Murine cytomegalovirus independently inhibits priming of helper and cytotoxic T lymphocytes.

Murine cytomegalovirus (MCMV) inhibits antigen-specific cytotoxic T lymphocyte (CTL) priming in vivo (Campbell et al., 1989). To address the mechanism of this immune suppression, two possibilities were considered: (1) MCMV directly interferes with in vivo priming of CTL precursors (CTLp), or (2) MCMV suppresses T helper cell functions necessary for CTL priming. We therefore quantitated T helper cell function in MCMV-infected, SV40-immune mice and assessed dependency of SV40-specific CTLp priming on T helper cell activity. MCMV infection of H-2b mice significantly suppressed the frequency of IL-2 producing T helper cells generated in SV40-immune mice. This suppression was not due to alterations in the number or percentage of CD4 lymphocytes. The helper cell deficiency correlated with suppressed SV40-specific CTL activity. However, CTLp priming in vivo was found to be independent of CD4 T helper cells and IL-2. Therefore, the suppressive effects of MCMV on helper and cytotoxic T cell functions are independent, implying that MCMV directly inhibits an event in lymphocyte priming common to both helper and cytotoxic T cells.

Murine cytomegalovirus-induced suppression of antigen-specific cytotoxic T lymphocyte maturation.

Antigen-specific cytotoxic T lymphocyte (CTL) maturation was inhibited in mice acutely infected with murine cytomegalovirus (MCMV). When immunization with Simian virus 40 (SV40) either preceded or followed infection with MCMV by 1 day, the frequency of SV40-specific CTL precursors among lymph node cells (LNC) was significantly reduced compared to noninfected mice. Replication of the herpesvirus in LNC could not be detected; however, MCMV rendered noninfectious by heat treatment was not suppressive to CTL development. Lymph node cells from nonimmunized, MCMV-infected mice contained a cell(s) present in low frequency which suppressed in vitro maturation of SV40 CTL in immune lymph nodes from mice not infected with MCMV. These suppressor cells did not affect the antigen- and interleukin-2-dependent growth nor cytotoxic activity of a mature, SV40-specific CTL clone. These results indicate that MCMV interferes with immunoregulatory functions required for development of antigen-specific CTL precursors to mature effector CTL. The immunosuppression is mediated at least in part by the MCMV-induced suppressor cell(s).

Induction of macrophage antitumor activity by acetylated low density lipoprotein containing lipophilic muramyl tripeptide.

A method has been developed for the selective delivery of lipophilic immunomodulators to macrophages, which results in the induction of antitumor activity. This method utilizes exhaustively acetylated low density lipoprotein (acetyl-LDL) to deliver the lipophilic immunomodulator, muramyl tripeptide phosphatidylethanolamine (MTP-PtdEtn; amide composed of N-acetylmuramyl-L-alanyl-D-isoglutamyl-L-alanine and dipalmitoyl phosphatidylethanolamine) to macrophages (M phi) by means of a scavenger lipoprotein receptor pathway. The binding of acetyl-LDL:MTP-PtdEtn to M phi showed specificity since minimal competition was observed in the presence of excess native LDL or phosphatidylcholine/cholesterol liposomes. Additional binding studies showed that acetyl-LDL may serve as a suitable delivery vehicle to a wide variety of M phi in different stages of activation. Cytostatic and tumoricidal activities by thioglycolate-elicited M phi against two tumor cell lines were examined in vitro following incubation with the acetyl-LDL:MTP-PtdEtn complex. Cytostatic activity against B16F10 melanoma cells was induced after the incubation of thioglycolate-elicited M phi with a minimum of 25 micrograms of acetyl-LDL protein containing 2.5 micrograms of bound MTP-PtdEtn (approximately equal to 40 molecules per particle of acetyl-LDL). The induction of cytostasis was not affected by liposome bilayers, which were also endocytosed by the M phi. In addition, tumoricidal activity against P815 mastocytoma cells was demonstrated at a 40:1 effector-to-target ratio using 18 micrograms of the acetyl-LDL:MTP-PtdEtn complex containing 3.6 micrograms of MTP-PtdEtn (approximately equal to 80 molecules per particle). These studies describe a method for the induction of antitumor activity by use of a chemically modified serum component, acetyl-LDL, to direct lipophilic immunomodulators to M phi.

Dissection of macrophage tumoricidal and protozoacidal activities using T-cell hybridomas and recombinant lymphokines.

Macrophage (M phi) phenotype and function can be modulated by various T-cell lymphokines (LK). The alteration of M phi phenotype is a result of LK concentration, duration of exposure, and the level of M phi activation when obtained from in vivo sources through elicitation by either sterile irritants or cellular immune mechanisms. To dissect M phi activation into discrete signals, we constructed T-cell hybridomas by fusing hypoxanthine-aminopterin-thymidine-sensitive BW5147 cells with nylon wool-purified, concanavalin A-stimulated T cells. The resulting T-cell hybrids were screened for their ability to (i) protect M phi from the cytopathic effect of Naegleria lysates, (ii) induce class II major histocompatibility complex gene product (Ia antigen) expression, (iii) increase tumoricidal and cytostatic activity, and (iv) alter ectoenzyme profiles on either resident or thioglycolate-elicited M phi. Two hybridomas (T-3 and T-9) were selected for further evaluation because of their activity patterns. Supernatants from T-3 and T-9 were compared with cloned gamma-interferon (IFN-gamma) for alterations of biological activities. Both T-3 and T-9 were able to protect resident-M phi cells from Naegleria lysate but had no protective effect on thioglycolate-induced M phi. T-9 supernatant had patterns of activity similar to IFN-gamma, whereas T-3 patterns were different. The addition of anti- IFN-gamma removed T-9 cytostatic activity while not affecting T-3-induced activity. The LK inducing protection from the cytopathic effect of Naegleria lysate is not IFN-gamma but another molecular moiety. We conclude that the activation of M phi for the destruction of tumor cells and amoebae may occur via different mechanisms.