Decline in Phytohemagglutinin Responsiveness of Spleen Cells from Aging Mice

Abstract

Recent studies (1, 2 ) with thymus-dependent antigens have conclusively confirmed earlier demonstrations (3, 4) that there is a dramatic decline in humoral immune competence during senescence. Systematic studies to determine whether a comparable decline in cell-mediated immunity occurs are needed, as previous studies are not in agreement [for review see Ref.(5) 1. More recently it has been postulated, and considerable interest has been generated in the concept, that agerelated failure of immune surveillance mechanisms may play a prominent role in carcinogenesis and other age-dependent disease processes (6-9). Conclusive demonstration of a decline in cell-mediated immunity is essential if this concept is to remain tenable. I t is now clearly established that thymusdependent lymphocytes (T cells) are an absolute requirement of cell-mediated immunity (10-12). I t is generally accepted that phytohemagglutinin (PHA) selectively stimulates T cells (13-15), and therefore PHA stimulation of lymphocytes is commonly used as a measure of cell-mediated immunity (16, 17). Thus, it seems apparent that the in vitro blastogenic response of mouse spleen cells to PHA should be useful in identifying age-related immunodeficiencies that might exist in the T-cell population of the spleen, although it is recognized that PHA responsiveness at best measures only one parameter of cellmediated immunity.Materials and Methods. Mice used for these experiments were a long-lived hybrid (C57BL/Cum X C3H anf/Cum $)F1(hereafter referred to as BC3F1) with a mean life-span of 31 months and a maximum life-span of 45 months and the shorter-lived BALB/c strain with a mean life-span of approximately 25 months. Spleens were removed under sterile conditions. Only spleens of normal size and free of pathology were used. Subsequent his tological examinat ion proved the spleens to be pathology free. Cell suspensions from individual spleens were prepared in Hanks' balanced salt solution, counted in a hemacytometer, and adjusted to contain 10 X lo6 leucocytes/ml. Routinely, the culture media consisted of 89% RPMI-1630 (Grand Island Biological Co., N. Y.), 10% fetal calf serum, and 1 % penicillin-streptomycin (5000 units penicillin and 5000 pg streptomycin/ml) . Five replicate cultures were prepared for each spleen cell suspension in screw-cap Falcon flasks (30 ml capacity with a bottom area of approximately 5.25 cm2) by adding 2.5 X lo6 spleen cells (0.25 ml) to each flask, which contained 3.75 ml of the complete culture medium. Cultures ~were loosely capped and incubated at 37" in a 5% C 0 2 atmosphere at 80% humidity.was reconstituted with sterile distilled water

Age-Related Changes in G₀-G₁ Transition and Proliferative Capacity of Mitogen-Stimulated Murine Spleen Cells

“…Age-related changes in mitogen responses have repeatedly been assessed, mostly by measuring the 3H-thymidine incorporation of lymphocytes [Hori et al, 1973;Mathies et al. 1973;Abraham et al, 1977], While with this method a marked decrease of these re sponses with age can be demonstrated, it does not allow any conclusion on the causes of such a decrease.…”

Study of Some Early Immunological Parameters in Aging Humans

“…Aging is associated with altered humoral and cellular immunity in experimental ani mals [ 1,6,11,21] as well as in man [10,15,[23][24][25], Although general agreement exists regarding the impairment in lymphocyte proliferative activity in aged humans, the same cannot be said for the precise nature of the T cell dysfunction. Contradictory results in this sense may be partially explained by the subjects studied: if these are not carefully selected, the influence of concurrent disease(s) can obscure proper evaluation of the immune function.…”

Impaired nuclear responsiveness to cytoplasmic signals in lymphocytes from elderly humans with depressed proliferative responses.

“…Lymphocytes from many elderly humans and animals exhibit significantly impaired proliferative responses to the plant lectin phytohemagglutinin (PHA),' as compared with lymphocytes from younger adults (1,2). The decrease in tritiated thymidine incorporation by the aged cells is partly due to an increased sensitivity to chromosomal damage and cell cycle arrest induced by the radioisotope (3,4).…”