RESUMEN
The host response to Mycobacterium tuberculosis includes macrophage activation, inflammation with increased immune effector cells, tissue necrosis, and cavity formation, and fibrosis, distortion, and bronchiectasis. To evaluate the molecular basis of the immune response in the lungs of patients with active pulmonary tuberculosis (TB), we used bronchoalveolar lavage to obtain cells at the site of infection. Affymetrix GeneChip microarrays and cDNA nylon filter microarrays interrogated gene expression in bronchoalveolar lavage (BAL) cells from 11 healthy controls and 17 patients with active pulmonary TB. We found altered gene expression for 69 genes in TB versus normal controls that included cell surface markers, cytokines, chemokines, receptors, transcription factors, and complement components. In addition, TB BAL cell gene expression patterns segregated into 2 groups: one suggestive of a T helper type 1 (Th1) cellular immune response with increased signal transducer and activator of transcription-4 (STAT-4), interferon-gamma (IFN-gamma receptor), and monokine induced by IFN-gamma (MIG) expression with increased IFN-gamma protein levels in BAL fluid; the other group displayed characteristics of Th2 immunity with increased STAT-6, CD81, and IL-10 receptor expression. We were able to demonstrate that a Th2 presentation could change to a Th1 pattern after anti-tuberculous treatment in 1 TB patient studied serially. These gene expression data support the conclusion that pulmonary TB produces a global change in the BAL cell transcriptome with manifestations of either Th1 or Th2 immunity.
Asunto(s)
Líquido del Lavado Bronquioalveolar/citología , Tuberculosis Pulmonar/genética , Adulto , Líquido del Lavado Bronquioalveolar/inmunología , Citocinas/análisis , Femenino , Expresión Génica , Humanos , Masculino , Análisis por Micromatrices , Mycobacterium tuberculosis/genética , Tuberculosis Pulmonar/inmunologíaRESUMEN
BACKGROUND: Pulmonary tuberculosis (TB) can present with polymorphonuclear neutrophil (PMN)-predominant alveolitis. TB accelerates acquired immunodeficiency syndrome by increasing human immunodeficiency virus type 1 (HIV-1) replication and mutation in alveolar macrophages. A 16-kDa CCAAAT/enhancer-binding protein beta (C/EBP beta ) isoform is a strong transcriptional repressor of the HIV long terminal repeat (LTR) in resting alveolar macrophages, leading to latent viral infection; its expression is lost during TB, derepressing the HIV LTR. METHODS: Lung segments were sampled from HIV/Mycobacterium tuberculosis-coinfected patients by means of bronchoalveolar lavage. In vitro coculture experiments defined the mechanism of induction of HIV-1 infection in macrophages by PMNs. RESULTS: Lung segments from patients with PMN-predominant TB had a markedly elevated viral load. Direct contact between activated PMNs and macrophages stimulated HIV-1 replication and LTR transcription and down-regulated inhibitory C/EBP beta . Isolated PMN membranes substituted for PMN contact, derepressing the HIV-1 LTR. The lipid raft fraction of PMN membranes expressed CD40 ligand (CD40L), CD28, and leukocyte function-associated antigen 1 (LFA-1 [i.e., CD11a and CD18]), and PMN activation increased lipid raft expression of CD40L and CD28. Blocking antibodies to CD40L, CD28, and LFA-1 inhibited PMN membrane-mediated HIV-1 LTR derepression. Alternately, cross-linking of macrophage receptors for CD40L, CD28, and LFA-1 (CD40, CD80/86, and intercellular adhesion molecule 1) abolished inhibitory C/EBP beta expression. CONCLUSION: PMN-macrophage contact derepresses the HIV-1 LTR and enhances HIV-1 replication in alveolar macrophages during pulmonary TB. Derepression is mediated through costimulatory molecule signaling.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/virología , Proteínas de Unión al ADN/metabolismo , VIH-1/fisiología , Leucocitos Mononucleares/inmunología , Mycobacterium tuberculosis/fisiología , Tuberculosis Pulmonar/fisiopatología , Síndrome de Inmunodeficiencia Adquirida/complicaciones , Lavado Broncoalveolar , ADN Viral , Humanos , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/virología , Mutación , Tuberculosis Pulmonar/complicaciones , Replicación ViralRESUMEN
