NF-kappaB Decoy Research Articles

Effects of decoy molecules targeting NF-kappaB transcription factors in Cystic fibrosis IB3–1 cells

One of the clinical features of cystic fibrosis (CF) is a deep inflammatory process, which is characterized by production and release of cytokines and chemokines, among which interleukin 8 (IL-8) represents one of the most important. Accordingly, there is a growing interest in developing therapies against CF to reduce the excessive inflammatory response in the airways of CF patients. Since transcription factor NF-kappaB plays a critical role in IL-8 expression, the transcription factor decoy (TFD) strategy might be of interest. In order to demonstrate that TFD against NF-kappaB interferes with the NF-kappaB pathway we proved, by chromatin immunoprecipitation (ChIP) that treatment with TFD oligodeoxyribonucleotides of cystic fibrosis IB3–1 cells infected with Pseudomonas aeruginosa leads to a decrease occupancy of the Il-8 gene promoter by NF-kappaB factors. In order to develop more stable therapeutic molecules, peptide nucleic acids (PNAs) based agents were considered. In this respect PNA-DNA-PNA (PDP) chimeras are molecules of great interest from several points of view: (1) they can be complexed with liposomes and microspheres; (2) they are resistant to DNases, serum and cytoplasmic extracts; (3) they are potent decoy molecules. By using electrophoretic mobility shift assay and RT-PCR analysis we have demonstrated that (1) the effects of PDP/PDP NF-kappaB decoy chimera on accumulation of pro-inflammatory mRNAs in P.aeruginosa infected IB3–1 cells reproduce that of decoy oligonucleotides; in particular (2) the PDP/PDP chimera is a strong inhibitor of IL-8 gene expression; (3) the effect of PDP/PDP chimeras, unlike those of ODN-based decoys, are observed even in the absence of protection with lipofectamine. These informations are of great impact, in our opinion, for the development of stable molecules to be used in non-viral gene therapy of cystic fibrosis.

Preconditioning Promotes Survival and Angiomyogenic Potential of Mesenchymal Stem Cells in the Infarcted Heart via NF-κB Signaling

We proposed that pharmacological manipulation of mesenchymal stem cells (MSCs) with diazoxide enhanced their survival and regenerative potential via NFkappaB regulation. MSCs preconditioned ((PC)MSCs) with diazoxide and later subjected to oxidant stress with 100 micromol/L H(2)O(2) either immediately or after 24 h exhibited higher survival (p < 0.01 vs nonpreconditioned MSCs; (Non-PC)MSCs) with concomitantly increased phosphorylation of PI3K, Akt, GSK3beta (cytoplasmic), and NF-kappaB (p65) (nuclear). Akt kinase activity was determined as a function of GSK3beta activity. Pretreatment of (PC)MSCs with Wortmannin (Wt), NEMO-binding domain (NBD), or NF-kappaB (p50) siRNA abolished NF-kappaB (p65) activity. Preconditioning increased NF-kappaB-dependent elevation of secretable growth factors associated with their paracrine effects. Inhibition of PI3K activity with Wt reduced (PC)MSCs viability at both early and 24 h time-points. However, inhibition of NF-kappaB reduced viability of (PC)MSCs only at 24 h time-point. For in vivo studies, DMEM without cells (group-1) or containing 1 x 10(6) male (Non-PC)MSCs (group-2), (PC)MSCs (group-3), (PC)MSCs pretreated with Wortmannin (group-4) or NF-kappaB decoy (group-5) were transplanted in a female rat model of acute myocardial infarction. Group-3 showed highest cell survival and growth factor expression, increased angiomyogenesis, and functional improvement. We conclude that activation of NF-kappaB by preconditioning promoted (PC)MSCs survival and angiomyogenic potential in the infarcted heart.

