Mourad Zerfaoui

University of Alabama at Birmingham, Birmingham, AL, United States

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Publications (23)98.05 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Minocycline protects against asthma independently of its antibiotic function and was recently reported as a potent PARP inhibitor. In an animal model of asthma, a single administration of minocycline conferred excellent protection against ovalbumin-induced airway eosinophilia, mucus hypersecretion, and Th2 cytokine production (IL-4/IL-5/IL-12(p70)/IL-13/GM-CSF) and a partial protection against airway hyperresponsiveness. These effects correlated with pronounced reduction in lung and sera allergen-specific IgE. A reduction in poly(ADP-ribose)-immunoreactivity in lungs of minocycline-treated/ovalbumin-challenged mice correlated with decreased oxidative DNA damage. Minocycline's effect on PARP may be indirect as the drug failed to efficiently block direct PARP activation in lungs of N-methyl-N'-nitro-N-nitroso-guanidine-treated mice or H2O2-treated cells suggesting. Minocycline blocked allergen-specific IgE production in B cells potentially by modulating T cell-receptor (TCR)-linked IL-4 production at the mRNA level but not through a modulation of the IL-4-JAK-STAT-6 axis, IL-2 production, or NFAT1 activation. Restoration of IL-4, ex vivo, rescued IgE production by minocycline-treated/ovalbumin-stimulated B cells. IL-4 blockade correlated with a preferential inhibition of the NF-κB activation arm of TCR but not GSK3, Src, p38 MAPK, or ERK1/2. Interestingly, the drug promoted a slightly higher Src and ERK1/2 phosphorylation. Inhibition of NF-κB was linked to a complete blockade of TCR-stimulated GATA-3 expression, a pivotal transcription factor for IL-4 expression. Minocycline also reduced TNF-α-mediated NF-κB activation and expression of dependent genes. These results show a potentially broad effect of minocycline but may block IgE production in part by modulating TCR function particularly by inhibiting the signaling pathway leading to NF-κB activation, GATA-3 expression, and subsequent IL-4 production.
    Journal of Biological Chemistry 11/2012; · 4.65 Impact Factor
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    ABSTRACT: The uptake and clearance of apoptotic cells by macrophages and other phagocytic cells, a process called efferocytosis, is a major component in the resolution of inflammation. Increased concentrations of extracellular histones are found during acute inflammatory states and appear to contribute to organ system dysfunction and mortality. In these studies, we examined the potential role of histones in modulating efferocytosis. We found that phagocytosis of apoptotic neutrophils or thymocytes by macrophages was significantly diminished in the presence of histones H3 or H4, but not histone H1. Histone H3 demonstrated direct binding to macrophages, an effect that was diminished by preincubation of macrophages with the opsonins growth arrest-specific gene 6 (Gas6) and milk fat globule-epidermal growth factor (EGF) 8 (MFG-E8). Incubation of histone H3 with soluble α(v)β₅ integrin and Mer, but not with α(v)β₃, diminished its binding to macrophages. Phagocytosis of apoptotic cells by alveolar macrophages in vivo was diminished in the presence of histone H3. Incubation of histone H3 with activated protein C, a treatment that degrades histones, abrogated its inhibitory effects on efferocytosis under both in vitro and in vivo conditions. The present studies demonstrate that histones have inhibitory effects on efferocytosis, suggesting a new mechanism by which extracellular histones contribute to acute inflammatory processes and tissue injury.
    Molecular Medicine 04/2012; 18(1):825-33. · 4.47 Impact Factor
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    ABSTRACT: Phagocytosis of apoptotic cells by macrophages, known as efferocytosis, is a critical process in the resolution of inflammation. High mobility group box 1 (HMGB1) protein was first described as a nuclear nonhistone DNA-binding protein, but is now known to be secreted by activated cells during inflammatory processes, where it participates in diminishing efferocytosis. Although HMGB1 is known to undergo modification when secreted, the effect of such modifications on the inhibitory actions of HMGB1 during efferocytosis have not been reported. In the present studies, we found that HMGB1 secreted by Toll-like receptor 4 (TLR4) stimulated cells is highly poly(ADP-ribosyl)ated (PARylated). Gene deletion of poly(ADP)-ribose polymerase (PARP)-1 or pharmacological inhibition of PARP-1 decreased the release of HMGB1 from the nucleus to the extracellular milieu after TLR4 engagement. Preincubation of macrophages or apoptotic cells with HMGB1 diminished efferocytosis through mechanisms involving binding of HMGB1 to phosphatidylserine on apoptotic cells and to the receptor for advanced glycation end products (RAGE) on macrophages. Preincubation of either macrophages or apoptotic cells with PARylated HMGB1 inhibited efferocytosis to a greater degree than exposure to unmodified HMGB1, and PARylated HMGB1 demonstrated higher affinity for phosphatidylserine and RAGE than unmodified HMGB1. PARylated HMGB1 had a greater inhibitory effect on Ras-related C3 botulinum toxin substrate 1 (Rac-1) activation in macrophages during the uptake of apoptotic cells than unmodified HMGB1. The present results, showing that PARylation of HMGB1 enhances its ability to inhibit efferocytosis, provide a novel mechanism by which PARP-1 may promote inflammation.
