Ilonna J. Rimm

In the absence of T cells, natural killer cells protect from mortality due to HSV-1 encephalitis

The importance of natural killer (NK) cells in the resistance to herpes simplex virus type 1 (HSV-1), a common infection of immunocompromised patients, is unclear. Previous data on the role of NK cells in murine HSV-1 infection has been contradictory. Adoptive transfer studies suggested that NK cells mediated resistance to HSV-1, but in vivo depletion approaches demonstrated that NK cells were not important. We studied the course of HSV-1 infection after intranasal (i.n.) inoculation of E26 mice (lacking NK and T cells), T cell knockout (T cell ko) mice (lacking T cells only), or normal control mice. The E26 mice showed greater mortality and an impaired ability to clear virus from lung and brain compared to T cell ko mice and control mice, and had severe necrotizing HSV-1 encephalitis. Therefore, the data support the hypothesis that NK cells play an important role in the natural defense of murine HSV-1 infection.

B cell-deficient mice have increased susceptibility to HSV-1 encephalomyelitis and mortality

We studied the susceptibility of B cell-deficient mice to encephalomyelitis following intraperitoneal inoculation of HSV-1. B cell-deficient mice developed striking CNS signs including tail atony, clumsy gait and limb paralysis after HSV-1 infection. In addition, B cell-deficient mice had decreased survival (LD50 = 2.2 x 10(7) PFU) compared to control C57BL/6 mice (LD50 = 2.3 x 10(8) PFU). B cell-deficient mice had encephalomyelitis and detectable virus in the brain 7 days post-infection while C57BL/6 mice did not. Passive transfer of hyperimmune sera protected B cell-deficient mice from death, suggesting a role for antibody in susceptibility to HSV-1 encephalomyelitis.

Antibody-dependent cellular cytotoxicity and natural-killer-like activity are mediated by subsets of activated T cells☆

Abstract Antibody-dependent cellular cytotoxicity (ADCC), cell-mediated lympholysis (CML), and natural killer (NK) activity are performed by several distinct human lymphoid cell types. Thus, in the circulating lymphocyte population, null cells mediate ADCC and NK while T cells perform CML. In contrast, upon activation, T cells can perform all three functions. Recently, monoclonal antibodies, anti-T4 and anti-T8, have been utilized to define the human inducer and cytotoxic/suppressor subsets, respectively. Earlier studies showed that the T4+ and T8+ subsets were nonoverlapping, phenotypically stable populations. In the present study, we determined whether ADCC, NK, and CML activities were restricted to either of these subsets of activated T cells. It was found that the activated T8+ subset effected virtually all of the CML, while in contrast, both activated T4+ and T8+ cells mediated ADCC and NK.

Natural killer-like activity mediated by activated T lymphocytes

Diverse types of lymphocytes mediate in vitro cytotoxic activity. In addition to CTLs (cytotoxic T lymphocytes) and NK (natural killer) cells which differ in their activation requirements, target specificities, and lytic mechanisms, a natural killer-like activity of activated cells (A-NK) has recently been described. The data presented here suggest that an activated T lymphocyte can mediate A-NK activity. A-NK activity can be separated from resting NK activity by its requirement for activation and an effector phenotype (T12+,Ia+,Mol-) which includes the presence of the T12 and Ia antigens and the absence of the Mol antigen. In contrast, resting NK activity is mediated by T12-,Ia-,Mol+ cells. Cells that mediate A-NK activity can be differentiated from CTLs by their differing kinetics of activation and susceptibility to inhibition by monoclonal antibodies. An additional distinguishing feature is the fact that A-NK cells are predominantly Ia+ and are derived from either the T4+ or T8+ T-cell subsets whereas CTLs generated under similar conditions are predominantly T8+,T4-,Ia-. The in vivo relevance of this newly defined T-cell cytolytic activity remains to be defined.

