Takeshi Yoshida

Serum migration inhibitory activity in patients with posttransplantation hepatic dysfunction

Abstract Macrophage migration inhibitory activity has been detected in the sera of six of eight patients with hepatic dysfunction following transplantation (renal or hepatic) and immunosuppression. Three of these individuals had detectable Australia antigen. Serum MIF activity bore an interesting temporal relation to parameters of hepatic disease, especially serum glutamic oxaloacetic transminase (SGOT) levels. Elevations of serum MIF preceded elevations of SGOT by approximately 5–10 days. This suggests that cell-mediated immune mechanisms may have not been beneficial, but rather may have contributed to the underlying hepatic pathology.

Serum migration inhibitory activity against macrophages and tumor cells

Abstract A factor that can inhibit the migration of tumor cells without cytotoxic effect (TMIF) is present in supernatants of activated lymphocytes. In the present experiments, we demonstrate that similar activity can be found in the serum of appropriately immunized animals. As is the case for macrophage migration inhibitory factor (MIF), TMIF activity appears only transiently after challenge with antigen. TMIF is also found in the serum of tumor-bearing animals. In this situation, activity persists from Day 7 postinoculation until the time of death. Although previous studies have demonstrated that TMIF and MIF are distinct, these results show that TMIF activity parallels serum migration inhibitory activity against macrophages. In the present experiments, the animals were given a lethal dose of tumor cells. However, it is likely that by virtue of its biologic properties, TMIF might have a protective effect under other experimental conditions.

Antigen-nonspecific and specific suppression of lymphokine production in desensitized guinea pigs

Doubly immunized guinea pigs may be desensitized with respect to delayed hypersensitivity reactions against both antigens (anergy) by injection of large doses of either one. This anergic response therefore has both a specific and nonspecific component. The specific component of desensitization persists longer than the nonspecific one. In the present study, we have explored the mechanism of both antigen-specific and antigen-nonspecific suppression during the later stages of desensitization. Guinea pigs immunized with two antigens, DNP-KLH and DNP-EA, were desensitized with DNP-EA. The lymph node cells obtained from the animals 1 day after desensitization were unable to produce MIF in the presence of either antigen. The cells obtained 3, 5, and 7 days after desensitization were able to generate MIF when stimulated with the non-specific antigen (DNP-KLH), but not with specific antigen (DNP-EA). It was shown that both T- and non-T-cell fractions obtained 1 day after desensitization had the capacity to antigen-nonspecifically suppress MIF production. In contrast, if the cells were obtained 3 or 5 days after desensitization, T cells could inhibit only the antigen-specific production of MIF, while non-T cells were still capable of suppressing antigen-specific and nonspecific MIF production. Interestingly, when these two populations were mixed back again, it was now only suppressive to the specific antigen-induced MIF production. This latter observation indicates that nonspecific suppressor non-T cells may themselves be regulated by suppressor T cells. Furthermore, antigen-specific suppressor T cells were shown to produce soluble factor(s) which inhibited the production of MIF.

Interaction of macrophages and lymphocytes for lymphokine production and cellular proliferation.

Abstract We have confirmed the requirement of macrophages in the antigen-induced T-lymphocyte proliferative response and in the generation of migration inhibition factor (MIF) by immune lymphocytes. Extending these observations, we have found that autologous and non-syngeneic, oil-induced peritoneal exudate macrophages were equally effective in restoring the proliferative response and MIF production by column-purified lymph node T cells. MIF activity was optimally restored when T cells were reconstituted with 1 to 40% exudate-derived macrophages whereas 10 to 30% macrophages were needed to optimally restore the T-cell proliferative response. Normal resident macrophages from the peritoneal cavity were also capable of restoring T-cell reactivity as were normal or BCG-activated pulmonary alveolar macrophages. It was also found that the addition of as few as 1.0% glycogen-elicited peritoneal exudate cells restored the production of MIF by T cells. Quantitative considerations demonstrated that the responsible cells in these preparations were polymorphonuclear cells rather than macrophages. In contrast, neither MIF production nor the proliferative response by T cells were restored by the addition of red blood cells. In these studies we were able to demonstrate that freeze-thawed macrophages could restore antigen-induced MIF production, but not antigen-induced cellular proliferation. The ability of freeze-thawed macrophages to stimulate T cells to produce MIF was apparently associated with the macrophage membranes and not with a soluble factor in the macrophage extracts. These results demonstrate that multiple sources of phagocytic cells may interact cooperatively with lymphocytes in reactions of cell-mediated immunity. Further, at least in the case of MIF production, this interaction involves a membrane-bound determinant that is effective even in the absence of viable macrophages.

The effect of heparin on chemotactic and migration inhibitory lymphokines

Abstract Heparin was studied in a dose range of 1 to 100 units/ml for its ability to interfere with migration inhibitory or chemotactic activity of lymphokine preparations. Heparin was found to inhibit macrophage and neutrophil chemotaxis but not macrophage migration inhibition. The dose range for these effects is similar to the in vivo concentrations necessary to suppress cutaneous reactions of delayed hypersensitivity. These results, taken in conjunction with previous studies, suggest that macrophage chemotactic factor may play a more significant role than macrophage migration inhibitory factor in the expression of cutaneous reactivity.