    Molecular Medicine 12/2011; 18(1):359-69. · 4.47 Impact Factor
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    ABSTRACT: Post-transcriptional modification of proteins is crucial for balancing protein structure and function in many biological processes. The addition of polymers of adenosine diphosphate (ADP)-ribose (PAR), which are synthesized by PAR polymerases (PARPs) from nicotinamide adenine dinucleotide (NAD), is one such distinctive post-translational modification. PARP-1, the best characterized of the 17-member PARP family, is considered a key isoform responsible for poly(ADP-ribosyl)ation of several nuclear proteins. ADP-ribose polymers add a highly negative charge to their target proteins, resulting in a modification of their activities and functions. PARPs not only participate in regulating cell survival and cell death programs, but are also involved in other biological functions with which novel members of the PARP family have been shown to be involved. Among such functions are transcription regulation, telomere cohesion and mitotic spindle formation during cell division, and intracellular energy metabolism. Recent work from our laboratory and others has highlighted the novel role of PARP-1 in regulating the intracellular trafficking of key cellular proteins such as p53 and nuclear factor-kappa B (NF-κB). Recent literature has revealed that ADP-ribosylation reactions may play important roles in cellular trafficking during inflammation, cell death, and DNA repair. This review will summarize recent findings and concepts linking the role of PARP enzymes and their poly-ADP-ribosylation activity in the regulation of intracellular transport processes. A special focus is placed on the proposed molecular mechanisms involved in such transport processes as the functional significance of PARylation of p53, NF-κB, and high-mobility group protein box 1.
    Cellular Signalling 08/2011; 24(1):1-8. · 4.47 Impact Factor
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    ABSTRACT: Previously, we demonstrated that inhibition of poly(ADP-ribose) polymerase (PARP) exerts protective effects against high-fat (HF) diet-induced atherogenesis in part by increasing tissue inhibitor of metalloproteinase (TIMP)-2 expression. Given that characteristics of dilated cardiomyopathy closely associate with atherosclerosis and are mediated by an imbalance between matrix metalloproteinases (MMPs) and TIMPs, we hypothesized that PARP-1 gene deletion may protect against HF-induced cardiac hypertrophy and dilatations by altering TIMP-2/MMPs balance in favor of a maintenance of tissue homeostasis. Hemodynamic parameters determined by echocardiography were similar in ApoE(-/-) mice and PARP-1-deficient ApoE(-/-) mice (DKO) fed a regular diet (RD). However, histological analysis revealed that cardiomyocytes of ApoE(-/-) mice on RD were hypertrophied, displaying an enlarged cell body and nucleus, traits that were absent in DKO animals. HF diet-fed ApoE(-/-) mice exhibited increased interventricular septum, left ventricular (LV) internal dimension, LV volume, and LV mass in addition to a separation of myocardial fibers suggestive of dilated cardiomyopathy. PARP-1 gene deletion protected against these degenerative changes. MMP activity was dramatically increased in hearts of ApoE(-/-) mice on HF diet and was accompanied by increased collagen degradation, mast cell degranulation, and increased myocyte cell death. PARP-1 gene knockout was associated with increased TIMP-2 expression antagonizing, as a result, the damaging effects of active MMPs. The present study demonstrates that PARP-1 gene deletion exerts protective effects against HF diet-induced dilated cardiomyopathy by maintaining increased expression of TIMP-2. With additional protective effects against cell death and inflammation, PARP-1 deficiency preserves cardiac tissue homeostasis.
