These platelet-induced alterations of the endothelial phenotype support chemotaxis, adhesion, and transmigration of monocytes to the site of inflammation (Figure 2).
Released from dense granules, -granules, lysosomes, the canalicular system, or the cytosol, platelets secrete or expose adhesion proteins (e.g., fibrinogen, fibronectin, vWF, thrombospondin, vitronectin, P-selectin, GPIIb/IIIa), growth factors (e.g., PDGF, TGF-ß, EGF, bFGF), chemokines (e.g., RANTES, platelet factor 4 [CXC chemokine ligand 4], epithelial neutrophil-activating protein 78 [CXC chemokine ligand 5]), cytokine-like factors (e.g., IL-1ß, CD40 ligand, ß-thromboglobulin), and coagulation factors (e.g., factor V, factor XI, PAI-1, plasminogen, protein S). These proteins act in a concerted and finely regulated manner to influence widely differing biological functions such as cell adhesion, cell aggregation, chemotaxis, cell survival and proliferation, coagulation, and proteolysis, all of which accelerate inflammatory processes and cell recruitment. For example, IL-1ß has been identified as a major mediator of platelet-induced activation of ECs (25, 26). The IL-1ß activity expressed by platelets appears to be associated with the platelet surface, and coincubation of ECs with thrombin-activated platelets induces IL-1ß–dependent secretion of IL-6 and IL-8 from ECs (26). Furthermore, incubation of cultured ECs with thrombin-stimulated platelets significantly enhances the secretion of endothelial monocyte chemoattractant protein-1 (MCP-1) in an IL-1ß–dependent manner (12). MCP-1 belongs to the CC family of chemokines and is thought to play a key role in the regulation of monocyte recruitment to inflamed tissue and in atherosclerosis (27, 28).

However, platelet IL-1ß does not only modify endothelial release of chemotactic proteins. IL-1ß additionally can increase endothelial expression of adhesion molecules. Surface expression of ICAM-1 and vß3 on ECs is significantly enhanced by activated platelets via IL-1ß (12). Both enhanced chemokine release and upregulation of endothelial adhesion molecules through platelet-derived IL-1ß act in concert and promote neutrophil and monocyte adhesion to the endothelium. IL-1ß–dependent expression of early inflammatory genes, such as MCP-1 or ICAM-1, involves the activation of the transcription factor NF-B. Transient adhesion of platelets to the endothelium initiates degradation of IB and supports activation of NF-B in ECs, thereby inducing NF-B–dependent chemokine gene transcription (29, 30). Likewise, platelet-induced NF-B activation was largely reduced by IL-1ß antagonists, which supports the notion that platelet IL-1ß is the molecular determinant of platelet-dependent activation of the transcription factor. Activation of NF-B involves a cascade of phosphorylation processes. One family of kinases that is involved in NF-B–dependent gene expression is the MAPKs, such as p38 MAPK. In a manner similar to that of recombinant human IL-1ß, activated platelets have the potential to induce phosphorylation of p38 MAPK. Correspondingly, transfection of a dominant-negative p38 mutant significantly reduced platelet-induced MCP-1 secretion in ECs (31).

Once recruited to the vascular wall, platelets may promote inflammation by chemoattraction of leukocytes through mediators such as platelet-activating factor and macrophage inflammatory protein-1, may stimulate smooth muscle cell proliferation (TGF-ß, PDGF, serotonin) (32), and may contribute to matrix degradation by secretion of MMP-2 (33).

