Immune responses during human schistosomiasis mansoni. XVI. Idiotypic differences in antibody preparations from patients with different clinical forms of infection

Antibodies were purified from pooled sera from patients with different clinical forms of schistosomiasis mansoni on immunoaffinity columns of schistosome soluble egg Ag (SEA). As previously reported, T lymphocytes in PBMC preparations from schistosomiasis patients (but not control subjects who have never been infected) proliferate when cultured in the presence of certain of these anti-SEA purified antibodies. We now show that PBMC from most patients with chronic schistosomiasis, regardless of the clinical form of their infection, respond to anti-SEA antibodies from sera of asymptomatic (intestinal) or hepatointestinal patients. In stark contrast, none responds to anti-SEA antibodies purified from sera of acute or hepatosplenic patients. All of these multiclonal anti-SEA antibody preparations were active in anti-SEA ELISA assays and gave comparable patterns of reactivity with SEA upon immunoblotting analysis. Immunization of rabbits with some of these anti-SEA antibody preparations, followed by absorption of the rabbit antisera on absorbents of normal Ig, produced specific anti-Id reagents. Use of these reagents in competitive ELISA systems demonstrated that the Id in stimulatory and nonstimulatory anti-SEA antibody preparations differ with regard to the proportion of the serologically defined Id expressed by each. It appears possible to screen patients' plasmas for the presence of shared Id by use of suitable Id/anti-Id competitive ELISA assays. Taken together these data indicate that only certain Id-positive preparations are stimulatory to patients' PBMC, and the expression of these T cell stimulatory, immunoregulatory Id on anti-SEA antibodies correlates with the clinical form of a patient's infection.     

Immune responses to a soluble schistosomal egg antigen preparation during chronic primary infection with Schistosoma mansoni.

Murine schistosomiasis mansoni is characterized by an intense, predominantly cell-mediated, anti-egg, granulomatous response to schistosomal egg antigens (SEA). Anti-SEA responses include lymphocyte blastogenesis, the production of the lymphokine eosinophil stimulation promoter (ESP), hemagglutinating antibody, heat-labile and heat-stable, 72-hr passive cutaneous anaphylaxis (PCA) antibodies, and pronounced peripheral blood eosinophilia. These responses were followed during the course of chronic (1 year) infection and analyzed with specific reference to the observed diminution of granuloma formation, in the presence of continued antigenic exposure, which occurs by 10 to 12 weeks after infection and persists during long-term schistosomiasis. Lymphocyte blastogenesis and peripheral blood eosinophilia were positive from the 8th week of infection until the 50th. Lymphokine production and circulating heat-labile PCA antibody were only positive for a few weeks after 8 weeks of infection. In contrast, hemagglutinating antibody and heat-stable, 72-hr PCA antibody increased during weeks 10 to 14 and remained high throughout chronic infection. The development and regression of these various immune responses to SEA indicate that there are several potential mechanisms that could explain the immunoregulatory interactions that result in specifically diminished lesion formation in this chronic infection.     

Immune responses during human schistosomiasis mansoni. XV. Anti-idiotypic T cells can recognize and respond to anti-SEA idiotypes directly

We previously have shown that former patients and patients with active cases of schistosomiasis mansoni have T lymphocytes in their PBMC that proliferate when exposed to immunoaffinity-purified antibodies against Schistosoma mansoni soluble egg antigens (SEA). These T cell anti-idiotypic responses required the participation of adherent cells, but the role of these cells in the response to the Id has been unclear. We now show that chloroquine does not interfere with Id-elicited stimulation of cells from former patients but completely inhibits their response to the SEA. F(ab')2 fragments of anti-SEA Id are stimulatory, and excess normal human IgG does not alter anti-Id responses. Soluble Id F(ab) fragments are not stimulatory, but rather inhibit stimulation by the intact Id from which they were made. Either intact Id or their F(ab')2 fragments can stimulate non-adherent T cells in the absence of adherent cells if an exogenous source of purified or recombinant human IL-1 is supplied. Nonstimulatory F(ab) fragments can stimulate nonadherent cells if they are bound first to Sepharose 4B and presented in conjunction with IL-1. Thus, T cells from former schistosomiasis patients can react with polyclonal anti-SEA-related Id directly. Under these conditions T cell proliferation requires receptor cross-linking and a source of IL-1 but does not require either "processing" of Id or MHC co-presentation.     

