The Journal of Experimental Medicine

Lewis, Robert A.; Wasserman, Stephen I.; Goetzl, Edward J.; Austen, K. Frank

doi: 10.1084/jem.140.5.1133pmid: 4378429

The capacity to extract slow-reacting substance of anaphylaxis (SRS-A) from human lung tissue or cells after immunologic activation, together with the measurement of SRS-A in both the extract and the surrounding fluid, permits study of total SRS-A generation. That the material extracted is SRS-A was established by both differential bioassay and purification. SRS-A accumulation was entirely intracellular after limited IgE-dependent direct or reversed anaphylactic activation. Intracellular accumulation also generally preceded release, with generation of SRS-A continuing well beyond a plateau in the cellular SRS-A level and the release of preformed mediators. The quantity of SRS-A generated after immunologic activation was modulated by the introduction of exogenous cyclic nucleotides, revealing a site of cyclic nucleotide action distinct from that on mediator release. The capacity to determine not only the release of preformed mediators but also the generation of a newly formed mediator, the sum of SRS-A in cells and supernate, adds an additional dimension to the analysis of the cellular events of immediate hypersensitivity. Footnotes Submitted: 8 July 1974

Watson, James; Riblet, Roy

doi: 10.1084/jem.140.5.1147pmid: 4138849

In vivo immune responses and in vitro mitogenic responses to bacterial lipopolysaccharides (LPS) have been compared in strains of C3H mice. C3H/HeJ spleen cultures did not support mitogenic responses to LPS and in vivo these mice produce low IgM responses to LPS. On the basis of these two responses, C3H/HeJ mice have been termed low LPS responders. All other strains of C3H mice tested (C3HeB/FeJ, C3H/DiSn, C3H/Str, CWB, CSW, and C3H/Sf and its H-2 congenics) are high LPS responders supporting large in vitro mitogenic and in vivo immune responses to LPS. The immune response difference between low and high LPS responders is a quantitative one. IgM responses are observed in C3H/HeJ mice in the range of 1.0–10 µg LPS. At lower and higher LPS concentrations, immune responses are not observed. In contrast, high LPS responders elicit LPS immune responses over a much wider dose range (0.1–200 µg). The ability to respond well to LPS is dominant as shown by the response of F 1 hybrid mice of low responder and high responder strains. The linkage relationships of mitogenic and immune responsiveness to LPS have been investigated in backcross (C3H/HeJ x CWB)F 1 x CWB mice. All mice that gave in vivo immune responses to LPS also supported mitogenic responses to LPS. The defect in C3H/HeJ mice that limits mitogenic and immune responsiveness to be due to a single autosomal gene which is not linked to the H-2 histocompatibility or heavy-chain allotype loci. Footnotes Submitted: 11 July 1974

Chieco-Bianchi, Luigi; Colombatti, Alfonso; Collavo, Dino; Sendo, Fujiro; Aoki, Tadao; Fischinger, Peter J.

doi: 10.1084/jem.140.5.1162pmid: 4608945

Adult AKR and C58 mice injected intramuscularly with murine sarcoma virus, Moloney isolate (M-MSV), developed high incidence of nonregressing local tumors. Histologically, these tumors revealed the typical pleomorphism of M-MSV sarcomas; in some cases, however, neoplastic tissue showed a nodular or diffuse growth of monomorphic myoblastlike cells, reminiscent of clonal aggregates. No depression of immune reactivity was found in M-MSV-injected mice as evaluated by direct hemolytic plaque-forming cells against SRBC and by virus-neutralizing antibody production. The MSV recovered from the induced tumors proved to be, by neutralization assay, a Gross (G)-MSV pseudotype. Moreover, tumor cell suspensions absorbed out cytotoxic antibody directed against G-cell surface antigens. Therefore, the conclusion was drawn that MSV with envelope characteristics of endogenous G leukemia virus had formed in vivo through a phenotypic mixing phenomenon. The failure of tumors to regress has been interpreted as mainly due to the partial unresponsiveness of host immune reactivity towards G-MuLV specified antigens. Since MSV-tumors arose in AKR mice after a very long latent period, the possibility was considered that this relative resistance might depend on immunologic mechanisms. In fact, M-MSV-injected AKR mice immunodepressed by goat antimouse lymphocyte serum or rendered partially tolerant by neonatal M-MuLV inoculation developed sarcomas with higher incidence and with a shorter latency. Furthermore, the MSV recovered from these early tumors proved to be the original Moloney pseudotype. Footnotes Submitted: 2 July 1974