Radial shock waves effectively introduced NF‐kappa B decoy into rat achilles tendon cells in vitro

The purpose of this study was to test if radial shock waves could enhance the introduction of nuclear factor-kappa B (NF-kappaB) decoy oligodeoxynucleotides, which is reported to markedly inhibit NF-kappaB activation and suppress pro-inflammatory cytokine gene expression, using rat Achilles tendon cells. In the presence of NF-kappaB decoy labeled with or without fluorescein isothiocyanate (FITC) in culture media, radial shock waves were applied to the tendon cells in variable conditions and cultivated for 24 h. The transfection rate was assessed by counting FITC-positive cells, and IL-1-induced NF-kappaB activation in the cells was assessed. Radial shock waves significantly enhanced introduction of NF-kappaB decoy-FITC into the tendon cells. IL-1-induced NF-kappaB activation was significantly inhibited by pretreatment of the cells with NF-kappaB decoy combined with radial shock wave exposure. The present study demonstrated the effectiveness of radial shock waves on introduction of NF-kappaB decoy into tendon cells. Radial shock wave treatment combined with local NF-kappaB decoy administration could be a novel therapeutic strategy for chronic tendinopathy.

Up-Regulation of Histamine H4Receptors Contributes to Splenic Apoptosis in Septic Mice: Counteraction of the Antiapoptotic Action of Nuclear Factor-κB

The histamine H(4) receptor is the most recently identified receptor and is considered to play a role in a variety of inflammatory diseases. Histamine levels in the plasma are known to be elevated in animal models of sepsis and in septic patients. The aim of this study was to test the hypothesis that the H(4) receptor may play a significant role in the pathophysiology of sepsis. Polymicrobial sepsis was induced by cecal ligation and puncture in BALB/c mice. Although the H(4) receptor gene was undetectable in normal peripheral key organs, with the exception of the spleen, the expression levels of this gene were highly up-regulated in all those organs of septic mice. In vivo transfection of nuclear factor-kappaB (NF-kappaB) decoy oligodeoxynucleotide, but not of its scrambled form, resulted in a great inhibition of sepsis-induced overexpression of the H(4) receptor gene. In septic mice, marked increases in caspase-3 activation and follicular lymphocyte apoptosis in spleens were strongly suppressed by systemic treatment with synthetic small interfering RNA (siRNA) targeted to the H(4) receptor. This was associated with the up-regulation of a number of antiapoptotic proteins. These antiapoptotic effects of H(4) receptor siRNA treatment were all inhibited by further application of NF-kappaB decoy oligonucleotide. Our results suggest that superinduction of the histamine H(4) receptor gene in peripheral key organs, including the spleen, that is promoted by sepsis is transcriptionally controlled by NF-kappaB, whereas stimulation of this receptor is involved in the development of sepsis-induced splenic apoptosis through counteraction of the antiapoptotic action of NF-kappaB.

NF-κB activation is required for apoptosis in fibrocystin/polyductin-depleted kidney epithelial cells

Autosomal recessive polycystic kidney disease (ARPKD) is caused by mutations in PKHD1, a gene encoding fibrocystin/polyductin (FC1), a membrane-associated receptor-like protein involved in the regulation of tubular cell adhesion, proliferation and apoptosis. Although it is generally accepted that apoptosis is implicated in ARPKD, the question of whether increased apoptosis is a normal response to abnormal cell proliferation or, instead, it is a primary event, is still subject to debate. In support of the latter hypothesis, we hereby provide evidence that apoptosis occurs in the absence of hyper-proliferation of FC1-depleted kidney cells. In fact, a decrease in cell proliferation, with a concomitant increase in apoptotic index and caspase-3 activity was observed in response to FC1-depletion by PKHD1 siRNA silencing in HEK293 and 4/5 tubular cells. FC1-depletion also induced reduction in ERK1/2 kinase activation, upregulation of the pro-apoptotic protein p53 and activation of NF-kappaB, a transcription factor which reduces apoptosis in many organs and tissues. Interestingly, selective inactivation of NF-kappaB using either an NF-kappaB decoy or parthenolide, a blocker of IKK-dependent NF-kappaB activation, reduced, rather then increased, apoptosis and p53 levels in FC1-depleted cells. Therefore, the proapoptotic function of NF-kappaB during cell death by FC1-depletion in kidney cells is evident.

