Josef W. Mannhalter

Effect of Polymeric IgG on Human Monocyte Functions

Fc and iC3b receptors are involved in various biological functions of phagocytic cells, such as immune adherence and phagocytosis of opsonized particles, degranulation and superoxide generation. In the present study we examined the expression of specific receptors for the Fc portion of IgG (FcR) and for iC3b (CR3), a cleavage product of the third complement component, on human monocytes following in vitro treatment with polymeric and monomeric IgG. Interaction of polymeric IgG (fluid phase) with the monocyte membrane led to a concomitant modulation of both Fc and iC3b receptors. Monomeric IgG, however, down modulated Fc receptor expression only if surface bound. Under these conditions, no concomitant modulation of the iC3b receptor could be observed. The down modulation of Fc and iC3b receptors induced by fluid-phase IgG polymers was also accompanied by a decrease in monocyte functions as expressed by reduced Fc receptor-mediated phagocytosis, decreased release of oxygen metabolites following stimulation by aggregated IgG and opsonized zymosan, as well as in impaired killing of bacteria. These data suggest that a down modulation of Fc and iC3b receptors might have important implications for host defense mechanisms, since interaction with these receptors is required for the proper elimination of many pathogens.

A functional defect in the early phase of the immune response observed in patients with hemophilia A

In search of a functional immunological defect in patients with hemophilia A, we investigated the early phase of the immune response and found a deficiency in the monocyte-T cell interaction after in vitro exposure to a bacterial antigen. T-cell proliferation in response to Escherichia coli-pulsed autologous monocytes was significantly (P less than 0.025) reduced in hemophiliacs [mean dpm +/- SEM (n = 25): 9036 +/- 2600] as compared to healthy controls [mean dpm +/- SEM (n = 24): 17,812 +/- 2985]. This functional defect was expressed in a severe form (antigen-induced monocyte-T cell interaction less than or equal to 2000 dpm) in both HTLV III antibody-positive and -negative patient populations. Significantly decreased T-helper/T-suppressor-cell ratios (mean dpm +/- SEM: patients 1.47 +/- 0.13; controls 2.04 +/- 0.11; P less than 0.001)--but without a concomitant reduction in the absolute number of T-helper cells--could also be observed.

Cellular and humoral immune responses in haemophiliacs after vaccination against tick-borne encephalitis

Summary. The primary immune response to a viral antigen (tick‐borne encephalitis, TBE) has been determined in haemophiliacs. Twelve HIV‐negative and four clinically asymptomatic, HIV‐positive haemophiliacs as well as 16 age‐matched healthy controls were included in the study. Antibody responses after TBE vaccination were comparable in HIV‐negative haemophiliacs and controls: however, antibody titres in HIV‐infected haemophiliacs were significantly lower after completion of the three‐dose vaccination schedule (geometric mean reciprocal antibody titres (SEM): controls. 193 (1.37), HIV‐positive haemophiliacs. 13 (2.18), P< 0.005). TBE vaccination failed to induce a T cell proliferative response in the HIV‐positive haemophiliacs. While in HIV‐negative patients the antigen‐specific lymphoproliferative responses after primary and one booster vaccination were comparable to those of the controls, cellular responses were decreased in HIV‐negative haemophiliacs following a second booster immunization 19 months after primary immunization (3H‐thymidine incorporation, delta dpm, mean ± SEM: controls, 34662 ± 7129, HIV‐negative haemophiliacs, 14339±7420, P<0.005). As the protective mechanisms for TBE infection are not yet completely understood, further work will be necessary to determine whether the decreased capacity to mount a sufficient long‐term cellular memory response in HIV‐negative haemophiliacs might be important for the protective effect of TBE vaccination in this population.

IMPAIRED TCR SIGNAL TRANSDUCTION, BUT NORMAL ANTIGEN PRESENTATION, IN A PATIENT WITH COMMON VARIABLE IMMUNODEFICIENCY

Summary. We describe a 27‐year‐old white man with common variable immunodeficiency (CVID) who has two healthy histoidentical brothers and one IgA‐deficient sister who shares one HLA haplotype with the patient. T cells from the patient with CVID showed an impaired response to recall antigens (tetanus toxoid, E. coli), whereas his IgA‐deficient sister and his two healthy histoidentical brothers responded normally. Cross‐mixing experiments using isolated monocytes and T cells from the CVID patient and one histoidentical brother revealed that the patients monocytes were fully functional in processing and presenting antigen to resting T cells of his brother, and provided normal accessory cell function for superantigen‐induced activation of his brothers resting T cells. In contrast, the patients T cells were unable to respond to antigen presented by the brothers monocytes and failed to respond with an increase in intracellular free Ca++ to stimulation with superantigen, which is known to bind to the TCR Vβ‐chain outside the antigen‐binding groove. However, stimulation with a combination of PMA and IM, directly activating protein kinase C and increasing intracellular free Ca++ by bypassing membrane receptors, induced normal Ca++ flux. These data indicate that the patient with CVID has a defect in TCR‐mediated signalling at the level of the T cells which is not present in his histoidentical healthy brothers or in his haploidentical IgA‐deficient sister.