Granulomatous hypersensitivity to Schistosoma mansoni egg antigens in human schistosomiasis. II. In vitro granuloma modulation induced by polyclonal idiotypic antibodies

We have previously reported on Id/anti-Id-receptor interactions in clinical human schistosomiasis. These findings support a hypothesis that anti-SEA cross-reactive Id develop in some patients during the course of a chronic infection and participate in regulation of anti-SEA cellular immune responses. We report here on experiments that extend those observations to the regulation of granulomatous hypersensitivity measured by an in vitro granuloma model. T cells from chronic intestinal schistosomiasis patients were stimulated in vitro with anti-SEA Id and assayed in an autologous in vitro granuloma assay for modulation of granuloma formation. These anti-SEA Id-reactive T cells were capable of regulating autologous in vitro granuloma formation. Both CD4 and CD8 T cells could be activated to regulate granuloma formation. This regulatory activity, initiated with stimulatory anti-SEA idiotypic antibodies, was antigenically specific and was dependent on the presence of intact F(ab')2 Ig molecules. The ability to elicit this regulatory activity appears to be dose dependent and is more easily demonstrated in chronically infected intestinal patients or SEA-sensitized individuals. These data support the hypothesis that anti-SEA cross-reactive Id are important in regulating granulomatous hypersensitivity in chronic intestinal schistosomiasis patients and these cross-reactive Id appear to play a major role in cell-cell interactions that result in the regulation of anti-SEA cellular immune responses.     

Immune responses and immunoregulation in relation to human schistosomiasis in Egypt. II. Cimetidine reversal of histamine-mediated suppression of antigen-induced blastogenesis

The effect of histamine on cell-mediated immune responses of chronic schistosomiasis patients was tested by peripheral blood mononuclear cell (PBMN) reactions to phytohemagglutinin-P (PHA) and soluble schistosomal antigenic preparations derived from eggs (SEA) or adult worms (SWAP). PBMN responses to PHA were suppressed by exogenous histamine (10(-5)M), and the addition of cimetidine (CIM) (10(-4)M), an H2-receptor antagonist, reversed this suppressive effect. Histamine primarily suppressed PBMN responses to suboptimal and optimal PHA concentrations. Exogenous histamine (10(-5)M) also suppressed PBMN responses of 27 schistosomiasis patients to SEA and SWAP, respectively. The addition of CIM (10(-4)M) to suppressed cultures reversed the effect of exogenous histamine. Most importantly, the addition of CIM to schistosomal antigen-induced cultures, without exogenous histamine, significantly increased patients' PBMN responses to SEA and SWAP. The mean optimal increase in SEA responses of 19 patients was 390%. With SWAP-induced responses of 21 patients this increase was 165%. The use of 10(-4)M diphenhydramine (DPH), an H1-receptor antagonist, resulted in general suppression of both PHA-induced and schistosomal antigen-induced PBMN responses. Lower concentrations of DPH lead to variable responses but did not result in consistent abrogation of the histamine-induced suppression. These data imply that an histamine-induced, H2-receptor-mediated suppressor circuit often helps modulate antigen-specific responsiveness of PBMN from patients with chronic schistosomiasis.     

