bluuebunny/crossref_metadata_2025 huggingface dataset:
Human monocyte-mediated tumor cytotoxicity. I. Demonstration of an oxygen-dependent myeloperoxidase-independent mechanism.
Authors: Mavier P , Edgington T S
Published: December 2022

The cytolytic capacity of monocytes per se and stimulated monocytes has been documented to only a limited extent, and when observed has been ascribed to the generation of a variety of cytolytic molecular entities. In the present study we have examined de novo human monocyte-mediated tumor cytotoxicity and that induced by the agent 12-O-tetradecanoylphorbol-13-acetate (TPA). Cytolytic function was analyzed by reference to the release of [111In] oxine from two prelabeled tumor cell lines, K562 and U937, in a 16-hr assay in the presence of serum to more closely mimic in vivo circumstances. Observed cytolysis was clearly related to TPA concentration and effector cell number. Maximal cytolysis was obtained with TPA at 5 ng/ml, at which specific releases were 43% +/- 6 and 18% +/- 5 (mean +/- 1 SEM) at an effector cell to target cell (E:T) ratio of 2.5:1 and 65% +/- 6, and 41% +/- 12 at an E:T ratio of 20:1, for K562 and U937, respectively. In contrast, unstimulated monocytes expressed minimal cytolytic activity, or at best a low cytotoxic effect at high cellular ratios. When TPA-stimulated monocyte-mediated cytolysis was examined, catalase (2750 U/ml) inhibited K562 and U937 cytolysis by 92% and 84%, respectively; superoxide dismutase (300 U/ml) only inhibited cytotoxicity by 17% and 24%, respectively, implicating a central role of H2O2 rather than superoxide ions. Sodium azide (1 mM), an inhibitor of myeloperoxidase, did not diminish cytolysis; in contrast, it increased K562 and U937 cytolysis by 34% and 57%. This increased cytotoxicity was observed for K562 at low levels of cytotoxicity. These data tend to dismiss an essential role of the H2O2-halide-myeloperoxidase pathway of cytolysis. The OH scavengers, histidine (20 mM) and ethanol (40 mM), did not affect K562 killing; mannitol (50 mM), another OH scavenger, had only a slight inhibitory effect (23%). Finally, H2O2 generated by a glucose-glucose oxidase system directly mediated K562 killing and, to a lesser extent, U937 lysis. These results point strongly towards the role of: 1) a myeloperoxidase-independent mechanism of cytotoxicity, with 2) H2O2 as a key mediator of the cytolytic mechanism, and 3) a limited role of O2.- in synergy with H2O2 in the cytolytic activity of monocytes, and suggest that significant cytolytic function requires an inductive event.
Thymectomized, irradiated, and bone marrow-reconstituted chimeras have normal cytolytic T lymphocyte precursors but a defect in lymphokine production.
Authors: Duprez V , Maziarz R , Weinberger O , Burakoff S J
Published: December 2022

A model system has been developed to study extrathymic T cell differentiation; mice have been thymectomized, lethally irradiated, and reconstituted with bone marrow cells depleted of Thy-1+ cells. After 8 wk, the spleen cells of these athymic, bone marrow-reconstituted chimeras contain Thy-1+ precytolytic T lymphocytes (CTL) that are able to respond to antigen only if supernatant from Con A-activated T cells is added to culture. The phenotype of these pre-CTL is similar to that of thymocytes, suggesting that they may be immature T cells. Initial evaluation of the CTL repertoire of these athymic mice demonstrated that the CTL generated to trinitrophenyl-modified syngeneic cells are H-2-restricted, and that the CTL generated to alloantigens have many of the cross-reactivities observed in normal mice but not in nude mice. In this report, we demonstrate a helper T cell defect in these thymectomized chimeras. These chimeras lack an Ly-1+ helper cell required for thymocytes to differentiate to CTL. Further studies revealed that when spleen cells from these thymectomized chimeras were stimulated with Con A, they produced normal levels of interleukin 2. However, these splenocytes were defective in the production of another factor needed for CTL differentiation.
Immunoregulatory activities of autoreactive T cells: an I-A-specific T cell clone mediates both help and suppression of antibody responses.
Authors: Clayberger C , Dekruyff R H , Cantor H
Published: December 2022

We have derived a Ly-1+, 2-3- T cell clone that is specific for autologous I-A on activated but not on resting cells. After activation, this clone produces factors that induce purified (B + adherent) cells to secrete antibody in response to sheep red blood cells and type 2 T-independent antigens. Regulation of plaque-forming cell responses by this clone is dose dependent: low numbers enhance the plaque-forming cell response, whereas high numbers suppress the response. The inhibition observed with high doses is associated with cytolysis of I-A+ cells, and this can be blocked by the addition of anti-I-A antibodies. The physiologic significance of this novel cell type in regulating immune responses is discussed.
Intraepithelial lymphocytes modulate Ia expression by intestinal epithelial cells.
Authors: Cerf-Bensussan N , Quaroni A , Kurnick J T , Bhan A K
Published: December 2022

