RESUMEN
In this study, we demonstrated the efficiency and feasibility of a gene therapy protocol against HIV infection using the antiviral effects of IFN-beta expression. Lentiviral vectors containing the human or the simian IFN-beta sequences under the influence of the murine moderate H2-kb promoter were constructed. To examine the capacity of IFN-beta to inhibit the replication of HIV in human CD4(+) cells, a transduction protocol permitting to efficiently transduce CD4(+) cells or PBMC (85+/-12% of CD4(+)-transduced cells) with a moderate expression of IFN-beta was developed. Results indicate that enforced expression of IFN-beta has no negative effects in terms of apoptosis and proliferation. In human CD4(+) cells, it drastically inhibits (up to 99.9%) replication after challenging with different strains of HIV-1. The expression of exogenous IFN-beta leads to an amplification of the CD4(+) cells (11-fold) and to a drastic decrease of the p24 protein. Micro-array analyses indicated that antiviral effect of IFN-beta could be due to a major regulation of the inflammatory response. These results are encouraging for the development of a clinical study of gene therapy against AIDS using IFN-beta.
Asunto(s)
Linfocitos T CD4-Positivos/metabolismo , Terapia Genética/métodos , Infecciones por VIH/terapia , VIH-1/efectos de los fármacos , Interferón beta/administración & dosificación , Replicación Viral/efectos de los fármacos , Animales , Antivirales/administración & dosificación , Antivirales/farmacología , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/trasplante , Proliferación Celular/efectos de los fármacos , Vectores Genéticos , Haplorrinos , Humanos , Inflamación/tratamiento farmacológico , Interferón beta/farmacología , Ratones , Transducción GenéticaRESUMEN
BACKGROUND: An attenuated immunodeficiency virus has been long considered innocuous. Nevertheless, converging data suggest that low levels of viral replication can still provoke AIDS. Pathogenesis of these attenuated infections is not understood. OBJECTIVES: To determine the pathogenicity of a long-term attenuated infection and to delineate T-cell dynamics during such an infection. METHODS: This is a cross-sectional study of 12 rhesus macaques infected with SIV Delta nef for 8 years. We evaluated apoptosis (annexin V), activation (HLA-DR, Ki67), and newly generated T cells (TCR excision circle: TREC). RESULTS: Infection with SIV Delta nef induced pathological CD4 T-cell depletion after 8 years of infection. Virus replication and CD8 T-cell activation positively correlated with the rate of disease progression. The frequency of TREC within CD8+CD45RA+ cells increased in SIV Delta nef-infected animals compared to age-matched non-infected controls. Moreover, in the cohort of infected animals, TREC+CD45RA+CD4+ T-cell counts correlated strongly with non-progression to AIDS. The animal with the lowest rate of disease progression exhibited a 115-fold increase in TREC+CD45RA+CD4+ T-cell counts compared to age-matched non-infected controls. In contrast, the animal showing the fastest rate of progression to AIDS displayed 600-fold lower TREC+CD45RA+CD4+ T-cell counts compared to age-matched non-infected controls. CONCLUSIONS: Our results suggest that the thymus plays a major role in the pathogenesis of an attenuated SIV infection and that a sustained thymic output could maintain CD4 T-cell homeostasis in the context of low viral loads.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida del Simio/virología , Virus de la Inmunodeficiencia de los Simios/fisiología , Animales , Apoptosis , Recuento de Linfocito CD4 , Relación CD4-CD8 , Linfocitos T CD8-positivos , Proliferación Celular , Progresión de la Enfermedad , Productos del Gen nef , Reordenamiento Génico de Linfocito T , Antígenos Comunes de Leucocito/análisis , Activación de Linfocitos , Macaca mulatta , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Factores de Tiempo , Carga Viral , Replicación ViralRESUMEN
We assessed the possible role in vivo CD4(+) CD8(+) T cells as a viral reservoir for simian immunodeficiency virus (SIV), in a macaque with 50% CD4(+) CD8(+) T cells in peripheral blood. During primary infection (day 14) of this rhesus macaque with the pathogenic SIVmac251 strain, proviruses were detected at similar frequencies in CD4(+) CD8(+) T cells (1/10) and CD4(+) T cells (1/10) and at a lower frequency in CD8(+) T cells (1/800). On day 235, no viral DNA was detected in CD8(+) cells, despite the persistent high viral load, indicating that CD8(+) cells do not constitute a reservoir during the chronic phase of SIV infection. Infection induced early lymphopenia of CD4(+), CD4(+) CD8(+), and CD8(+) cells; only the CD8(+) cell population returned to initial levels and expanded further. We found that CD4(+) CD8(+) T cells expressed the costimulatory CD28 molecule less and were more prone to die in vitro after phytohemagglutinin/interleukin 2 stimulation than were CD4(+) T cells. Taken together, massive death of CD4(+) CD8(+) T cells during acute stages of SIV infection may explain why CD8(+) T cells did not represent a major reservoir for SIV at the onset of infection.