Maria Luisa Veronese

B-cell activation during HIV-1 infection. III. Down-regulating effect of mitogens.

Spontaneous in vitro production of HIV-1-specific antibodies, a hallmark of infected subjects, is often down-regulated by the addition of pokeweed mitogen. We observed that a decrease in such ongoing anti-HIV-1 antibody synthesis could also be induced in cultures from most patients by addition of phytohemagglutinin and Concanavalin A, but not by Epstein-Barr virus, a selective B-cell mitogen. In most cases, this down-regulatory effect of mitogens was evident within the first 24 h of culture. The observed mitogen-associated decrease in spontaneous antibody synthesis was prevented by treating peripheral blood mononuclear cells with agents inhibiting non-major histocompatibility complex-restricted cytotoxic activity or by adding third-party cells to the cultures. In most cases, the mitogen-induced effect was also counteracted by removal of T lymphocytes or CD8+ T-cell sub-population. These findings recall a similar phenomenon observed in normal subjects following intentional immunization, and indicate that mitogen-induced down-regulation of spontaneous in vitro anti-HIV-1-antibody production most probably occurs through a lectin-dependent cytotoxic effect on activated B cells.

B cell activation and human immunodeficiency virus infection. V: Phenotypic and functional alterations in CD5+ and CD5- B cell subsets

B cell dysregulation is a hallmark of human immunodeficiency virus infection. Since B lymphocytes comprise two distinct subpopulations, CD5+ and CD5− cells, we addressed their individual phenotypic and functional behavior. Seropositive patients with both limited and advanced disease progression had an increased percentage of peripheral blood CD5+ B cells, compared to seronegative controls (20.1±2.1 and 22.7±5.7, respectively, vs 17.0±3.4 in controls); however, due to the lymphopenia and reduced number of circulating B cells in infected individuals, the absolute number of CD19+ CD5+ lymphocytes was actually reduced. Although HIV-specific antibodies were synthesized spontaneouslyin vitro only by CD5− B cells, a 10-fold lower degree of spontaneous, non-HIV-specific activation was also displayed by unstimulated CD5+ B cells. These findings indicate that B cell dysregulation during HIV infection involves both the CD5− and the CD5+ B cell compartments; moreover, in view of the putative role of CD5+ B cells in autoimmune phenomena and IL-10 production, these data reinforce the possibility that B cell dysfunction might be causally involved in AIDS pathogenesis.

Standardization of in vitro synthesis and detection of HIV-1-specific antibodies

Optimal conditions for in vitro anti-human immunodeficiency virus type 1 (HIV-1) antibody (Ab) synthesis and detection were re-appraised. Western blot (WB) and radioimmunoassay (RIA) could detect about 1 and 10 ng, respectively, of HIV-1-specific Ab (HIV-Ab), while the sensitivity of an enzyme-linked immunosorbent assay (ELISA) was much lower. Optimal HIV-Ab recovery was obtained by culturing 2.5 x 10(6) peripheral blood mononuclear cells (PBMC)/ml from seropositive subjects for 16 days in the absence of mitogens; at higher cell concentrations, background levels were unacceptably high. The background of non-de novo synthesized HIV-Ab was due to insufficient PBMC washing and/or cytophilic immunoglobulin (Ig); a particular washing procedure, as well as 24 h peripheral blood mononuclear cells (PBMC) pre-culture, might help in limiting this phenomenon. However, results should be compared with those obtained in cultures containing puromycin especially in infants, where a higher CD16 antigen expression in lymphocytes is likely responsible for increased amounts of cytophilic Ig released in culture supernatants, compared to adults.

Immune dysfunction in the elderly: Effect of thymic hormone administration on several in vivo and in vitro immune function parameters

The effects of short- term thymic hormone administration on age- associated immune function were evaluated. Two groups of individuals > 65 years of age were treated for 30 days with thymic extracts (TP1) or placebo; before and after this treatment a panel of in vitro and in vivo parameters was determined according to a very rigorous experimental protocol. In most individuals, TP1 treatment was associated with an improvement in cutaneous delayed- type response to PPD. Moreover, an increase in a circulating T cell subpopulation bearing the CD45R surface antigen (“virgin” T cells), and in NK cell cytotoxic activity was also observed in some subjects. Finally, lymphocyte responsiveness to PHA tended to increase, while no effect on lymphocyte ability to produce IL- 2 following mitogen stimulation was observed. These findings suggest that TP1 treatment may influence age- related alterations in immune function parameters in some subjects. (Again 2: 347–355, 1990)

Dominance of a single Epstein-Barr virus strain in SCID-mouse tumors induced by injection of peripheral blood mononuclear cells from healthy human donors

Severe Combined Immune Deficiency mouse tumors, induced by inoculating peripheral blood mononuclear cells from 11 healthy human donors (hu-PBMC-SCID tumors), were used to analyse Epstein-Barr virus (EBV) type and strain variations. PCR analysis of EBNA 2- and EBNA 3C-specific sequences showed that EBV type A was present in SCID-mouse tumors induced by PBMC from all donors but one, while, using amplimers for a highly polymorphic region within the latent membrane protein (LMP) coding sequence, 5 different strains could be detected among the samples examined. The same LMP fragment was present in different tumors arising in the same animal, as well as in different mice injected with PBMC from any donor. Compared to B95.8 and AG876 prototype viruses, sequence analysis of LMP variants disclosed a higher homology to the latter, with 33 bp additional repetitions and a few point mutations in specific sites. This study confirms and extends previous data on the presence of a single EBV type and strain in the peripheral blood of most normal healthy subjects using the SCID-mouse system.

B Cell Activation and HIV-1 Infection

The human immunodeficiency virus type 1 (HIV-1), the causative agent of the acquired immunodeficiency syndrome (AIDS) in man, shows tropism for CD4+ T cells mostly, and this accounts for the intense impairment in cellular immunity function (for review, see Rosenberg and Fauci 1989); however, B cell function is also severely deranged by HIV-1 infection (for review, see Amadori and Chieco-Bianchi 1990). Although the biologic properties of HIV-1, and its life cycle in the host are well understood, a comprehensive view of the pathogenesis of AIDS is still lacking, and it is still debated whether the T cell deficiency depends only on the virus’ cytopathic effect, or whether other mechanisms also come into play. Among the pathways proposed as co-factors in generating AIDS, the possibility that B cell deregulation might be involved is intriguing. This article reviews current knowledge on the features characterizing B cell function during HIV-1 infection, and addresses the possible participation of the humoral compartment in the pathogenesis of AIDS and associated disorders.