Ischaemia/reperfusion induced cardiac stem cell homing to the injured myocardium by stimulating stem cell factor expression via NF‐κB pathway

Ischaemia/reperfusion (I/R) is a major cause of heart failure. Recently cardiac stem cells (CSCs) were proposed as the most appropriate cell type for heart disease therapy. However, it is still unclear whether I/R can stimulate the CSCs homing to the injured myocardium. Male Sprague-Dawley rats were subjected to a 30-min ischaemia followed by reperfusion of different intervals. RT-PCR, western blotting and immunohistochemistry were performed to detect stem cell factor (SCF) expression at mRNA and protein levels respectively. Activation of nuclear factor-kappaB (NF-kappaB) was determined by electrophoretic mobility shift assay. To assess the homing of CSCs in vivo, BrdU-labelled CSCs were injected into AV-groove before induction of ischaemia and examined by immunofluorescent staining in the injured myocardium after I/R. From day 3 to day 6 after reperfusion, the accumulation of CSCs was significantly elevated in the injured area, which was matched with the increased SCF expression during I/R. Pretreatment of rats with NF-kappaB inhibitor, N-acetyl-L-cysteine (NAC) not only suppressed NF-kappaB activation induced by I/R but also attenuated SCF expression. Further analysis revealed that I/R induced phosphorylation of IkappaBalpha after 15 min of reperfusion, and the raised phosphor-IkappaBalpha returned to the basal level at 2 h of reperfusion. In simulated I/R(SI/R) in vitro, it enhanced NF-kappaB activation and SCF expression in cultured neonatal rat cardiomyocytes, which was markedly inhibited by NF-kappaB decoy oligodeoxynucleotide or NAC. Taken together, our results demonstrated that I/R induced CSCs homing to the injured myocardium by stimulating myocardial SCF expression via activation of NF-kappaB.

Nuclear factor-kappa B decoy suppresses nerve injury and improves mechanical allodynia and thermal hyperalgesia in a rat lumbar disc herniation model

Nuclear factor-kappa B (NF-kappaB) is a gene transcriptional regulator of inflammatory cytokines. We investigated the transduction efficiency of NF-kappaB decoy to dorsal root ganglion (DRG), as well as the decrease in nerve injury, mechanical allodynia, and thermal hyperalgesia in a rat lumbar disc herniation model. Forty rats were used in this study. NF-kappaB decoy-fluorescein isothiocyanate (FITC) was injected intrathecally at the L5 level in five rats, and its transduction efficiency into DRG measured. In another 30 rats, mechanical pressure was placed on the DRG at the L5 level and nucleus pulposus harvested from the rat coccygeal disc was transplanted on the DRG. Rats were classified into three groups of ten animals each: a herniation + decoy group, a herniation + oligo group, and a herniation only group. For behavioral testing, mechanical allodynia and thermal hyperalgesia were evaluated. In 15 of the herniation rats, their left L5 DRGs were resected, and the expression of activating transcription factor 3 (ATF-3) and calcitonin gene-related peptide (CGRP) was evaluated immunohistochemically compared to five controls. The total transduction efficiency of NF-kappaB decoy-FITC in DRG neurons was 10.8% in vivo. The expression of CGRP and ATF-3 was significantly lower in the herniation + decoy group than in the other herniation groups. Mechanical allodynia and thermal hyperalgesia were significantly suppressed in the herniation + decoy group. NF-kappaB decoy was transduced into DRGs in vivo. NF-kappaB decoy may be useful as a target for clarifying the mechanism of sciatica caused by lumbar disc herniation.