    Cardiovascular pathology: the official journal of the Society for Cardiovascular Pathology 01/2011; 20(2):e57-68. · 1.63 Impact Factor
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    ABSTRACT: We recently showed that poly(ADP-ribose)polymerase-1 (PARP-1) may play a role in allergen (ovalbumin)-induced airway eosinophilia, potentially through a specific effect on IL-5 production. We also reported that while IL-5 replenishment promotes reversal of eosinophilia in lungs of PARP-1(-/-) mice, IL-4 or Immunoglobulin E replenishment do not, suggesting a potentially significant regulatory relationship between PARP-1 and IL-5. To explore the mechanism by which PARP-1 regulates IL-5 production and to determine how PARP-1 inhibition blocks allergen-induced eosinophilia. This study was conducted using a murine model of allergic airway inflammation and primary splenocytes. PARP-1 knockout-associated reduction in IL-5 upon allergen exposure occurs at the mRNA level. Such an effect appears to take place after IL-4 receptor activation as PARP-1 inhibition exerted no effect on JAK1/JAK3 activation. Signal transducer and activator of transcription-6 (STAT-6) protein was severely downregulated in spleens of PARP-1(-/-) mice without any effect on mRNA levels, suggesting an effect on protein integrity rather than gene transcription. Interestingly, the degradation of STAT-6 in PARP-1(-/-) mice required allergen stimulation. Additionally, PARP-1 enzymatic activity appears to be required for STAT-6 integrity. The downregulation of STAT-6 coincided with mRNA and protein reduction of GATA-binding protein-3 and occupancy of its binding site on the IL-5 gene promoter. IL-4 was sufficient to induce STAT-6 downregulation in both PARP-1(-/-) mice and isolated splenocytes. Such degradation may be mediated by calpain, but not by proteasomes. These results demonstrate a novel function of PARP-1 in regulating IL-5 expression during allergen-induced inflammation and explain the underlying mechanism by which PARP-1 inhibition results in IL-5 reduction.
    Allergy 01/2011; 66(7):853-61. · 5.88 Impact Factor
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    ABSTRACT: The DNA binding activity of NF-κB is critical for VCAM-1 expression during inflammation. DNA-dependent protein kinase (DNA-PK) is thought to be involved in NF-κB activation. Here we show that DNA-PK is required for VCAM-1 expression in response to TNF. The phosphorylation and subsequent degradation of I-κBα as well as the serine 536 phosphorylation and nuclear translocation of p65 NF-κB were insufficient for VCAM-1 expression in response to TNF. The requirement for p50 NF-κB in TNF-induced VCAM-1 expression may be associated with its interaction with and phosphorylation by DNA-PK, which appears to be dominant over the requirement for p65 NF-κB activation. p50 NF-κB binding to its consensus sequence increased its susceptibility to phosphorylation by DNA-PK. Additionally, DNA-PK activity appeared to increase the association between p50/p50 and p50/p65 NF-κB dimers upon binding to DNA and after binding of p50 NF-κB to the VCAM-1 promoter. Analyses of the p50 NF-κB protein sequence revealed that both serine 20 and serine 227 at the amino terminus of the protein are putative sites for phosphorylation by DNA-PK. Mutation of serine 20 completely eliminated phosphorylation of p50 NF-κB by DNA-PK, suggesting that serine 20 is the only site in p50 NF-κB for phosphorylation by DNA-PK. Re-establishing wild-type p50 NF-κB, but not its serine 20/alanine mutant, in p50 NF-κB(-/-) fibroblasts reversed VCAM-1 expression after TNF treatment, demonstrating the importance of the serine 20 phosphorylation site in the induction of VCAM-1 expression. Together, these results elucidate a novel mechanism for the involvement of DNA-PK in the positive regulation of p50 NF-κB to drive VCAM-1 expression.
    Journal of Biological Chemistry 10/2010; 285(52):41152-60. · 4.65 Impact Factor
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    ABSTRACT: The role of inducible NO synthase (iNOS) in allergic airway inflammation remains elusive. We tested the hypothesis that iNOS plays different roles during acute versus chronic airway inflammation. Acute and chronic mouse models of OVA-induced airway inflammation were used to conduct the study. We showed that iNOS deletion was associated with a reduction in eosinophilia, mucus hypersecretion, and IL-5 and IL-13 production upon the acute protocol. Such protection was completely abolished upon the chronic protocol. Interestingly, pulmonary fibrosis observed in wild-type mice under the chronic protocol was completely absent in iNOS(-/-) mice despite persistent IL-5 and IL-13 production, suggesting that these cytokines were insufficient for pulmonary fibrosis. Such protection was associated with reduced collagen synthesis and indirect but severe TGF-beta modulation as confirmed using primary lung smooth muscle cells. Although activation of matrix metalloproteinase-2/-9 exhibited little change, the large tissue inhibitor of metalloproteinase-2 (TIMP-2) increase detected in wild-type mice was absent in the iNOS(-/-) counterparts. The regulatory effect of iNOS on TIMP-2 may be mediated by peroxynitrite, as the latter reversed TIMP-2 expression in iNOS(-/-) lung smooth muscle cells and fibroblasts, suggesting that the iNOS-TIMP-2 link may explain the protective effect of iNOS-knockout against pulmonary fibrosis. Analysis of lung sections from chronically OVA-exposed iNOS(-/-) mice revealed evidence of residual but significant protein nitration, prevalent oxidative DNA damage, and poly(ADP-ribose) polymerase-1 activation. Such tissue damage, inflammatory cell recruitment, and mucus hypersecretion may be associated with substantial arginase expression and activity. The results in this study exemplify the complexity of the role of iNOS in asthma and the preservation of its potential as a therapeutic a target.