Hämmerling, Günter J.; McDevitt, Hugh O.

doi: 10.1084/jem.140.5.1180pmid: 4547372

125 I(T,G)-A--L-binding T cells have been studied in mice whose ability to mount an immune response to (T,G)-A--L is under control of the H-2 -linked Ir-1A gene. Nonimmunized high and low responder mice have approximately the same frequency of T-ABC. Following immunization, T-ABC proliferated only in high responders, but not in low responders, indicating expression of Ir-1A in T cells. When, for comparison, 125 Iarsanyl-mouse serum albumin binding B and T cells were investigated in mice whose antibody response to the hapten arsanyl is controlled by an allotype-linked Ir gene, it was found that following immunization the number of B-ABC increased only in high responders. In contrast, T-ABC proliferated to the same extent in both high and low responders, suggesting exclusive expression of the allotype-linked Ir gene in the B-cell line. Preliminary studies indicate that anti-Ia sera inhibit neither B-ABC nor T-ABC. Footnotes Submitted: 15 July 1974

Fournié, Gilbert J.; Lambert, Paul H.; Miescher, Peter A.

doi: 10.1084/jem.140.5.1189pmid: 4607609

The present data demonstrate the induction of antisingle-stranded (SS) DNA and antidouble-stranded DNA antibodies in various strains of mice, including athymic C57BL/6 nude mice, after the injection of bacterial lipopolysaccharide (LPS). This anti-DNA response is dose dependent and varies quantitatively according to the strain of the injected mice. It is not correlated to the H-2 histocompatibility locus nor to the immune response to LPS. The lipid A fraction appears to be the active part of the LPS molecule for this particular effect. In addition, it was found that DNA is released in circulating blood a few hours after the injection of LPS. Most of the DNA released has physicochemical and immunochemical characteristics of SS DNA. Therefore, the anti-DNA response induced by injections of LPS may be the result of a release of DNA in a particularly immunogenic form at a time when the immune system, in particular the B lymphocytes, is rendered capable by LPS of developing an immune response to such a soluble antigen. These effects of LPS may account for the triggering or the exacerbation of ante-DNA antibodies during infections with gram-negative bacteria, and a similar mechanism may be involved in the pathogenesis of systemic lupus erythematosus. Footnotes Submitted: 8 July 1974

Unanue, Emil R.; Karnovsky, Morris J.

doi: 10.1084/jem.140.5.1207pmid: 4547538

Capping of surface Ig by anti-Ig antibodies involves a membrane perturbation requiring an energy-dependent step. Lymphocytes treated with anti-Ig are stimulated to move. Previously, we had shown that movement was not essential for capping, although it influenced the localization of the cap. We have investigated the role of cell movement and of microtubular proteins in this phenomenon. Treatment of B lymphocytes with colchicine does not affect capping of Ig nor does it affect the increase in translational movement produced by anti-Ig antibodies. Treatment of lymphocytes with cytochalasin B stops translational movement and may affect capping to some degree under appropriate circumstances. Lymphocytes treated with both drugs are impaired in capping. We surmise that there may be two cytoplasmic events regulating directly or indirectly capping: one associated with the process of translational movement, the other associated with the activity of microtubules. Lymphocytes treated with concanavalin A do not cap Ig. Colchicine reverses this inhibition. Certain experimental procedures antagonize the colchicine effect, the most striking of which is the use of cytochalasin B. Colchicine appears to increase movement of the Con A-treated lymphocyte, and this increased movement appears responsible for the accumulation of complexes to the posterior part of the cell. Con A inhibits patching of Ig by anti-Ig, and this is not reversed by colchicine. Footnotes Submitted: 5 June 1974

