Benvenuto Pernis

Role of CD8+ T Cells in Murine Experimental Allergic Encephalomyelitis

The course of experimental allergic encephalomyelitis (EAE), an animal model for multiple sclerosis, is affected by immunoregulatory T lymphocytes. When animals are immunized with encephalitogenic peptide of myelin basic protein and recover from the first episode of EAE, they become resistant to a second induction of this disease. Animals depleted of CD8+ T cells by antibody-mediated clearance were used to examine the role of CD8+ T cells in EAE. These cells were found to be major participants in the resistance to a second induction of EAE but were not essential for spontaneous recovery from the first episode of the disease.

Murine CD8+ T cells that specifically delete autologous CD4+ T cells expressing Vβ8 TCR: a role of the Qa-1 molecule

Interactions mediated by TCRs expressed on different T cell subsets may play a role in immunoregulation. To investigate this idea, we studied the regulation of superantigen-induced TCR V beta-restricted responses. We asked whether the in vivo regulation of CD4+ V beta 8+ T cells following SEB injection is controlled by CD8+ T cells. We found that in mice deficient in CD8+ T cells, the down-regulation of CD4+ V beta 8+ T cells below baseline is not observed. Moreover, following SEB administration, CD8+ T cells emerge that preferentially kill subpopulations of activated CD4+ V beta 8+ but not CD4+ V beta 8- T cells in vitro. This TCR V beta-specific cytotoxicity is dependent on beta 2-microglobulin and is inhibited by antisera specific for Qa-1 but not by antibody to MHC class Ia. These data suggest the idea that the specificity of immune regulation may involve CD8+ T cell recognition of TCR V beta determinants and Qa-1 molecules expressed on CD4+ T cells.

Internalization of lymphocyte membrane components

The functions of lymphocytes, in particular their participation in immune responses, depends on the expression and properties of several molecules that are present on their membranes at different steps in their development and activation. One way to probe this complex and various set of events is to follow the movement of the membrane molecules in different lymphocytes and in different functional conditions and, more specifically, to study their internalization and subsequent fate. In this article, Benvenuto Pernis summarizes the main facts concerning internalization of lymphocyte membrane components, either spontaneous or induced by cross-linking agents, and discusses what they tell us about the physiology of lymphocytes.

Human CD8+ T lymphocyte clones specific for T cell receptor Vβ families expressed on autologous CD4+ T cells

CD8+ T cells control immune responses, and recent studies suggest that this regulation is, in part, specifically directed towards TCR structures expressed by CD4+ cells. To develop a system to study the role of the TCR in regulatory interactions, we isolated clones of CD4+ cells expressing identified TCR V beta chains. These CD4+ clones were used to stimulate and expand autologous CD8+ cells, which kill the inducing CD4+ clone as well as independently isolated autologous CD4+ clones sharing the same TCR V beta as the inducing cell but not CD4+ T cells expressing different V beta TCRs. This V beta-specific cytotoxicity is dependent on the state of activation of the target cells and is not inhibited by an anti-class I monoclonal antibody, W6/32. We envision that V beta-specific CD8+ T cells of this type may regulate immune responses by direct interaction with antigen-activated CD4+ cells.

Cellular localization of rheumatoid factor idiotypes.

the stimulation of lymphocytes from rheumatoid arthritis patients with pokeweed mitogen produces a large number of plasma cells that express the dominant cross-reactive idiotype previously found on monoclonal IgM anti-gamma-globulins from patients with mixed cryoglobulinemia. Similar experiments with the cells of normal individuals show a much lower percentage of these cells with a lower intensity of staining with the fluorescent reagents utilized. Efforts to demonstrate rheumatoid factor in the same cells by fluorescent staining with aggregated gammaglobulin were entirely unsuccessful. This also proved to be the case for pokeweed mitogen-stimulated cells from the mixed cryoglobulinemic patients with large amounts of rheumatoid factor in the serum, despite high percentages of cells expressing the cross-reactive idiotype and also the individual idiotype. On the other hand, native plasma cells from synovial tissue of rheumatoid arthritis patients showed some cells with both the cross-reactive idiotype and aggregate staining. The exact reason for the failure to demonstrate rheumatoid factor by aggregate staining in pokeweed mitogen-stimulated cultures remains to be determined despite considerable effort to resolve the problem. The most likely possibility is that these plasma cells are relatively immature and have not accumulated polymeric IgM in their cytoplasm to the degree seen in synovial tissue plasma cells. The monomeric forms are readily recognized by the antiidiotypic antibodies and these reagents appear to be of particular value for cellular studies of this type.

Studies on the cytotoxic action of silica dusts on macrophages in vitro.