A finely regulated functional interaction of platelets with chemokines has also been implicated in atherogenesis (34). Activated platelets can release chemokines and can induce the secretion of chemokines in various cells of the vascular wall; in turn, certain chemokines can enhance platelet aggregation and adhesion in combination with primary agonists and can trigger monocyte recruitment (35). One such candidate for monocyte recruitment is RANTES, which has been shown to trigger monocyte arrest on inflamed and atherosclerotic endothelium (35). Deposition of platelet-derived RANTES induces monocyte recruitment mediated by P-selectin (36, 37). Another platelet-derived chemokine is platelet factor 4 (PF4), the most abundant protein secreted by activated platelets. First, PF4 acts as a chemoattractant for monocytes promoting their differentiation into macrophages (38). Second, PF4 may directly aggravate the atherogenic actions of hypercholesterolemia by promoting the retention of lipoproteins. Sachais and colleagues have recently shown that PF4 can facilitate the retention of LDL on cell surfaces by inhibition of its degradation by the LDL receptor (39). In addition, PF4 markedly enhances the esterification and uptake of oxidized LDL by macrophages (40). The fact that PF4 has been found in human atherosclerotic lesions and was found associated with macrophages in early lesions and with foam cells in more advanced lesions (41) supports the concept that PF4 released from locally activated platelets enters the vessel wall and promotes vascular inflammation and atherogenesis.

Furthermore, release of platelet-derived CD40 ligand (CD40L, CD154) induces inflammatory responses in endothelium. Henn et al. (42) showed that platelets store CD40L in high amounts and release it within seconds after activation in vitro. Ligation of CD40 on ECs by CD40L expressed on the surface of activated platelets increased the release of IL-8 and MCP-1, the principal chemoattractants for neutrophils and monocytes (42). In addition, platelet CD40L enhanced the expression of endothelial adhesion receptors including E-selectin, VCAM-1, and ICAM-1, all molecules that mediate the attachment of neutrophils, monocytes, and lymphocytes to the inflamed vessel wall (42). Moreover, CD40L induces endothelial tissue factor expression (43). Hence, like IL-1ß, CD40L expressed on platelets induces ECs to release chemokines and to express adhesion molecules, thereby generating signals for the recruitment of leukocytes in the process of inflammation. CD40 ligation on ECs, smooth muscle cells, and macrophages initiates the expression and release of matrix-degrading enzymes, the MMPs. These enzymes, which degrade ECM proteins, significantly contribute to destruction and remodeling of inflamed tissue. Activated platelets release MMP-2 during aggregation (33, 44). Furthermore, adhesion of activated platelets to ECs results in generation and secretion of MMP-9 and of the protease receptor urokinase-type plasminogen activator receptor (uPAR) on cultured endothelium (45). The endothelial release of MMP-9 is dependent on both the fibrinogen receptor GPIIb/IIIa and CD40L, since inhibition of either mechanism resulted in reduction of platelet-induced matrix degradation activity of ECs. Moreover, GPIIb/IIIa ligation results in substantial release of CD40L in the absence of any further platelet agonist (45, 46) (Figure 2). These results suggest that the release of platelet-derived proinflammatory mediators like CD40L is dependent on GPIIb/IIIa–mediated adhesion. This mechanism may be pathophysiologically important to localize platelet-induced inflammation of the endothelium at a site of firm platelet-endothelium adhesion.

Dectin-1 J Exp Med. 2003 May 5;197(9):1119-24.

Dectin-1 Mediates the Biological Effects of　β-Glucans

Gordon D. Brown, Jurgen Herre, David L. Williams, Janet A. Willment, Andrew S. J. Marshall, and Siamon Gordon

Abstract
The ability of fungal-derived-glucan particles to induce leukocyte activation and the production of inflammatory mediators, such as tumor necrosis factor α, is a well characterized
phenomenon. Although efforts have been made to understand how these carbohydrate polymers
exert their immunomodulatory effects, the receptors involved in generating these responses　are unknown. Here we show that Dectin-1 mediates the production of TNF-α in response　to zymosan and live fungal pathogens, an activity that occurs at the cell surface andrequires the cytoplasmic tail and immunoreceptor tyrosine activation motif of Dectin-1 as well as Toll-like receptor (TLR)-2 and Myd88. This is the first demonstration that the inflammatory response to pathogens requires recognition by a specific receptor in addition to the TLRs. Furthermore, these studies implicate Dectin-1 in the production of TNF-α in response to fungi, a critical step required for the successful control of these pathogens.

