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Immunoregulatory states induced by i.p. inoculation with the metazoan parasite Echinococcus multilocularis in the murine system were investigated. Proliferative responses and IL-2 production induced by Con A in spleen cells from BALB/c mice were significantly depressed at an early stage after infection with E. multilocularis protoscoleces (PSC). Addition of plastic-adherent cells from normal syngeneic mice to the nonadherent spleen cells from infected mice did not restore the depressed Con A responsiveness. On the other hand, exogenous IL-2 reconstituted completely the proliferative responses to Con A. Flow cytometry analysis revealed that CD4- CD8+ cells with a low density of CD8 Ag (CD8dull cells) increased in spleens from infected mice 2 weeks after inoculation. Addition of the spleen cell subpopulation containing the CD8dull cells, but not that depleted of the CD8dull cells, to normal spleen cells resulted in marked suppression of the Con A responses. These findings suggest that the CD8dull cells detected in spleens of mice inoculated with E. multilocularis PSC may play a key role in the suppressive regulation of immune responses. The relevance of the immune suppression seen in the early stages of experimental infection with E. multilocularis PSC to the eventual establishment of a host-parasite relationship is discussed.

A synthetic 17 amino acid peptide (CKS-17) homologous to a highly conserved region of human and animal retroviral transmembrane proteins was investigated for its influence on the generation of murine alloantigen-specific CTL activity in vitro. CKS-17 coupled to a carrier protein, BSA or human serum albumin, inhibited the generation of anti-allo CTL in a dose-dependent manner. Controls consisting of BSA and human serum albumin, which had undergone the coupling procedure or neurotensin, an unrelated peptide, coupled to BSA in an identical manner as CKS-17 showed no such inhibitory action. The suppression was not restricted to the Ag specificity of the CTL activity. CKS-17 exerted inhibitory effects on the early afferent phase of CTL induction. Kinetic studies showed that the suppressive activity occurred when CKS-17 was introduced to the immunologically stimulating culture concomitant with or up to 48 h after initiation of culture. Analysis of the frequency of CTL precursor cells using limiting-dilution assays revealed that CKS-17 did act to reduce the number of precursor cells. Abrogation of the inhibition of CTL activity was observed when IL-2 was introduced to the culture together with the stimulator cells. Other lymphokines, such as IL-4, exerted a similar influence to counteract this suppression.

Cats were classified into 4 categories by immunofluorescence antibody assay for the presence of feline leukemia virus (FeLV) and histologically as a) normal, FeLV-, b) normal, FeLV+, c) lymphosarcoma (LSA), FeLV+, and d) LSA, FeLV-. Determinations by Raji cell radioimmunoassay modified for cat serum revealed circulating immune complex (CIC) levels of healthy cats to be less than or equal to 50 micrograms equivalent aggregated cat immunoglobulin/ml (microgram/ml). In contrast, sera of cats in groups b, c, and d all contained significantly higher CIC levels (up to 12,000 micrograms/ml) associated with marked hypocomplementemia and C activation occurring via the classical pathway. Sera from FeLV+, LSA+ cats with high levels of CIC and sera of healthy cats were fractionated according to size and bouyant density by centrifugation through 10 to 40% sucrose gradients. Analysis of fractions from LSA+, FeLV+ sera revealed that both immune complexes (ICs), FeLV reverse transcriptase (RT), and IgG were present in fractions corresponding to a bouyant density of 1.15 to 1.18 g/ml. The CIC containing fractions activated C by the classical pathway. Sera from normal cats did not have CIC or RT and none of the fractions activated the classical pathway. These data suggest that vital antigen-antibody complexes are present in sera of viremic cats with LSA and these complexes activate the C system.

A synthetic 17 amino acid peptide (CKS-17) homologous to a highly conserved region of human and animal retroviral transmembrane proteins was investigated for its influence on the in vitro production of IFN-gamma from human peripheral mononuclear cells. The results showed that CKS-17 coupled to a carrier protein, BSA, inhibited production of IFN-gamma in a dose-dependent manner. Controls, consisting of BSA, which had undergone the coupling procedure or neurotensin coupled to BSA in an identical manner as CKS-17, showed no such inhibition. Reduction in IFN-gamma production could not be attributed to decreased viability of cells, delay of IFN-gamma production or to involvement of suppressor cells. Moreover, inhibition of IFN-gamma production was not related to the inhibition of DNA synthesis. The inhibition appeared to be a direct effect of CKS-17 on IFN-gamma-producing cells. Kinetic studies revealed that this suppression occurred when CKS-17 was introduced to the culture concurrent with or within 48 h after introduction of IFN inducers. Preincubation experiments showed that the presence of CKS-17 in the culture medium was not necessary to exert its inhibitory effect. These results suggest that a portion of retroviral envelope proteins possess important immunomodulatory actions.