Croce, Carlo M.; Koprowski, Hilary

doi: 10.1084/jem.140.5.1221pmid: 4371646

Fusion of mouse peritoneal macrophages with SV40-transformed human cells, deficient in hypoxanthine guanine phosphoribosyltransferase, resulted in the formation of transformed somatic cell hybrids which contained, without exception, the human chromosome 7 carrying the SV40 genome. It is postulated that the hybridization of mouse nondividing cells with human cancer cells could permit the identification of the human "oncogenic" chromosome(s) present in human cancer cells, since such chromosome(s) should be retained by the totality of the mouse-human hybrid cells. Footnotes Submitted: 4 June 1974

Theofilopoulos, Argyrios N.; Wilson, Curtis B.; Bokisch, Viktor A.; Dixon, Frank J.

doi: 10.1084/jem.140.5.1230pmid: 4138539

Cells from a human lymphoblastoid cell line (Raji), with B-cell characteristics, and having receptors for human IgG Fc, C3b, and C3d, were used in an immunofluorescence test as in vitro detectors of immune complexes in animal and human sera. By this test, as little as 200–300 ng aggregated human gamma globulin or immune complexes per ml serum could be detected. The receptors for IgG Fc on the Raji cells were shown to be inefficient in detecting aggregated human gamma globulin and binding of aggregates to these receptors was inhibited by physiologic concentrations of 7S human IgG. Enhancement of aggregated human gamma globulin binding and binding of immune complexes formed in vitro to Raji cells was observed when the receptors for complement on these cells were used. By using the receptors for complement on Raji cells, circulating immune complexes were detected in rabbits with acute serum sickness, in mice with acute lymphocytic choriomeningitis virus infection, and in humans with immune complex type glomerulonephritis. The Raji cell test may be useful in detecting complement fixing immune complexes in different disease states, in monitoring circulating complexes in patients with immune complex diseases and in identifying the antigen(s) responsible for the induction of pathogenic immune complexes in humans and animals. Footnotes Submitted: 8 July 1974

Werdelin, Ole; Brændstrup, Otto; Pedersen, Eskild

doi: 10.1084/jem.140.5.1245pmid: 4138693

We have studied the physical interaction between macrophages and lymphocytes during the immune response to purified protein derivative of tuberculin (PPD) in vitro. Mixtures of peritoneal macrophages and lymph node lymphocytes from guinea pigs immunized with tubercle bacilli formed cell clusters during 20 h of culture with PPD. The number of clusters produced was correlated to the number of immune lymphocytes in the cultures. Peritoneal macrophages which had been pulsed with PPD and untreated lymph node lymphocytes produced cell clusters in the absence of free PPD in numbers equivalent to those produced by the same cells in the presence of free PPD. In cultures containing a mixture of PPD-pulsed macrophages, not-pulsed macrophages, and immune lymphocytes with no free PPD, cell clusters developed mainly between the antigen-pulsed macrophages and lymphocytes. Cluster formation was antigen-specific with the specificity residing in the lymphocytes, mainly or exclusively in the T lymphocytes. These data indicate that in the process of cell cluster formation macrophages serve as antigen-binding (or -processing) cells, while a subpopulation of lymphocytes interact physically and specifically with the macrophages. Footnotes Submitted: 25 June 1974

Nielsen, Morten H.; Jensen, Henning; Brændstrup, Otto; Werdelin, Ole

doi: 10.1084/jem.140.5.1260pmid: 4138694

Macrophage-lymphocyte clusters are formed when lymph node cells and autologous peritoneal exudate cells from guinea pigs immunized with tubercle bacilli are cultured in the presence of purified protein derivative of tuberculin (PPD) for 20 h. We have studied the ultrastructure of these clusters employing transmission and scanning electron microscopy. The most simple macrophage-lymphocyte cluster consisted of one macrophage, one large central lymphocyte with a blastoid appearance attached to the macrophage with a broad area of contact, and from a few to more than 20 small peripheral lymphocytes attached to the central lymphocyte by their uropods. Some clusters were of more complex type, containing two or three macrophages or one macrophage with more than one central lymphocyte attached to the surface, but even in these clusters each peripheral lymphocyte was attached only to one central lymphocyte. By morphological criteria the peripheral lymphocytes were T lymphocytes. Footnotes Submitted: 25 June 1974