We have demonstrated that although intestinal epithelial cells in fetuses and young rats do not express Ia antigens, in adult rats intestinal epithelial cells do express Ia antigens, as indicated by immunoperoxidase staining with monoclonal antibodies. Ia expression by intestinal epithelial cells appeared to be related to an increase in the number of intraepithelial lymphocytes (IEL). Most of the IEL were T cells and expressed the phenotype associated with cytotoxic/suppressor T cells, and a large number contained cytoplasmic granules. To directly study a possible modulating effect of IEL on intestinal epithelium, an Ia-negative intestinal epithelial cell line (IEC 17) of rat origin was cultured in the presence of supernatants obtained from Con A- or PHA-stimulated lymphocytes. IEL, as well as spleen cells but not bone marrow cells, were able to secrete a factor(s) capable of inducing Ia antigens on IEC 17 cells, as judged by immunoperoxidase staining and radioimmunoassay. Ia-positive IEC 17 cells were detectable after 12 hr and maximum Ia expression was obtained by 48-hr incubation. Persistence of Ia expression by intestinal epithelial cells required the continued presence of Ia-inducing factor in the medium. Lymphocyte proliferation was not essential for the secretion of the Ia-inducing factor(s). The characteristics and the kinetics of secretion of the Ia-inducing factor were similar to that of an interferon-like activity, but not of interleukin 2. Con A-induced supernatants from IEL and spleen cells were also capable of suppressing the growth of IEC 17 cells. The results of this study indicate that IEL, because of their close association with intestinal epithelial cells, may be involved in modulating a variety of epithelial cell functions, including the expression of Ia antigens. This leads us to speculate that Ia-positive epithelial cells, like Ia-positive macrophages and dendritic cells, may be involved in antigen presentation to T lymphocytes.
Role of antigen-specific B cells in the induction of SRBC-specific T cell proliferation.
Authors: Malynn B A , Wortis H H
Published: December 2022

In this study, we ask whether antigen presentation can be effected by antigen-activated B cells. Antigen-dependent in vitro proliferation of T cells from mice primed with SRBC or HoRBC occurs in the presence of B cells primed to the relevant antigen. B cells prepared from lymph nodes of mice primed with irrelevant antigens are not effective antigen-presenting cells for RBC-specific T cell proliferation over a wide range of SRBC doses. This is true even when both RBC and the antigen to which the B cells are primed are included in the culture. In contrast, B cells specific for a hapten determinant coupled to SRBC are able to support proliferation of T cells specific for SRBC determinants. We conclude from these data that antigen-specific B cells play a role in the induction of T cell proliferative responses to SRBC and HoRBC antigens. Two models are proposed: either B cells, upon antigen interaction with surface antibody, are able to act as accessory cells to induce Ia-dependent proliferation of immune T cells; or B cells augment the T cell proliferative response by secretion of antibody, leading to opsonization of the antigen for macrophage uptake and presentation.
Expression of the transferrin receptor in murine peritoneal macrophages is modulated in the different stages of activation.
Authors: Hamilton T A , Weiel J E , Adams D O
Published: December 2022

A specific high affinity binding site for the serum glycoprotein transferrin was identified on murine peritoneal macrophages. The binding reached equilibrium within 60 min and was reversible, saturable, and specific for transferrin. Although the presence of this receptor was detected on the cell surface by studies carried out using intact cells, the majority (70 to 90%) of the binding activity was detectable only in detergent extracts of such cells. This suggests that a substantial portion of the binding activity is localized within the macrophage. The association constant (Ka) for binding to intact cells (6 to 9 X 10(8) M-1) was comparable to values reported for transferrin receptors described on other cell types. The expression of transferrin binding activity was examined in macrophages exhibiting qualitatively and quantitatively different degrees of functional activation. Resident peritoneal macrophages, exudate macrophages primed by elicitation with pyran copolymer, and activated macrophages induced by chronic infection of mice with bacillus Calmette-Guerin (BCG) or elicitation with heat killed Propionibacterium acnes (P. acnes) had low numbers of binding sites (1000 to 5000 total sites/cell). Macrophages elicited by sterile inflammatory agents (thioglycollate broth, fetal bovine serum, or casein) all exhibited a greater number of transferrin receptors (15,000 to 20,000 total sites/cell). This modulation did not appear to result from differential shifts between surface and internal loci. Our results suggest that the expression of the transferrin receptor may be a useful marker of the responsive stage of macrophage functional activation and the membrane changes that accompany activation.
Human suppressor cell induction in vitro: preferential activation by class I MHC antigen.
Authors: Goeken N E
Published: December 2022

Human peripheral blood lymphocytes heated at 45 degrees C for 1 hr were found to continue to express all the serologically detected class II MHC antigens (HLA DR, MT, MB) but not to stimulate proliferation in primary or secondary MLR. Such cells did, however, stimulate the formation of potent suppressor cells. Three additional stimulator cell models for the presentation of either class I antigen only (purified platelets and purified T cells) or class I antigen plus nonimmunogenic class II antigen (D/DR-compatible cells) gave identical results. Supernatants from cultures stimulated with any of these cell types had significantly reduced IL 2 activity when compared to control MLR. The suppressor cells generated in such cultures were not restricted to the class I or class II MHC antigen of the original stimulator. These data are interpreted to mean that 1) the class II epitopes detected by alloantisera and the epitopes that serve as lymphocyte-activating determinants are metabolically or conformationally distinct, and 2) that presentation of class I MHC antigen alone or in conjunction with nonimmunogenic class II MHC antigen preferentially stimulates the formation of suppressor cells. It is hypothesized that the latter may be an additional mechanism that contributes to the efficacy of matching for class II determinants in human renal transplantation.
Immunoactive products of placenta. III. Suppression of natural killing activity.
Authors: Kolb J P , Chaouat G , Chassoux D
Published: December 2022

