The Journal of Experimental Medicine

Harris, Susanna; Harris, T. N.; Farber, Miriam B.

doi: 10.1084/jem.108.4.411pmid: 13575675

In earlier studies it was shown that if rabbit lymph node cells were incubated with Shigella -trypsin filtrate and transferred to other, recipient, rabbits, agglutinins to Shigella appeared subsequently in the sera of the recipients. However, if leucocytes from the donor rabbits were injected into the recipients at a suitable interval prior to lymph node cell transfer, agglutinins to Shigella did not appear after cell transfer. In the present study, a number of experiments were done which bear on the immunologic character of this suppressive effect on the transferred cells, the "pre-injection effect." X-irradiation of the recipients before the injection of donor leucocytes led to a decrease in the extent of the pre-injection effect, i.e . to the appearance of higher agglutinin liters, in the recipient animals. It was found possible to transfer the pre-injection effect by cells of rabbit lymph nodes draining sites of injection of rabbit leucocytes. No such effect was obtained with heated aliquots of these cell suspensions or with lymph node cells from sheep-erythrocyte-injected donors. It was also possible to transfer the pre-injection effect passively by serum. Suppression of transferred lymph node cells was observed regularly after injection of serum pooled from groups of rabbits which had been injected with leucocytes pooled from the blood of 60 to 70 rabbits. The active material in anti-leucocyte serum could be precipitated in the globulin fraction of the serum and could be removed by absorption with rabbit lymph node cells. In the course of experiments on the passive transfer of the pre-injection effect by antisera of individual rabbits to leucocytes of individual donor rabbits, evidence was obtained of the existence in the sera of some normal rabbits of antibodies to some of the rabbit leucocyte antigens. Footnotes Submitted: 24 April 1958

Johnstone, Douglas E.; Howland, Joe W.

doi: 10.1084/jem.108.4.431pmid: 13575676

The effect of leukopenia on the susceptibility of rabbits to the localized Shwartzman reaction was studied in those receiving whole body irradiation and nitrogen mustard respectively. Of animals receiving nitrogen mustard in which the total polymorphonuclear leukocyte count was less than 100 cells/c.mm. only 17 per cent developed positive local Shwartzman reactions whereas 75 per cent of irradiated animals with total polymorphonuclear leukocyte counts less than 100 cells/c.mm. had positive reactions. A local Shwartzman reaction occurred in 60 of 71 control rabbits (85 per cent). Animals of both the treated groups showed positive Shwartzman reactions at a time when their total polymorphonuclear leukocyte counts were less than 500 cells/c.mm. From the findings, it is concluded ( a ) that the presence of normal numbers of circulating polymorphonuclear leukocytes is not an obligatory prerequisite condition to the localized hemorrhagic necrosis of the Shwartzman phenomenon, and ( b ) the mechanisms of action of nitrogen mustard and whole body irradiation on body tissues differ in relation to the Shwartzman reaction. Footnotes Submitted: 6 May 1958

Gatling, Robert R.

doi: 10.1084/jem.108.4.441pmid: 13575677

A series of experiments are reported which illustrate and characterize an altered state of reactivity of the skin of the hypersensitive rabbit to epinephrine (and norepinephrine). The requirements for the production of the lesion are: a state of hypersensitivity, circulating antigen and antibody and focal deposition of epinephrine or norepinephrine. The size and intensity of the reaction appear to be directly related to the amount of epinephrine injected (within certain limits) and to the degree of hypersensitivity as measured by the Arthus reaction. Although circulating antigen is required, the precise quantity does not seem to be as critical as does the amount of circulating antibody; a certain minimum amount is necessary, however. Experimental evidence is given which indicates the utilization of antigen in the production of the lesion. Data tending to identify this lesion with the Arthus reaction are given. The possibility of an analogous relationship between the mechanism of this experimental lesion and the necrotizing arteritis of the collagen diseases is postulated. Footnotes Submitted: 29 December 1957

Finger, Irving; Kabat, Elvin A.

doi: 10.1084/jem.108.4.453pmid: 13575678

The limits of sensitivity of three gel diffusion methods are compared and their utilization in the detection of small amounts of antibody to antigens present in traces in a preparation is illustrated with the diphtheria toxin-human antitoxin system. The Preer modification of the Oakley-Fulthorpe technique and the Oudin tube method were found more sensitive than the Ouchteriony plate method, and permitted the detection of as little as 3 µg. antibody N/ml. of serum. Antisera from eight Schick-negative individuals immunized with purified diphtheria toxoid have all been shown to contain, in addition to antitoxin, antibodies to substances present as impurities in the purified toxoid injected. The amounts of these antibodies in a serum and a partial characterization of their antigen-antibody curves have been determined through the combined use of quantitative precipitin and gel diffusion methods. Different amounts of antibody have been precipitated by toxin and toxoid from individual sera. Evidence is presented that this may have been due to slight differences in antigenic specificity. A serum, Hu, which had been held to contain no precipitating antibody, has now been shown, by the Preer and Oudin techniques, to contain at least 12 µg. of precipitating antibody N (per ml. serum) against an impurity in the toxoid preparation. This estimate has been confirmed by quantitative precipitin determinations. The presence of antibodies to impurities in all human antitoxins examined in the present work brings into question the assumption that human antitoxin as such has a skin-sensitizing capacity. Footnotes Submitted: 19 May 1958

