Immune activation during HIV-1 infection : implication for B cell dysfunctions and therapy monitoring

Sammanfattning: Aberrant activation of cells of the immune system plays a major role in the pathogenesis of HIV infection. The major aims of the thesis were: (a) to measure the effects of current antiretroviral therapy on immune activation through the analysis of soluble activation markers; (b) to evaluate the extent of T and B cell immune activation in relation to immunological and virological parameters; (c) to analyse the T-B cell molecular pathway leading to hyperactivation of B cells and loss of B cell function. The goals of highly active antiretroviral therapy (HAART) are to suppress viral replication and to reduce immune activation, thereby stimulating recovery of immune functions We evaluated two soluble molecules (Fas and CD27) as markers of immune activation during HAART. In a longitudinal study of 69 patients we observed a minimal decrease of the sFas plasma levels after one year of therapy. No significant changes in sFas were observed between virological responders and non-responders. These findings suggested that long-term HAART may be required to norm~ the dysregulation of the Fas pathway. Analysis of plasma sCD27 was done on 26 HIV- 1-infected patients undergoing HAART during 24 months of therapy. HAART induced a significant reduction, but not a normalization, of plasma sCD27 after 24 months. A full normalization of plasma sCD27 was observed in virological responders and in patients with moderate immunodeficiency at baseline. Baseline sCD27 levels were predictive of a greater increase in CD4+ T cell count during the follow-up. Discontinuation of therapy resulted in a rapid increase of sCD27 plasma levels associated with viremia rebound and drop in CD4+ T cell count. Our findings suggested that plasma sCD27 may represent an alternative and simple marker to monitor immune activation during potent antiretroviral therapy. Polyclonal B cell activation is a major hallmark of the B cell dysfunction in HIV-1-infected patients. The phenotype and function of different B cell subsets were analysed. The population of memory (CD27+) B lymphocytes was dramatically reduced in HIV-1-infected patients. A positive correlation was found between the memory B cells and the CD70+ T cells, suggesting that T-B cell interaction through CD70-CD27 is upregulated and may chronically lead to terminal differentiation of memory B cells. Both naive and memory B lymphocytes from HIV-1-infected patients expressed high levels of Fas and Fas ligand although this pathway did not appear to be involved in apoptosis of memory B cells during HIV infection. Detection of NGF, a survival factor for memory B cells, was significantly reduced in the patients with low memory B cells. Furthermore, addition of NGF in culture resulted in decreased cell death of memory B cells from 2 of 4 infected subjects, indicating that deprivation of NGF may partly account for the loss of memory B cells. Despite the loss of memory B cells, total B lymphocytes from infected individuals showed a normal to high responsiveness to in vitro stimulation since they produced high amount of IgG upon activation with either agonistic anti- CD40 mAb or co-culture with a CD70transfected cell line. We also evaluated the relationship between specific humoral immunity and loss of memory B cells in infected patients. Among the HIV-1-infected subjects, we found a decrease in antibody levels to tetanus, measles and HIV-1 in patients with very low memory B cells. Despite the selective loss of antibodies, hypergammaglobulinemia, was detected in all patients. These observations suggest that polyclonal B cell activation is accompanied by loss of specific antibodies during HIV-1 infection. Together with the loss of memory B cells, these defects might account for the increased susceptibility to secondary/opportunistic infections and to poor response to immunisation against non HIV antigens observed in HIV-1-infected subjects. We also analysed the expression of CD70 and LAIR-1 on naive and memory B cells from HIV-1- infected and uninfected subjects. We observed that in the HIV-1-infected individuals naive B cells showed increased levels of CD70 that is otherwise barely detectable in healthy subjects. Since only CD70+ B cells produce antibodies in a T-cell dependent fashion, activated naive B cells might represent one possible source of antibody production in HIV-1 infection. On the other hand, memory B cells from infected subjects showed a significantly reduced expression of LAIR-1, whose expression is normally downregulated during terminal differentiation.