P. Jenkins

Immune recognition of Echinococcus granulosus. 1. Parasite-activated, primary transformation by normal murine lymph node cells

Summary Culture of murine lymph node cells together with living protoscolices of Echinoccus granulosus is described. The presence of the parasite induced potent blastic transformation in lymphocytes of unimmunized mice as indicated by tritiated thymidine incorporation. The response was markedly reduced by killing the parasite immediately prior to culture. No blastogenic activity was detectable in supernatants from living parasites cultured alone. Protoscolices from artificially infected syngeneic mice were effective stimuli, as were protoscolices from naturally infected horse and sheep. Stimulation was not detectably reduced by maintenance of the parasite in vitro for 72 h at 37°C or for 46 days at 4°C prior to culture. It is concluded that unprimed lymphocytes are stimulated to transform by surface contact with a stimulator synthesized, but not secreted, by the parasite. The biological significance of the reaction and its possible contribution to immunosuppression are discussed.

Regulation of macrophage-mediated larvicidal activity in Echinococcus granulosus and Mesocestoides corti (Cestoda) infection in mice

Killing of metacestodes by normal or post-infection macrophages and the regulation of this activity by cytokines were studied in vitro. The protoscolecidal activity of normal macrophages against Echinococcus granulosus was inhibited by a product of naive T-enriched lymphocytes co-cultured with protoscoleces (PSC). By contrast, supernates from co-cultures of Mesocestoides corti tetrathyridia (MCT) and T-enriched or B-enriched normal lymphocytes increased killing of MCT by normal macrophages. Larvicidal activity (against both PSC and MCT) was enhanced by high concentrations of macrophage-activating factors produced by Con A-stimulated rat lymphocytes (Con A-LK), but was reduced by low concentrations of these factors. Activation by synergism between Con A-LK and recombinant interferon-gamma(r. IFN-gamma) was demonstrated in macrophage-mediated killing of MCT at high effector to target ratio. Cytokine-activation of normal or post-MCT infection macrophages was compared. Macrophages from both 8 and 20 week post-infection mice were refractory to lymphokines from lymphocyte-MCT cultures and displayed greatly reduced killing of MCT. Macrophage activation by Con A-LK and r.IFN-gamma was also impaired, implying a general defect in the ability of these post-infection macrophages to respond to macrophage activating signals. The data indicate that two different mechanisms may exist by which metacestodes regulate potentially larvicidal effector mechanisms. E. granulosus can elicit the production of lymphokines suppressive for PSC killing, whereas M. corti appears directly to induce a refractory state in effector macrophages.

A study of immunoregulation of BALB/c mice by Echinococcus granulosus equinus during prolonged infection

Two models of intraperitoneal infection with E. granulosus equinus by protoscolices and by cyst passage in BALB/c mice were used to provide mesenteric lymph node cells for adoptive cell transfer into syngeneic recipient normal responder mice. The cell transfer inocula were shown to have depleted Thy‐1 cells, but to be highly suppressive to the normal sheep erythrocyte response of the recipients. The nature of the depletion and non‐specific suppression, and the infectious nature of the latter, are discussed in relation to other examples of mitogenic stimulation resulting in non‐specific T cell suppressor activity. The functions of Ly‐2,3+ cells, not only as suppressor, but as alloreactive cytotoxic cells, are discussed as a possible, autoimmune explanation for the longevity of the parasite within the mouse model, in contradistinction to the predictable early rejection of analogous xenografts.

Echinococcus granulosus infection in mice: host responses during primary and secondary infection

The reaction of Balb/c mice to primary and secondary subcutaneous infection with Echinococcus granulosus protoscoleces (PSC) is described. From 3 to 14 days following primary exposure to PSC, draining lymph nodes increase in weight and there is expansion of T and B lymphocyte populations, enhancement of in vitro lymphocyte transformational responses and production of PSC-specific IgM and IgE antibodies. Despite the persistence of viable PSC in host tissues, lymphocyte responses decline to pre-infection values over the period 3-8 weeks post-infection. Secondary exposure to PSC immediately induces lymphoproliferation, enhancement of transformational responses, production of IgE antibody and encapsulation of PSC by inflammatory cells. Although specific antibody levels remain high until at least 8 weeks after challenge infection, lymphocyte activity begins to decline after 4 days and is profoundly suppressed by 10 days. Parasite viability appears to be significantly reduced in secondary, as opposed to primary, infection and is associated with the accumulation of large numbers of eosinophils, mast cells and macrophages in infected tissues.

Macrophage modifying factor secreted by the tetrathyridia of Mesocestoides corti (Cestoda): monoclonal antibody to the modifying factor antagonizes its immunological activity

Immunomodulation of macrophage activity by in vitro secretions of Mesocestoides corti has been previously demonstrated. The modifying activity secreted by M. corti had the effect of reducing the normal accessory function of macrophages in a Con‐A‐activated lymphocyte proliferation assay. This paper describes the purification of the modifying activity by FPLC techniques and the generation of a monoclonal antibody (MoAb) to this molecule in mice. The MoAb bound immunomodulatory FPLC fractions of M. corti in an ELISA. When MoAb was applied in conjunction with immunomodulatory parasite secretions to macrophages in vivo or in vitro, the modifying effect of the secretions was abolished. This profound effect of the MoAb should help to elucidate the mechanisms by which metacestode parasites avoid host immune responses and may enable therapeutic intervention.