J. Clin. Invest. 2005 115: 2625-2632

Review series

IKK/NF-B signaling: balancing life and death – a new approach to cancer therapy
Jun-Li Luo, Hideaki Kamata and Michael Karin
Laboratory of Gene Regulation and Signal Transduction, Department of Pharmacology and Cancer Center, School of Medicine, UCSD, La Jolla, California, USA.

Address correspondence to: Michael Karin, Laboratory of Gene Regulation and Signal Transduction, Department of Pharmacology and Cancer Center, School of Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA. Phone: (858) 534-1361; Fax: (858) 534-8158; E-mail: karinoffice@ucsd.edu.

IB kinase/NF-B (IKK/NF-B) signaling pathways play critical roles in a variety of physiological and pathological processes. One function of NF-B is promotion of cell survival through induction of target genes, whose products inhibit components of the apoptotic machinery in normal and cancerous cells. NF-B can also prevent programmed necrosis by inducing genes encoding antioxidant proteins. Regardless of mechanism, many cancer cells, of either epithelial or hematopoietic origin, use NF-B to achieve resistance to anticancer drugs, radiation, and death cytokines. Hence, inhibition of IKK-driven NF-B activation offers a strategy for treatment of different malignancies and can convert inflammation-induced tumor growth to inflammation-induced tumor regression.

J. Clin. Invest. 2005 115: 2640-2647

Review

Mitochondria: pharmacological manipulation of cell death
Lisa Bouchier-Hayes, Lydia Lartigue and Donald D. Newmeyer
La Jolla Institute for Allergy and Immunology, Department of Cellular Immunology, San Diego, California, USA.

Address correspondence to: Donald D. Newmeyer, La Jolla Institute for Allergy and Immunology, Department of Cellular Immunology, 10355 Science Center Drive, San Diego, California 92121, USA. Phone: (858) 558-3539; Fax: (858) 558-3526; E-mail: don@liai.org.

Cell death by apoptosis or necrosis is often important in the etiology and treatment of disease. Since mitochondria play important roles in cell death pathways, these organelles are potentially prime targets for therapeutic intervention. Here we discuss the mechanisms through which mitochondria participate in the cell death process and also survey some of the pharmacological approaches that target mitochondria in various ways.

J. Clin. Invest. 115:2277-2286 (2005)

こういう僕にとって自明のことが論文になると複雑な気持ちです。


Research Article

Intensive insulin therapy protects the endothelium of critically ill patients
Lies Langouche1, Ilse Vanhorebeek1, Dirk Vlasselaers1, Sarah Vander Perre1, Pieter J. Wouters1, Kristin Skogstrand2, Troels K. Hansen3 and Greet Van den Berghe1
1Department of Intensive Care Medicine, Katholieke Universiteit Leuven, Leuven, Belgium.
2Department of Clinical Biochemistry, Statens Serum Institut, Copenhagen, Denmark.
3Immunoendocrine Research Unit, Medical Department M, Aarhus University Hospital, Aarhus, Denmark.

Address correspondence to: Greet Van den Berghe, Department of Intensive Care Medicine, Katholieke Universiteit Leuven, Herestraat 49, B-3000 Leuven, Belgium. Phone: 32-16-34-40-21; Fax: 32-16-34-40-15; E-mail: greta.vandenberghe@med.kuleuven.be.

Received for publication April 18, 2005, and accepted in revised form June 7, 2005.

The vascular endothelium controls vasomotor tone and microvascular flow and regulates trafficking of nutrients and biologically active molecules. When endothelial activation is excessive, compromised microcirculation and subsequent cellular hypoxia contribute to the risk of organ failure. We hypothesized that strict blood glucose control with insulin during critical illness protects the endothelium, mediating prevention of organ failure and death. In this preplanned subanalysis of a large, randomized controlled study, intensive insulin therapy lowered circulating levels of ICAM-1 and tended to reduce E-selectin levels in patients with prolonged critical illness, which reflects reduced endothelial activation. This effect was not brought about by altered levels of endothelial stimuli, such as cytokines or VEGF, or by upregulation of eNOS. In contrast, prevention of hyperglycemia by intensive insulin therapy suppressed iNOS gene expression in postmortem liver and skeletal muscle, possibly in part via reduced NF-B activation, and lowered the elevated circulating NO levels in both survivors and nonsurvivors. These effects on the endothelium statistically explained a significant part of the improved patient outcome with intensive insulin therapy. In conclusion, maintaining normoglycemia with intensive insulin therapy during critical illness protects the endothelium, likely in part via inhibition of excessive iNOS-induced NO release, and thereby contributes to prevention of organ failure and death.