Summary The synthesis of C1q by tissues of pig embryos during gestation was studied using specific incorporation of 14C-labeled amino acids into C1q in vitro. C1q synthessi was first demonstrated in intestinal tissue (on the 48th gestational day) and this was the most active site of synthesis of C1q at all times. Significant synthesis of C1q subsequently was demonstrated in lymph nodes and spleen of older embryos (by gestational days 66 to 79) and at still later stages in tissues from other highly lymphoid organs such as liver and lung (gestational day 90 and later). We propose that there is a special relationship between development of synthesis of C1q and development of the lymphoid tissues, which suggests that the lymphoid cell is the site of C1q synthesis.

Pretreatment of sheep erythrocytes (E) with supernatants of short-term cultures of mouse spleen or thymus cells induces the following interactions with guinea pig complement: 1) Supernatant pretreated EAlim (S.EAlim) are less susceptible to lysis by excess complement. S.EAC1̄4 yield a lower C2 titer, have an increased Tmax, but an unchanged C2 decay when compared with medium-pretreated EAC1̄4. Lysis of already formed EAC1̄4̄2̄is not inhibited. Therefore the inhibition affects C2 activation. 2) E, pretreated with supernatant (S.E), are able to fix C1̄, as detected by transfer to EAC4: In order to generate the same number of C1̄-fixing sites by using IgM hemolysins, five to six times the optimal amount is required. Yet S.EC1̄ cannot be lysed by the other components added in excess. S.EC1̄ can remove C4 from the fluid phase with the same efficiency as EIgMC1̄ but S.EC1̄ treated with C4 cannot remove C2. 3) Native C1̄ binds to S.E: S.E and EIgM remove C1 from the fluid phase with the same efficiency; fixation of 1̄ on S.E is decreased after incubation with native C1̄

The feline leukemia virus (FeLV) is a contagiously spread oncornavirus that causes malignant transformation of lymphoid cells. Feline cells transformed by FeLV produce a new cell surface antigen, the Feline Oncornavirus Associated Cell Membrane Antigen (FOCMA), which is a tumor-specific cellular antigen specified by, but not part of, the virion (Hardy, in press). An indirect immunofluorescent antibody test for FeLV antigens in leukocytes of peripheral blood indicates that approximately 90% of cats with lymphosarcoma (LSA) are infected with FeLV. In response to infection the cat may produce FeLV neutralizing and/or FOCMA antibodies. In the present study functional levels of total complement (TCH50), C4, C2, and Factor B were determined in cat sera. Protein levels of C1q were estimated by radial immunodiffusion using monospecific antiserum against cat C1q. A total of 60 animals were studied, 32 of which were normal and FeLV negative.

Linkage between MHC (major histocompatibility complex) in man and genes responsible for some C components (C2, C4, Factor B) has been described. In contrast, independent segregation has been established for genetic variants of C3 and C6. We report here linkage studies of two families (CC, MF) with C1r deficiency. The following markers were studied: C1r levels (family CC), HL-A, Factor B and MLC (mixed lymphocyte reactivity). In the family CC, as in family MF, C1r° gene was found to be inherited in a straight-forward Mendelian fashion. Linkage to the HL-A region, suggested by the C1r-deficient homozygote CC which was homozygous for HL-A2,W10, MLC and BfT, was ruled out by the independent segregation of 2,W10;T haplotype from the C1r° gene. MLC studies were consistent with the HL-A and Bf typing. Bf typing enabled the differentiation of two HL-A2,W10 serologically identical HL-A haplotypes, one of which was linked to BfT and the other to BfS.

Our interest in complement and leukemia has emerged from studies by Kassel et al. (J. Exp. Med., 138:925, 1973) and Hardy et al. (unpublished results) that infusion of normal plasma in mice, cats, and dogs causes massive dstruction of leukemic cells. Evidence in the mouse indicates that the anti-leukemic factor is a component of the complement system, C5. We have therefore begun to assay C and C component levels in cat serum. With functionally pure C1 and C2 and intermediate cells (EAC1, EAC4, EAC1,4) prepared from cat serum, the measurement of C1, C4 and C2 can be accomplished when the source of terminal C components, C3 to C9 is guinea pig serum in EDTA. With cat serum in EDTA, lysis does not occur. The blocking effect appears to be due to a potent inhibitor present in cat serum. This inhibitor is anti-complementary to guinea pig and human serum.

In immunodeficiency patients undergoing severe graft-vs-host (GVH) disease after bone marrow transplantation, a fall in the early C components was observed. To investigate this further, we induced a systemic GVH disease in (LxBN) F1 hybrid rats (70 g) by giving them a 200 × 106 or 400 × 106 Lewis spleen cell suspension i.v. Clinical GVH disease was present on day 5 and most of the animals died by 2 weeks of infection. Six of eight rats in the experimental group had a fall in the levels of serum C2 and C4 concomitant with the onset of the GVH reaction. C1 was variable in that only three of eight had diminished levels. One of six control rats had a significant fall in early C components. In a subsequent experiment (F344xBN)F1 hybrid rats (60 g) were injected i.v. with a 400 × 106 F344 spleen cell suspension or 200 × 106 F344 Ficoll/Hypaque (F/H) separated lymphocytes.