    The Journal of Immunology 09/2010; 185(5):3076-85. · 5.52 Impact Factor
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    ABSTRACT: The role of NF-kappaB in the expression of inflammatory genes and its participation in the overall inflammatory process of chronic diseases and acute tissue injury are well established. We and others have demonstrated a critical involvement of poly(ADP-ribose) polymerase (PARP)-1 during inflammation, in part, through its relationship with NF-kappaB. However, the mechanism by which PARP-1 affects NF-kappaB activation has been elusive. In this study, we show that PARP-1 inhibition by gene knockout, knockdown, or pharmacologic blockade prevented p65 NF-kappaB nuclear translocation in smooth muscle cells upon TLR4 stimulation, NF-kappaB DNA-binding activity, and subsequent inducible NO synthase and ICAM-1 expression. Such defects were reversed by reconstitution of PARP-1 expression. PARP-1 was dispensable for LPS-induced IkappaBalpha phosphorylation and subsequent degradation but was required for p65 NF-kappaB phosphorylation. A perinuclear p65 NF-kappaB localization in LPS-treated PARP-1(-/-) cells was associated with an export rather an import defect. Indeed, whereas PARP-1 deficiency did not alter expression of importin alpha3 and importin alpha4 and their cytosolic localization, the cytosolic levels of exportin (Crm)-1 were increased. Crm1 inhibition promoted p65 NF-kappaB nuclear accumulation as well as reversed LPS-induced p65 NF-kappaB phosphorylation and inducible NO synthase and ICAM-1 expression. Interestingly, p65 NF-kappaB poly(ADP-ribosyl)ation decreased its interaction with Crm1 in vitro. Pharmacologic inhibition of PARP-1 increased p65 NF-kappaB-Crm1 interaction in LPS-treated smooth muscle cells. These results suggest that p65 NF-kappaB poly(ADP-ribosyl)ation may be a critical determinant for the interaction with Crm1 and its nuclear retention upon TLR4 stimulation. These results provide novel insights into the mechanism by which PARP-1 promotes NF-kappaB nuclear retention, which ultimately can influence NF-kappaB-dependent gene regulation.
    The Journal of Immunology 08/2010; 185(3):1894-902. · 5.52 Impact Factor
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    ABSTRACT: We reported that PARP-1 exhibits differential roles in expression of inflammatory factors. Here, we show that PARP-1 deletion was associated with a significant reduction in inflammatory cell recruitment to mouse airways upon intratracheal administration of LPS. However, PARP-1 deletion exerted little effect in response to TNF exposure. LPS induced massive neutrophilia and moderate recruitment of macrophages, and TNF induced recruitment of primarily macrophages with smaller numbers of neutrophils in the lungs. Following either exposure, macrophage recruitment was blocked severely in PARP-1(-/-) mice, and this was associated with a marked reduction in MCP-1 and MIP-1alpha. This association was corroborated partly by macrophage recruitment in response to intratracheal administration of MCP-1 in PARP-1(-/-) mice. Surprisingly, although neutrophil recruitment was reduced significantly in LPS-treated PARP-1(-/-) mice, neutrophil numbers increased in TNF-treated mice, suggesting that PARP-1 deletion may promote a macrophagic-to-neutrophilic shift in the inflammatory response upon TNF exposure. Neutrophil-specific chemokines mKC and MIP-2 were reduced significantly in lungs of LPS-treated but only partially reduced in TNF-treated PARP-1(-/-) mice. Furthermore, the MIP-2 antagonist abrogated the shift to a neutrophilic response in TNF-exposed PARP-1(-/-) mice. Although CXCR2 expression increased in response to either stimulus in PARP-1(+/+) mice, the DARC increased only in lungs of TNF-treated PARP-1(+/+) mice; both receptors were reduced to basal levels in treated PARP-1(-/-) mice. Our results show that the balance of pro-neutrophilic or pro-macrophagic stimulatory factors and the differential influence of PARP-1 on these factors are critical determinants for the nature of the airway inflammatory response.
