1-2-dielaidoylphosphatidylethanolamine and Lupus-Erythematosus--Systemic

To investigate the ability of systemic lupus erythematosus (SLE) autoantigen- and Sjögren's syndrome (SS) autoantigen-associated U1 small nuclear RNA (U1 snRNA) and hY1RNA to induce interferon-alpha (IFNalpha) production.. In vitro-transcribed U1 snRNA or hY1RNA and lipofectin were added to peripheral blood mononuclear cell (PBMC) cultures. Purified U1 snRNP particles and IgG from SLE patients (SLE-IgG) were added to cultures of PBMCs, enriched monocytes, or natural interferon-producing cells (NIPCs); the latter are also known as plasmacytoid dendritic cells (pDC). Cells were double-stained for IFNalpha and either blood dendritic cell antigen 2 (NIPCs/pDC) or CD14 (monocytes) and then analyzed by flow cytometry. In some experiments, RNase or inhibitors of Fc gamma receptor IIa (Fc gammaRIIa) (specific antibodies), endocytosis (chloroquine, bafilomycin A), or Toll-like receptors (TLRs; oligodeoxynucleotide 2088) were used. The produced IFNalpha was measured by immunoassay.. Lipofected U1 snRNA and hY1RNA both induced IFNalpha production in monocytes, but not in NIPC/pDC. In contrast, U1 snRNP combined with SLE-IgG induced IFNalpha production only in NIPCs/pDC, and this response was decreased by RNase treatment or inhibition of the Fc gammaRIIa, the endocytosis pathways, or the TLRs.. Our finding that U1 snRNA and hY1RNA have IFNalpha-inducing capacity indicates that immune complexes containing such RNA, for example U1 snRNP particles, can be at least partly responsible for the ongoing IFNalpha production seen in SLE and SS. These results may help to explain the molecular mechanisms behind the pathogenesis of these and other autoimmune diseases in which autoantibodies to RNA-binding proteins occur.

Topics: Antigen-Antibody Complex; Antigens, CD; Autoantigens; Cells, Cultured; Flow Cytometry; Humans; Immunoglobulin G; Interferon-alpha; Lupus Erythematosus, Systemic; Monocytes; Phosphatidylethanolamines; Receptors, IgG; RNA, Small Nuclear; Sjogren's Syndrome

The oligodeoxyribonucleotide (ODN) 5'-TTTTCAATTCGAAGATGAAT-3' (ODN H), identified in systemic lupus erythematosus (SLE) serum, induced the production of interferon (IFN)-alpha in human peripheral blood mononuclear cells (PBMC) when combined with lipofectin. Flow cytometric analysis with staining for surface antigens and intracellular IFN-alpha, showed that the IFN-alpha-producing cells (IPC) were the natural IPC, also termed type 2 dendritic cell precursors (pDC2) or plasmacytoid monocytes. The importance of unmethylated CpG dinucleotides for the interferogenic activity of ODN was studied. Methylation of CpG impaired the activity of single-stranded (ss) ODN H, but increased that of the complementary ssODN I. Furthermore, CpG-methylated double-stranded (ds) ODN Hmet-Imet lost, but hemimethylated dsODN H-Imet retained interferogenic activity. Inversion of the CpG to GpC had no effect on the interferogenic activity of ssODN H, increased that of ssODN I, however abolished the activity of dsODN H-I. Alteration of the CpG in ODN H to ApG and in the ODN I to CpT destroyed their activity. The induction of IFN-alpha is therefore sequence-specific, but unmethylated CpGs are not always required, especially not in ssODNs. Interferogenic DNA sequences could therefore be more frequent in eukaryotic genomes than previously thought and their capacity to activate natural IPC may have implications for immune responses to microbial antigens and nuclear autoantigens.

Topics: Antigens; Autoantigens; Base Sequence; CpG Islands; Humans; In Vitro Techniques; Interferon Inducers; Interferon-alpha; Leukocytes, Mononuclear; Lupus Erythematosus, Systemic; Oligodeoxyribonucleotides; Phosphatidylethanolamines