NF-κB Decoy Oligonucleotides Suppress RANTES Expression and Monocyte Chemotactic Activity via NF-κB Inactivation in Stromal Cells of Ectopic Endometrium

The nuclear factor kappa B (NF-kappaB) pathway is a critical mediator of regulated on activation, normal T cell expressed and secreted (RANTES) gene regulation and therefore represents a potential target for therapy of endometriosis-associated symptoms. The objective of this study was to investigate the effect of NF-kappaB decoy oligonucleotides (ODNs) on NF-kappaB activation, RANTES expression, and monocyte chemotactic activity in ectopic endometrial stromal cells in vitro. A specific sandwich enzyme-linked immunosorbent assay (ELISA) was used to quantify RANTES expression in ectopic and normal endometrial stromal cells stimulated by interleukin (IL)-1beta. Four hours after transfection of NF-kappaB decoy ODNs, 10 ng/ml IL-1beta was added to induce the ectopic endometrial stromal cells to secrete RANTES. The NF-kappaB activation, RANTES expression, and monocyte chemotactic activity in ectopic endometrial stromal cells were respectively evaluated by electrophoretic mobility shift assay, ELISA, and Boyden chambers. IL-1beta induced significantly higher levels (P < 0.05) of RANTES expression in a time-dependent manner in ectopic endometrial stromal cells compared with IL-1beta-untreated ectopic and normal endometrial stromal cells. The RANTES accounts for the majority (68%) of the monocyte chemotactic activity in conditioned media of ectopic endometrial stromal cells. In vitro transfection of NF-kappaB decoy ODNs dramatically decreased (P < 0.05) the NF-kappaB activation, RANTES expression, and monocyte chemotactic activity in IL-1beta-induced ectopic endometrial stromal cells. NF-kappaB decoy ODNs may exert anti-inflammatory effects in ectopic endometrial stromal cells via the suppression of NF-kappaB activation, RANTES expression, and monocyte chemotactic activity.

Application of NF.KAPPA.B inhibitor for arthritis

Recent progress in DNA technologies has provided the strategies to regulate the transcription of disease-related genes in vivo using antisense oligodeoxynucleotide (ODN). Transfection of cis-element double-stranded oligodeoxynucleotides (decoy ODNs) has been reported as a new therapeutic tool of anti-gene strategies for gene therapy. In the field of arthritis, decoy ODNs strategies have been significant therapeutic potential. The concept of regulation the disease related gene expression at the level of transcriptional factor may be more therapeutic effects compared with monotherapy in arthritis. Injection of NFkappaB decoy ODN into the affected joint resulted in marked suppression of joint destruction in CIA models. In vitro studies demonstrated that the inhibitory effect on inflammatory cytokines and matrix metalloproteinase production from stimulated synovial cells derived from rheumatoid arthritis patients. NFkappaB decoy ODN inhibited induction of osteoclasts and bone resorption ability. Parthenolide is one of the main sesquiterpense lactones responsible for the bioactivities of feverfew and recently reported to inhibit NFkappaB activation. Parthenolide has ameliorated the severity of joint destruction in experimental animal model. Based upon these findings, NFkappaB may be one of important therapeutic target for arthritis.

Development and Application of Glycosylated Particulate Carriers for Delivery of Nucleic Acid Medicine

Recently several systems including viral and non-viral carriers that can be used to transfer foreign genetic material into cells have been developed with the aim of enhancing gene transfer in vivo. Non-viral vectors are relatively easy to produce in clinically relevant quantities, and associated with fewer safety concerns. Furthermore, synthetic non-viral vectors provide flexibility in formulation design and can be tailored to interact efficiency with DNA cargo and the specific route of vector injection, and can enhance delivery to specific tissues or cells through incorporation of a targeting ligand. Applying cell-specific targeting technology to liposomes would improve in vivo gene transfection efficacy and reduce any unexpected side-effects. Among the various receptors, asialoglycoprotein receptors and mannose receptors are the most promising for gene targeting since they exhibit high affinity and are rapidly internalized. Receptor-mediated delivery systems are able to introduce foreign DNA into specific cell types in vivo. Our group succeeded in the development of glycosylated cationic liposomes for cell-selective targeting of plasmid DNA, siRNA, and NFkappaB decoy based on the optimization of physicochemical properties of glycosylated liposome complex.