Immune responses during human Schistosoma mansoni. XVII. Recognition by monoclonal anti-idiotypic antibodies of several idiotopes on a monoclonal anti-soluble schistosomal egg antigen antibody and anti-soluble schistosomal egg antigen antibodies from patients with different clinical forms of infection

A monoclonal human anti-soluble schistosomal egg Ag(SEA) antibody (E5) that stimulates anti-Id T cells and is idiotypically represented in pools of immunoaffinity-purified human anti-SEA antibodies from chronic, generally asymptomatic, intestinal (INT) patients (AM1 and AM5) was used to raise several monoclonal anti-Id: 1C2, 1C6, 4A8, 4F9, and 2A7. Cross-inhibition between these anti-Id identified distinct idiotopes on E5. Anti-SEA preparations from schistosomiasis patients (AM1, AM5, and others) were tested for their inhibition of the E5/monoclonal anti-Id reactions, in competitive ELISA. In either the E5/4A8 or E5/1C6 ELISA system, anti-SEA from INT (AM1 or AM5) or hepatointestinal (HI) (AM7) patients were able to inhibit these reactions. However, anti-SEA antibodies from acute (AM9) or hepatosplenic (HS) (AM3 or AM8) patients did not express Id that were inhibitory in these systems. These results suggest that a relatively high proportion of INT and HI anti-SEA antibodies express a dominant cross-reactive idiotope (CRI) recognized by 1C6/4A8. This CRI is also easily detected in plasmas from individual INT patients. Anti-Id 1C2 reacted strongly with an Id in AM1, AM5, or AM7, but one which also occurred, to a lesser extent, in AM3, AM8, and AM9. Monoclonal anti-Id 4F9 and 2A7 reacted weakly with idiotopes expressed by antibodies from all patients, regardless of the clinical form of their infection. These observations indicate that anti-SEA antibodies from INT and HI, but not acute or HS patients express dominant, CRI that are identified by 1C6, 4A8, or 1C2 and are also expressed on the INT-derived anti-SEA mAb E5.     

In vitro responses of rat lymphocytes following adult thymectomy. II. Increased inhibition by splenic adherent cells of responses to phytohemagglutinin

Adult thymectomy in rats results in a marked fall of spleen cell responsiveness to PHA over a period of days to weeks. Examination of dose-response curves showed that, with high PHA dose or ≥ 10⁶ cells/ml, there is a profound inhibition of the response with spleen cells from thymectomized animals compared with cells of matched sham-operated controls. However, when adherent cells are removed from the cell suspensions, the remaining nonadherent cells give an almost linear dose-response relationship with increasing PHA similar to that exhibited by the nonadherent cells of the controls or sometimes a slightly decreased response. Similarly, when increasing numbers of spleen cells from these animals are cultured (with admixed thymocytes to make a constant total of 2 × 10⁶ cells/ml) with PHA, the linear portion of the doseresponse curve can be extrapolated to give a similar value for the maximal potential response, which again is the same as or somewhat less than the corresponding value for sham-operated controls. A difference in inhibitory capacity is also shown in mixtures of the two spleen cell populations with LNC or with purified spleen cells. It is concluded that adult thymectomy results in increased “suppressor” activity in the spleen within a few days and may reduce slightly the number of T lymphocytes in the spleen reactive with PHA.     

In vitro responses of praziquantel-resistant and -susceptible Schistosoma mansoni to praziquantel

The resistance status of five praziquantel-susceptible and five praziquantel-resistant isolates was confirmed by chemotherapy in CD(1) mice with 3 x 200mg/kg micronised praziquantel. Micronised praziquantel had higher efficacy than two other praziquantel formulations (prepared without milling). The five resistant isolates were less responsive to praziquantel than the five susceptible isolates (59-74% reduction in worm burden in resistant isolates compared with 92-100% in susceptible isolates). Observations were made on the in vitro responses of different stages of 10 isolates to praziquantel. There were different in vitro responses to praziquantel at the egg, miracidial, cercarial and adult stages of Schistosoma mansoni between praziquantel-resistant and praziquantel-susceptible isolates. There were differences in the response of resistant and susceptible isolates following exposure of freshly hatched miracidia to 10(-6)M praziquantel for 1 min and observing the percent change in shape. Using this test it should be possible to determine whether failed therapy in patients infected with S. mansoni is due to the presence of praziquantel-resistant worms. Similarly, by exposing freshly shed cercariae to 4 x 10(-7)M praziquantel and observing the percent of tail shedding over 80 min it should be possible to monitor for the presence of praziquantel-resistant worms in snails collected in the field.     

Inhibition of lymphocyte transformation responses to phytohemagglutinin by normal human plasma.

