The Journal of Experimental Medicine

Lee, Chue Shue; Mauer, S. Michael; Brown, David M.; Sutherland, David E. R.; Michael, Alfred F.; Najarian, John S.

doi: 10.1084/jem.139.4.793pmid: 4273909

Immunoglobulins and complement are deposited in the glomerular mesangium of rats with progressive glomerulosclerosis secondary to chemically induced diabetes mellitus. Isotransplantation of a kidney from a rat diabetic for 6 mo into a normal recipient results within 2 mo in the disappearance of IgG, IgM, and ß 1 C from the mesangium and arrest or reversal of the light microscopic glomerular lesions. Kidneys isotransplanted from normal donors into diabetic rats developed mesangial matrix thickening and deposition of IgG, IgM) and ß 1 C in the mesangium. No glomerular changes occur upon transplantation of a normal kidney into a normal rat. These findings indicate that diabetic glomerular changes in the rat are reversible and are secondary to the diabetic state rather than to the inducing agent. Footnotes Submitted: 14 January 1974

Holmgren, J.; Lindholm, L.; Lönnroth, I.

doi: 10.1084/jem.139.4.801pmid: 4361243

The interaction of cholera toxin and a number of toxin derivatives, containing different proportions of light and heavy toxin-composing subunits (L and H), with mouse lymphocytes was studied. Experiments with 125 Itoxin showed that a single cell can rapidly, within minutes, bind up to 40,000 molecules of toxin, the association constant was estimated to 7 ± 4 x 10 8 liters/mol, and binding was found to be very similar at 37°C and 5°C. Immunofluorescence studies revealed that the toxin attachment is located on the cell surface, and that purified L subunit but not H subunit binds to the cells. A natural cholera toxoid, built up by aggregated L subunits, showed almost identical binding properties as toxin to the cells. Pure G M1 ganglioside, the proposed membrane receptor structure for toxin, prevented entirely the cellular binding of both toxin and toxoid. Cholera toxin in concentrations down to approximately 5 x 10 –11 mol/liter (corresponding to 10 bound molecules/cell) inhibited thymus cells from being stimulated to DNA synthesis by concanavalin A (con A), and spleen cells from such stimulation by phytohemagglutinin. The G M1 ganglioside but not a series of other pure structurally related gangliosides and neutral glycosphingolipids neutralized this toxin activity. Toxin derivatives which, in similarity with toxin, possessed H as well as L subunits but in other proportions, were potent inhibitors of con A-induced thymocyte stimulation, whereas the natural toxoid (aggregated L subunits), purified toxin L subunit and purified toxin H subunit were up to 300-fold less active on a weight basis. The capacity of cholera proteins to inhibit con A-induced thymocyte stimulation correlated well with their activity in the rabbit intradermal toxicity assay. The inhibitory action of cholera toxin on con A-induced thymocyte stimulation did not depend on decreased cell viability from the toxin treatment, nor was it caused by a reaction between toxin and con A. 125 Icon A bound equally well to the cells when toxin was present as when it was absent, which proves that the toxin did not compete for cellular con A receptors. Nor did the toxin seem to disturb the general mobility of membrane receptors or their ability to accumulate in caps. It is concluded that the L type of subunit confers rapid and firm binding of cholera toxin to lymphocyte membranes, probably to G M1 ganglioside receptors. For biologic activity the additional presence of H subunit is important. One manifestation of toxin action on lymphocytes is inhibition of lectin-induced DNA synthesis; probably this effect relates to the ability of cholera toxin to raise the levels of intracellular cyclic 3'5'-adenosine monophosphate. Footnotes Submitted: 28 November 1973

Willenborg, David O.; Prendergast, Robert A.

