The Journal of Experimental Medicine

Broberger, Ove; Perlmann, Peter

doi: 10.1084/jem.117.5.705pmid: 14015587

By means of immunofluorescent methods it has been shown that sera from children with ulcerative colitis contain antibodies which react with fetal colon cells in tissue culture. 5 out of 13 sera from patients reacted positively when tested for staining antibodies while 12 sera from healthy individuals yielded negative results. The specificity of the staining reactions was confirmed by inhibition experiments. The staining capacity of various sera was correlated to their hemagglutinating titer when tested against phenol-water extracts of human colon. The presence of blood group substances of the ABO system on fetal colon cells in tissue culture could be demonstrated by application of fluorescent H agglutinins from eel. Cross-inhibition experiments indicated that the H agglutinins stained colon antigens which were different from those reacting with the antibodies of ulcerative colitis sera. The reactivity of cultured fetal colon cells with the antibodies in ulcerative colitis sera was retained for up to 12 days, with optimal staining at 4 to 5 days. Reactivity with H agglutinins was present for a longer period, sometimes more than 20 days. Although antigen could be shown to be present on fetal colon cells in tissue culture, exposure of the culture, in the presence of fresh guinea pig serum, to sera from patients with ulcerative colitis did not lead to any visible cytotoxic damage. In order to investigate the possible cytotoxic effect of the sera with a more sensitive technique, freshly explanted fetal colon was dispersed by trypsinization and the cells labeled with 32 P-orthophosphate. Subsequently, these cells were exposed to sera, in a final concentration of 30 per cent, from patients or healthy controls in the presence of fresh guinea pig serum (final concentration 15 per cent). Approximately 20 per cent of the cellular isotope was released into the medium within 150 minutes of incubation, but the release was the same in the samples treated either with patients' sera or normal control sera. Thus, under the present conditions, the patients' sera did not exert any specific cytotoxic action on colon cells. Footnotes Submitted: 1 November 1962

Perlmann, Peter; Broberger, Ove

doi: 10.1084/jem.117.5.717pmid: 13942482

Freshly isolated fetal human colon cells were labeled with 32 P-orthophosphate or 14 C-amino acids and exposed to white blood cells from children with ulcerative colitis or from healthy controls. Exposure of the colon cells to patients' white cells led to a rapid isotope release, significantly higher than that obtained with normal white cells. After 150 minutes of incubation, 75 per cent of the total isotope present was found in the media of the colitis samples but only 40 per cent in those of the controls. Consistent results were obtained with white blood cells from 14 patients and 18 healthy individuals. Similar results were obtained with either fresh white cells or with white cells aged for 12 to 18 hours and consisting to 60 to 70 per cent of lymphocytes and to 20 to 30 per cent of large mononuclear cells. No specific cytotoxic activity could be conferred onto normal white cells by pretreating them with patients' serum containing antibodies against colon antigen. The cytotoxic action of the patients' white cells was immunologically specific, since no difference from the controls was found in the isotope release when cells from other organs or animals were similarly treated. Preliminary experiments suggested that the patients' white cells could be desensitized by pretreating them with colon extract. For obtaining a significant cytotoxic effect of the patients' white cells, the presence of 10 to 20 per cent of fresh guinea pig or human serum in the incubation medium was required. Footnotes Submitted: 1 November 1962

Notkins, Abner Louis; Shochat, Stephen J.