Placental and trophoblast-enriched cells from iso or allopregnant mice (14 to 17 days of gestation) strongly suppress natural killing by mouse splenocytes. The suppression is not strain restricted and acts at the level of the effector cell. It was not reduced by pretreatment of trophoblast cells with RNA and protein synthesis inhibitors, nor by prostaglandin synthetase inhibitors. However, treatment with trypsin abolished the suppressive activity. Because CTL and ADCC cytotoxicity were also inhibited by trophoblast cells at the effector stage, we postulate that trophoblast cells in vivo are endowed with the ability to inactivate various types of cytotoxic cells in their vicinity that are potentially harmful for the conceptus.
Preferential elimination of NK and CTL functions by anti-D44 monoclonal antibody.
Authors: Calvo C F , Boumsell L , Kolb J P , Laffy B , Bernard A , Senik A
Published: December 2022

Monoclonal antibodies reactive with T cells at various stages of maturation were used in negative selection experiments to study their effects on NK function in the presence of complement. Anti-D47 and anti-A50, respectively, directed against corticothymocytes and mature peripheral E+ cells were without effect. Anti-D66 reactive with an epitope of the T cell E receptor inhibited up to 60% of NK activity. Anti-D44, which primarily recognizes corticothymocytes and 60 to 80% of the E(+)-PBL was found to abrogate NK activity together with alloreactive CTL reactivity but to leave intact most of the MLR and PHA proliferative responses. Therefore D44 appears as a discrete antigen allowing preferential elimination of NK cells and CTL from PBL.
Characteristics of human NK clones: target specificity and phenotype.
Authors: Allavena P , Ortaldo J R
Published: December 2022

Clones derived from purified human large granular lymphocytes (LGL) of three different donors were expanded in culture medium supplemented with interleukin 2 (IL 2). Their cytotoxic activity was tested in a 51Cr-release cytotoxicity assay against a panel of three to five NK-susceptible tumor cell lines. Of 196 LGL clones tested, only 44 (22.4%) displayed significant cytotoxic activity. A heterogeneous pattern of reactivity was seen; 26 clones (59%) killed all the targets within the panel tested, whereas 18 clones (41%) had a more restricted specificity. Among these 18 clones, 12 lysed only one target (K562, six clones; ADCC, three clones; Daudi, two clones; MOLT-4, one clone), whereas the other six killed two different targets (ADCC and A1ab, one clone; K562 and MOLT-4, five clones). Clones derived from LGL preselected for positive reactivity with the monoclonal antibodies (MoAb) alpha OKM1, alpha OKT10 and alpha B73.1 also demonstrated either broad or restricted patterns of cytotoxicity. The LGL reactive with MoAb alpha B73.1 gave a high percentage of cytotoxic clones. Phenotype analysis showed that clones could express both antigens associated with T cells (i.e., OKT3, OKT4, and OKT8) and antigens shared by LGL (i.e., OKM1, OKT10, and B73.1). The pattern of surface markers varied considerably among the clones; however, no clear correlation between the pattern of antigenic phenotype and cytotoxic activity was seen. These data show that clones derives from purified preparations of LGL present different functional and antigenic characteristics, and support the hypothesis that the heterogeneity of the entire NK population is attributable, at least in part, to a mixture of clones that vary substantially in their target specificities and phenotypes.
Partial purification and characterization of a BCGFII from EL4 culture supernatants.
Authors: Dutton R W , Wetzel G D , Swain S L
Published: December 2022

Two B cell growth factor activities have been previously described. One activity, present in the culture supernatants of PMA-induced EL4 is active in a co-stimulator assay with normal B cells and anti-immunoglobulin. The other activity is present in the culture supernatants of the alloreactive T cell line C.C3.11.75 and can be assayed in a co-stimulator assay with normal B cells and dextran sulfate or with BCL1 in vivo line B cell tumor. We have termed the first activity BCGFI and the second BCGFII. We have now shown that a very similar BCGFII activity can be obtained from EL4 culture supernatants induced by PMA. This (EL4)BCGFII has an apparent m.w. of 55,000, is eluted from DEAE Sephacel at 0.05 M NaCl, and has a pI of 5.5, which is clearly distinct from the properties of (EL4)BCGFI activity. (EL4)BCGFII activity is similar to but not identical to (DL)BCGFII. It differs from (DL)BCGFII in chromatographic behavior and in the kinetics of the response of BCL1 to the two factors. (EL4)BCGFII activity can be detected in 18 to 24 hr by virtue of its ability to cause in vitro proliferation of in vivo BCL1 tumor B cells.
Interaction of C3 with antigen-antibody complexes in the process of solubilization of immune precipitates.
Authors: Takata Y , Tamura N , Fujita T
Published: December 2022

The binding properties of activated C3 to immune complexes were studied by using solubilization phenomenon as a model system. IgG or F(ab')2 immune precipitates were solubilized by the six isolated alternative pathway proteins, and the solubilized complexes were analyzed by SDS-PAGE. As a result of solubilization, we observed some high m.w. bands. Under reducing conditions, the bands with m.w. of 150,000 and 115,000 appeared in the case of IgG and F(ab')2 complexes, respectively. Two-dimensional SDS-PAGE revealed that hydroxylamine treatment resulted in the dissociation of the 150,000-m.w. polypeptide into the C3 alpha-65 and the heavy chain of IgG. Similarly, the 115,000-m.w. polypeptide was dissociated into the C3 alpha-65 and the Fd chain. Therefore, it is likely that iC3b binds covalently to the Fd region of the heavy chain of IgG via an ester bond. Under nonreducing conditions, iC3b-IgG and iC3b-F(ab')2 complexes had apparent m.w. of 340,000 and 270,000, respectively, corresponding to one iC3b molecule bound to one antibody molecule. In addition, a considerable amount of iC3b also binds to antigen molecules via an ester bond. The findings that C3 binds to the F(ab')2 molecules and bovine serum albumin, which contain only a small amount of carbohydrate, suggest that C3 may not bind to the carbohydrate moiety of antibody molecules. Indeed, various carbohydrate molecules did not inhibit the solubilization even at high concentrations. In contrast, acetyl tyrosine having an aromatic ring and a hydroxyl group produced the best inhibition of the solubilization. Furthermore, we demonstrated that generation of C3b in the presence of 3H-tyrosine resulted in covalent binding of the tyrosine specifically to the C3 alpha' chain, indicating that the inhibition of solubilization may be due to the competition between tyrosine and immune complexes for the covalent binding of C3. Thus, it could be concluded that C3 binds covalently to the amino acid residues of antigen and antibody molecules during solubilization.
Human B lymphocytes show greater susceptibility to H2O2 toxicity than T lymphocytes.
Authors: Farber C M , Liebes L F , Kanganis D N , Silber R
Published: December 2022