doi: 10.1084/jem.139.4.820pmid: 4205878

Mice treated with sea star factor (SSF), a protein extracted from sea star coelomocytes, became highly susceptible to infection with a normally sublethal dose of Listeria monocytogenes . This was in contrast to the expected result of increased resistance originally postulated because of the macrophage-activating properties of SSF. Enhanced susceptibility was seen when SSF was given from 96 h before to 48 h after infection with Listeria . Mice pretreated with SSF failed to develop immunity to Listeria when given a dose of organisms capable of immunizing nontreated mice. Treatment of immune mice with SSF did not alter their immune status. In addition, incubation of immune lymphocytes with SSF in vitro did not alter their ability to adoptively transfer immunity to normal recipients. Immune lymphocytes treated with SSF and then incubated with anti-SSF and C did, however, lose the ability to transfer immunity. These results suggest that SSF enhances infection by binding to T lymphocytes, inhibiting their replication upon contact with Listeria antigen and thus preventing the generation of a population of sensitized lymphocytes capable of effecting anti- Listeria immunity. Footnotes Submitted: 28 November 1973

Unkeless, Jay C.; Gordon, Saimon; Reich, E.

doi: 10.1084/jem.139.4.834pmid: 4816302

Cultured thioglycollate-stimulated peritoneal macrophages synthesize, accumulate, and continuously release high levels of plasminogen activators for at least 4 days whereas cultures of unstimulated macrophages do not; the higher specific catalytic activity of released vs. cell-associated enzyme suggests that the plasminogen activators are actively secreted. The major macrophage plasminogen activator is a serine protease of mol wt 48,000, and thus resembles the comparable enzyme released by virally transformed fibroblasts. Macrophages release a second plasminogen activator of mol wt 28,000 that is also a serine enzyme. The secretion products released by stimulated and unstimulated macrophages have been compared by SDS-polyacrylamide gel electrophoresis after chemical labeling with 3 H-DFP or biosynthetic labeling with 14 C-amino acids. These procedures show that some proteins are formed in both cultures, whereas others are uniquely secreted by each type of macrophage. The serine enzymes released by the two kinds of macrophages differ in specificity and electrophoretic mobility. Footnotes Submitted: 1 January 1974

Plate, Janet M. D.

doi: 10.1084/jem.139.4.851pmid: 4131511

Cellular responses in vitro to H-2D region histocompatibility antigens were demonstrated to be under the genetic control of two or three ( P = 0.013) independently segregating loci. The H-2 region itself accounts for one of these loci, however, its activity appears to be dependent upon an association with other non- H-2 -associated genetic information. The ability to stimulate a response and to respond to that stimulus are two separate genetic functions in certain MLR combinations. The stimuli in our studies were products of the H-2D region and cell donors must differ at that region in order for a response to occur. The control of the level of responses was determined by other genetic material. Differences at these "response loci" were not necessary for the induction of a proliferative response in the mixed lymphocyte cultures. Footnotes Submitted: 1 January 1974

Vitetta, Ellen S.; Grundke-Iqbal, Inge; Holmes, Kathryn V.; Uhr, Jonathan W.

doi: 10.1084/jem.139.4.862pmid: 4544585

Lymphoid cells from the spleen, lymph nodes, and thoracic duct of axenic and control mice were incubated with 3 Htyrosine and synthesis and secretion of protein and Ig studied. It was found that only IgM was synthesized by cells from axenic mice whereas cells from control mice also synthesized IgG. Splenocytes from both axenic and control mice had 8S IgM on their surface. Radiolabeled splenocytes from axenic mice were incubated to determine the kinetics of release of 125 I-labeled cell surface IgM and 3 Htyrosine-labeled IgM. Cell surface IgM was shed as 8S with an initial half-life of release of 5–8 h whereas 3 Htyrosine-labeled Ig was secreted as 19S with an initial half-life of 2–3 h. These findings suggest that two independent pathways are involved. It is suggested that small lymphocytes shed 8S IgM and plasma cells secrete 19S IgM. It was observed that lymphoid cells from axenic mice synthesize a higher proportion of IgM relative to total protein. Electron microscopic examination of splenocytes from such mice revealed a markedly higher proportion of plasma cells and a paucity of lymphoblasts compared to controls. It was suggested, therefore, that axenic mice lack a population of stimulated T cells which can induce a switch from IgM to IgG synthesis and which is capable of suppressing IgM synthesis. Lymphoid cells from axenic mice synthesize and secrete less protein that coprecipitates with antigen-antibody complexes. Footnotes Submitted: 1 January 1974