doi: 10.1084/jem.117.5.735pmid: 13939042

The procedure used to determine the infective titer of the LDH agent, the reproducibility of this assay, and the relationship between virus dose and plasma enzyme activity were described. Multiplication of the LDH agent began within 6 hours after infection and reached 10 10.8 ID 50 /ml of plasma within 24 hours. The titer rapidly decreased over the next 72 hours but viremia persisted for at least 16 months with titers as high as 10 5.2 ID 50 /ml. The appearance of the LDH agent in the circulation preceded the first noticeable rise in plasma LDH activity by close to 24 hours. After 10 months, when the plasma titer of the LDH agent had decreased nearly one millionfold, the plasma enzyme LDH had decreased by less than 50 per cent. The LDH agent is inactivated by ether but withstands lyophilization, and freezing and thawing. It is stable at low temperatures. Ultracentrifugation at 105,000 G for 2 hours leaves less than 0.1 per cent of the LDH agent in the supernatant fluid and filtration through gradocol membranes suggesst that the upper size of the LDH agent is about 55 mµ. Spread of the LDH agent from infected to uninfected mice kept in the same cage and transmission from mothers (infected prior to mating) to their offspring was relatively low. Footnotes Submitted: 18 December 1962

Shepro, David; Kula, Nora; Halkett, James A. E.

doi: 10.1084/jem.117.5.749pmid: 13977141

The cheek pouch of the hamster is alymphatic. Molecules, too large to penetrate the vascular endothelium, reach general circulation by slowly diffusing through the cheek pouch membrane to vessels found in the connective tissues in the neck and these vessels drain primarily into the superficial cervical node. The tissues of the cheek pouch membrane limit the diffusion of large particles and it is this "barrier" which explains, in part, the "privilege" conferred by the cheek pouch. Footnotes Submitted: 6 January 1963

Paterson, Philip Y.; Harwin, S. Martin

doi: 10.1084/jem.117.5.755pmid: 13941827

Rats regularly develop evidence of allergic encephalomyelitis (AE) 2 to 3 weeks following sensitization to nervous tissue plus adjuvant. Independent of the severity of AE which occurs, gradual recovery is the rule and by the 6th to 9th week after sensitization rats appear clinically well and microscopic lesions of AE have virtually disappeared. Pooled serum collected from rats 3 or 6 weeks after sensitization contains complement-fixing (CF) antibrain antibodies. Such pooled serum exerts a striking suppressive influence on development of AE when passively administered to rats actively sensitized to nervous tissue. Serum pools which contain CF antibrain antibody suppress the disease. Serum pools lacking CF antibody do not suppress the disease. Serum containing CF antibrain antibody after treatment with 2-mercaptoethanol no longer fixes complement with brain antigen in vitro and no longer suppresses AE in vivo . The data suggest that transfer of protection against AE by passively administered antibrain rat serum is due to an antibrain antibody, possibly the CF antibodies. The meaning of these findings is discussed in terms of the role(s) of circulating antibrain antibody in the pathogenesis of AE. Footnotes Submitted: 1 January 1963

Najarian, John S.; Feldman, Joseph D.

doi: 10.1084/jem.117.5.775pmid: 13937309

Passive transfer of tritiated thymidine-labeled lymphoid cells sensitized to the simple chemical DNFB into homologous guinea pigs resulted in positive contact skin reactions 24 hours after skin testing with DNFB. Labeled sensitized cells were found to accumulate at these sites, whereas, labeled nonsensitized lymphoid cells did not appear non-specifically in contact skin reaction sites. The labeled cells were small and large lymphocytes and immature cells of the lymphoid series. The maximum reactions were obtained at 24 hours, with an average of 3.4 per cent of the infiltrating mononuclear cells showing a label. At 48 hours, the macro- and microscopic reactions were similar to the 24 hour reactions but diminished in intensity, and the number of labeled cells in the infiltrates had decreased to 1 per cent of the total infiltrating mononuclear cells. ¼ to ⅓ of the labeled cells were found within the epidermis in the test skin sites. These data have indicated that contact sensitivity, like tuberculin sensitivity, required the sensitized cell to initiate the skin reaction and that the majority of the cellular infiltrate was the result of non-specific host response to injury. Footnotes Submitted: 24 January 1963

Kantoch, M.; Warwick, A.; Bang, F. B.