Lymphocytes from patients with chronic lymphocytic leukemia (CLL) and from normal subjects were incubated with a glucose-glucose oxidase hydrogen peroxide (H2O2) generating system to study the effect of oxidant stress on these cells. Within 4 hr, 90% of normal but only 21% of CLL lymphocytes remained viable. When normal and CLL preparations enriched in B or T cells were exposed to H2O2, B lymphocytes from both groups were highly susceptible to oxidative damage while T lymphocytes were relatively resistant. The H2O2 scavenger catalase prevented the cytotoxicity. The present work identifies the human B lymphocyte as a cell that should be a suitable target for selective killing by H2O2-generating systems.
Immunologically induced generation of tetraene and pentaene leukotrienes in the peritoneal cavities of menhaden-fed rats.
Authors: Leitch A G , Lee T H , Ringel E W , Prickett J D , Robinson D R , Pyne S G , Corey E J , Drazen J M , Austen K F , Lewis R A
Published: December 2022

The generation of sulfidopeptide leukotrienes and leukotriene B (LTB) in response to an IgG-mediated immune complex reaction in the peritoneal cavities of rats fed either a menhaden oil-supplemented diet or a beef tallow-supplemented diet for 9 to 10 wk was determined with the combined techniques of radioimmunoassay (RIA) and reverse-phase high performance liquid chromatography. Rats on the fish fat diet (FFD) incorporated eicosapentaenoic acid (EPA) into pulmonary and splenic tissues with an EPA:arachidonic acid ratio of approximately 2:1, whereas rats on the beef fat diet (BFD) showed no detectable EPA. The estimated total quantities of immunoreactive sulfidopeptide leukotrienes generated by each group of rats were similar, ranging from 70 to 99 ng/ rat in the FFD groups and 65 to 109 ng/rat in the BFD groups; for rats on the FFD this total included the pentaene products LTC5, LTD5, and LTE5 in quantities ranging from 24 to 39 ng/rat. The total quantities of immunoreactive LTB generated in the two groups of rats were similar, being 6 to 29 ng LTB4/rat for the BFD groups and the sum of LTB4 and LTB5 of 8 to 36 ng/rat for the FFD groups. There was a two- to seven-fold preferential generation of immunoreactive LTB5 over LTB4 in the FFD rats. LTC5 was equipotent with LTC4 in contracting guinea pig pulmonary parenchymal strips and ileal tissues. In contrast, LTB5 was 1/30 to 1/60 as potent and did not reach the same maximum as LTB4 in eliciting neutrophil chemotaxis. The finding that FFD favors the immunologic generation of LTB5, which has attenuated biologic activity when compared to LTB4, suggests that EPA-enriched tissues may produce less pro-inflammatory activity than tissues that are EPA-poor.
Stimulation of natural killer cells in two random-bred strains of athymic rats by interferon-inducing pyrimidinone.
Authors: Lotzová E , Savary C A , Khan A , Stringfellow D A
Published: December 2022

We tested the effect of 2-amino-5-bromo-6 phenyl-4 pyrimidinol (ABPP), an interferon (IFN) inducer, on NK cell cytotoxicity in various tissues of athymic and euthymic rats. Significant augmentation of NK cell cytotoxicity was observed in SP, PB, PE, lung, and liver of adult rats. Cytotoxic potential was also induced by ABPP in young rats, not exhibiting cytotoxicity before the treatment. Cytotoxic cells were detected in nylon wool-filtered fraction, and were not removed by carbonyl iron treatment. Separation on Percoll gradient indicated that ABPP-stimulated cytotoxic SPC resided in LGL-enriched fractions and some of them exhibited less dense characteristics than SPC from untreated rats. The ABPP-mediated cytotoxicity was thymus-independent; both athymic and euthymic rats were equally augmented. The observation that only tissues displaying NK cell augmentation produced significant levels of IFN suggest possible involvement of IFN in ABPP-mediated augmentation.
An outer membrane-disorganizing peptide PMBN sensitizes E. coli strains to serum bactericidal action.
Authors: Vaara M , Viljanen P , Vaara T , Mäkelä P H
Published: December 2022