Characterization of human thymic epithelial cells grown in serum free medium

Thymic epithelial cells have a critical influence on T-cell differentiation. In order to characterize these cells in humans, a serum-free growth medium was developed for their long-term culture. Important components of this medium included transferrin, epidermal growth factor, prostaglandin E1, and selenious acid. The presence of a keratin cytoskeleton, tonofilaments, and desmosomes confirmed the epithelial nature of these cells. Indirect immunofluorescence study of these epithelial cells demonstrated the presence of Ia and B-2 microglobulin antigens. The availability of highly enriched thymic epithelial cultures should simplify the functional characterization of this cell.

Impact of Vβ8+ / + T cells on the development of increased airway reactivity and IgE production in SJL mice

SJL mice have been extensively characterized as “low‐responder” animals in terms of IgE‐dependent immediate‐type hypersensitivity responses. Since these mice are genetically deficient in certain TCR Vβ gene segments, we asked whether this might be the reason for the “low‐responder” status. Specifically in H‐2 d mice the TCR‐Vβ8.2 gene element has been shown to play an important role in Th2 immune responses to ovalbumin (OVA). Utilizing a TCR Vβ8.2‐transgenic SJL (SJL Vβ8+ / + ) mouse, we examined whether the H‐2s ‐bearing “low‐responder” mouse could be converted into a “high‐responder” animal. Remarkably, non‐sensitized SJL Vβ8+ / + mice demonstrated strongly elevated levels of total IgE antibody. Mitogen‐stimulated T cells from these mice released high amounts of IL‐4 as compared to SJL wild‐type (wt) mice. In addition, sensitization to OVA via the airways resulted in the development of increased airway responsiveness in SJL Vβ8+ / + mice, but not in SJL wt animals. The results indicate that the capacity to produce IgE and IL‐4 and to develop increased airway responsiveness can be restored in SJL wt mice by introducing the Vβ8.2 gene segment into the TCR repertoire.

Detection of rare apoptotic T cells in vivo.

The flow cytometric analysis of apoptosis in lymphocytes from in vivo samples has been difficult because of the low frequency of apoptotic events. To overcome this obstacle, many investigators have relied on in vitro incubations to increase the number of apoptotic cells before analysis. In this report, we show that an adaptation of the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick-end labeling (TUNEL) assay for use in flow cytometry can be used to detect rare apoptotic lymphocytes from freshly harvested LN suspensions. This approach is both specific and extremely sensitive. This method also is amenable to multiparameter analyses and allows a phenotypic analysis of these rare apoptotic cells. However, we observed that some monoclonal antibodies can stain apoptotic-but not viable-cells nonspecifically. Therefore, the specificity of all antibodies to stain apoptotic cells was confirmed in competition assays.

T cell receptor (β chain) transgenic mice have selective deficits in γδ T cell subpopulations

Abstract TCR-β (T cell receptor-β-chain) transgenic mice have altered lymphocyte development. TCR-β transgenic mice are hyporesponsive to alloantigens in vivo and are deficient in γδ T cells. In order to begin a study of the relationship between a deficiency of alloreactive γδ cells and the defective function of in vivo alloantigen recognition, we analysed the γδ T cell development in TCR-β mice. The presence of the TCR-Vβ8.2 chain transgene is associated with inhibition of γ chain gene rearrangement. In order to determine how the presence of the TCR-β transgene affects γδ T cell development, γδ T cells were studied in the skin, intestine and spleen. TCR-β mice have dramatically reduced numbers of γδ T cells in the spleen and moderately reduced numbers of γδ T cells among intestinal intraephithelial lymphocytes. In contrast, these mice have normal numbers of γδ dendritic epidermal cells (DEC). These selective deficits could be due to the developmental regulation of transgene transcription during fetal life. We examined transcription of the TCR-β transgene in the fetal thymus and found that the TCR-β transgene is first transcribed at high levels on day 16 of fetal life, after DEC have already migrated from the thymus to the epidermis. Furthermore, mRNA from the transgene was detected in DEC, ruling out the formal possibility that DEC bear a γδ receptor only because they are incapable of expressing the transgene. The expression of the transgene is temporally associated with inhibition of further TCR gene rearrangement and this developmental sequence may be responsible for the selective tissue deficits of γδ T cells.