RESUMEN
Interleukin 6 (IL-6) is considered a proliferation and survival factor for B cells. To assess the role of IL-6 in Kaposi sarcoma-associated herpesvirus (KSHV) latency, KSHV latency locus-transgenic mice (referred to as latency mice) lacking IL-6 were evaluated. IL-6(-/-) latency mice had the same phenotypes as the latency mice, i.e., increased frequency of marginal zone B cells, hyperplasia, and hyperglobulinemia, indicating that the KSHV latency locus, which includes all viral microRNAs (miRNAs), can compensate for lack of IL-6 in premalignant B cell activation.
Asunto(s)
Linfocitos B/virología , Herpesvirus Humano 8/fisiología , Interleucina-6/metabolismo , Activación de Linfocitos , Latencia del Virus , Animales , Interleucina-6/deficiencia , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones TransgénicosRESUMEN
To maintain tolerance, autoreactive B cells must regulate signal transduction from the BCR and TLRs. We recently identified that dendritic cells and macrophages regulate autoreactive cells during TLR4 activation by releasing IL-6 and soluble CD40 ligand (sCD40L). These cytokines selectively repress Ab secretion from autoreactive, but not antigenically naive, B cells. How IL-6 and sCD40L repress autoantibody production is unknown. In this work, we show that IL-6 and sCD40L are required for low-affinity/avidity autoreactive B cells to maintain tolerance through a mechanism involving receptor cross-talk between the BCR, TLR4, and the IL-6R or CD40. We show that acute signaling through IL-6R or CD40 integrates with chronic BCR-mediated ERK activation to restrict p-ERK from the nucleus and represses TLR4-induced Blimp-1 and XBP-1 expression. Tolerance is disrupted in 2-12H/MRL/lpr mice where IL-6 and sCD40L fail to spatially restrict p-ERK and fail to repress TLR4-induced Ig secretion. In the case of CD40, acute signaling in B cells from 2-12H/MRL/lpr mice is intact, but the chronic activation of p-ERK emanating from the BCR is attenuated. Re-establishing chronically active ERK through retroviral expression of constitutively active MEK1 restores tolerance upon sCD40L, but not IL-6, stimulation, indicating that regulation by IL-6 requires another signaling effector. These data define the molecular basis for the regulation of low-affinity autoreactive B cells during TLR4 stimulation; they explain how autoreactive but not naive B cells are repressed by IL-6 and sCD40L; and they identify B cell defects in lupus-prone mice that lead to TLR4-induced autoantibody production.
Asunto(s)
Autoanticuerpos/biosíntesis , Subgrupos de Linfocitos B/inmunología , Subgrupos de Linfocitos B/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Nefritis Lúpica/metabolismo , Receptor Cross-Talk/inmunología , Receptor Toll-Like 4/fisiología , Animales , Subgrupos de Linfocitos B/patología , Antígenos CD40/metabolismo , Ligando de CD40/metabolismo , Núcleo Celular/enzimología , Núcleo Celular/metabolismo , Células Cultivadas , Técnicas de Cocultivo , Femenino , Tolerancia Inmunológica/genética , Nefritis Lúpica/enzimología , Nefritis Lúpica/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos MRL lpr , Ratones Transgénicos , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/metabolismo , Transporte de Proteínas/genética , Transporte de Proteínas/inmunología , Receptores de Antígenos de Linfocitos B/fisiología , Receptores de Interleucina-6/metabolismo , Transducción de Señal/genética , Transducción de Señal/inmunologíaRESUMEN
Antimicrobial peptides, such as cathelicidin and beta defensins, directly kill microbes and have been detected in human sebaceous glands and cell lines. Despite the presence of several such peptides, the apparent abundance of these is insufficient for direct killing of most skin pathogens. In this study, we sought to determine which molecules provide the majority of antimicrobial peptide activity in human sebocytes. Acid-soluble protein extracts of SEB-1 sebocytes were separated by reverse-phase high-performance liquid chromatography and were assayed for their capacity to inhibit the growth of Staphylococcus aureus. Antimicrobial activity was isolated in a single major fraction and identified to be histone H4 by mass spectrometry and western blot analysis. The importance of histone H4 in the antimicrobial activity of sebocytes was confirmed by a specific neutralizing antibody and by direct demonstration that recombinant histone H4 had antimicrobial activity against S. aureus and Propionibacterium acnes. In addition, histone H4 enhanced the antimicrobial action of free fatty acids in human sebum. Taken together, these results indicate that the release of histone H4 by holocrine secretion from the sebaceous gland may play an important role in innate immunity.