Experiments were performed in order to investigate the site of cellular damage induced by silica psrticles in vitro to peritoneal guinea pig macrophages. Considerable amounts of cytoplasmic enzymes, lactate dehydrogenase and aldolase, were released into the medium a short time after addition of silica to the cultures, whereas lysosomal enzymes, acid phosphatase and RNAase, were detected only after more prolonged incubation and in lower quantities. Correspondingly, loss of fluorochromasia from macrophages occurred in a matter of minutes after contact with silica particles. When coarse, nonphagocytabte silica particles were used, a rapid loss of fluorochromasia and early release of cytoplasmic enzymes were also seen.

Interaction of Baboon Anti-α-Galactosyl Antibody with Pig Tissues

As barriers to xenotransplantation are surmounted, such as suppression of hyperacute rejection allowing improved graft survival, it becomes important to define longer-term host-xenograft interactions. To this end we have prepared in baboons high titer anti-α-Galactosyl (αGal) and anti-porcine aortic endothelial cell antibodies, similar to human natural xenoantibodies and reactive with epitopes of thyroglobulin, laminin, and heparan sulfate proteoglycans. When injected into pigs with a protocol similar to that used in the rat to show the nephritogenic potential of heterologous anti-laminin and anti-heparan sulfate proteoglycan antibodies, baboon immunoglobulins bound first to renal vascular endothelium, and later to interstitial cells, especially fibroblasts and macrophages, and to antigens in basement membranes and extracellular matrix, where they colocalized with laminin- and heparan sulfate proteoglycan-antibodies, and with bound Griffonia simplicifolia B4. A similar binding was observed in other organs. The pigs did not develop an acute complement-dependent inflammation, but rather chronic lesions of the basement membranes and the extracellular matrix. Incubation of renal fibroblasts with baboon anti-α-Galactosyl antibodies resulted in increased synthesis of transforming growth factor-β and collagen, suggesting a possible basis for the fibrotic response. The results demonstrate that in this experimental model a consequence of αGal antibody interaction with porcine tissues, is immunoreactivity with αGal on matrix molecules and interstitial cells, priming mechanisms leading to fibrosis resembling that in chronic allograft rejection. The possibility that similar lesions may develop in long-surviving pig xenografts is discussed.

Acidification of internalized class I major histocompatibility complex antigen by T lymphoblasts

It has previously been shown that activated murine T lymphocytes express intracellular vesicles containing the class I major histocompatibility complex (MHC) antigen H-2K. Evidence has also been provided that such vesicles may be part of a cellular pathway of spontaneous H-2K antigen internalization and recycling, which is specific to T-lymphoid cells. Dual fluorescence flow cytometry has now been used to establish that H-2K antigen is acidified upon internalization in concanavalin A-stimulated but not lipopolysaccharide-stimulated murine splenocytes, thus providing further support that in T lymphoblasts this class I MHC antigen may travel intracellular routes similar to those reported for other cell surface receptors.

Anti-Immunoglobulins and Their Idiotypes: Are They Part of the Immune Network?

The anti-gamma globulins represent a very heterogeneous group of proteins with widely different specificities that might be considered a portion of the immune network. The hypothesis is presented that at least some of these proteins have other specificities, with the anti-gamma globulin reactivity being a secondary property. Anti-idiotypic antibodies are possible candidates. This is based on the low binding affinity for gamma globulin of many of these proteins and the results of CRI and sequence studies. All the proteins of the major CRI group have VKIIIb light chains, and these have a dominant but not total influence on this CRI. This has been evident from studies with rabbit antibodies and recently also with monoclonal hybridoma antibodies. Sequence studies have also demonstrated the similarity in light chains that, however, are not greater than with other VKIIIb chains; the heavy chains show little similarity except possibly in the J segment. The possibility is discussed that antibodies with secondary anti-gamma globulin binding properties might have selective advantages.

Idiotype expression in rheumatoid synovial plasma cells

SummaryThe majority of the polyclonal plasma cells identified in the synovial tissues of rheumatoid arthritis patients contain immunoglobulins that express a common idiotype related to that of the monoclonal cryoglobulins of the Wa group (RCRI). In these same tissues, there are twice as many plasma cells which are marked by a common idiotype yet do not show binding of aggregated IgG. These plasma cells are members of an idiotypically parallel set relative to those which produce rheumatoid factor. Direct comparison of rabbit polyclonal RCRI+ plasma cells with murine monoclonal RCRI+ plasma cells using two color fluorescent counterstains shows that the epitope recognized by the monoclonal anti-RCRI exists among a set of epitopes present in the RCRI. Since all the monoclonal RCRI+ plasma cells are not also positive with the polyclonal anti-RCRI antisera, the epitope characterized by the monoclonal antibody Glo 86.3 is not included among the set of epitopes the polyclonal antiserum identifies. A suitably absorbed anti-idiotypic polyclonal antiserum is superior to monoclonal antibodies for the purpose of detection of molecules (and cells) that are part of a polyclonal reaction involved in a given immune response.