    Journal of leukocyte biology 10/2009; 86(6):1385-92. · 4.99 Impact Factor
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    ABSTRACT: Hypercholesterolemia is increasingly considered the basis for not only cardiovascular pathologies but also several complications affecting other organs such as lungs. In this study, we examined the effect of hypercholesterolemia on lung integrity using a mouse model (ApoE(-/-)) of high-fat (HF) diet-induced atherosclerosis. A 12-week HF diet regimen induced systemic production of TNF-alpha, IFN-gamma, GMC-SF, RANTES, IL-1alpha, IL-2 and IL-12 with TNF-alpha as the predominant cytokine in ApoE(-/-) mice. Concomitantly, TNF-alpha, IFN-gamma and MIP-1alpha were detected in brochoalveolar lavage (BAL) fluids of these mice, coinciding with lung inflammation consisting primarily of monocytes/macrophages. Such lung inflammation correlated with marked collagen deposition and an increase in matrix metalloproteinase-9 activity in ApoE(-/-)mice without mucus production. Although TGF-beta1 was undetectable in the BAL fluid of ApoE(-/-) mice on HF diet, it showed a much wider tissue distribution compared with that of control animals. Direct exposure of smooth muscle cells to oxidized-LDL, in vitro, induced a time-dependent expression of TNF-alpha. Direct intratracheal TNF-alpha-administration induced a lung inflammation pattern in wild-type mice that was strikingly similar to that induced by HF diet in ApoE(-/-) mice. TNF-alpha administration induced expression of several factors known to be critically involved in lung remodeling, such as MCP-1, IL-1beta, TGF-beta1, adhesion molecules, collagen type-I and TNF-alpha itself in the lungs of treated mice. These results suggest that hypercholesterolemia may promote chronic inflammatory conditions in lungs that are conducive to lung remodeling potentially through TNF-alpha-mediated processes.
    Laboratory Investigation 09/2009; 89(11):1243-51. · 3.96 Impact Factor
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    ABSTRACT: We recently showed that poly(ADP-ribose) polymerase (PARP) is activated within atherosclerotic plaques in an animal model of atherosclerosis. Pharmacological inhibition of PARP or reduced expression in heterozygous animals interferes with atherogenesis and may promote factors of plaque stability, possibly reflecting changes in inflammatory and cellular factors consistent with plaque stability. The current study addresses the hypothesis that pharmacological inhibition of PARP promotes atherosclerotic plaque regression. Using a high-fat diet-induced atherosclerosis apolipoprotein E(-/-) mouse model, we demonstrate that administration of the potent PARP inhibitor, thieno[2,3-c]isoquinolin-5-one (TIQ-A), when combined with a regular diet regimen during treatment, induced regression of established plaques. Plaque regression was associated with a reduction in total cholesterol and low-density lipoproteins. Furthermore, plaques of TIQ-A-treated mice were highly enriched with collagen and smooth muscle cells, displayed thick fibrous caps, and exhibited a marked reduction in CD68-positive macrophage recruitment and associated foam cell presence. These changes correlated with a significant decrease in expression of monocyte chemoattractant protein-1 and intercellular cell adhesion molecule-1, potentially as a result of a robust reduction in tumor necrosis factor expression. The PARP inhibitor appeared to affect cholesterol metabolism by affecting acyl-coenzymeA/cholesterol acyltransferase-1 expression but exerted no effect on cholesterol influx or efflux as assessed by an examination of the ATP-binding cassette transporter-1 and the scavenger receptor-A expression levels in the different experimental groups. In accordance, PARP inhibition may prove beneficial not only in preventing atherogenesis but also in promoting regression of preexisting plaques.