Stollerman, G. H.; Kantor, F. S.; Gordon, B. D.

doi: 10.1084/jem.108.4.475pmid: 13575679

The bloods of two apparently healthy human beings, of 25 studied, failed to produce a strong bactericidal test for type-specific antibody to the M protein of group A streptococci under in vitro conditions wherein most human blood leukocytes rapidly phagocytize and destroy virulent organisms in the presence of anti-M antibody and accessory plasma factors. The defect in bactericidal activity of these two individuals is associated with the plasma rather than with the blood leukocytes. Leukocytes suspended in these atypical plasmas showed a characteristic delay in the rate of activation of phagocytosis. Although previously the bloods of laboratory animals (except monkeys) had been reported to be much less active than human blood in this system, occasional exceptions were encountered in rabbits in this study. Two rabbits were found whose bloods were as strongly bactericidal against streptococci, in the presence of type-specific antibody, as the blood of the average "normal" human being. The atypical behavior of some human and rabbit bloods in the bactericidal test may be explained by variations in accessory plasma factors that are as yet unidentified and that influence the rate of phagocytosis of virulent streptococci in vitro in the presence of type-specific antibody. Footnotes Submitted: 13 June 1958

Alexander, Hattie E.; Koch, Gebhard; Mountain, Isabel Morgan; Van Damme, Olga

doi: 10.1084/jem.108.4.493pmid: 13575680

Ribonucleic acid prepared by the method of Gierer and Schramm from concentrated and partially purified types I and II polioviruses has been demonstrated to be infectious for HeLa and human amnion cells in monolayers. In areas of cytopathogenic action resulting from invasion of cells by RNA, intact poliovirus, of the type from which the RNA had been prepared, is present. The infectivity of the RNA was completely inactivated by a 2 minute exposure to purified ribonuclease or to whole normal monkey serum shown to contain measurable concentrations of this enzyme. Whole virus infectivity was not influenced by RNAase or whole normal monkey serum. Normal and polio-immune globulin, desoxyribonuclease, lysozyme, proteolytic enzymes, and bovine albumin failed to inactivate the infectivity of RNA. The degree of infectivity of isolated RNA from poliovirus for cells in monolayer was greatly influenced by the ionic strength of the environment. The experimental evidence suggests that isolated poliovirus RNA is the carrier of the biological activity responsible for infection of cells and for transmission of genetic information which controls type specificity. Footnotes Submitted: 3 June 1958

Tsaltas, Theodore T.

doi: 10.1084/jem.108.4.507pmid: 13575681

Some biochemical aspects of the collapse of the rabbit ears produced by the intravenous injection of papain have been studied. A marked depletion of chondromucoprotein (M.C.S.) and a reduction of the S 35 content of cartilage matrix were found to coincide with the gross and histologic changes in the cartilage. At the same time there was a marked increase in the amount of S 35 in the serum and an increase of S 35 and glucuronic acid excreted in the urine. Alteration in the composition of the M.C.S. remaining in the cartilage of the papain-injected animals was detected. The findings indicate that the collapse of the rabbit ears is due to loss of chondromucoprotein from cartilage and reduction of chondroitin sulfate in the chondromucoprotein that remains. All these changes were reversed in recovery. Footnotes Submitted: 9 June 1958

Nelson, Robert A.

doi: 10.1084/jem.108.4.515pmid: 13575682

Evidence is presented that phenomena ascribed to the properdin system may be explained in terms of classical antibody (Ab) in combination with three of the components of complement (C'), i.e ., C' 1 , C' 4 , and C' 2 respectively. The results suggest that the complex of properdin (P) with zymosan (Z) represents a mixture of Z·Ab, Z·Ab·C' 1, 4 , Z·Ab-C' 1, 4, 2 , and Z·Ab·C' 1, 4, 2 in a decayed state. The purported preferential reactivity of Z with the third component of C' (C' 3 ) is not supported by the present experiments in which Z was reacted with C' in both guinea pig and human sera. Approximately the same number of reactive units of C' 1, 4 and of C' 3 were inactivated by Z, as well as by D. pneumoniae and a washed specific precipitate of bovine albumin-anti-albumin. The latter evidence that small amounts of antigen-antibody complex fix significant amounts of C' 3 stands in contrast with the classical concept that C' 3 is not fixed in ordinary complement fixation reaction. The observed reactivity of C' 3 is explained on the basis of the present use of essentially undiluted serum as a source of C' 3 and of 37° as the temperature for fixation. Ancillary data indicate that purified properdin contains Ab. In the presence of a chelating agent and at 0° properdin agglutinated Z granules. Measurements of nitrogen (N) uptake by Z suggest that 0.25 µg. N were contributed by solutions stated to contain 1 unit of properdin. The broad spectrum of reactivity or cross-reactivity of the Ab in normal serum is likely due to the wide distribution in nature of closely related polysaccharides. Further immunochemical studies are necessary to establish definitively the origin and mode of action in "natural resistance" of antibodies reactive with these polysaccharides. Footnotes Submitted: 21 May 1958