僕が5年前に考えていて，いくつかデータを持っているものがこうやってきちんと論文になって発表されると，すごいなあと思うとともに，どことなく悔しいです。しかし研究をするひとだけが知っている大変な苦労があるのだと思います。1日は24時間しかないので，やはり，共同研究者を増やしていくしかないように思っています。

J. Clin. Invest. 2005 115: 2128-2138.

Research Article

PI3K rescues the detrimental effects of chronic Akt activation in the heart during ischemia/reperfusion injury

Tomohisa Nagoshi1, Takashi Matsui1, Takuma Aoyama1, Annarosa Leri2, Piero Anversa2, Ling Li1, Wataru Ogawa3, Federica del Monte1, Judith K. Gwathmey4,5, Luanda Grazette1, Brian Hemmings6, David A. Kass7, Hunter C. Champion7 and Anthony Rosenzweig1
1Program in Cardiovascular Gene Therapy, Cardiovascular Research Center, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA.
2Department of Medicine, New York Medical College, Valhalla, New York, USA.
3Department of Clinical Molecular Medicine, Division of Diabetes, Digestive, and Kidney Diseases, Kobe University Graduate School of Medicine, Kobe, Japan.
4Gwathmey Inc., Cambridge, Massachusetts, USA.
5Harvard Medical School, Boston, Massachusetts, USA.
6Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.
7Division of Cardiology, The Johns Hopkins Hospital, Baltimore, Maryland, USA.

Address correspondence to: Anthony Rosenzweig, Program in Cardiovascular Gene Therapy, Massachusetts General Hospital, 114 16th Street, Room 2600, Charlestown, Massachusetts 02129, USA. Phone: (617) 726-8286; Fax: (617) 724-7387; E-mail: arosenzweig@partners.org.

Received for publication August 18, 2004, and accepted in revised form March 29, 2005.

Acute activation of the serine-threonine kinase Akt is cardioprotective and reduces both infarction and dysfunction after ischemia/reperfusion injury (IRI). However, less is known about the chronic effects of Akt activation in the heart, and, paradoxically, Akt is activated in samples from patients with chronic heart failure. We generated Tg mice with cardiac-specific expression of either activated (myristoylated [myr]) or dominant-negative (dn) Akt and assessed their response to IRI in an ex vivo model. While dn-Akt hearts demonstrated a moderate reduction in functional recovery after IRI, no function was restored in any of the myr-Akt–Tg hearts. Moreover, infarcts were dramatically larger in myr-Akt–Tg hearts. Biochemical analyses demonstrated that chronic Akt activation induces feedback inhibition of PI3K activity through both proteasome-dependent degradation of insulin receptor substrate–1 (IRS-1) and inhibition of transcription of IRS-1 as well as that of IRS-2. To test the functional significance of these signaling changes, we performed in vivo cardiac gene transfer with constitutively active PI3K in myr-Akt–Tg mice. Restoration of PI3K rescued function and reduced injury after IRI. These data demonstrate that PI3K-dependent but Akt-independent effectors are required for full cardioprotection and suggest a mechanism by which chronic Akt activation can become maladaptive.

さすがにすばらしい研究です。

J. Clin. Invest. 2005 115: 2508-2516.

Research Article

Ets-1 is a critical regulator of Ang II-mediated vascular inflammation and remodeling
Yumei Zhan1,2, Courtney Brown1,2, Elizabeth Maynard1,2, Aleksandra Anshelevich1,2, Weihua Ni1,2, I-Cheng Ho3 and Peter Oettgen1,2
1Division of Cardiology, Beth Israel Deaconess Medical Center, and
2Department of Medicine and New England Baptist Bone and Joint Institute, Harvard Institutes of Medicine, Boston, Massachusetts, USA.
3Division of Rheumatology, Allergy, and Immunology, Brigham and Women’s Hospital, Boston, Massachusetts, USA.