    Journal of Pharmacology and Experimental Therapeutics 02/2009; 329(1):150-8. · 3.89 Impact Factor
  • PLoS ONE 01/2009; 4(10). · 3.53 Impact Factor
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    ABSTRACT: The aims of this study were to investigate the role of poly(ADP-ribose) polymerase (PARP)-1 in dyslipidemia-associated vascular dysfunction as well as autonomic nervous system dysregulation. Apolipoprotein (ApoE)(-/-) mice fed a high-fat diet were used as a model of atherosclerosis. Vascular and autonomic functions were measured in conscious mice using telemetry. The study revealed that PARP-1 plays an important role in dyslipidemia-associated vascular and autonomic dysfunction. Inhibition of this enzyme by gene knockout partially restored baroreflex sensitivity in ApoE(-/-) mice without affecting baseline heart-rate and arterial pressure, and also improved heart-rate responses following selective blockade of the autonomic nervous system. The protective effect of PARP-1 gene deletion against dyslipidemia-induced endothelial dysfunction was associated with preservation of eNOS activity. Dyslipidemia induced PARP-1 activation was accompanied by oxidative tissue damage, as evidenced by increased expression of iNOS and subsequent protein nitration. PARP-1 gene deletion reversed these effects, suggesting that PARP-1 may contribute to vascular and autonomic pathologies by promoting oxidative tissue injury. Further, inhibition of this oxidative damage may account for protective effects of PARP-1 gene deletion on vascular and autonomic functions. This study demonstrates that PARP-1 participates in dyslipidemia-mediated dysregulation of the autonomic nervous system and that PARP-1 gene deletion normalizes autonomic and vascular dysfunctions. Maintenance of eNOS activity may be associated with the protective effect of PARP-1 gene deletion against dyslipidemia-induced endothelial dysfunction.
    PLoS ONE 01/2009; 4(10):e7430. · 3.53 Impact Factor
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    ABSTRACT: Inducible nitric oxide synthase (iNOS) inhibition was recently shown to exert no effect on allergen challenge in human asthma, raising serious concerns about the role of the protein in the disease. The present study investigated the role of iNOS in ovalbumin-induced eosinophilia from the perspective of its relationship with poly(ADP-ribose) polymerase-1 (PARP-1) and oxidative DNA damage. A mouse model of ovalbumin-induced eosinophilia was used to conduct the studies. iNOS-associated protein nitration and tissue damage were partially responsible for allergen-induced eosinophilia. iNOS expression was required for oxidative DNA damage and PARP-1 activation upon allergen challenge. PARP-1 was required for iNOS expression and protein nitration, and this requirement was connected to nuclear factor-kappaB. PARP-1 was an important substrate for iNOS-associated by-products after ovalbumin-challenge. PARP-1 nitration blocked its poly(ADP-ribosyl)ation activity. Interleukin-5 re-establishment in ovalbumin-exposed PARP-1(-/-) mice reversed eosinophilia and partial mucus production without a reversal of iNOS expression, concomitant protein nitration or associated DNA damage. The present results demonstrate a reciprocal relationship between inducible nitric oxide synthase and poly(ADP-ribose) polymerase-1 and suggest that expression of inducible nitric oxide synthase may be dispensable for eosinophilia after interleukin-5 production. Inducible nitric oxide synthase may be required for oxidative DNA damage and full manifestation of mucus production. Such dispensability may explain, in part, the reported ineffectiveness of inducible nitric oxide synthase inhibition in preventing allergen-induced inflammation in humans.
    European Respiratory Journal 11/2008; 33(2):252-62. · 6.36 Impact Factor
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    ABSTRACT: The G protein-coupled serotonin 5-hydroxytryptamine (5-HT)(2A) receptor is primarily recognized for its role in brain neurotransmission, where it mediates a wide variety of functions, including certain aspects of cognition. However, there is significant expression of this receptor in peripheral tissues, where its importance is largely unknown. We have now discovered that activation of 5-HT(2A) receptors in primary aortic smooth muscle cells provides a previously unknown and extremely potent inhibition of tumor necrosis factor (TNF)-alpha-mediated inflammation. 5-HT(2A) receptor stimulation with the agonist (R)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane [(R)-DOI] rapidly inhibits a variety of TNF-alpha-mediated proinflammatory markers, including intracellular adhesion molecule 1 (ICAM-1), vascular adhesion molecule 1 (VCAM-1), and interleukin (IL)-6 gene expression, nitric-oxide synthase activity, and nuclear translocation of nuclear factor kappaB, with IC(50) values of only 10 to 20 pM. It is significant that proinflammatory markers can also be inhibited by (R)-DOI hours after treatment with TNF-alpha. With the exception of a few natural toxins, no current drugs or small molecule therapeutics demonstrate a comparable potency for any physiological effect. TNF-alpha-mediated inflammatory pathways have been strongly implicated in a number of diseases, including atherosclerosis, rheumatoid arthritis, psoriasis, type II diabetes, depression, schizophrenia, and Alzheimer's disease. Our results indicate that activation of 5-HT(2A) receptors represents a novel, and extraordinarily potent, potential therapeutic avenue for the treatment of disorders involving TNF-alpha-mediated inflammation. Note that because (R)-DOI can significantly inhibit the effects of TNF-alpha many hours after the administration of TNF-alpha, potential therapies could be aimed not only at preventing inflammation but also treating inflammatory injury that has already occurred or is ongoing.