Cord blood macrophages present bacterial antigen (Escherichia coli) to paternal T cells

Cord-blood monocytes (cord-blood M phi s) pulsed with Escherichia coli or tetanus toxoid had the ability to present antigen and to induce proliferation of antigen-specific paternal T cells. The magnitude of the proliferative response was comparable (86 +/- 14%) to that obtained after co-culturing paternal T cells with antigen-pulsed autologous M phi s. Cord-blood M phi s were surface Ia positive (54 +/- 12% as compared to 78 +/- 6% of adult M phi s), which correlated well with their ability to present antigen, but the experimental design was not suitable to show quantitative relationships. Compared to M phi s from the peripheral blood of adults, cord-blood M phi s released equal or larger amounts of interleukin 1 upon stimulation with E. coli. Immunoadsorption studies showed the capacity of cord-blood M phi s to specifically remove E. coli-reactive T cells from a paternal T-cell pool (65% of antigen reactivity could be removed). This indicated that cord-blood M phi s were able to express bacterial antigen in their membrane. These results provide evidence that human mononuclear phagocytes are mature at birth with respect to surface Ia expression, release of interleukin 1, and presentation of bacterial antigen.

Defective macrophage—T-cell interaction in common varied immunodeficiency

Abstract Macrophage—T-cell interaction (MTI) was studied in 9 patients with common varied immunodeficiency (CVID) and in 23 healthy control individuals. Adherent cells of patients and of controls were pulsed with Escherichia coli and subsequently cultured with autologous T-cell-enriched populations (TCE). The proliferative response of the TCE population was estimated by measuring [3H]thymidine incorporation after a 7-day culture period. Mean values in controls were 22,852 dpm ± 3263, in patients 3284 ± 1420. respectively (P

An HLA-DRα promoter DNA-binding protein is expressed ubiquitously and maps to human chromosomes 22 and 5

The class II major histocompatibility complex antigens are a family of integral membrane proteins whose expression is tissue-specific and developmentally regulated. Three consensus sequences, X1, X2, and Y, separated by an interspace element, is found upstream from all class II genes. Deletion of each of these sequences eliminates expression of class II genes in vitro or in transgenic mice. Here we further characterize the expression of a cDNA encoding a DNA binding protein (human X-box binding protein, hXBP-1) which, like the proteins in whole nuclear extract, recognizes both the X2 promoter element of the human DRα and DPβ and mouse Aα genes. The hXBP-1 cDNA hybridizes to human RNA species of approximately 2.2 kilobases (kb) and 1.6 kb, which are expressed in class II negative as well as class II positive cells. hXBP-1 transcripts are present in several class II deficient mutant B cell lines, although in one such line, 6.1.6, levels were somewhat reduced. Chromosome mapping studies demonstrate that hXBP-1 arises from a small gene family, two of whose members map to human chromosomes 5 and 22. Taken together, these data suggest a high degree of complexity in the transcriptional control of the class II gene family.

Humoral and cellular immunity induced by antigens adjuvanted with colloidal iron hydroxide

The immunopotentiating activities of colloidal iron hydroxide, a novel, experimental mineral adjuvant, and of aluminium hydroxide. the licensed adjuvant for human vaccines, were compared. Our studies revealed that colloidal iron hydroxide and aluminium hydroxide behaved comparably with respect to supporting induction of an antibody response to tetanus toxoid. Furthermore, mice immunized with both, the experimental vaccine (tick-borne encephalitis virus (TBEV) antigen adsorbed to colloidal iron hydroxide) or with a commercially available TBEV vaccine (adjuvanted with aluminium hydroxide), developed long-lasting antibody responses which protected the animals from TBEV infection even one year after vaccination. The use of colloidal iron hydroxide as adjuvant had the additional advantage to reproducibly support induction of HIV-1 envelope-specific cytotoxic T lymphocytes (CTL), when used as adjuvant for a HIV-1 env-carrying recombinant fowlpox virus and being applied via the subcutaneous route. Aluminium hydroxide was much less active in this respect. Non-adjuvanted recombinant fowlpox elicited CTLs only when given intravenously or intraperitoneally, vaccination routes considered not to be suitable for routine use in humans. Further studies to evaluate the use of colloidal iron as possible alternative and/or supplement for routinely used mineral adjuvants may therefore be warranted.

Vaccination against tick-borne encephalitis virus, a flavivirus, prevents disease but not infection, although viremia is undetectable.

By adoptive transfer of sera or immunoglobulin preparations, vaccine-induced protection against TBEV has been demonstrated to be mediated by antibodies to the surface protein of TBEV, glycoprotein E. Nevertheless, the mechanism of vaccine-induced protection against TBEV remains unclear. Protection by E antibodies without in vitro neutralization was shown by one group, whereas others found a correlation between protection in vivo and neutralization in vitro. Here, the authors confirm in a mouse model of tick-borne encephalitis (TBE) that immunization with the whole-killed virus vaccine protects mice against a subsequent challenge with a highly lethal dose of virus, i.e. 250 LD50 doses. Vaccine-induced immunity, however, is not completely neutralizing as demonstrated by the development of immune responses to a non-structural virus protein absent from the vaccine, yet expressed in the course of virus replication. Antibodies specific for the non-structural protein 1 (NS1) and cytotoxic T-cells could be detected after, but not prior to, virus challenge of vaccinated animals, establishing that protection by this highly effective vaccine is not equivalent with complete neutralization of the challenge virus.

Method for the isolation of biologically active monomeric immunoglobulin A from a plasma fraction

A purification method for immunoglobulin A (IgA) yielding monomeric IgA with a purity of over 97% has been developed. This procedure uses ethanol-precipitated plasma (Cohn fraction III precipitate) as the starting material and includes heparin-Sepharose adsorption, dextran sulfate and ammonium sulfate precipitation, hydroxyapatite chromatography, batch adsorption by an anion-exchange matrix and gel permeation. Additional protein G Sepharose treatment leads to an IgA preparation of greater than 99% purity. The isolated IgA presented with an IgA subclass distribution, equivalent to IgA in unfractionated plasma, and was biologically active, as was shown by its ability to down-modulate Haemophilus influenzae-b-induced IL-6 secretion of human monocytes.