Prostaglandin E1 treatment of NZB/W F1 hybrids—Induction of in vitro and in vivo cell-mediated immune responses to DNA☆

Abstract Female NZB/W F1 hybrid mice, 4 months of age or older, demonstrate excessive humoral immune responses to native DNA while the expression of cell-mediated immunity (CMI) toward the same antigen is weak. In this study we examined the ability of cells from NZB/W mice to respond with CMI reactions after in vivo or in vitro stimulation with native DNA. In vivo experiments demonstrated that NZB/W mice did not respond to intradermal injections of native DNA with classic delayed hypersensitivity (DH) reactions. Additionally, their splenic lymphocytes did not produce migration inhibitory factor (MIF) following in vitro stimulation with DNA. Significantly, treatment of these animals twice daily with 200 μg Prostaglandin E (PGE1) markedly altered their CMI responsiveness. PGE1-treated mice were capable of making delayed hypersensitivity skin reactions to DNA. Histological evaluation of the reaction sites revealed a mononuclear infiltrate in the cutis and subcutis, typical of classic DH reactions. In vitro, a majority (58%) of the spleen cell cultures derived from treated animals released MIF activity following DNA stimulation. Diaflo ultrafiltration of the MIF-containing supernatants showed that the MIF was not due to antigen-antibody complexes, but rather to a true lymphokine. The results of this study provide clear evidence that PGE1 treatment of NZB/W mice significantly alters the immunological reactivity of these animals toward DNA, one of the antigens implicated in the pathogenesis of systemic lupus erythematosus.

Desensitization V: Suppression of MIF production by lymphokine-activated macrophages☆

The systemic injection of high doses of antigen into a previously immunized animal results in a state of transient anergy with respect to cell-mediated immune reactions. This phenomenon is known as desensitization. We have previously shown that desensitization is a multistage process. The initial 24-hr period is characterized by excessive lymphokine production with a failure to express delayed hypersensitivity reactions due to abolition of local chemotactic gradients. Subsequent stages of desensitization involve failure of lymphokine production in vivo. The results presented here demonstrate that lymphocytes obtained from immunized and desensitized animals later than 24 hr after desensitization are markedly suppressed in their ability to produce MIF. In addition, it was found that lymphokine-activated macrophages can suppress in vitro MIF production by lymphocytes from immune, nondesensitized animals. In vitro and in vivo activation of macrophages were equally effective. Thus, it is likely that at least one mechanism for the inhibition of lymphokine production in the post-24-hr period of desensitization, involves activation of a population of suppressor macrophages by lymphokines produced during the initial 24-hr period.

Differential susceptibility of human and guinea pig lymphokines to reduction and alkylation

Abstract Guinea pig and human macrophage migration inhibitory factors (MIF) were modified by reduction and alkylation. Guinea pig MIF lost activity when reduced and alkylated in the presence of urea or guanidine. Irreversible sulfhydryl modification was dependent upon the denaturation of the MIF molecule, suggesting that sulfhydryl bonds within the native molecule may contribute to biologic activity. Alkylation alone in guanidine had no effect on biologic activity, suggesting that free sulfhydryl groups are not essential for migration inhibitory activity. In contrast, human MIF was unaffected by any of the procedures used. Since previous studies have shown that human MIF can dissociate during extensive dialysis, these observations suggest that sulfyhydryl bonds are not important in the expression of human migration inhibitory activity. We have also shown that skin reactive factor (SRF), another lymphokine activity present in the activated lymphocyte culture supernates studied, has a pattern of susceptibility to reduction-alkylation procedures which is similar to that of MIF.

REGULATION OF LYMPHOK1NE FUNCTION

Although we have focused on desensitization as an experimental model, the findings obtained suggest mechanisms by which physiological control of cell-mediated immunity is attained. Thus, such regulation may occur either at the stage of expression of lymphokine activity or at the level of lymphokine production itself. The former may involve unique suppressor molecules, but as indicated above may be due to the inflammatory lymphokines themselves. The latter appears to involve suppressor systems similar to those operative in antibody formation. Over and above any specific model for regulation, the findings presented here suggest that local fluctuations in lymphokine distribution can modulate immunologically induced inflammatory responses, either through alterations in chemotactic gradients, or by other mechanisms as yet unknown.

Partial Characterization of a Low Molecular Weight Phagocytosis Inhibitory Factor Obtained from Human Erythrocyte Membranes

Phagocytosis Inhibitory Factor (PIF), a small (< 3000 D) molecule, was partially purified from human red blood cell membranes. This factor inhibits latex phagocytosis by monocytic cells. PIF is not toxic under the experimental conditions employed and the phagocytosis inhibitory activity is reversible since removal of this factor restores the phagocytic capability of cells. The phagocytic activity of murine macrophages was not affected by PIF.