An initial case of suppressed restenosis with nuclear factor‐kappa B decoy transfection after percutaneous coronary intervention

Nuclear factor-kappa B (NF-kappaB) is well known for playing a pivotal role in restenosis after percutaneous coronary intervention (PCI). This is the first report to demonstrate an effect of NF-kappaB decoy oligodeoxynucleotides (ODN) to prevent restenosis after PCI after a 4-year observation using a coronary computed tomography (CT) scan. We showed that the decoy treatment suppressed neointimal formation after stent implantation compared to that in the same artery. Thus, for the first time, we demonstrate the clinical usefulness of the CT scan to reveal the effects of NF-kappaB decoy ODN transfer after PCI.

Transcription Factor Oligodeoxynucleotides to NF-κB Inhibit Transcription of IL-8 in Bronchial Cells

Chronic pulmonary inflammation in patients affected by cystic fibrosis (CF) is characterized by massive bronchial infiltrates of neutrophils, which is sustained by the interaction of pathogens (e.g., Pseudomonas aeruginosa) with surface bronchial cells. To explore new treatment options focused on the reduction of neutrophil chemotaxis, we applied the transcription factor (TF) decoy approach, based on the intracellular delivery of double-stranded oligodeoxynucleotides (ODNs) causing inhibition of the binding of TF-related proteins to the different consensus sequences in the promoter of specific genes. In CF bronchial IB3-1 cells, P. aeruginosa induced transcription of the neutrophil chemokines IL-8 and GRO-gamma, of the adhesion molecule intercellular adhesion molecule (ICAM)-1, and of the cytokines IL-1beta and IL-6. Since consensus sequences for the TF, NF-kappaB, are contained in the promoters of all these genes, IB3-1, CuFi-1, Beas-2B, and CaLu-3 cells were transfected with double-stranded TF "decoy" ODNs mimicking different NF-kappaB consensus sequences. IL-8 NF-kappaB decoy ODN partially inhibited the P. aeruginosa-dependent transcription of IL-8, GRO-gamma, and IL-6, whereas decoy ODNs to both HIV-1 long terminal repeat and Igk produced a strong, 80 to 85% inhibition of transcription of IL-8, without reducing that of GRO-gamma, ICAM-1, IL-1beta, and IL-6. In conclusion, intracellular delivery of "decoy" molecules aimed to compete with the TF, NF-kappaB, is a promising strategy to obtain inhibition of IL-8 gene transcription.

Development of Cell-selective Targeting Systems of NFκB Decoy for Inflammation Therapy

NFkappaB regulate several inflammatory related molecules and evoke immune and inflammatory response by several stimuli, therefore inhibition of NFkappaB activation would be a novel therapeutic strategy. To date, there are many conventional drugs including nonsteroldal or steroldal anti-inflammatory drugs or immune suppressors etc. were known to inhibit NFkappaB activation, however, several side effects were also reported. Recently, double stranded oligonucleotide including NFkappaB binding sequence, called NFkappaB decoy, was developed to prevent NFkappaB activation, which is powerful tool in a new class of anti-gene strategy for molecular therapy with low side effect. However, NFkappaB decoy is easily degraded by nuclease and rapidly excreted to urine, therefore it is necessary to develop carrier for NFkappaB decoy therapy. Here, we shall review delivery system for NFkappaB decoy and introduce our cell-selective delivery system for NFkappaB decoy using sugar decorated cationic liposomes.