Address correspondence to: Peter Oettgen, Harvard Institutes of Medicine, 4 Blackfan Circle, Boston, Massachusetts 02115, USA. Phone: (617) 667-3390; Fax: (617) 975-5299; E-mail: joettgen@caregroup.harvard.edu.

Ang II is a central mediator of vascular inflammation and remodeling. The transcription factor Ets-1 is rapidly induced in vascular smooth muscle and endothelial cells of the mouse thoracic aorta in response to systemic Ang II infusion. Arterial wall thickening, perivascular fibrosis, and cardiac hypertrophy are significantly diminished in Ets1–/– mice compared with control mice in response to Ang II. The induction of 2 known targets of Ets-1, cyclin-dependent kinase inhibitor p21CIP and plasminogen activator inhibitor–1 (PAI-1), by Ang II is markedly blunted in the aorta of Ets1–/– mice compared with wild-type controls. Expression of p21CIP in VSMCs leads to cellular hypertrophy, whereas expression of p21CIP in endothelial cells is associated with cell cycle arrest, apoptosis, and endothelial dysfunction. PAI-1 promotes the development of perivascular fibrosis. We have identified monocyte chemoattractant protein–1 (MCP-1) as a novel target for Ets-1. Expression of MCP-1 is similarly reduced in Ets1–/– mice compared with control mice in response to Ang II, which results in significantly diminished recruitment of T cells and macrophages to the vessel wall. In summary, our results support a critical role for Ets-1 as a transcriptional mediator of vascular inflammation and remodeling in response to Ang II.

J. Clin. Invest. 115:2341-2350 (2005)

Research Article

Organ-specific roles for transcription factor NF-B in reovirus-induced apoptosis and disease
Sean M. O’Donnell1,2, Mark W. Hansberger2,3, Jodi L. Connolly2,3, James D. Chappell2,4, Melissa J. Watson1,2, Janene M. Pierce5, J. Denise Wetzel1,2, Wei Han6, Erik S. Barton2,3, J. Craig Forrest2,3, Tibor Valyi-Nagy2,4, Fiona E. Yull6, Timothy S. Blackwell6, Jeffrey N. Rottman6, Barbara Sherry7 and Terence S. Dermody1,2,3
1Department of Pediatrics,
2Elizabeth B. Lamb Center for Pediatric Research,
3Department of Microbiology and Immunology,
4Department of Pathology,
5Department of Surgery, and
6Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.
7Department of Molecular Biomedical Sciences, North Carolina State University, Raleigh, North Carolina, USA.

Address correspondence to: Terence S. Dermody, Elizabeth B. Lamb Center for Pediatric Research, D7235 Medical Center North, Vanderbilt University School of Medicine, 1161 21st Avenue South, Nashville, Tennessee 37232, USA. Phone: (615) 343-9943; Fax: (615) 343-9723; E-mail: terry.dermody@vanderbilt.edu.

Reovirus induces apoptosis in cultured cells and in vivo. In cell culture models, apoptosis is contingent upon a mechanism involving reovirus-induced activation of transcription factor NF-B complexes containing p50 and p65/RelA subunits. To explore the in vivo role of NF-B in this process, we tested the capacity of reovirus to induce apoptosis in mice lacking a functional nfkb1/p50 gene. The genetic defect had no apparent effect on reovirus replication in the intestine or dissemination to secondary sites of infection. In comparison to what was observed in wild-type controls, apoptosis was significantly diminished in the CNS of p50-null mice following reovirus infection. In sharp contrast, the loss of p50 was associated with massive reovirus-induced apoptosis and uncontrolled reovirus replication in the heart. Levels of IFN-ß mRNA were markedly increased in the hearts of wild-type animals but not p50-null animals infected with reovirus. Treatment of p50-null mice with IFN-ß substantially diminished reovirus replication and apoptosis, which suggests that IFN-ß induction by NF-B protects against reovirus-induced myocarditis. These findings reveal an organ-specific role for NF-B in the regulation of reovirus-induced apoptosis, which modulates encephalitis and myocarditis associated with reovirus infection.