The small cationic outer membrane-disorganizing peptide PMBN sensitized four smooth, encapsulated strains of Escherichia coli (serotypes 02:K1, 04:K12, 018:K1, and 018:K5) to the lethal action of serum. The concentrations of PMBN required were low (0.3 to 1.0 microgram/ml). One E. coli strain (IH 11030; 075:K5) remained virtually resistant to serum and also to anti-075 hyperimmune serum plus complement (C) even in the presence of PMBN. This strain was nevertheless sensitive to the outer membrane permeability-increasing action of PMBN. In the bactericidal system, PMBN could be replaced by high concentrations of lysine20 or protamine but not lysine4. The PMBN-dependent bactericidal activity of GPS was abolished by heating or zymosan treatment that inactivate its C but not by lack of the action of the classical pathway of the C in C4-deficient GPS. PMBN formed a bactericidal system also with normal rabbit, rat, and human serum but not with mouse serum. The bactericidal system against E. coli 018:K1 and its derivative EH 817 (018:K1-) was found to require a factor that can be removed from normal sera by absorption with a rough E. coli strain. This factor could be replaced by specific anti-018 antibodies. The bactericidal activity of fetal calf serum plus PMBN against E. coli 018:K1 was enhanced by normal rabbit or anti-E. coli 018 hyperimmune serum. We suggest that PMBN unshields the deep structures and the hydrophobic membrane milieu of the outer membrane and facilitates the insertion of the membrane attack complex of the C into this milieu.
Demonstration of species-specific and cross-reactive components of the adult microsomal antigens from Schistosoma mansoni and S. japonicum (MAMA and JAMA).
Authors: Tsang V C , Hancock K , Maddison S E , Beatty A L , Moss D M
Published: December 2022

Analysis of human serum reactivities to the Schistosoma mansoni adult microsomal antigens (MAMA) showed that S. japonicum and S. haematobium infection sera, as a rule, did not react as well to MAMA as did the homologous S. mansoni infection sera. The degree of species specificity, although not absolute, was quite pronounced. Purification of the corresponding microsomal antigens from S. japonicum adults (JAMA) and subsequent assays with both homologous and heterologous infection sera show a distinct and reciprocating species specificity between S. mansoni and S. japonicum microsomal antigens. The specificities of these antigens were quantitated by k-ELISA. Qualitative analysis of active antigenic components for both JAMA and MAMA involved assay by the "Western blot" or enzyme-linked immunoelectrotransfer blot (EITB). The EITB patterns of both antigens, after resolution by SDS-PAGE, show species-specific reactive bands at the 16,000 to 35,000 m.w. region. S. japonicum-specific antigens are located at the 18,000 to 35,000 m.w. region whereas S. mansoni-specific antigens were associated with m.w. components of 16,000 to 29,000. High m.w. antigen components (greater than 40,000) of both JAMA and MAMA are recognized by both heterologous and homologous infection sera and are thus not species specific. The demonstration of the clear separation of species-specific antigen bands of JAMA and MAMA by physical size offers a unique opportunity to isolate and characterize the species specificities of antibody-antigen reactions in these parasitic infections.
Variation in V lambda genes in the genus Mus.
Authors: Scott C L , Potter M
Published: December 2022

The complement of Ig V lambda genes in nine species of feral mice representing the four extant subgenera of the genus Mus was examined and compared with that of BALB/c inbred mice. Although all inbred strains examined have two V lambda genes, there is variation in the number of copies of V lambda genes in the wild mice. All feral representatives of M. musculus domesticus, from which inbred strains are derived, have at least three V lambda genes, indicating that a V lambda gene may have been lost during the inbreeding process. At least three V lambda genes are also found in representatives of three other M. musculus subspecies, including the stock of M. musculus musculus "Czech II" shown to have at least 12 C lambda genes. In comparing the complement of V lambda and C lambda genes in these animals, evidence is found that supports a mechanism of lambda gene reiteration involving duplication of a unit containing a V lambda and two C lambda genes. However, the possibility that C lambda gene amplification occurred independent of V lambda gene evolution cannot be ruled out. M. spicelegus and M. spretus, species that are semifertile with M. musculus, have one to three V lambda genes. Species more distantly related to M. musculus, such as M. cookii and M. platythrix, appear to have more (four to six) V lambda genes. Greater V lambda gene heterogeneity is also found in these animals. We propose that the ancestors of the subgenus Mus had more V lambda genes than are seen in modern species and that the paucity of V lambda genes in M. musculus, M. spicelegus, and M. spretus may be the result of V lambda gene deletion events that occurred since the divergence of the ancestor of these three species and those of the distantly related species.
Recovery of myelin basic protein reactive T cells from spinal cords of Lewis rats with autoimmune encephalomyelitis.
Authors: Burns J , Rosenzweig A , Zweiman B , Moskovitz A , Lisak R
Published: December 2022

Mononuclear cells were isolated from the spinal cords of 13 Lewis rats with clinical EAE induced by immunization with MBP in complete Freund's adjuvant. MBP-reactive lymphocytes were recovered from nine of these 13 SC-MNC populations by in vitro culture with MBP, followed by propagation of responding lymphocytes in IL 2-containing medium. T cell subset phenotype determination indicated a predominance of helper phenotype T cells in each of the two cultures examined. Clinical EAE was induced in naive Lewis rats by adoptive transfer of MBP-reactive lymphocytes recovered from three of these cultures.
Regulation of the primary in vitro response to TNP-polymerized ovalbumin by T suppressor cells induced by ovalbumin feeding.
Authors: Cowdery J S , Johlin B J
Published: December 2022

