IgG subclasses of FVIII inhibitors in an argentine cohort of severe hemophilia A patients: Analysis by flow cytometry.

Introduction: FVIII inhibitors consist of a polyclonal population of antibodies. Previous studies have demonstrated different distribution of IgG subclasses. IgG4 was associated to high level of FVIII inhibitors and failure of immune tolerance induction (ITI) treatment.

Common Variable Immunodeficiency and Circulating TFH.

CD4+ T follicular helper cells (TFH) were assessed in adult patients with common variable immune deficiency (CVID) classified according to the presence of granulomatous disease (GD), autoimmunity (AI), or both GD and AI (Group I) or the absence of AI and GD (Group II). TFH lymphocytes were characterized by expression of CXCR5 and PD-1. TFH were higher (in both absolute number and percentage) in Group I than in Group II CVID patients and normal controls (N).

Paroxysmal nocturnal haemoglobinuria. Experience over a 10 years period.

Introduction: Paroxysmal nocturnal hemoglobinuria (PNH) is a hemolytic, clonal and acquired disorder of the hematopoietic stem cell with a deficiency of all glycophosphatidyl-inositol (GPI) linked proteins. The aim of this retrospective study was to analyse haematological and biochemical data from 152 patients referred to our laboratory for diagnosis of PNH by flow cytometry (FC).

Methods: Patients and healthy donor (152 and 99 respectively) were studied.

CD4+ T Lymphocytes with follicular helper phenotype (T(FH)) in patients with SH2D1A deficiency (XLP).

Peripheral blood mononuclear cells with T(FH) phenotype from two asymptomatic XLP patients were studied. Normal/high numbers of CXCR5+, CD4+ T cells coexpressing PD-1 were demonstrated. Peripheral blood mononuclear cells (PBMC) from these patients responded to sub-optimal PHA/IL-2 stimulation upregulating ICOS and CD40L and increasing intracellular expression of IL-10, IL-21 and IL-4 by CD4+ T(FH) cells.

Severe haemophilia A patients have reduced numbers of peripheral memory B cells.

The development of inhibitors is a complication of replacement treatment in Haemophilia. Loss of factor VIII-specific memory B cells in the spleen is associated with down regulation of antibodies in mice treated with high doses of FVIII, but changes in B cell memory have not been described in haemophilic patients. The aim of this study was to evaluate the phenotype of circulating lymphocytes in severe haemophilia A.

A flow cytometry evaluation of anti-FVIII antibodies: correlation with ELISA and Bethesda assay.

In this study, we describe a flow cytometry (FC) system for detecting antibodies to factor VIII (FVIII) and compare its results with those of enzyme-linked immunosorbent assay (ELISA) that detects both inhibitory (I-Ab) and non-inhibitory (NI-Ab) antibodies and the Nijmegen modification of the Bethesda method, detecting I-Ab. FC was set up in our laboratory. Recombinant FVIII (rFVIII) was coupled to microspheres (FVIII-m) and reacted with different plasma dilutions.

[Neutrophils without CD16b receptors].

Occurrence of the rare CD16b deficiency ("null" phenotype) in neutrophils from two female patients (41 and 15 years old) is reported. The first case was referred with a diagnosis of anemia related to paroxistic nocturnal hemoglobinuria and the second case, with presumptive diagnosis of immune neutropenia. In both cases, absence of CD16b expression was determined by flow cytometry without deficiencies of other neutrophil alloantigens or defects of membrane anchorage through glycosil phosphatydil inositol (GPI) linkage.

Increased lymphocyte viability after non-stimulated peripheral blood mononuclear cell (PBMC) culture in patients with X-linked lymphoproliferative disease (XLP).

Survival of lymphocytes after prolonged culture was studied in two asymptomatic XLP patients. Viability of XLP PBMC after 30 days of non-stimulated culture was higher than that of normal controls (N), mainly due to the persistence of CD8 memory lymphocytes. IFNgamma high CD8 T lymphocytes remained higher in XLP than in N after 30 days.

Loss of circulating CD27+ memory B cells and CCR4+ T cells occurring in association with elevated EBV loads in XLP patients surviving primary EBV infection.

Detailed longitudinal studies of patients with X-linked lymphoproliferative disease (XLP) may increase our understanding of the immunologic defects that contribute to the development of lymphoma and hypogammaglobulinemia in XLP. We describe progressive changes observed in immunoglobulin concentrations, lymphocyte subsets, and Epstein-Barr virus (EBV) loads occurring in a 2-year period in a newly infected, but otherwise healthy, carrier (patient 9). We compare these findings with those observed in the patient's brother, who had hypogammaglobulinemia and XLP (patient 4).

Interleukin-2 restores natural killer activity inhibited by sera from HIV+ hemophilic patients.

Natural killer (NK) activity is impaired in patients with positive serology for the human immunodeficiency virus (HIV). We previously found an inhibitory effect of sera from hemophilic (He) HIV+ patients on normal NK activity. In the present study, we have further characterized this effect by studying its reversibility, temperature and time incubation dependence.

Increased CD4-positive monocytes in HIV-infected haemophilic patients.

The monocyte-macrophage system is known to play a central role in HIV infection, and expression of CD4 on the surface of monocytes/macrophages is important, since this molecule is a key factor for the entrance of HIV into susceptible cells. In this paper we evaluated the expression of CD4 in monocytes of haemophilic patients (He) who had been infected with HIV (HIV + He) through transfusion of contaminated plasma concentrates. Thirty seropositive patients (HIV + He), 10 seronegative He patients (HIV-He) and 20 voluntary normal blood donors were studied.

Inhibition of normal natural killer cytotoxicity by sera from hemophilic patients.

In this study we searched for circulating antibodies or other serum factors that could account for the natural killer (NK) defect observed in hemophiliacs (He) infected with the human immunodeficiency virus (HIV). We analyzed the effect of negative or positive sera for HIV from He on normal NK activity. We showed that sera from He interfered with normal NK cytotoxicity.

HIV infection and natural killer cytotoxicity in hemophilic patients.

In this study we analyzed the ability of peripheral blood mononuclear cells (PBMC) from hemophilic patients (He) with negative or positive serology for the human immunodeficiency virus (HIV), to increase natural killer (NK) cytotoxicity upon stimulation with physiological and non physiological agents. Purified interleukin-2 (IL-2), the interferon (IFN)-inducer polyinosinic polycytidylic acid (PIC), recombinant alpha- and gamma-IFN and the protein kinase activator phorbol myristate acetate (PMA) were used as stimulatory agents. The NK functional response was correlated with the presence of PBMC bearing phenotypic markers of activated cells (IL-2 receptor, IL-2R) and of different NK cell maturation stages.

Effect of lymphokines on natural killer cytotoxicity in patients with high risk of developing the acquired immune deficiency syndrome.

Regulation of natural killer (NK) activity of peripheral blood mononuclear cells (PBMC) from patients with the acquired immune deficiency syndrome (AIDS) and from individuals at high risk of developing AIDS (R-AIDS) was studied. The response of untreated PBMC to the interferon inducer polyinosinic polycytidilic acid (Poly I:C) was lower in AIDS and R-AIDS than in normal controls and PBMC from R-AIDS were more susceptible to stimulation with lymphokine rich supernatants (Con A-SN, PHA-SN, lectin free IL-2) than AIDS and normal controls. To determine the role of the different T lymphocyte subsets in the regulation of NK activity, PBMC were selectively treated with monoclonal non-cytotoxic anti-Leu 2a and anti-Leu 3a antibodies and then stimulated with lymphokine rich supernatants.