J. Clin. Invest. 114:1531-1537，2004

Review

Regulation of immunity by lysosphingolipids and their G protein–coupled receptors
Edward J. Goetzl1 and Hugh Rosen2
1Departments of Medicine and Microbiology-Immunology, UCSF, San Francisco, California, USA.
2Department of Immunology, The Scripps Research Institute, La Jolla, California, USA.

J. Clin. Invest. 115:1111-1119，2005

Review

Inflammation, stress, and diabetes
Kathryn E. Wellen and Gökhan S. Hotamisligil
Department of Genetics & Complex Diseases, Harvard School of Public Health, Boston, Massachusetts, USA.

面白く読みました。

J. Clin. Invest. 115:1111-1119，2005 インスリン抵抗性

全身性炎症における高血糖の機序・インスリン抵抗性

As discussed above, it is now apparent that obesity is associated with a state of chronic, low-grade inflammation, particularly in white adipose tissue. How do inflammatory cytokines and/or fatty acids mediate insulin resistance? How do the stresses of obesity manifest inside of cells? In recent years, much has been learned about the intracellular signaling pathways activated by inflammatory and stress responses and how these pathways intersect with and inhibit insulin signaling.

Insulin affects cells through binding to its receptor on the surface of insulin-responsive cells. The stimulated insulin receptor phosphorylates itself and several substrates, including members of the insulin receptor substrate (IRS) family, thus initiating downstream signaling events (32, 33). The inhibition of signaling downstream of the insulin receptor is a primary mechanism through which inflammatory signaling leads to insulin resistance. Exposure of cells to TNF- or elevated levels of free fatty acids stimulates inhibitory phosphorylation of serine residues of IRS-1 (34-36). This phosphorylation reduces both tyrosine phosphorylation of IRS-1 in response to insulin and the ability of IRS-1 to associate with the insulin receptor and thereby inhibits downstream signaling and insulin action (35, 37, 38).

Recently it has become clear that inflammatory signaling pathways can also become activated by metabolic stresses originating from inside the cell as well as by extracellular signaling molecules. It has been demonstrated that obesity overloads the functional capacity of the ER and that this ER stress leads to the activation of inflammatory signaling pathways and thus contributes to insulin resistance (39-41). Additionally, increased glucose metabolism can lead to a rise in mitochondrial production of ROS. ROS production is elevated in obesity, which causes enhanced activation of inflammatory pathways (42, 43).

Several serine/threonine kinases are activated by inflammatory or stressful stimuli and contribute to inhibition of insulin signaling, including JNK, inhibitor of NF-B kinase (IKK), and PKC- (44). Again, the activation of these kinases in obesity highlights the overlap of metabolic and immune pathways; these are the same kinases, particularly IKK and JNK, that are activated in the innate immune response by Toll-like receptor (TLR) signaling in response to LPS, peptidoglycan, double-stranded RNA, and other microbial products (45). Hence it is likely that components of TLR signaling pathways will also exhibit strong metabolic activities.

JNK. The 3 members of the JNK group of serine/threonine kinases, JNK-1, -2, and -3, belong to the MAPK family and regulate multiple activities in development and cell function, in large part through their ability to control transcription by phosphorylating activator protein–1 (AP-1) proteins, including c-Jun and JunB (46). JNK has recently emerged as a central metabolic regulator, playing an important role in the development of insulin resistance in obesity (47). In response to stimuli such as ER stress, cytokines, and fatty acids, JNK is activated, whereupon it associates with and phosphorylates IRS-1 on Ser307, impairing insulin action (36, 39, 48). In obesity, JNK activity is elevated in liver, muscle, and fat tissues, and loss of JNK1 prevents the development of insulin resistance and diabetes in both genetic and dietary mouse models of obesity (47). Modulation of hepatic JNK1 in adult animals also produces systemic effects on glucose metabolism, which underscores the importance of this pathway in the liver (49). The contribution of the JNK pathway in adipose, muscle, or other tissues to systemic insulin resistance is currently unclear. In addition, a mutation in JNK-interacting protein–1 (JIP1), a protein that binds JNK and regulates its activity, has been identified in diabetic humans (50). The phenotype of the JIP1 loss-of-function model is very similar to that of JNK1 deficiency in mice, with reduced JNK activity and increased insulin sensitivity (51). Interestingly, the JNK2 isoform plays a significant nonredundant role in atherosclerosis (52), though apparently not in type 2 diabetes. Recent studies in mice demonstrate that JNK inhibition in established diabetes or atherosclerosis might be a viable therapeutic avenue for these diseases in humans (52, 53).