    Journal of Pharmacology and Experimental Therapeutics 09/2008; 327(2):316-23. · 3.89 Impact Factor
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    ABSTRACT: This study determines that vascular smooth muscle cell (VSMC) signaling through extracellular signal-regulated kinase (ERK) 1/2-mitogen-activated protein (MAP) kinase, alphavbeta(3)-integrin, and transforming growth factor (TGF)-beta1 dictates collagen type I network induction in mesenteric resistance arteries (MRA) from type 1 diabetic (streptozotocin) or hypertensive (HT; ANG II) mice. Isolated MRA were subjected to a pressure-passive-diameter relationship. To delineate cell types and mechanisms, cultured VSMC were prepared from MRA and stimulated with ANG II (100 nM) and high glucose (HG, 22 mM). Pressure-passive-diameter relationship reduction was associated with increased collagen type I deposition in MRA from HT and diabetic mice compared with control. Treatment of HT and diabetic mice with neutralizing TGF-beta1 antibody reduced MRA stiffness and collagen type I deposition. Cultured VSMC stimulated with HG or ANG II for 5 min increased ERK1/2-MAP kinase phosphorylation, whereas a 48-h stimulation induced latent TGF-beta1, alphavbeta(3)-integrin, and collagen type 1 release in the conditioned media. TGF-beta1 bioactivity and Smad2 phosphorylation were alphavbeta(3)-integrin-dependent, since beta(3)-integrin antibody and alphavbeta(3)-integrin inhibitor (SB-223245, 10 microM) significantly prevented TGF-beta1 bioactivity and Smad2 phosphorylation. Pretreatment of VSMC with ERK1/2-MAP kinase inhibitor (U-0126, 1 microM) reduced alphavbeta(3)-integrin, TGF-beta1, and collagen type 1 content. Additionally, alphavbeta(3)-integrin antibody, SB-223245, TGF-beta1-small-intefering RNA (siRNA), and Smad2-siRNA (40 nM) prevented collagen type I network formation in response to ANG II and HG. Together, these data provide evidence that resistance artery fibrosis in type 1 diabetes and hypertension is a consequence of abnormal collagen type I release by VSMC and involves ERK1/2, alphavbeta(3)-integrin, and TGF-beta1 signaling. This pathway could be a potential target for overcoming small artery complications in diabetes and hypertension.
    AJP Heart and Circulatory Physiology 08/2008; 295(1):H69-76. · 4.01 Impact Factor
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    ABSTRACT: The aim of this study was to take a combination of animal and cell culture approaches to examine the individual responses of vascular cells to varying inflammatory factors in order to gain insights on the mechanism(s) by which poly(ADP-ribose) polymerase (PARP) inhibition promotes factors of plaque stability. Apolipoprotein (ApoE(-/-)) mice fed a high-fat diet were used as a model of atherosclerosis. Primary endothelial cells, smooth muscle cells (SMCs), and ex-vivo generated foam cells (FCs) were used in our in vitro studies. PARP inhibition significantly decreased the markers of oxidative stress and caspase-3 activation and increased smooth muscle actin within plaques from ApoE(-/-) mice fed a high-fat diet. PARP inhibition protected against apoptosis and/or necrosis in SMCs and endothelial cells in response to H(2)O(2) or tumour necrosis factor (TNF). Remarkably, PARP inhibition in FCs resulted in significant sensitization to 7-ketocholesterol (7-KC) by increasing cellular-toxic-free cholesterol, potentially through a down-regulation of acyl-CoA:cholesterol acyltransferase-1 (ACAT-1) expression. 7-KC induced necrosis exclusively in endothelial cells, which was, surprisingly, unaffected by PARP inhibition indicating that PARP inhibition does not prevent all forms of necrotic cell death. In SMCs, PARP-1 inhibition by gene deletion conferred protection against 7-KC or TNF, potentially by reducing caspase-3-like activation, preventing induction of c-Jun N-terminal protein kinase phosphorylation, and inducing extracellular signal-regulated kinase phosphorylation independently of PARP classical enzymatic activity. These data present PARP-1 as an important player in the death of cells constituting atherosclerotic plaques contributing to plaque dynamics. PARP inhibition may be a protective, a neutral, or a sensitizing factor. Additionally, PARP-1 may be a novel factor that can alter lipid metabolism. These novel functions of PARP not only challenge the current understanding of the role of the enzyme in cell death but also provide insights on the intricate contribution of PARP in cellular responses to predominant inflammatory factors within atherosclerotic plaques, presenting additional evidence for the viability of PARP inhibition as a therapeutic strategy for atherosclerosis.