Marked variability in lymphocyte transformation responses to a suboptimal phytohemagglutinin concentration (0.1 μg/ml) was observed when peripheral blood mononuclear cells of normal subjects were cultured in media containing 15% autologous plasma. Low responses were related to the plasma and were caused by direct inhibition rather than an inability to support the response. This inhibitory activity varied greatly among different plasma specimens and was also found in human and animal serum. It appeared to be specific for suboptimal concentrations of phytohemagglutinin and could be overcome by increasing the mitogen concentration. The inhibitory activity was heat stable, was not dialyzable, and appeared to affect a very early stage of the response since a delay in addition of the inhibitory plasma reversed its effect. Our interpretation of these results is that human plasmas vary greatly with respect to their ability to bind phytohemagglutinin so that the addition of different plasmas to lymphocyte cultures stimulated by a limiting concentration of phytohemagglutinin results in marked variability of the results obtained.     

Modulation of nonspecific defence mechanisms and specific immune responses after suppression induced by xenobiotics

Summary In the present study the possible modulation of nonspecific defence mechanisms and specific immune responses after suppression induced by oxytetracycline, oxolinic acid, and lindane – an organochlorine pesticide, were analysed in carp. Modulation was attempted using a natural immunostimulant, dimerized lysozyme (KLP-602, Nika Health Products, USA). Five groups of fish were given intraperitoneal injections of selected doses of the drugs on days 7, 6 and 4 before immunization with Yersinia ruckeri vaccine. At each injection, group I was also given 10 mg kg⁻¹ oxytetracycline, group II 10 mg kg⁻¹ oxolinic acid, group III 10 mg kg⁻¹ lindane, group IV was only immunized, and group V was injected with PBS. The immunostimulant was added in food before and after immunization. The study showed the immunotoxic effects of oxytetracycline, oxolinic acid and lindane; dimerized lysozyme corrected the defence mechanisms suppressed by these drugs and chemicals. The immunostimulatory effects of dimerized lysozyme were better when added before rather than after fish immunization.     

Immune responses during human Schistosomiasis mansoni. XI. Immunologic status of patients with acute infections and after treatment

Sixteen patients, 8 to 30 yr of age, with acute (toxemic) phase schistosomiasis mansoni were studied immunologically within 2 to 3 mo of their exposure to Schistosoma mansoni cercariae, and were monitored after chemotherapy. Total leukocyte levels and peripheral blood eosinophilias were higher in these patients than in similar individuals with chronic schistosomiasis mansoni. In contrast to chronic patients, the eosinophilias of the acute cases were decreased rather than elevated upon treatment. Total lymphocyte population (T and B cell) percentages were not altered during acute infection. Lymphoid subset (T3+, T4+, and T8+) analysis revealed elevated levels of both T4+ and T8+ cells. In vitro blastogenic responses of peripheral blood mononuclear cells (PBMN) to heterogeneous schistosome-derived antigens (eggs, SEA; adult worms, AW; and cercariae, CERC) were evaluated. SEA responsiveness was considerably higher than that of patients with chronic S. mansoni infections. The ratios of SEA to AW responses in acute cases gave a mean of 2.0, as opposed to 0.5 for a comparable group of chronically infected patients. The sera of most acute patients already contained suppressive factors that specifically decreased schistosomal antigen-induced PBMN blastogenesis. Chemotherapy of acute cases lead to a diminution of PBMN responsiveness to SEA and CERC. Treatment of patients with chronic infections lead to the elevation of such responses. PBMN from patients with acute infections produced lymphokine leukocyte inhibition factor upon exposure of the cells to SEA but not AW. A similar pattern was true for production of the lymphokine activity mitogenic factor. Levels of antibody in sera of acutely infected patients against SEA, CERC, and AW were considerably higher than levels in sera of chronically infected patients matched for age and intensity of their infections. These high antibody titers persisted for at least 6 mo after treatment, and were unrelated to the intensity of infection. The immunologic status of these patients with acute schistosomiasis mansoni differed considerably from patients with chronic infections. These findings re-emphasize the immunoregulatory events that apparently develop upon continued exposure to schistosomes and their products during chronic infection.     

