title: Adenovirus-mediated expression of a dominant negative mutant of p65/RelA inhibits proinflammatory gene expression in endothelial cells without sensitizing to apoptosis.
abstract: We hypothesized that blocking the induction of proinflammatory genes associated with endothelial cell (EC) activation, by inhibiting the transcription factor nuclear factor kappaB (NF-kappaB), would prolong survival of vascularized xenografts. Our previous studies have shown that inhibition of NF-kappaB by adenovirus-mediated overexpression of I kappaB alpha suppresses the induction of proinflammatory genes in EC. However, I kappaB alpha sensitizes EC to TNF-alpha-mediated apoptosis, presumably by suppressing the induction of the NF-kappaB-dependent anti-apoptotic genes A20, A1, manganese superoxide dismutase (MnSOD), and cellular inhibitor of apoptosis 2. We report here that adenovirus mediated expression of a dominant negative C-terminal truncation mutant of p65/RelA (p65RHD) inhibits the induction of proinflammatory genes, such as E-selectin, ICAM-1, VCAM-1, IL-8, and inducible nitric oxide synthase, in EC as efficiently as does I kappaB alpha. However, contrary to I kappaB alpha, p65RHD does not sensitize EC to TNF-alpha-mediated apoptosis although both inhibitors suppressed the induction of the anti-apoptotic genes A20, A1, and MnSOD equally well. We present evidence that this difference in sensitization of EC to apoptosis is due to the ability of p65RHD, but not I kappaB alpha, to inhibit the constitutive expression of c-myc, a gene involved in the regulation of TNF-alpha-mediated apoptosis. These data demonstrate that it is possible to block the expression of proinflammatory genes during EC activation by targeting NF-kappaB, without sensitizing EC to apoptosis and establishes the role of c-myc in controlling induction of apoptosis during EC activation. Finally, these data provide the basis for a potential approach to suppress EC activation in vivo in transgenic pigs to be used as donors for xenotransplantation.
authors.affiliation: Immunobiology Research Center, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: M P
authors.initials: MP
authors.lastname: Soares
authors.forename: A
authors.initials: A
authors.lastname: Muniappan
authors.forename: E
authors.initials: E
authors.lastname: Kaczmarek
authors.forename: K
authors.initials: K
authors.lastname: Koziak
authors.forename: C J
authors.initials: CJ
authors.lastname: Wrighton
authors.forename: F
authors.initials: F
authors.lastname: Steinhäuslin
authors.forename: C
authors.initials: C
authors.lastname: Ferran
authors.forename: H
authors.initials: H
authors.lastname: Winkler
authors.forename: F H
authors.initials: FH
authors.lastname: Bach
authors.forename: J
authors.initials: J
authors.lastname: Anrather
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D002478:Cells, Cultured; D003546:Cysteine Endopeptidases; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D005822:Genetic Vectors / Q000235:genetics; D018398:Homeodomain Proteins; D006801:Humans; D020672:I-kappa B Proteins; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D015778:Minor Histocompatibility Antigens; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000502:physiology; D009687:Nuclear Proteins; D009928:Organ Specificity; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000502:physiology; D051818:Replication Protein C; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D013482:Superoxide Dismutase / Q000096:biosynthesis / Q000235:genetics; D013552:Swine; D051996:Transcription Factor RelA; D014158:Transcription, Genetic; D014162:Transfection; D019076:Transgenes; D000072598:Tumor Necrosis Factor alpha-Induced Protein 3
chemical_list: C096816:BCL2-related protein A1; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D020672:I-kappa B Proteins; D047908:Intracellular Signaling Peptides and Proteins; C096063:MATA1 protein, S cerevisiae; D015778:Minor Histocompatibility Antigens; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D009687:Nuclear Proteins; D011506:Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D051996:Transcription Factor RelA; D000072000:NF-KappaB Inhibitor alpha; D013482:Superoxide Dismutase; C076529:TNFAIP3 protein, human; D000072598:Tumor Necrosis Factor alpha-Induced Protein 3; D003546:Cysteine Endopeptidases; C513261:Tnfaip3 protein, mouse; D051818:Replication Protein C