Binz, Hans; Lindenmann, Jean; Wigzell, Hans

doi: 10.1084/jem.139.4.877pmid: 4544586

Antialloantibodies were prepared in F 1 hybrid rats by immunization with alloantibodies from one parent raised against antigens of the other parent. The Ig fraction of such antialloantibodies was iodinated and used to investigate the nature of idiotype-carrying normal lymphoid cells. Lymphoid cell populations of the proper genotype fixed radioactive antialloantibody in proportion to their B-cell content. Activated T-cell populations, when depleted of B cells, did not fix significant amounts of radioactivity. Idiotype-carrying cells were sensitive to rabbit antirat Ig serum lysis and to antialloantiserum lysis, but not to rabbit anti-T-serum lysis. Also, the normal idiotype-containing B cell could be shown to have the expected antigen-binding specificity of its receptor. This was shown by the use of fibroblast cell monolayers that adsorbed specifically those cells capable of fixing the proper antialloantibody. Footnotes Submitted: 16 December 1973

Kimura, Arthur K.

doi: 10.1084/jem.139.4.888pmid: 4131512

The present study describes a method for the production of a specific anti-T-cell receptor antiserum, and characteristics of its ability to block specific cell-mediated cytotoxicity in vitro. Immunization and antiserum adsorption procedures were designed to select for idiotypic differences in the recognition units of C3H lymphocytes immune to two different strains of mouse cells, such that the reactivity of only one population of effector cells is inhibited by this antiserum. Both in vivo and in vitro sensitized effector T cells are subject to this inhibition. That the site of the antiserum blockade is clearly on the effector cell and not on the target cell is demonstrated. Footnotes Submitted: 16 December 1973

Ahmed, Aftab; Strong, Douglas M.; Sell, Kenneth W.; Thurman, Gary B.; Knudsen, Richard C.; Wistar, Richard; Grace, William R.

doi: 10.1084/jem.139.4.902pmid: 4593239

Conflicting reports on the immune responsiveness of patients with subacute sclerosing panencephalitis (SSPE) have been reported. This report shows that the leucocytes from four SSPE patients exhibited strong sensitivity to both measles and SSPE virus preparations as measured by the macrophage migration inhibition test, mixed lymphocyte virus infected cell culture test, and the lymphotoxin assay. Earlier suggestions that a factor may be operating to suppress cellular reactivity are confirmed by the demonstration that the response of lymphocytes from SSPE patients could be blocked by the addition of SSPE spinal fluid or plasma. It was determined that the blocking factor was stable at –20°C, heat labile at 56°C for 30 minutes, trypsin and neuraminadase sensitive, and had a mol wt greater than 150,000 as determined by Sephadex G-200 gel chromatography. The blocking factor appeared to be specific for SSPE virus and did not block the response of lymphocytes to nonspecific mitogenic agents and other viral and bacterial agents. Footnotes Submitted: 26 October 1973

Yoshiki, Takashi; Mellors, Robert C.; Hardy, William D.; Fleissner, Erwin

doi: 10.1084/jem.139.4.925pmid: 4131513

The indirect membrane immunofluorescence test and the absorption analysis of rabbit anti-FeLV, rabbit anti-FeLVp 30, and rabbit anti-MuLVp 30 antisera yielded the following conclusions. An antigen shared by mammalian (murine and feline) C-type RNA leukemia and sarcoma viruses was detected on the surface of cells infected or transformed by C-type viruses. The antigen was characterized as membrane-bound gs antigen bearing two determinants, membrane-bound gs-1, intraspecies-specific antigenic determinant, and membrane-bound gs-3, interspecies-specific antigenic determinant. Membrane-bound gs antigen was located on the cell surface, frequently near the site of virus budding but not on the envelope of murine C-type RNA virus. Footnotes Submitted: 9 January 1974