Marked regression of liver metastasis by combined therapy of ultrasound‐mediated NFkB‐decoy transfer and transportal injection of paclitaxel, in mouse

Nuclear factor-kappaB (NF kappaB) plays a pivotal role in cancer progression. In this study, we developed a decoy cis-element oligo-deoxyribonucleic acid against NF kappaB-binding site (NF kappaB-decoy), which effectively inhibits NF kappaB activity, and tested the effect of combined therapy comprising local transfection of NF kappaB-decoy into the liver and transportal injection of paclitaxel on cancer growth and metastasis using an orthotopic murine model of colon cancer liver metastasis. For NF kappaB-decoy transfection, we employed a novel approach using ultrasound exposure with an echocardiographic contrast agent, Optison. We examined the influence of NF kappaB-decoy transfer on susceptibility to paclitaxel in cancer cells and the mechanism involved using several in vitro analysis systems. We then studied the in vivo effect of combined NF kappaB-decoy transfer and paclitaxel in preventing cancer progression using a murine model of liver metastasis created by splenic injection of a human colon cancer cell line, HT29. In vitro experiments, including MTT-assay, fluorescence-activated cell sorter and cDNA array analysis, revealed that NF kappaB-decoy transfer significantly increased the susceptibility of cancer cells to paclitaxel, and that decreased expression of anti-apoptotic genes along with increased expression of genes relevant to the apoptosis-promotor may be involved. In vivo experiments showed that local transfection of NF kappaB-decoy into the liver followed by portal injection of paclitaxel effectively induced cancer cell apoptosis in the liver metastasis, and significantly prolonged animal survival compared to controls, without notable side effects. In conclusion, a combination of local NF kappaB-decoy transfer into the liver and transportal injection of paclitaxel may be a safe and effective new therapy for liver metastasis.

Increase in nuclease resistance and incorporation of NF‐κB decoy oligodeoxynucleotides by modification of the 3′‐terminus

For the development of molecular therapy based on oligodeoxynucleotides (ODN), ODN have to be stable against nucleases and be specific to the target transcription factor. To decrease non-specific binding and degradation from the 3'-terminus of ODN, we designed partially annealed ODN by binding the extremities of two single strands, resulting in a ribbon-shaped ODN, so called ribbon-type decoy ODN (R-ODN). We evaluated the efficiency in the process of enzymatic ligation of R-ODN, the binding activity to nuclear factor-kappaB (NF-kappaB), and the stability against Exonuclease III and nucleases present in serum. The functional activity of R-ODN to inhibit NF-kappaB in vitro was evaluated in human aortic smooth muscle cells (VSMC): TNF-alpha-induced proliferation rate and MMP-9 expression were assessed after R-ODN transfection. Although R-ODN have a phosphodiester backbone, their physical conformation was designed to provide nuclease resistance without interfering with their binding activity. As expected, R-ODN showed more resistance to exonucleases and stability in 100% serum than non-modified decoy ODN (N-ODN). Importantly, the R-ODN construction did not interfere with its binding activity to NF-kappaB, similar to N-ODN. Transfection of R-ODN significantly inhibited the expression of MMP-9 induced by TNF-alpha in VSMC as assessed by real-time polymerase chain reaction (PCR), and R-ODN also inhibited the proliferation of VSMC induced by TNF-alpha (10 ng/ml), similar to phosphorothioate decoy ODN. Overall, the development of ribbon NF-kappaB decoy ODN could provide a useful tool for basic and clinical research.

