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B Decoys Suppress Endotoxin-Induced Lung Injury
The Institute of Cardiovascular Sciences, St. Boniface General Hospital Research Centre, and the Department of Physiology, Faculty of Medicine, University of Manitoba, Winnipeg, Manitoba, Canada
Received for publication January 19, 2005.
Accepted for publication January 25, 2005.
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.
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The NF- B Signaling Pathway and Inflammation
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 Top The NF-{kappa}B Signaling... Role NF-{kappa}B in Acute...Decoys as Therapeutic Tools...References |
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B (NF-B) is a prominent nuclear transcription factor suggested to be a central regulator of genes and end- effectors of the host's inflammatory response. NF-B was first identified as a key regulatory molecule required for B-lymphocyte proliferation and maturation (Miyamoto and Verma, 1995
). Since these original observations, it has become appreciated that NF-B is a ubiquitously expressed dimeric transcription factor involved in wide range of biological processes that include inflammation, cell adhesion and cell survival (for review, see Karin, 1999).
NF-B belongs to a family of highly conserved transcription factors that include Rel-A, c-Rel, v-Rel, and Drosophila melanogaster dorsal proteins. The major form of NF-B in cells is a heterodimeric complex composed of 50- and 65-kDa (p50/p65) protein subunits; minor complexes of p50/p50 homodimers have also been reported (Urban and Baeuerle, 1991; Hansen et al., 1992). The p65 subunit is important for gene transcription (Schmitz and Baeuerle, 1991). In unstimulated cells, NF-B is retained as an inactive complex bound to the cytoplasmic inhibitor protein of IB (Baeuerle and Baltimore, 1988; Urban and Baeuerle, 1990; Nolan and Baltimore, 1992; Zabel et al., 1993). A key step in the activation of NF-B involves phosphorylation and degradation of IB (Fig. 1). Insight into the signaling mechanisms that led to IB phosphorylation has revealed a kinase complex composed of IB kinases (IKK) (Scherer et al., 1995; Verma et al., 1995; McKinsey et al., 1996; Van Antwerp and Verma, 1996), IKK, IKK
, and IKK
(Karin, 1999; O'Mahony et al., 2000; Ghosh and Karin, 2002). IKK and IKK are catalytic subunits, whereas IKK is a regulatory subunit (for review, see Karin, 1999; Ghosh and Karin, 2002). It is noteworthy that IKK seems to be crucial for activating NF-B (Li et al., 1999) because it phosphorylates critical serine residues 32 and 36 of IB (Maniatis, 1999), resulting in the ubiquitination and subsequent degradation of IB by the 26S proteasome (for review, see Nabel and Verma, 1993; Karin, 1999). The degradation of IB permits NF-B to translocate to the nucleus and affect gene transcription. Several genes associated with the inflammatory process, including iNOS, cyclo-oxygenase-2, tumor necrosis factor and others, contain putative NF-B binding sites within their promoters, thus highlighting the importance of NF-B as a key regulator of inflammatory gene activation and identifying it as prime candidate for targeted inactivation. Indeed, several strategies to ameliorate or suppress the inflammatory process associated with different pathologies have been directed toward blocking the biological actions of NF-B (Scheinman et al., 1995; Brand et al., 1996; Baeuerle and Baichwal, 1997; Desmet et al., 2004; Tanaka et al., 2005).
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Role NF-B in Acute Endotoxic Lung Injury
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TopThe NF-{kappa}B Signaling...Role NF-{kappa}B in Acute...Decoys as Therapeutic Tools...References |
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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 (Fig. 1).
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.
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Decoys as Therapeutic Tools For Disease |
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TopThe NF-{kappa}B Signaling...Role NF-{kappa}B in Acute...Decoys as Therapeutic Tools...References |
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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.
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Footnotes |
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Please see the related article on page 1018.
ABBREVIATIONS: iNOS, nitric-oxide synthase; NF-B, nuclear factor B; IKK, IB kinases; ODN, oligonucleotides; HVJ, Hemagglutinating virus of Japan.
Address correspondence to: Dr. Lorrie A. Kirshenbaum, Institute of Cardiovascular Sciences, St. Boniface General Hospital Research Centre Rm. 3016, 351 Taché Avenue, Winnipeg, Manitoba, Canada, R2H 2A6. E-mail: lorrie{at}sbrc.ca
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References |
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