Publisher Summary This chapter discusses many observations indicating a decline in immune function with aging both in experimental animals and in humans. It is evident that a decline in T cell function gradually occurs and this affects the entire immune system. Antibody formation and B cell function appear to be secondarily altered. Various autoantibodies are increased in older experimental animals and humans that may relate to the observed decrease in suppressor cells. Cell-mediated immunity has been analyzed in vivo and in vitro. Various T cell-dependent functions decline with age. Normal immune functions can begin to decline as early as when an individual reaches sexual maturity. The decline is due to changes in the immune cells and their milieu. Cell loss, shift in the proportion of subpopulations, and qualitative cellular changes, the three possible types of changes that can cause the decline, have all been detected.

Age influence on the immune system.

Publisher Summary This chapter reviews the evidence for cell-mediated immune reactions against tumors in animals and man, particularly, the reactions that can lead to destruction of neoplastic cells cultivated in vitro. Various mechanisms by which tumor cells can escape from immunological destruction have been discussed, most notably one involving blocking factors, present in the serum. Findings analogous to those obtained when studying tumor immunity (coexistence of cell-mediated reactivity and blocking serum activity) have been summarized from three other areas (pregnancy, allografting, and chimeras). The studies reviewed in the chapter have given a relatively large amount of evidence for various cell-mediated and humoral immunological reactions against growing tumors, and at least some of the reactions observed in vitro appear to be able to influence tumor growth in vivo .

Lymphocyte-Mediated Cytotoxicity And Blocking Serum Activity To Tumor Antigens

Publisher Summary This chapter discusses the nature of endotoxins and their interactions with cells of the immune system. The direct interaction of endotoxin with B lymphocytes leading to the formation of antibodies to endotoxin as well as a spectrum of other immunoglobulin molecules is discussed. In addition, the interactions of endotoxins with T cells and macrophages and the associated immunoregulatory effects are discussed. The central role of the lipid A portion of the endotoxin molecule in cellular activation and the molecular events occurring at the cell surface in the course of stimulation are presented. The potential of endotoxins as therapeutic manipulators of the immune response in man is also evaluated. Major direct interactions of endotoxins with B lymphocytes have been documented, leading to synthesis and secretion of antibody directed not only against antigenic determinants on the endotoxin molecules themselves, but also with specificities characteristic of the complete repertoire of variable region gene products. Knowledge of the immunobiology of endotoxins has prompted investigations in their therapeutic uses in man as both antigenically distinct and cross-reactive immunogens in protection against gram-negative infections.

Bacterial endotoxins and host immune responses

THERE is an obvious need for a marker that will differentiate one type of lymphocyte from another. The need has become urgent in view of recent evidence suggesting that there are at least two populations of lymphocytes, one thymus-derived and one bone marrow-derived, which participate in different ways in the immune response1. The theta (θ) isoantigen (θ is determined by a single locus with two alleles: θAKR found in AKR and RF mice and θC3H present in most other inbred strains of mice tested), described by Reif and Allen2,3, which is found chiefly in thymus lymphocytes and brain, and to a lesser extent in peripheral lymphocytes in mice, seemed a possible antigenic marker of thymus-derived lymphocytes. To establish that θ is such a marker, it is necessary to demonstrate that there is a discrete population of peripheral lymphocytes which carry the antigen and that these cells are thymus-dependent.

Theta isoantigen as a marker of thymus-derived lymphocytes in mice (Reprinted from Nature vol 224, pg 378-379, 1969)