In Vivo Reduction of the Nuclear Factor-κB Activity Using Synthetic cis-Element Decoy Oligonucleotides Suppresses Intimal Hyperplasia in the Injured Carotid Arteries in Rabbits

Nuclear factor-kappaB (NF-kappaB) plays a critical role in inflammation-related reactions, and is also found in the injured arterial wall. The purpose of this study was to introduce synthetic double-stranded cis-element "decoy" oligonucleotides (ODNs) into the arterial wall using the hemagglutinating virus of Japan (HVJ) liposome, and to investigate the inhibitory potential of decoy ODN against balloon injury-induced intimal hyperplasia by reducing NF-kappaB activity. Fluorescein isothiocyanate (FITC)-labeled decoy ODNs using the HVJ-liposome method were tranfected in balloon-injured rabbit carotid arteries. We then performed electrophoretic mobility shift assay to examine NF-kappaB activity using balloon-injured arteries, and we introduced NF-kappaB decoy into balloon-injured arteries. Transfection of FITC-labeled decoy ODNs by using the HVJ-liposome method demonstrated highly efficient protein expression with diffuse, frequent, and widespread nuclear signals over the entire medial layer, while the same amount of naked ODNs showed much less efficiency with scattered distribution of fluorescence in balloon-injured carotid arteries. Electrophoretic mobility shift assay showed activation of NF-kappaB in balloon-injured arteries. In vivo transfection of decoy ODNs mediated by HVJ liposome abolished the NF-kappaB activity in injured arteries with specific binding affinity to NF-kappaB protein. Intimal hyperplasia of carotid artery after balloon injury was reduced by approximately 50% by NF-kappaB decoy transfection compared with buffer treatment or scrambled decoy transfection. Our results demonstrated involvement of NF-kappaB in intimal formation after arterial injury, and indicated that NF-kappaB can be an appropriate molecular target for anti-restenosis therapy.

The potential role of fucosylated cationic liposome/NFκB decoy complexes in the treatment of cytokine-related liver disease

Cytokine production by Kupffer cells, which is regulated by NFkappaB, causes severe liver injury in endotoxin syndrome. NFkappaB decoy has been reported to inhibit NFkappaB-mediated transcription. The purpose of this study is to inhibit LPS-induced cytokine production by Kupffer cell-targeted delivery of NFkappaB decoy using fucosylated cationic liposomes (Fuc-liposomes). Cholesten-5-yloxy-N-{4-[(1-imino-2-L-thiofucosyl-ethyl)-amino] butyl-}formamide (Fuc-C4-Chol) was synthesized to prepare Fuc-liposomes. Tissue accumulation, intrahepatic distribution and serum cytokine concentrations were investigated after intravenous injection of Fuc-liposomes/NFkappaB decoy complexes. Intravenously injected Fuc-liposome complexes rapidly and highly accumulated in the liver while little naked NFkappaB decoy accumulated in the liver. An intrahepatic distribution study showed that Fuc-liposome complexes are mainly taken up by non-parenchymal cells. The liver accumulation of Fuc-liposome complexes was inhibited by GdCl(3) pretreatment, which selectively inhibited Kupffer cell uptake. This result suggested that Kupffer cells contribute to liver accumulation. TNFalpha, IFNgamma, ALT and AST serum levels in LPS-infected mice were significantly attenuated by treatment with Fuc-liposome complexes compared with naked NFkappaB decoy. Fuc-liposome complexes also reduced the amount of activated NFkappaB in the liver nuclei. Fuc-liposomes would be a useful carrier for Kupffer cell-selective delivery of NFkappaB decoy by intravenous injection.

Blockade of Experimental Atopic Dermatitis via Topical NF-κB Decoy Oligonucleotide