PKC and IKK. Two other inflammatory kinases that play a large role in counteracting insulin action, particularly in response to lipid metabolites, are IKK and PKC-. Lipid infusion has been demonstrated to lead to a rise in levels of intracellular fatty acid metabolites, such as diacylglycerol (DAG) and fatty acyl CoAs. This rise is correlated with activation of PKC- and increased Ser307 phosphorylation of IRS-1 (54). PKC- may impair insulin action by activation of another serine/threonine kinase, IKKß, or JNK (55). Other PKC isoforms have also been reported to be activated by lipids and may also participate in inhibition of insulin signaling (56).

IKKß can impact on insulin signaling through at least 2 pathways. First, it can directly phosphorylate IRS-1 on serine residues (34, 57). Second, it can phosphorylate inhibitor of NF-B (IB), thus activating NF-B, a transcription factor that, among other targets, stimulates production of multiple inflammatory mediators, including TNF- and IL-6 (58). Mice heterozygous for IKKß are partially protected against insulin resistance due to lipid infusion, high-fat diet, or genetic obesity (59, 60). Moreover, inhibition of IKKß in human diabetics by high-dose aspirin treatment also improves insulin signaling, although at this dose, it is not clear whether other kinases are also affected (61). Recent studies have also begun to tease out the importance of IKK in individual tissues or cell types to the development of insulin resistance. Activation of IKK in liver and myeloid cells appears to contribute to obesity-induced insulin resistance, though this pathway may not be as important in muscle (62-64).

Other pathways. In addition to serine/threonine kinase cascades, other pathways contribute to inflammation-induced insulin resistance. For example, at least 3 members of the SOCS family, SOCS1, -3, and -6, have been implicated in cytokine-mediated inhibition of insulin signaling (65-67). These molecules appear to inhibit insulin signaling either by interfering with IRS-1 and IRS-2 tyrosine phosphorylation or by targeting IRS-1 and IRS-2 for proteosomal degradation (65, 68). SOCS3 has also been demonstrated to regulate central leptin action, and both whole body reduction in SOCS3 expression (SOCS3+/–) and neural SOCS3 disruption result in resistance to high-fat diet–induced obesity and insulin resistance (69, 70).

Inflammatory cytokine stimulation can also lead to induction of iNOS. Overproduction of nitric oxide also appears to contribute to impairment of both muscle cell insulin action and ß cell function in obesity (71, 72). Deletion of iNOS prevents impairment of insulin signaling in muscle caused by a high-fat diet (72). Thus, induction of SOCS proteins and iNOS represent 2 additional and potentially important mechanisms that contribute to cytokine-mediated insulin resistance. It is likely that additional mechanisms linking inflammation with insulin resistance remain to be uncovered.

Mol Pharmacol 2005 67: 977-979

Delphine Baetz, James Shaw, and Lorrie A. Kirshenbaum
(Perspective) Nuclear Factor-B Decoys Suppress Endotoxin-Induced Lung Injury
僕の論文の評価と展望を書いています。