Spleen cells from DBA/2 mice that received a single feeding of 20 mg of ovalbumin (OVA) 7 days previously were specifically hyporesponsive to primary in vitro challenge with the thymic-dependent antigen TNP-polymerized ovalbumin (TNP-POL-OVA). The tolerance observed in spleen cells from OVA-fed animals was dependent upon OVA-specific T suppressor cells, because splenic T cells from OVA-fed mice suppressed the primary response to TNP-POL-OVA of cultures containing normal T and B cells. The tolerance and suppression was OVA specific, because spleen cells from OVA-fed animals responded well to other antigens (including TNP on another carrier), and splenic T cells from OVA-fed mice did not affect the response of normal T and B cells to sheep erythrocytes. These data confirm the existence of T suppressor cells after OVA feeding and provide a direct means of assaying their activity in a primary in vitro response.
Expression of Ia antigens on macrophages is reduced after stimulation with homologous C3b.
Authors: Mauer U , Burger R , von Steldern D , Bitter-Suermann D , Hadding U
Published: December 2022

The complement cleavage product C3b was examined for its ability to influence the expression of Ia antigens on macrophages. Monoclonal anti-Ia antibodies were applied for quantitation of Ia antigens by using FACS analysis. Albumin-elicited guinea pig peritoneal macrophages were cultured for 48 hr in the presence of C3b. C3b at final concentrations varying from 10 to 80 micrograms/ml reduced the proportion of Ia-positive macrophages from 87 to 25%. The C3b-stimulated loss of Ia antigens from the macrophage surface was mediated by metabolites of the cyclooxygenase pathway as shown by the inhibitory effect of indomethacin. Exposure of macrophages to C3b for 5 min was sufficient to induce Ia reduction, seen after subsequent 48-hr incubation. C3b generated in vivo in the course of inflammatory reactions might therefore influence the interaction of immunocompetent cells by modulating macrophage Ia expression.
Analysis of the binding of fluorescent C5a and C3a to human peripheral blood leukocytes.
Authors: van Epps D E , Chenoweth D E
Published: December 2022

Fluorescein-labeled human C5a and C3a were prepared and utilized to analyze the binding of C5a and C3a to human neutrophils and mononuclear cells. The fluorescein derivatives of C5a (Fl-C5a) and C3a (Fl-C3a) contained approximately one fluorescein molecule per molecule of protein. Fl-C5a retained biologic activity as determined by neutrophil O2- production, enzyme release, receptor binding, and reaction with rabbit anti-C5a antibody. Fl-C3a was biologically active as measured by contraction of guinea pig ileal strips, and maintained 87% of its antigenic character when reacted with rabbit anti-human C3a. The binding of Fl-C5a and Fl-C3a to human neutrophils and mononuclear cells was assessed with the use of flow cytometry. Fl-C5a bound to greater than 90% of neutrophils, with an average ED50 ranging from 2.8 to 6.8 nM, depending on the method of analysis. Fl-C5a binding to neutrophils was specific and was not inhibited by the presence of formyl-methionyl-leucyl-phenylalanine (f-MLP), C3a, or casein. Fl-C5a binding was totally blocked by an excess of C5a. C5a des arg partially inhibited the binding of Fl-C5a to neutrophils, but was 1000-fold less effective than C5a. Similar experiments with mononuclear cells showed that Fl-C5a was bound by monocytes but not by lymphocytes. Fl-C5a binding to monocytes was blocked totally by C5a but not by C3a or f-MLP. Comparative binding studies with neutrophils, monocytes, and lymphocytes showed that Fl-C5a was bound by an average of 93% +/- 4 of neutrophils, 68% +/- 9 of monocytes, and 6% +/- 3 of lymphocytes. Fl-C3a did not show significant binding to neutrophils, monocytes, or lymphocytes. These studies demonstrate that fluorescein derivatives of C5a and C3a can be prepared with retention of biologic activity, and provide a means to evaluate the binding of C5a to individual cells.
Human lung macrophages enhance and inhibit lymphocyte proliferation.
Authors: Liu M C , Proud D , Schleimer R P , Plaut M
Published: December 2022

To evaluate the effector functions of human lung macrophages, cell preparations containing 70 to 95% macrophages were obtained from surgically resected lungs of cancer patients and were co-cultured with allogeneic or autologous peripheral blood mononuclear cells (PBMC) and Con A. In contrast to previous reports of either marked stimulatory or inhibitory effects of human lung macrophages on lymphocyte function, the present results demonstrate that the proliferative response was a complex function of both the numbers of PBMC and macrophages. In the presence of low numbers of PBMC, small numbers of macrophages enhanced proliferation, whereas larger numbers inhibited proliferation; in the presence of larger numbers of PBMC, macrophages only inhibited. Inhibition was not mediated by cyclo-oxygenase products of arachidonic acid metabolism, because indomethacin did not reverse it. The enhancing effect of macrophages was greater when tested with PBMC depleted of monocytes. Lung macrophages were 10-fold more potent in mediating enhancement than corresponding numbers of peripheral blood monocytes. Both the enhancing and the inhibitory activities could be reproduced by lung macrophage lysates or supernatants derived from macrophages cultured in serum-free medium. Macrophages cultured at high density yielded inhibitory supernatants, which on dilution resulted in enhancing activity. The enhancing activity appeared in supernatants maximally after 24 hr, and its appearance was not inhibited by culturing macrophages in the presence by culturing macrophages in the presence of cycloheximide. Sephacryl S-200 chromatography of such supernatants yielded two peaks of enhancing activity, with apparent m.w. of 160,000 and 40,000, which we call lung macrophage-derived lymphocyte-activating factors (LM-LAF). Fractions with LM-LAF activity contained no IL 1 activity (assessed by augmentation of mitogen-induced proliferation of mouse thymocytes), but IL 1 activity was present in a peak of m.w. of 15,000. The 15,000 m.w. fraction did not enhance the proliferation of human PBMC. These results demonstrate that human lung macrophages are potent modulators of lectin-mediated proliferation of human PBMC. The effects are mediated in part by the release of soluble, pre-formed factors that appear to be distinct from previously described monokines.
Studies of congenitally immunologically mutant New Zealand mice. IX. Age-related microenvironmental effects on autoantibody production in NZB and NZB.Xid mice studied by transplantation.
Authors: Bray K R , Gershwin M E , Skelly R R , Ahmed A , Kincade P W
Published: December 2022