Opportunistic infections such as pulmonary tuberculosis (TB) increase local HIV-1 replication and mutation. As AIDS progresses, alteration of the HIV-1 gp120 V3 sequence is associated with a shift in viral coreceptor use from CCR5 (CD195) to CXCR4 (CD184). To better understand the effect of HIV/TB coinfection, we screened transcripts from bronchoalveolar lavage cells with high density cDNA arrays and found that CXCR4 mRNA is increased in patients with TB. Surprisingly, CXCR4 was predominately expressed on alveolar macrophages (AM). Mycobacterium tuberculosis infection of macrophages in vitro increased CXCR4 surface expression, whereas amelioration of disease reduced CXCR4 expression in vivo. Bronchoalveolar lavage fluid from TB patients had elevated levels of CCL4 (macrophage inflammatory protein-1beta), CCL5 (RANTES), and CX3CL1 (fractalkine), but not CXCL12 (stromal-derived factor-1alpha). We found that M. tuberculosis infection of macrophages in vitro increased viral entry and RT of CXCR4-using [corrected] HIV-1, but not of CCR5-using [corrected] HIV-1. Lastly, HIV-1 derived from the lung contains CD14, suggesting that they were produced in AM. Our results demonstrate that TB produces a permissive environment for replication of CXCR4-using virus by increasing CXCR4 expression in AM and for suppression of CCR5-using HIV-1 by increasing CC chemokine expression. These changes explain in part why TB accelerates the course of AIDS. CXCR4 inhibitors are a rational therapeutic approach in HIV/TB coinfection.
Asunto(s)
Quimiocinas CC/biosíntesis , Quimiocinas CX3C/biosíntesis , VIH-1/inmunología , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/virología , Mycobacterium tuberculosis/inmunología , Receptores de Quimiocina/biosíntesis , Replicación Viral/inmunología , Secuencia de Aminoácidos , Líquido del Lavado Bronquioalveolar/inmunología , Líquido del Lavado Bronquioalveolar/virología , Línea Celular Tumoral , Células Cultivadas , Quimiocinas CC/genética , Quimiocinas CC/fisiología , Quimiocinas CX3C/genética , Quimiocinas CX3C/fisiología , Regulación de la Expresión Génica/inmunología , VIH-1/aislamiento & purificación , VIH-1/patogenicidad , Humanos , Macrófagos Alveolares/metabolismo , Datos de Secuencia Molecular , Infecciones Oportunistas/inmunología , Infecciones Oportunistas/metabolismo , Infecciones Oportunistas/virología , ARN Mensajero/biosíntesis , Receptores CCR4 , Receptores de Quimiocina/fisiología , Especificidad de la Especie , Tuberculosis Pulmonar/inmunología , Tuberculosis Pulmonar/microbiología , Tuberculosis Pulmonar/virología , Regulación hacia Arriba/inmunologíaRESUMEN
Gamma interferon (IFN-gamma) is critical in the immune response against Mycobacterium tuberculosis. In an ongoing trial of aerosol IFN-gamma in conjunction with standard drug therapy, we have observed activation of IFN signaling in bronchoalveolar lavage (BAL) cells from tuberculosis (TB) patients. We hypothesized that aerosol IFN-gamma treatment of pulmonary TB would increase expression of genes important for the control of TB. We investigated the expression of downstream genes by measuring inducible nitric oxide synthase (iNOS) and the chemokine IFN-inducible 10-kDa protein (IP-10) by real-time quantitative reverse transcription-PCR. In vitro, M. tuberculosis induced IP-10, and IFN-gamma stimulated this further, with no effect on iNOS expression. We studied 21 patients with pulmonary TB and 7 healthy subjects. Similar to the in vitro model, IP-10 mRNA was increased in BAL cells from TB patients and was augmented after treatment with aerosolized IFN-gamma. TB was also associated with elevated iNOS mRNA, but aerosolized IFN-gamma did not further enhance expression. Genomic analysis identified 1,300 of 4,058 genes expressed in BAL cells from six TB patients before and after 1 month of therapy, including aerosolized IFN-gamma. However, only 15 genes were differentially regulated by IFN-gamma. We conclude that iNOS and IP-10 mRNA expression is increased in TB but that aerosol IFN-gamma treatment increases expression of few genes in the human lung.