Lymphoreticular responses to metacestodes: Taenia multiceps (Cestoda) can modify interaction between accessory cells and responder cells during lymphocyte activation

This study was designed to test the accessory function of macrophages after activation with products of Taenia multiceps coenuri. Activation was carried out by intraperitoneal injection of mice with coenurus fluid or protoscolex culture supernatant, and function was assessed by adding these macrophages in progressively increasing numbers to macrophage-depleted lymphocyte cultures transforming under the influence of plant mitogens or coenurus-fluid mitogen. In contrast to normal macrophages, which have a progressively enhancing action on the above reactions, parasite-activated macrophages at similar concentrations were progressively inhibitory. However, low concentrations of the activated macrophages enhanced mitosis as well as, or better than, normal. Lymph node cells from injected mice showed abnormal response to macrophage-derived signals. In particular there was subnormal reaction to macrophages in the presence of coenurus mitogen. These results suggest that T. multiceps coenuri may survive in the host because of their ability to reduce effective interaction between lymphocytes and accessory cells.

Modification of cellular immunity by Taenia multiceps (Cestoda): accessory macrophages and CD4 + lymphocytes are affected by two different coenurus factors

Taenia multiceps coenurus fluid was analysed by fast protein liquid chromatography in order to separate the factors responsible for previously reported modification of immunological activity in macrophages and T-cells. One factor, F7, was found to be mitogenic for murine L3T4+ T-cells, to be macrophage dependent, to require macrophage compatibility at the I region of the H2 complex, to increase the sensitivity of T-cells to regulatory signals from macrophages and to increase the rate of generation of splenic rosette-forming cells (RFC) against sheep red cells. A second factor, F24, was found to alter macrophages so as to render them suppressive, rather than stimulatory, for parasite-activated and Con A-activated lymphocyte transformation, to depress the rate of generation of RFC and to antagonize the mitogenic effect of F7. The combined actions of these two factors are, therefore, sufficient to explain the known immunomodulatory effects of the metacestode.

The effects of tumour necrosis factor on host-parasite relations in murine Mesocestoides corti (Cestoda) infection.

The regulatory role of tumour necrosis factor (TNF) was investigated in murine infection with tetrathyridia of Mesocestoides corti. Recombinant TNF alpha reduced macrophage larvicidal activity in vitro. M. corti primed mice for TNF release in response to bacterial lipopolysaccharide (LPS) in vivo. TNF activity was amplified 100-fold at 14 days post-infection (p.i.), with a further rise at day 28 p.i. Maximal inflammatory reaction was observed histologically in the liver at the height of TNF activity. Hepatic necrosis was located within inflammatory foci, but not within the vicinity of the parasite itself, suggesting that TNF may contribute to the pathogenesis of infection. Peritoneal cells from infected mice, when stimulated with tetrathyridia in vitro, showed a 4-fold increase in TNF alpha activity at day 14 p.i. However, when peritoneal cells were stimulated with LPS in vitro, a marked increase in TNF alpha secretion was observed at 2 months post-infection followed by a slow decline. It is suggested that impaired macrophage effector function, previously attributed to endogenous endotoxin, which gains access to peritoneal macrophages through an inability of the liver to detoxify endotoxin, may be mediated through TNF alpha.

Differential regulation of murine Mesocestoides corti infection by bacterial lipopolysaccharide and interferon-γ

Many liver-invasive parasites cause extensive liver damage which may result in an impaired ability to catabolize endotoxin. The influence of endogenous endotoxin on the progress of liver-invasive parasitic diseases has been investigated in murine Mesocestoides corti infection. Invasion of liver tissue by tetrathyridia resulted in extensive parenchymal destruction with fibrosis. In association with this, undetoxified endotoxin, in potentially biologically active concentration, was found on peritoneal macrophages, 5 months post-M, corti infection. Host susceptibility was influenced by the Lps gene for responsiveness to lipopolysaccharide (LPS). The parasite burden of LPS-responsive (C3H/HeN) mice was significantly increased in the livers of these mice when compared to LPS-resistant (C3H/HeJ) mice. LPS reduced the ability of normal peritoneal macrophages to kill tetrathyridia, when co-cultured in vitro. LPS also abrogated the ability of recombinant interferon-gamma (r.IFN-gamma) to enhance macrophage larvicidal activity. These in vitro findings were confirmed in vivo. Daily intraperitoneal administration of LPS, at low concentration, caused a 4-fold increase in parasite burden in the liver, while r.IFN-gamma at optimal concentration reduced parasite burden by 57%. Post-infection macrophages have previously been shown to be refractory to cytokine-activation for larval killing. In this report, we conclude that (1) this refractoriness may be due to the presence of undetoxified endotoxin on post-infection macrophages and (2) endotoxin may reduce host resistance by abrogating effector macrophage response to IFN-gamma.

Modification of macrophage —T cell interaction during infection of mice with Mesocestoides corti (Cestoda)

Peritoneal macrophages from Mesocestoides corti-infected mice showed a marked and progressive loss of ability to act as accessory cells for syngeneic Con A-stimulated mesenteric lymph node lymphocytes. The same effect on the macrophages could be induced by intraperitoneal injection of M. corti culture supernatant, despite a concurrent increase in numbers of peritoneal adhesive macrophages. The findings are used to compare and contrast the known immunomodulatory effects of M. corti and taeniid metacestodes, the latter differing chiefly in their potential for modifying T-cell as well as macrophage behaviour.