Spleens from Moloney sarcoma virus (MSV) tumor-bearing C57BL/6N mice contained four times the normal number of mononuclear cells and displayed a markedly elevated "spontaneous" (mitogen-independent) DNA synthesis on a per cell basis. The number of macrophages were increased three-fold while there was a slight reduction in the percentage of T lymphocytes. The phytohemagglutinin (PHA) response on a per cell basis of spleens from tumor-bearing mice was decreased about 90% when compared with normal control mice. The primary in vitro immune response to sheep red blood cells was also suppressed to levels of less than 10% of normals. The PHA response could be restored by purification of MSV spleen cells by rayon adherence columns and by removal of phagocytic cells by an iron/magnet technique. The activity of suppressor cells in MSV spleens was demonstrated in mixtures with syngeneic normal spleen cells where a marked impairment of the PHA response was observed. Spleen cells from tumor-free nude mice and normal spleen cells treated by anti-theta serum plus guinea pig complement (C'), both totally unreactive to PHA, had no such effect. The inhibitor cell in MSV spleens was shown to be insensitive to inactivation by anti-theta plus C', but could be removed by the adherence columns and the iron/magnet technique. These data suggest that this suppressor cell is a cell of the monocyte/macrophage series. Suggestive evidence was also presented that the suppressor cells belong to a proliferating population in MSV spleens. Similar suppressor cells have been previously demonstrated in spleens of mice during a variety of immune responses. Our data show, that a tumor, although stimulating the immune system, nevertheless may be suppressive on certain immune functions through the activation of suppressor cells.

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Evidence of suppressor cell activity in spleens of mice bearing primary tumors induced by moloney sarcoma virus

The media and culture conditions required for in vitro stimulation of mouse lymphoid cells are described. The medium was arginine-rich and contained heat-inactivated human serum. A component of the human sera necessary for stimulation of the cells was a natural mouse cell agglutinin, which affected both background stimulation and the degree of induced stimulation with phytohemagglutinin (PHA). Absorption of the agglutinin from the human serum rendered the medium incapable of sustaining DNA synthesis in the presence of PHA.

The response to PHA of mouse spleen and thymus cells was age-dependent and, although this response was not present at birth, it rapidly rose to adult levels. Spleen cells from mice immunized with bacillus Calmette-Guerin (BCG) or sheep erythrocytes (SRBC) showed increased in vitro reactivity to added purified protein derivative (PPD) or SRBC stroma, dependent on the time of immunization.

The dose response curve for the SRBC stroma stimulated, immune spleen cells is compatible with a theory of cell to cell interaction being necessary for an in vitro reaction to antigen.

The possible role of the mouse cell agglutinin (AMLG) is discussed.

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Cellular recognition by mouse lymphocytes in vitro i. definition of a new technique and results of stimulation by phytohemagglutinin and specific antigens

The in vitro stimulation of mouse spleen cells by foreign histocompatibility antigens is described. Spleen cells from mice less than 3 weeks of age could not be stimulated in mixed lymphocyte cultures. They were also deficient in their ability to stimulate adult allogeneic cells in mixed lymphocyte cultures. The ability of the cells to be stimulated by histocompatibility antigens was correlated in time with their ability to stimulate allogeneic cells. Stimulation of mouse spleen cells was accomplished by allogeneic spleen cells with differences at the H-2 locus or at the H-3, H-13 loci, and with soluble H-2 antigen. F 1 hybrid spleen cells were stimulated by parental strain spleen cells or by parental strain red blood cell stroma. Immunization with allogeneic spleen cells was found to transiently affect the degree of stimulation obtained in mixed lymphocyte cultures of spleen cells from the immunized mice mixed with mitomycin-treated cells of the strain used for immunization. The cells from immunized animals also showed an increased degree of stimulation when reacted in MLC with allogeneic cells sharing some of the H-2 antigens of the strain used for the immunization. Immunization did not affect the degree of stimulation obtained in MLC of cells from strains with weak histocompatibility differences. The possible explanations of in vitro lymphocyte stimulation by histocompatibility antigens of allogeneic or semi-allogeneic cells are discussed.