The mechanism of polyclonal expansion of B cells and subsequent autoantibody production in New Zealand mice remains a critical question. We have been studying the requirements for autoantibody production both in NZB mice as well as NZB mice congenic with the Xid gene of CBA/N mice. In this study, we have attempted to alter the immunologic phenotype of NZB.Xid mice by transfer of cells from young and old NZB mice. There was little difficulty in restoring normal levels of serum IgM, IgG3, splenic Lyb-5 cells, and response to DNP-Ficoll in young NZB.Xid mice that were injected with young NZB bone marrow cells. Although such animals had an almost immediate change in their immune profile to values characteristic of NZB mice, they required, much like unmanipulated NZB mice, a latency period of an additional 6 mo before autoantibodies were detected. In contrast, adult NZB.Xid mice, who likewise developed an immune profile similar to NZB after transfer of bone marrow cells from young NZB mice, began to express autoantibodies immediately without any latency period. NZB.Xid mice who were recipients of adult NZB bone marrow cells did not show sustained autoantibody production, reflecting the limited state of B cell precursors in adult NZB mice. Thus, the age of both donor cells and the age of recipient mice are critical factors for determining the latency period and the age at which autoantibodies will appear. Similarly we attempted to alter the production of autoantibodies in NZB mice that were irradiated and injected with bone marrow cells from NZB.Xid animals. NZB mice had a major amelioration of disease when they received cell transfers from young NZB.Xid mice. This amelioration, which included the acquisition of the immune profile of NZB.Xid animals, was not seen in adult NZB mice that were recipient of young NZB cells. We suggest that although Lyb-5 cells may be the effective mechanism for autoantibody production, there are other interacting influences that may selectively turn on or turn off autoantibodies and that are required and are responsible for the latency period.
Accessory function and interleukin 1 production by human leukemic cell lines.
Authors: Wakasugi H , Harel A , Dokhelar M C , Fradelizi D , Tursz T
Published: December 2022

Highly purified T lymphocytes do not proliferate in response to mitogens, unless adherent HLA-DR-positive monocytes are added to the culture. This accessory function (AF) of monocytes requires the release of interleukin 1 (IL 1). Cells from three human leukemic cell lines, K562, HL60, and U937, could very efficiently replace monocytes in a 72-hr mitogen-induced T cell proliferation assay. The AF was clearly related to precise maturational stages of these cells; the hematopoietic precursor K562 cells spontaneously exerted high AF, but lost this property when treated with differentiation inducers. On the contrary, the promyelocytic HL60 cells and the "histiocytic" U937 cells exhibited no spontaneous AF, but acquired this property when induced to differentiate along the granulocytic and/or monocytic pathway. Three leukemic cells could not only stimulate T cells to proliferate and produce IL 2 in the presence of mitogens, but also under appropriate culture conditions these cells could produce IL 1, which could not be distinguished from normal human monocyte derived IL 1 by gel filtration and isoelectric focusing. Moreover, analysis of phenotypic markers revealed that AF and production of IL 1 could be demonstrated in different cell types and therefore are not restricted to the monocytic lineage. No HLA-DR antigen could be detected on K562 and HL60 cells. Thus, the expression of the DR antigens is not required for AF and IL 1 production in response to mitogens. These three human leukemic cell lines will provide convenient sources of human IL 1.
Immunologic characterization and molecular profile of carcinoembryonic antigen detected by monoclonal antibodies.
Authors: Imai K , Moriya Y , Fujita H , Tsujisaki M , Kawaharada M , Yachi A
Published: December 2022

Four distinct monoclonal antibodies, which reacted with CEA preparations but not with nonspecific cross-reacting antigen or with nonspecific cross-reacting antigen 2, were established. Except for monoclonal antibody AS001 , all of these monoclonal antibodies immunoprecipitated molecular forms of 200K and 180K daltons that are not bridged by disulfide bonds. Immunodepletion experiments and sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis revealed that these monoclonal antibodies recognized the same antigenic structure when 125I-CEA preparation was used. Monoclonal antibody AS001 is of particular interest, because this antibody reacted only with a 200K dalton molecule which is a part of the molecules recognized by the other three monoclonal antibodies. The rosette inhibition assay and the immunoprecipitation experiments suggest that each monoclonal antibody recognizes a different antigenic determinant. The antigenic determinants recognized by monoclonal antibodies YK013 and AS001 may be peptides in nature, whereas the determinants recognized by antibodies YK024 or AS005 might be carbohydrate. The radioimmunoassay with monoclonal antibody AS001 was established, and the results clearly indicate that the incidence of positivity for the sera from digestive tract cancer patients and from lung cancer patients obtained by monoclonal antibody AS001 was higher than that obtained by the polyclonal antibody. Monoclonal antibody AS001 was able to detect the corresponding antigen in the sera, which the polyclonal antibody failed to detect. This study therefore suggests that monoclonal antibodies may enhance and improve the diagnostic value in cancer patients with undetectable or lower CEA levels detected by conventional anti-CEA antibodies.
Altered surface distribution of both C3b receptors and Fc receptors on neutrophils induced by anti-C3b receptor or aggregated IgG.
Authors: Jack R M , Fearon D T
Published: December 2022