Over millions of years, cells have evolved an elaborate defense systems to ward off and destroy foreign invaders. A key step in the host cells' defense repertoire against bacterial or viral infection includes an inflammatory response mediated by certain cells of the immune system. Once activated, such cells (predominantly T lymphocytes) undergo extensive clonal expansion, resulting in the production and liberation of pro-inflammatory factors such as tumor necrosis factor , nitric oxide (NO), and others that underlie the sequelae of the inflammatory process. Though crucial for cell survival, excess or uncontrolled activation of inflammatory pathways has been linked to a number of human pathologies including autoimmune diseases, cardiovascular diseases, such as atherosclerosis and myocarditis, and end-organ failure from endotoxic injury. For example, excessive levels of circulating NO from sustained activation of inducible nitric-oxide synthase (iNOS) is believed to account for the increased vascular permeability and vasodilatory responses associated with sepsis-induced lung injury commonly seen in patients with adult respiratory distress syndrome. Interventions designed to modulate immune cell activation and the inflammatory process more generally may have therapeutic advantage in treating patients with systemic inflammatory diseases.

To test whether functional inactivation of NF-B could suppress endotoxin-induced lung injury, in this issue of Molecular Pharmacology, Matsuda et al. (2005) evaluate the effects of "decoy" `cis'-acting oligonucleotides (ODN) directed against NF-B on inflammatory gene expression and pulmonary function in a cecal-ligation puncture model of sepsis. In this report, the authors delivered ODN in vivo using the hemagglutinating virus of Japan-envelope (HVJ), which had been previously reported as an effective vehicle for delivering small ODN into cells in vivo (Morishita and Kaneda, 2002). The premise behind this technique lies in the ability of the NF-B ODN decoy to suppress simultaneously the expression of several inflammatory genes that contain NF-B binding elements. The authors found that intravenous injection of ODN significantly reduced the increase of NF-B activity during sepsis, as indicated by electromobility shift analysis. Moreover, NF-B decoy markedly reduced the expression levels of iNOS, COX-2, histamine H1-receptor, platelet-activating factor receptor, and bradykinin B1 and B2 receptors in the septic lung tissue. It is noteworthy that animals treated with NF-B ODN displayed an improved outcome with a significant reduction in sepsis-induced lung injury compared with control animals or animals treated with scrambled ODN.

The strength of the ODN decoy technique lies in its ability to compete for the cis-NF-B binding elements, thereby blocking the actions of NF-B. As shown in Fig. 1, the NF-B decoy inactivates NF-B activity by competing with the endogenous NF-B cis elements at the level of the DNA, thereby interfering with NF-B gene transcription and gene activation. Therefore, such a decoy approach has the potential to block the inflammatory response mediated by one or more genetic pathways. Several reports document the utility of a "decoy" strategy to treat disease manifestation. For example, NF-B decoys have been shown to reduce the incidence of myocardial cell and neuronal cell damage after ischemic injury (Morishita et al., 1997; Ueno et al., 2001) and to decrease hypoxia-induced apoptosis of human aortic endothelial cells (Matsushita et al., 2000). Furthermore, in vivo transfection of NF-B decoy has been shown to attenuate the development of autoimmune myocarditis (Yokoseki et al., 2001). This strategy may also be adapted to treat other disease entities involving de-regulated activation of NF-B including proliferative disorders such as cancer (Biswas et al., 2003, 2004).

Although many studies support the use of "decoy" technology to inactivate certain genes or genetic pathways to "treat" a given pathology, several considerations must be kept in mind. First, given that many genes undergo alternative splicing, thereby resulting in gene products with different functions within the cell, the complete inactivation of a given genetic element and/or pathway may have detrimental cellular consequences. Therefore, it may be important to design decoy molecules that could be selectively "turned on" or "turned off" to prevent untoward effects. Second, specificity of the decoy ODN in question may be an issue because overlapping or cryptic sites within DNA may result in nonspecific effects from inactivation of members of one or more gene family. Third, safety issues regarding vector type and administration technique must be considered, especially given that the ultimate goal is to modulate gene expression in humans.

Nevertheless, given that the inflammatory process represents a major cause of morbidity in patients with endotoxin-induced sepsis, the elegant work of Matsuda et al. (2005) provides new compelling evidence that oligonucleotide decoys directed against the transcription factor NF-B may be an effective treatment for averting, or at least attenuating, the inflammatory response at the genetic level.