Asunto(s)
Quimiocinas CXC/genética , Interferón gamma/farmacología , Óxido Nítrico Sintasa/genética , Tuberculosis Pulmonar/terapia , Adulto , Aerosoles , Secuencia de Bases , Líquido del Lavado Bronquioalveolar/citología , Líquido del Lavado Bronquioalveolar/inmunología , Estudios de Casos y Controles , Quimiocina CXCL10 , ADN Complementario/genética , Femenino , Expresión Génica/efectos de los fármacos , Humanos , Técnicas In Vitro , Interferón gamma/administración & dosificación , Pulmón/efectos de los fármacos , Pulmón/inmunología , Masculino , Persona de Mediana Edad , Óxido Nítrico Sintasa de Tipo II , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Recombinantes , Tuberculosis Pulmonar/genética , Tuberculosis Pulmonar/inmunologíaRESUMEN
Tuberculosis leads to immune activation and increased human immunodeficiency virus type 1 (HIV-1) replication in the lung. However, in vitro models of mycobacterial infection of human macrophages do not fully reproduce these in vivo observations, suggesting that there are additional host factors. Surfactant protein A (SP-A) is an important mediator of innate immunity in the lung. SP-A levels were assayed in the human lung by using bronchoalveolar lavage (BAL). There was a threefold reduction in SP-A levels during tuberculosis only in the radiographically involved lung segments, and the levels returned to normal after 1 month of treatment. The SP-A levels were inversely correlated with the percentage of neutrophils in BAL fluid, suggesting that low SP-A levels were associated with increased inflammation in the lung. Differentiated THP-1 macrophages were used to test the effect of decreasing SP-A levels on immune function. In the absence of infection with Mycobacterium tuberculosis, SP-A at doses ranging from 5 to 0.01 micro g/ml inhibited both interleukin-6 (IL-6) production and HIV-1 long terminal repeat (LTR) activity. In macrophages infected with M. tuberculosis, SP-A augmented both IL-6 production and HIV-1 LTR activity. To better understand the effect of SP-A, we measured expression of CAAT/enhancer binding protein beta (C/EBPbeta), a transcription factor central to the regulation of IL-6 and the HIV-1 LTR. In macrophages infected with M. tuberculosis, SP-A reduced expression of a dominant negative isoform of C/EBPbeta. These data suggest that SP-A has pleiotropic effects even at the low concentrations found in tuberculosis patients. This protein augments inflammation in the presence of infection and inhibits inflammation in uninfected macrophages, protecting uninvolved lung segments from the deleterious effects of inflammation.
Asunto(s)
Inflamación/etiología , Macrófagos/inmunología , Proteína A Asociada a Surfactante Pulmonar/fisiología , Tuberculosis/inmunología , Proteína beta Potenciadora de Unión a CCAAT/fisiología , Duplicado del Terminal Largo de VIH , Humanos , Interleucina-6/biosíntesis , Proteína A Asociada a Surfactante Pulmonar/análisisRESUMEN
Tuberculosis is the seventh leading cause of morbidity and mortality in the world, with eight million cases per year. Animal and human studies demonstrate an enrichment of CD4 cells at sites of disease, with a more favorable clinical course when there is a Th1 response with the presence of gamma interferon (IFN-gamma). We previously treated patients who had multidrug-resistant tuberculosis with recombinant IFN-gamma (rIFN-gamma) in aerosol form and were able to convert smear-positive cases to smear negative with 12 treatments over 1 month. We hypothesized that rIFN-gamma would induce signal transducer and activator of transcription (STAT) and interferon regulatory factor (IRF) binding activity in alveolar macrophages (AM). AM treated in vitro showed clear upregulation of STAT-1 and IRF-1 by rIFN-gamma. STAT-1 was not activated and IRF-1 was only weakly induced after 1 day of infection by Mycobacterium tuberculosis TN913. In bronchoalveolar lavage (BAL) cells obtained from 10 of 10 tuberculosis patients 10 +/- 2 days post-antituberculosis treatment, there was no detectable STAT-1 or IRF-1 DNA-binding activity. After 4 weeks of treatment with rIFN-gamma aerosol in addition to the antituberculosis drugs, 10 of 10 patients had increased STAT-1, IRF-1, and/or IRF-9 DNA-binding activity in BAL cells from lung segments shown radiographically to be involved and in those shown to be uninvolved. Symptoms and chest radiographs improved, and amounts of macrophage inflammatory cytokines and human immunodeficiency virus type 1 (HIV-1) viral loads (in five of five HIV-1-coinfected patients) declined in the second BAL specimens. rIFN-gamma aerosol induces signal transduction and gene expression in BAL cells and should be evaluated for efficacy in a randomized, controlled clinical trial.