Omelan A. Lukasewycz

Lymphocytes from copper-deficient mice exhibit decreased mitogen reactivity

Abstract Copper deficiency was produced in C58 mice during perinatal development. Splenocytes from six-week old copper-deficient and copper-sufficient mice were evaluated for their ability to respond to various mitogens. When compared to cells from copper-supplemented control mice, splenic lymphocytes from copper-deficient animals showed a markedly decreased response to stimulation by the B cell mitogen, Lipopolysaccharide, and to the T cell mitogens Phytohemagglutinin and Concanavalin A, as well as to Pokeweed mitogen, a non-specific stimulator of both B and T cells. This decreased response was evident both in terms of [ 3 H]-thymidine incorporation and in terms of stimulation index. The diminution of mitogenic reactivity to Concanavalin A was related to the degree of copper deficiency as evaluated by plasma ceruloplasmin activity. Chronic copper deficiency results in an impairment of splenocyte T cell and B cell responsiveness.

Effects of Copper Deficiency on the Immune System

Copper is an essential metal for proper functioning of all living systems. Biochemical mechanisms have evolved that result in homeostatic balance of copper. This ensures that adequate but not toxic levels are absorbed, transported, utilized, and excreted. Throughout the biological kingdom copper expresses its function through specific ligands as free copper ion is rapidly complexed. These ligands are usually specific cuproenzymes. Knowledge of these cuproenzymes forms the basis of our current understanding of the biochemical function of copper (Prohaska, 1988).

Splenocytes from copper-deficient mice are low responders and weak stimulators in mixed lymphocyte reactions

Abstract Dietary copper deficiency was produced in C58 and C67BL mice to investigate the role of copper in lymphocyte function. Mice fed a diet low in copper (-Cu) developed signs of copper deficiency, anemia and hypoceruloplasminemia, compared to control mice fed the same diet with supplemental copper in the drinking water (+Cu). Splenocytes from eight-week-old mice, -Cu and +Cu, were compared as to their ability to respond to and to stimulate allogeneic cells in one-way mixed lymphocyte reactions (MLR). Splenocytes from -Cu C58 mice demonstrated a significantly decreased MLR response when stimulated by splenocytes from either C57BL or BALB/c mice. A novel observation was made when testing the ability of inactivated lymphocytes as stimulators in MLR. Splenocytes from -Cu mice, both C58 and C57BL, stimulated allogeneic controls less effectively (38% and 24%, respectively) compared to corresponding splenocytes from +Cu mice. These results suggest that chronic dietary copper deficiency decreases lymphocyte reactivity. Furthermore, these data support the hypothesis that copper deficiency has altered cell surface determinants associated with MLR reactivity.

Biochemical and immunological changes in mice following postweaning copper deficiency

Weanling albino male mice rapidly develop biochemical signs of copper deficiency when fed a purified diet containing 0.5 mg Cu/kg. Plasma ceruloplasmin activity of copper-deficient (−Cu) mice was 5% of that of copper-adequate (+Cu) control mice after only 3 d on the diet. More gradual loss of organ (liver, spleen, and thymus) cytochromec oxidase activity was observed during the next 4 wk. Body weight was equivalent between +Cu and −Cu mice, but thymus weight dropped faster in −Cu mice than +Cu mice. The number of antibody producing cells to sheep erythrocytes was lower in −Cu mice compared to +Cu mice after 17 d on the diet. Spleen cytochrome oxidase activity of −Cu mice was 50% of that of +Cu mice by 10 d on the diet. Mitogenic response of splenic and thymic lymphocytes to concanavalin A (con A) was not greatly different between +Cu and −Cu mice. Splenocytes from −Cu mice had a 3-fold higher thymidine incorporation rate in the absence of mitogen compared to +Cu mice. The depressed antibody and high mitogenic background responses of −Cu mice were similar to previous work with another strain (C58) of mice that had been started on copper-deficient treatment from birth. However, the normal proliferative response to con A stimulation in postweaning copper deficiency differs from the previous model. Mice of both studies were very copper-deficient as judged by liver copper levels. Timing of the copper-deficient treatment influences the manner, in which copper deficiency alters the immune response.

Immunological Consequences of Copper Deficiency in Mice

Dietary copper deficiency was produced in C58 mice by feeding dams a purified diet, low in copper (-Cu), from parturition. Some dams were given CuSO4 in their drinking water and served as controls (+Cu). Offspring from these dams were continued on their respective treatments until 6-9 weeks of age. Serum ceruloplasmin activity and blood hemoglobin levels indicated that copper deficiency was evident in the -Cu mice, but to varying degrees. Humoralmediated immunity was evaluated by the hemolytic plaqueforming cell assay of Jerne-Nordin. The number of cells from -Cu spleens producing antibodies against sheep erythrocytes was greatly reduced compared to the +Cu spleens. Furthermore, regression analysis demonstrated that the degree of reduction in antibody production in -Cu splenocytes was highly correlated with the degree of functional copper deficiency. Cell-mediated immunity was studied, in vivo, by investigating T-cell dependent immunity to lineb syngeneic malignant lymphocytes. Twenty of twenty-two -Cu male mice died of leukemia following an immunizing dose of cells, whereas no mortality was observed in ten +Cu mice. In vitro studies with isolated splenocytes also verified a T-cell impairment, as -Cu preparations were approximately one-tenth as active as +Cu preparations to blast transformation by concanavalin A. These studies indicate that copper deficiency in mice suppresses both humoral and cellular immunity. (Supported, in part, by American Cancer Society grant IN-13-T-23.)