doi: 10.1084/jem.117.5.781pmid: 14030664

Using peritoneal macrophage cultures it was found that both PRI mice and their macrophages in culture were susceptible to mouse hepatitis virus and that C 3 H mice and macrophages were resistant. All F 1 macrophages and some back-cross cell cultures were susceptible. The degeneration of F 1 and back-cross macrophages obtained either from adult mouse peritoneal exudate or newborn mouse liver, occurred more slowly than PRI macrophages. Segregation of susceptibility occurred in the first back-cross generation. Tests of three back-cross generations from susceptible mice yielded about one-quarter of the mice shown to be susceptible either by direct test or test of their macrophages. A clear correlation between susceptibility in vivo and in vitro was established both in the test of the percentage segregation and in tests of individual back-cross mice. A small series of tests, however, indicated that 50 per cent of the back-cross mice had the genetic capacity to transmit susceptibility. Thus a hypothesis of two genes for susceptibility, although not excluded, may yield to a hypothesis of a single dominant gene, incompletely expressed. Resistant cells were converted into susceptible cells by ingestion of a relatively large particle containing a heat-stable substance. This susceptibility, although complete, was temporary. The nature of the factor causing the change has been discussed. Footnotes Submitted: 8 January 1963

Silverstein, Arthur M.; Uhr, Jonathan W.; Kraner, Keith L.; Lukes, Robert J.

doi: 10.1084/jem.117.5.799pmid: 13992961

The fetal lamb in utero is able to form large amounts of specific antibody in response to antigenic stimulus as early as the 66th to 70th day of the 150 day gestation period. Among the several antigens employed, the fetal lamb responded earliest, and with the highest titers, to bacteriophage φX. Slightly less effective as an antigen was horse ferritin, while ovalbumin proved to be a weak antigen, especially in younger fetuses. Ineffective in stimulating an antibody response at any time during fetal or early neonatal life were diphtheria toxoid, Salmonella typhosa , and BCG. Thus, it may not be feasible to fix precisely the time of onset of immunologic responsiveness in a species, inasmuch as it appears to differ so greatly from one antigen to another. The quantity of antibody found 10 days after φX immunization was not significantly different in fetuses injected at 60 to 120 days of gestation. The earliest anti-phage antibody produced by the lamb fetus is a macroglobulin sensitive to the action of 2-mercaptoethanol. Only in older fetuses with longer lasting stimuli were appreciable amounts of 7S γ-globulin antibodies formed. The conformity of these observations to theories on the ontogenesis of the immune response is discussed. Footnotes Submitted: 3 February 1963

Cotran, Ramzi S.

doi: 10.1084/jem.117.5.813pmid: 14023347

Rats with retrograde proteus pyelonephritis were treated with antibiotics until their kidneys became sterile. Using fluorescent antibody techniques, specific P. mirabilis antigen was found in some sterile pyelonephritic kidneys 20 weeks after cessation of treatment and presumed renal sterilization. Persistent antigen was associated with interstitial chronic inflammation but not with acute inflammation or progressive scarring. Rat gamma globulin and proteus antibody were localized in plasma cells of the renal inflammatory infiltrates. It is suggested that persistent antigen in chronic pyelonephritis may lead to the continued local appearance of antibody-producing cells. Footnotes Submitted: 30 October 1962

Amundsen, Egil; Gustafsson, Bengt E.

doi: 10.1084/jem.117.5.823pmid: 14012683

Experimental low ileal strangulation obstruction has been produced in germfree and conventional rats. The mean survival time was 240 hours in the germfree rats and 44 hours in the conventional controls. 4 of the 10 germfree rats survived 15 or more days, whereas the 10 conventional animals were all dead within 2½ days. The strangulation obstruction fluid from the germfree animals was sterile and non-toxic when injected into mice even after a fourfold concentration. The same fluid from the conventional animals contained a great number of microorganisms and caused death within 24 hours when injected intraperitoneally into mice. Footnotes Submitted: 1 January 1963