Cellular Recognition by Mouse Lymphocytes in Vitro II. Specific Stimulation by Histocompatibility Antigens in Mixed Cell Culture

Summary The composition of spleen cell populations and the response of spleen cells in vitro to phytohemagglutinin (PHA) were studied in mice bearing methylcholanthrene-induced sarcomas and mice immunized with membrane antigen. With the use of discontinuous albumin density gradients to separate spleen cell suspensions, it was found that tumor-bearing mice and immunized mice had a shift in their spleen cell population, with a depletion of the dense, small lymphocyte population and a majority representation of a less dense, large mononuclear cell population. The PHA reactivity of the spleen cells from the tumor-bearing and immunized mice was less than normal. This finding correlated with the shift in the spleen cell population because the less dense cell types respond minimally to PHA. In mice in which the tumors were excised, the PHA response of the whole spleen cell population returned to normal. Our findings with the tumor-bearing mice suggest that a poor response to PHA may not be indicative of an immune deficiency but rather may demonstrate the possibility that the mice are undergoing an active immune response to their tumors. In human cancer patients, a poor peripheral lymphocyte response to PHA and an inability to demonstrate delayed hypersensitivity reactions have been interpreted as indication of an immune deficiency. However, it could be speculated that these findings in these patients may also suggest the presence of an active immune response to their tumors, rather than an immune deficiency. At present, this speculation provides a different working hypothesis for the evaluation of the immune status of cancer patients.

Phytohemagglutinin Unresponsiveness in Mouse Spleen Cells Induced by Methylcholanthrene Sarcomas

The response of spleen cells from three mouse strains, C57BL/6 CBA and A/J, to alloantigenic stimulation in mixed cell cultures was studied throughout the first year of life. The degree of stimulation by alloantigen of spleen cells from each of the strains and the stimulation pattern in relation to the age of the animal were both variable. Spleen cells from the three strains had no reactivity to alloantigen at birth. Following a period of 1 to 4 weeks, the reactivity increased rapidly. Peak values for all strains occurred between 14 and 20 weeks of age. In the C57BL/6 and CBA mice the responsiveness of the spleen cells to alloantigen declined markedly and was minimal at a year of age. However, in the A/J mice the reactivity appeared to remain at near peak levels throughout the first year of life.

Effect of age upon primary alloantigen recognition by mouse spleen cells.

The effects of thymectomy and thymus graft restoration upon the in vitro primary responses to alloantigens and PHA have been studied. It has been found that neonatal thymectomy substantially eliminates both PHA reactivity and responsiveness to alloantigens assayed in vitro in host spleen cell populations. Analysis of albumin density gradient-separated subpopulations of the spleen and thymus in such animals was also performed. It was found that the total and proportional representation of the individual density subpopulations was identical in neonatally thymectomized, in normal, and in thymectomized and thymus graft-restored animals. Therefore, thymectomized mice appear to retain a nonfunctioning, small, dense, lymphocyte population. Reconstitution of thymic-dependent in vitro reactivity was nearly complete when syngeneic, but not allogeneic or semisyngeneic thymus was employed. Occasional partial restoration did occur when F1 thymus was employed, but never when allogeneic thymus was grafted. The grafted thymus contained PHA and alloantigen-reactive cells in a large, less dense B layer subpopulation, whereas the restored animals, as in the case of normals, showed these reactivities to be a property of a small, more dense cell population.

https://rupress.org/jem/article-pdf/133/1/63/1083962/63.pdf

Cellular recognition in vitro by mouse lymphocytes : effects of neonatal thymectomy and thymus graft restoration on alloantigen and pha stimulation of whole and gradient-separated subpopulations of spleen cells

Tomoo Takiguchi

Papers

Cellular recognition by mouse lymphocytes in vitro i. definition of a new technique and results of stimulation by phytohemagglutinin and specific antigens

Cellular Recognition by Mouse Lymphocytes in Vitro II. Specific Stimulation by Histocompatibility Antigens in Mixed Cell Culture

Phytohemagglutinin Unresponsiveness in Mouse Spleen Cells Induced by Methylcholanthrene Sarcomas

Effect of age upon primary alloantigen recognition by mouse spleen cells.

Cellular recognition in vitro by mouse lymphocytes : effects of neonatal thymectomy and thymus graft restoration on alloantigen and pha stimulation of whole and gradient-separated subpopulations of spleen cells