Atopic dermatitis (AD) is a common chronic skin inflammatory disease. Long-term use of topical corticosteroids in skin inflammation poses risks of systemic and local side effects. The NF-kappaB transcription factor family plays a central role in the progression and maintenance of AD. This study explores the possibility of using topical NF-kappaB Decoy as a novel therapeutic alternative for targeting Th1/Th2-driven skin inflammation in experimental AD. A high-affinity, topical NF-kappaB Decoy developed for human efficacy demonstrates: (i) efficient NF-kappaB Decoy penetration in pig skin, (ii) NF-kappaB Decoy nuclear localization in keratinocytes and key immune cells, and (iii) potent "steroid-like" efficacy in a chronic dust-mite antigen skin inflammation treatment model. NF-kappaB Decoy exerts its anti-inflammatory action through the effective inhibition of essential regulators of inflammation and by induction of apoptosis of key immune cells. Unlike betamethasone valerate (BMV), long-term NF-kappaB Decoy treatment does not induce skin atrophy. Moreover, topical NF-kappaB Decoy, in contrast to BMV, restores compromised stratum corneum integrity and barrier function. Steroid withdrawal causes rapid rebound of inflammation, while the NF-kappaB Decoy therapeutic benefit was maintained for weeks. Thus, topical NF-kappaB Decoy provides a novel mechanism of reducing chronic skin inflammation with improved skin homeostasis and minimal side effects.

Regulation of NF‐κB Signaling by Decoy Oligodeoxynucleotides

The nuclear factor-kappa B (NF-kappaB) plays a critical role in regulating expression of genes responsible for a wide range of cellular processes, including innate and adoptive immune responses, and pathways related to cell survival and proliferation. Based on its property as a transcription factor, NF-kappaB has been considered as a good target for the treatment of inflammatory immune diseases. To regulate gene expression, "decoy" oligodeoxynucleotides (ODNs) are ideal tools to interfere with binding of transcription factors to promoter regions in genes. Herein we review the application of NF-kappaB decoy ODNs to control inflammatory disorders and transplantation tolerance. In addition, we provide information about induction of antigen-specific peripheral tolerance by their topical application. Regulation of NF-kappaB signaling by NF-kappaB decoy ODNs might be an effective tool to control a variety of disorders caused by inflammatory immune responses.

The effects of nuclear factor-kappaB decoy oligonucleotides on dextran sulphate sodium-induced colitis: experiment with mice

To investigate the effects of nuclear factor-kappaB (NF-kappaB) decoy oligonucleotide (ODN) on dextran sulphate sodium (DSS)-induced colitis. Nine female BABL/C mice underwent infusion of 0.15 ml normal saline into the distant colon and used as controls (Group 1). Twenty-seven female BABL/C mice were made into DSS-induced colitis models and then randomly divided into 3 groups: Group 2 (underwent infusion of 0.15 ml normal saline into the distant colon), Group 3 (infused with NF-kappaB decoy ODN 25 nmol solved in 0.15 ml), and Group 4 (infused with NF-kappaB scrambled decoy ODN 25 nmol solved in 0.15 ml). Disease active index (DAI) was observed every day. Nine days later the mice were killed and their colons were taken out to undergo histological examination. The tumor necrosis factor (TNF)-alpha level of the colon mucosa was measured by enzyme linked immunosorbent assay (ELISA). NF-kappaB expression was determined by immunohistochemical staining. The distribution of NF-kappaB decoy ODN was investigated by confocal laser microscopy. (1) The DAI scores, histological scores and TNF-a level in the colon mucosa of Groups 2 - 4 were all significantly higher than those of Group 1 (all P < 0.05). The DAI scores, histological scores and TNF-a level in the colon mucosa of Group 3 were all significantly lower than those of Groups 2 and 4 (all P < 0.01). (2) In the tissue sections NF-kappaB p65 was positive mainly in the nucleus in the 3 DSS-treated groups without significant differences among these 3 groups, and was mainly positive in the cytoplasm in the control group. (3) Confocal laser microscopy showed that NF-kappaB decoy ODN could be ingested efficiently into the mucosa and submucous layer of colon. (4) There were no significant differences in the liver function, kidney function, and blood glucose among all groups. NF-kappaB pathway is associated with the pathogenesis of DSS-induced colitis which is very similar to human UC. Blockade of NF-kappaB pathway by NF-kappaB decoy ODN shows protective effect on the mice with DSS-induced colitis.