Henle, Gertrude; Deinhardt, Friedrich; Bergs, Victor V.; Henle, Werner

doi: 10.1084/jem.108.4.537pmid: 13575683

Inoculation of the MCN and Lung-To lines of human cells in continuous culture with Newcastle disease (NDV), mumps, or 6-6 viruses led to slight cytopathic effects (CPE) if the multiplicity of infection exceeded one. On second passage or with smaller initial inocula no CPE became apparent. The viruses multiplied, however, as determined by titrations in HeLa cultures or chick embryos. Indeed, persistently infected sublines of MCN and Lung-To were readily established without resort to special manipulations and some of these have been carried now for over 18 months on the same media and schedules as the uninfected parent strains. The viruses were found to be associated mainly with the cells and only 1, or at most 10 per cent of it was detectable in the media. The titers obtained were always low in relation to the available cell population. Reduction or even omission of the horse serum component in the media or ultraviolet irradiation of the cultures did not increase the yield of virus, and CPE became apparent only when similarly treated, uninfected cultures were, likewise, affected by the manipulations. The persistently infected cultures differed from their uninfected counterparts in that they exhibited ( a ) decreased cellular growth rates and ultimate yields; ( b ) increased aerobic glycolysis; and ( c ) a high degree of resistance to cytopathogenic viruses, influenza A (PR8), herpes simplex and, especially vesicular stomatitis (VSV) viruses. Prolonged treatment of persistently infected cultures by addition of specific antiviral immune sera to the media reduced significantly the amount of virus present and the degree of resistance to VSV. However, upon removal of the sera after as many as 187 days of treatment the viruses reappeared in all but one instance. The cured culture, on reinfection, became again persistently infected. No evidence was obtained to indicate genetic inhomogeneity of the cell populations. Of 50 cloned MCN lines none was destroyed by NDV and all became persistently infected. None were initially resistant to VSV but all after establishment of persistent NBV infection. All 39 cloned lines derived from MCN NDV cultures in the presence of anti-NDV serum, were free of virus and susceptible to VSV, and all acquired persistent infections and with it resistance to VSV following inoculation of NDV. NDV maintained in MCN cultures differed from the parent, chick embryo-adapted strain with respect to its plaque morphology. Whereas the former yielded only plaques on monolayers of chick embryo fibroblasts which were of pin-point size and hazy, those obtained with the latter were rarely of this type and mostly large and clear. This apparent selection of virus particles did not alter significantly their behavior with respect to cytopathogenicity for uninfected MCN cultures. Footnotes Submitted: 24 May 1958

Bergs, Victor V.; Henle, Gertrude; Deinhardt, Friedrich; Henle, Werner

doi: 10.1084/jem.108.4.561pmid: 13575684

Efforts were made to elucidate the nature of the resistance to vesicular stomatitis virus (VSV) observed in MCN cultures persistently infected with Newcastle disease, mumps, or 6-6 viruses (MCN NDV , MCN Mps and MCN 6-6 , respectively). Cells derived from persistently infected cultures adsorbed VSV to the same extent as their uninfected counterparts. Only a fraction of the adsorbed virus could be recovered from the cells indicating that it enters into an eclipse in all of the cell types. While propagation of VSV in MCN cells is largely inhibited at low pH levels, the resistance of persistently infected cultures could not be ascribed to their increased lactic acid formation. Resistance was not absolute in that a few cells in persistently infected cultures apparently supported VSV reproduction. Furthermore resistance of the cultures was found to be transitory in that the VSV infection gradually gained the upper hand after 2 to 4 weeks of incubation. Addition of ultraviolet-inactivated NDV to MCN cultures induced resistance to VSV as long as the equivalent of at least one ID 50 (for chick embryos) of inactivated virus was provided per cell. Establishment of resistance required some time and its duration depended upon whether or not the free inactivated NDV was removed or neutralized after given adsorption periods. The transitory nature of resistance in persistently infected cultures, or in MCN cells following adsorption of inactivated NDV, is most likely explained by the fact that the cells continue to divide and that the daughter cells are, at least in part, susceptible to VSV. The results are compatible with the conclusion that the resistance observed represents another example of interference between 2 viruses. Footnotes Submitted: 24 May 1958