Asunto(s)
Antiinfecciosos/metabolismo , Histonas/metabolismo , Glándulas Sebáceas/metabolismo , Secuencia de Aminoácidos , Antiinfecciosos/farmacología , Línea Celular , Células Cultivadas , Histonas/análisis , Histonas/farmacología , Humanos , Inmunidad Innata/fisiología , Datos de Secuencia Molecular , Propionibacterium acnes/efectos de los fármacos , Glándulas Sebáceas/citología , Staphylococcus aureus/efectos de los fármacosRESUMEN
The accelerated development of atherosclerosis with increased risk of cardiovascular disease in systemic lupus erythematosus (SLE) patients is not well understood. An appropriate mouse model would greatly help to understand the mechanisms of this association. We have therefore combined the ApoE(-/-) model of atherosclerosis with three different murine models of SLE. We found that induction of cGVH in B6.ApoE(-/-) mice, breeding a Fas null gene onto the B6.ApoE(-/-) mice, and breeding the ApoE(-/-) defect onto MRL/lpr mice all caused a modest increase of atherosclerosis at 24 weeks of age compared to B6.ApoE(-/-) controls. B cells in B6.ApoE(-/-) mice had certain phenotypic differences compared to congenic C57BL/6 mice, as indicated by high expression of MHC II, Fas, CD86, and by increased number of cells bearing marginal zone phenotype. Furthermore, B6ApoE(-/-) mice had significant titers of anti-oxLDL and anti-cardiolipin autoantibodies compared to their B6 counterparts. Our studies also indicate that, following induction of cGVH, marginal zone B cells in B6.ApoE(-/-) are depleted, and there is considerable increase in anti-oxLDL and anti-cardiolipin abs along with secretion of lupus-specific autoantibodies, such as anti-dsDNA and anti-chromatin abs. Histological sections showed that cGVH and/or Fas deficiency could exacerbate atherosclerosis. The production of anti-oxLDL and anti-cardiolipin in ApoE(-/-) mice was also increased. These observations define a connection between induction of lupus-like symptoms and development of severe atherosclerosis in ApoE deficient lupus mouse models.
Asunto(s)
Apolipoproteínas E/deficiencia , Aterosclerosis/patología , Lupus Eritematoso Sistémico/patología , Animales , Anticuerpos Antinucleares/inmunología , Anticuerpos Antifosfolípidos/inmunología , Complejo Antígeno-Anticuerpo/inmunología , Aorta/inmunología , Apolipoproteínas E/genética , Apolipoproteínas E/inmunología , Aterosclerosis/inmunología , Aterosclerosis/metabolismo , Linfocitos B/inmunología , Colesterol/sangre , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Histocitoquímica , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos MRL lpr , Ratones Noqueados , Bazo/citología , Bazo/inmunologíaRESUMEN
Nuclear autoantigens in systemic lupus erythematosus are thought to derive primarily from apoptotic cells, yet there is no direct evidence that interfering with apoptosis impairs the generation of lupus autoantibodies. Here we use a mouse model that lacks the endonuclease caspase-activated DNase (CAD), resulting in an absence of chromatin and nuclear fragmentation during apoptotic cell death. We show that in this mouse, production and release into circulation of chromatin is impaired after exposure to several apoptotic triggers, but that the absence of CAD does not interfere with upstream steps of apoptosis or immune system function. Finally we show that in CAD-mutant mice, impaired lupus autoimmunity is skewed toward known cytoplasmic components, and autoimmunity toward membrane autoantigens is preserved, while autoimmunity toward chromatin and other lupus nuclear targets is severely impaired or absent. We also show, as control, that the induction of experimental autoimmune encephalomyelitis is not affected by the absence of CAD. Thus, our work in vivo strongly suggests that apoptotic molecular steps during cell death generate nuclear autoantigens to sustain the specific autoimmune response in systemic lupus erythematosus.
Asunto(s)
Anticuerpos Antinucleares/biosíntesis , Cromatina/inmunología , Fragmentación del ADN , Lupus Eritematoso Sistémico/inmunología , Animales , Anticuerpos Antinucleares/inmunología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Enfermedades Autoinmunes/inmunología , Células Cultivadas , Cromatina/metabolismo , Desoxirribonucleasas/genética , Modelos Animales de Enfermedad , Rayos gamma , Inmunoglobulina G/inmunología , Lipopolisacáridos/administración & dosificación , Lupus Eritematoso Sistémico/inducido químicamente , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfolípidos/inmunología , Terpenos/administración & dosificaciónRESUMEN
Autoreactive B cells may become activated in a T-independent manner via synergistic engagement of the BCR and TLRs. Using the VH3H9 Ig H chain transgene to track anti-chromatin B cells, we demonstrate that VH3H9/Vlambda1 anti-chromatin B cells proliferate in response to stimulatory oligodeoxynucleotides containing CpG motifs, suggesting that these autoreactive B cells are responsive to TLR9 signaling. Strikingly, some VH3H9 B cells, but not the well-characterized VH3H9/Vlambda1 B cells, proliferate spontaneously in culture medium. This proliferation is blocked by inhibitory CpG oligodeoxynucleotides, implicating the TLR9 (or possibly TLR7) pathway. Most hybridomas generated from the proliferating cells are polyreactive, and one exhibits binding to nuclear Ags but not to the other Ags tested. Thus, B cells carrying autoreactive and/or polyreactive specificities may be susceptible to T cell-independent activation via dual engagement of the BCR and TLRs.