Human neutrophils to which monospecific Fab' or F(ab')2 anti-C3b receptor had been bound at 0 degrees C were incubated for timed intervals at temperatures ranging from 0 degrees C to 37 degrees C, after which the cells were labeled with TRITC -conjugated second antibody. Neutrophils bearing Fab' anti-C3b receptor and incubated for up to 30 min at 37 degrees C, and cells bearing F(ab')2 anti-C3b receptor and incubated at 0 degrees C, exhibited diffusely distributed punctate clusters of receptors. Neutrophils bearing the bivalent anti-receptor and incubated at 30 degrees C or 37 degrees C for 5 min had redistributed C3b receptors into caps and patches that were associated with subplasmalemmal accumulations of myosin. The redistribution of cross-linked C3b receptors was inhibited by pretreatment of the neutrophils with either cytochalasin D or chlorpromazine. On approximately one-half of the cells demonstrating capped C3b receptors there was a corresponding redistribution of Fc receptors, as demonstrated by subsequent binding of FITC-aggregated IgG (FITC agg-IgG). In contrast, capping of C3b receptors did not alter the diffuse distribution of HLA-A on these cells. Cross-linking of Fc receptors on neutrophils by FITC agg-IgG also induced temperature-dependent capping of these receptors that was inhibited by cytochalasin D and chlorpromazine. In approximately one-half of the cells demonstrating capped Fc receptors, subsequent labeling of C3b receptors revealed a similar redistribution of these receptors. Thus, the neutrophil responds to cross-linking of either C3b receptors or Fc receptors by a cytoskeletal-dependent rearrangement of both receptors that causes their overlapping topographic distribution, demonstrating a form of cooperative interaction between these two types of receptors that are involved in the phagocytic reactions of these cells.
The role of lipopolysaccharides in the action of the bactericidal/permeability-increasing neutrophil protein on the bacterial envelope.
Authors: Weiss J , Muello K , Victor M , Elsbach P
Published: December 2022

The killing of gram-negative bacteria by the bactericidal/permeability-increasing protein ( BPI ) of neutrophils requires surface binding, and is accompanied by a discrete increase in outer membrane permeability to small hydrophobic substances. This outer membrane alteration appears to be related to perturbation of outer membrane lipopolysaccharides (LPS). BPI causes extracellular release of LPS, but only at supra-saturating doses. Nevertheless, because the organization of LPS in the outer membrane is altered by pretreatment of bacteria with saturating doses of BPI (producing maximal bactericidal and permeability-increasing effects), the amount of LPS released during Tris-EDTA treatment is reduced by 80%. BPI markedly (approximately 50%) and selectively stimulates biosynthesis of LPS, suggesting an attempt by BPI -killed bacteria to repair outer membrane damage. The removal of surface-bound BPI by 40 mM Mg2+ initiates time- and temperature-dependent repair of the outer membrane permeability barrier and a further increase (approximately 170% of control) in LPS synthesis, even though the bacteria are no longer viable. Mg2+-induced repair is blocked when: 1) a temperature-sensitive mutant (Salmonella typhimurium HD50 ) with a conditional defect in LPS synthesis is incubated at the nonpermissive temperature (42 degrees C); and 2) LPS synthesis is selectively inhibited by a diazaborine derivative (Sandoz drug No. 84474). In contrast, repair is normal by the mutant at permissive temperatures (30 degrees C) and by the parent strain (S. typhimurium AG701 ) at both 30 degrees C and 42 degrees C. Inhibition (greater than 85%) of protein synthesis by chloramphenicol has little or no effect on repair. These findings indicate that the repair of the permeability barrier after the removal of BPI from the surface requires newly made LPS, but apparently no biosynthesis of other outer membrane constituents, which strongly suggests that the effects of BPI on LPS are mainly responsible for the break-down of the outer membrane permeability barrier.
Passive immunization against murine malaria with an IgG3 monoclonal antibody.
Authors: Majarian W R , Daly T M , Weidanz W P , Long C A
Published: December 2022

Spleen cells of BALB/c mice that were immune to the 17X strain of P. yoelii were fused with P3X63Ag8 myeloma cells. Two hundred fifty-three of 1053 hybrid cells produced antibodies reactive with disrupted 17X parasites in a solid phase radioimmunoassay. One of these antibodies, McAb 302, reacted with the merozoites of the 17X (nonlethal) and 17XL (lethal) variants of P. yoelii. Of greater significance, McAb 302 passively protected mice against challenge infection with the lethal variant. Mice treated with this antibody before infection developed low-grade parasitemia (less than 0.3%) of short duration when challenged with P. yoelii 17XL . In contrast, control mice that had been untreated or injected with ascites fluid lacking McAb 302 uniformly died with fulminating malaria upon challenge with the same parasite. In other experiments, McAb 302 was shown capable of controlling blood parasite levels when administered to mice with patent P. yoelii 17XL infections. Although all control mice died, mice protected with a single dose of McAb 302 ultimately cleared their infections. Regardless of how passive immunization was performed, mice given McAb 302 were resistant to subsequent challenge with P. yoelii 17XL , indicating they had developed significant immunity during their initial controlled infections. McAb 302 also showed pronounced passive protective activity against the nonlethal 17X strain of P. yoelii, which is a parasite of reticulocytes. The protection afforded by McAb 302 was specific, because mice passively immunized with this antibody died when challenged with the unrelated P. vinckei. McAb 302 was shown to possess the IgG3 isotype and precipitated a 230-kd protein plus several smaller polypeptides from metabolically labeled parasite antigen preparation derived from both variants of P. yoelii. It did not react with similar preparations of other murine plasmodial species.