Induction of granuloma modulation in murine schistosomiasis mansoni by enteric exposure to schistosome eggs

Enteric administration of antigen can induce systemic tolerance. In murine schistosomiasis mansoni, blood flukes produce eggs which enter the intestine. An immunologic phenomenon associated with this disease is a spontaneous diminution in the intensity of the granulomatous response in the liver, lungs, and colonic mucosa with chronic infection, which is termed modulation. It was determined whether modulation of liver granulomas could be induced by enteric immunization with schistosome eggs. Mice infected for 4 wk were immunized by injection of 25,000 eggs into cecal pouches. This induced modulation of liver granulomas by the eighth week of infection. Neither cecal injection of normal saline nor i.p. or subcutaneous injection of eggs could induce the modulatory process. Modulation could be adoptively transferred from enterically immunized donors by injection of spleen cells into infected recipients or into uninfected recipients with synchronous liver granulomas induced by the hepatic embolization of schistosome eggs. Spleen cells treated with anti-Thy-1.2 or anti-Lyt-1.1 and complement could no longer adoptively transfer modulation. These data show that enteric immunization with schistosome eggs can induce modulation of the liver granuloma by a cellular mechanism similar to that described for the natural infection.     

Legionella pneumophila-induced suppression of early blastogenic responsiveness of murine spleen cells to specific and nonspecific stimulation in vitro

Legionella pneumophila whole cells, including viable organisms or a killed vaccine, early after injection into mice suppressed the blastogenic responses of mouse spleen cells to both specific (i.e.,Legionella) and nonspecific (i.e., plant mitogen andEschericia coli lipopolysaccharide) stimulators. Mice given injections of sublethal numbers of viableLegionella or of a killed vaccine evidenced 3–4 weeks thereafter a marked increase in blastogenic sensitivity of their spleen cells to theLegionella antigen, either whole cells or soluble antigen, but no increase in responsiveness to nonspecific mitogens (i.e., concanavalin A, phytohemagglutinin, andE. coli lipopolysaccharide) was evident. In contrast, during the first week or so after injection of mice with either viable or killedLegionella, marked suppression of blastogenic responsiveness of spleen cells toLegionella antigens was evident. Concomitant suppression also occurred to concanavalin A and phytohemagglutinin, as well as toE. coli lipopolysaccharide. However, by the second week after injection of the animals with live or killedLegionella, such suppression disappeared. The importance of such early specific suppression of a cellular immune response early after exposure toLegionella antigen, in contrast with the early and sustained rise in specific antibody formation is being further investigated.     

In vitro viability of lymphoid cells from lines of mice genetically selected for high or low responsiveness to phytohemagglutinin

The kinetics of viability of lymph node and spleen cells of mice genetically selected for "high" or "low" in vitro lymphocyte responsiveness to PHA were studied in PHA or PPD-stimulated short-term cultures. Lo/PHA cells were found to be less viable than Hi/PHA cells in unstimulated control cultures. PHA improved the viability of Lo/PHA cells while inducing proliferation of Hi/PHA cells with the appearance of more and larger lymphoblasts in the latter. PPD only improved the viability of spleen cell cultures, more so for the Hi/PHA line. The interline difference in thymidine uptake was smaller after PPD than after PHA stimulation. Modifications of culture conditions designed to decrease the interline difference in cell viability lessened but did not abolish the separation between the two lines for the PHA response as measured by thymidine uptake.     

Cellular immune aspects of the human fetal maternal relationship. II. In vitro response of gravida lymphocytes to phytohemagglutinin

At optimal doses of phytohemagglutinin, human gravida lymphocytes showed no depression of responses when compared to nonpregnant female control lymphocytes. Compared to the control cells, gravida lymphocytes also seemed to have a higher spontaneous incorporation, a higher response at suboptimal PHA concentrations, and a lower peak dose response. These findings were more evident in the latter third of the pregnancy and suggest that maternal lymphocytes by this test are apparently fully reactive, and that they are undergoing low-level stimulation.