    Cardiovascular Research 07/2008; 78(3):429-39. · 5.81 Impact Factor
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    ABSTRACT: Identifying therapeutic drugs that block the release or effects of T-helper type 2 (Th2) cytokines after allergen exposure is an important goal for the treatment of allergic inflammatory diseases including asthma. We recently showed, using a murine model of allergic airway inflammation, that poly(ADP-ribose) polymerase (PARP) plays an important role in the pathogenesis of asthma-related lung inflammation. PARP inhibition, by single injection of a novel inhibitor, thieno[2,3-c]isoquinolin-5-one (TIQ-A), before ovalbumin (OVA) challenge, prevented airway eosinophilia in C57BL/6 mice with concomitant suppression of Th2 cytokine production and mucus secretion. To evaluate the efficacy of the drug when it is given after OVA challenge for its possible therapeutic potential. This study was conducted using a murine model of allergic airway inflammation. A single injection of TIQ-A (6 mg/kg) one or 6 h post-allergen challenge conferred similar reduction in OVA challenge-induced eosinophilia. More significantly, post-allergen challenge administration of the drug exerted even better suppression on the production of IL-4, IL-5, IL-13, and IgE and prevented airway hyperresponsiveness to inhaled-methacholine. The significant decrease in IL-13 was accompanied by a complete absence of airways mucus production indicating a potential protection against allergen-induced airway remodelling. The coincidence of the inflammation trigger and the time of drug administration appear to be important for the drug's more pronounced protection. The observed time window for efficacy, 1 or 6 h after allergen challenge may be of great clinical interest. These findings may provide a novel therapeutic strategy for the treatment of allergic airway inflammation, including asthma.
    Clinical & Experimental Allergy 06/2008; 38(5):839-46. · 4.79 Impact Factor
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    ABSTRACT: Although nuclear translocation of NF-kappaB and subsequent binding to promoters of ICAM-1 and VCAM-1 have been shown to be decisive for their expression, a number of discrepancies in the expression patterns of these adhesion molecules have been reported in both cell culture systems and disease settings, including atherosclerosis, asthma, and autoimmune diseases. Here we show that while p65 NF-kappaB nuclear translocation in TNF-treated smooth muscle cells (SMCs) was sufficient for the expression of VCAM-1, expression of ICAM-1 showed a critical requirement for PARP-1. I-kappaBalpha phosphorylation and subsequent degradation were virtually identical in both TNF-treated wild-type and PARP-1-/- SMCs. VCAM-1 expression in TNF-treated PARP-1-/- SMCs was completely inhibited by the NF-kappaB inhibitor, pyrrolidine dithiocarbamate, confirming that VCAM-1 expression was indeed NF-kappaB-dependent. The expression of both VCAM-1 and ICAM-1 was associated with a transient interaction between PARP-1 and p65 NF-kappaB when examined in the fibroblastic cell line, COS-7, and in the airway epithelial cell line, A549. Such interactions were confirmed using florescence resonance energy transfer analysis. Protein acetylation activity, mediated by p300/CBP, was required for both VCAM-1 and ICAM-1 expression in TNF-treated SMCs; however, the interaction of PARP-1 with p300/CBP was dispensable for VCAM-1 expression. These findings indicate that p65 NF-kappaB nuclear translocation may be sufficient for certain genes (e.g., VCAM-1) while insufficient for others (e.g., ICAM-1), thus providing a novel insight into the role of NF-kappaB in driving target gene expression. Furthermore, the data suggest a differential requirement for PARP-1 expression in inflammatory processes.
    Cellular Signalling 02/2008; 20(1):186-94. · 4.47 Impact Factor

Publication Stats

333 Citations
98.05 Total Impact Points

Institutions

  • 2011
    • University of Alabama at Birmingham
      • Department of Medicine
      Birmingham, AL, United States
  • 2009–2011
    • Tulane University
      • • Department of Urology
      • • Department of Physiology
      New Orleans, LA, United States
  • 2008–2011
    • Louisiana State University Health Sciences Center New Orleans
      • Department of Pharmacology and Experimental Therapeutics
      Baton Rouge, LA, United States