The Journal of Experimental Medicine

Gallily, Ruth; Warwick, Anne; Bang, Frederik B.

doi: 10.1084/jem.125.4.537pmid: 4289738

Adult or weanling C 3 H mice were found to be genetically resistant to a strain of mouse hepatitis virus. Infant C 3 H mice, however, developed infection and died from mouse hepatitis virus when minimal infectious doses of virus were given to them. There was a delay in the time of death compared to that of the genetically susceptible strain, and the virus recovered from these mice had increased pathogenicity for C 3 H mice. The ontogeny of resistance to hepatitis in the C 3 H mice thus progresses from delayed susceptibility to complete resistance as the age of the host increases. It is reflected in increased resistance of macrophages derived in vitro from liver cultures of infant mice of different ages. This increase in resistance with age was reduced by maintaining the cultures for a longer period of time before inoculation, or by increasing the number of explants in a given culture. Resistant cells were uniformly furnished by mice age 16 days, or more. It is concluded that a process of maturation of resistance of the cells takes place after the mice are born, but that this does not continue under in vitro conditions, and that it may be modified by the environment of the cells. Footnotes Submitted: 15 August 1966

Davies, D. A. L.; Boyse, E. A.; Old, L. J.; Stockert, Elisabeth

doi: 10.1084/jem.125.4.549pmid: 6020006

Mouse H-2 histocompatibility antigen has been extracted, solubilized, and partly purified from the cells of an A strain spontaneous leukemia carrying TL (thymus-leukemia) antigens. H-2 and TL. 1, 2, 3 activities were measured by inhibition of the cytotoxic effect of the corresponding isoantibodies. TL activity was associated with the H-2 active fraction obtained by solubilization and fractionation by gel filtration. TL specificity was largely separated from H-2 antigen by subsequent chromatography on DEAE Sephadex as an adjacent component in a series of fractions. The soluble H-2 antigen prepared from the leukemia cells was tested for most of the specificities determined by H-2 a with no exceptional results. TL. 1, 2, 3 activities, measured as each component separately, were located in approximately the same position; there is no clear indication yet whether the three TL specificities are separable from one another. It appears that in addition to the close genetic linkage between the H-2 and TL loci, and their reciprocal interaction in producing H-2 and TL antigens, these antigens exhibit some similarity at the chemical level. Footnotes Submitted: 27 November 1966

Smith, Thomas J.; Wagner, Robert R.

doi: 10.1084/jem.125.4.559pmid: 4163879

Interferon is produced in cultures of rabbit leukocytes in response to infection with Newcastle disease virus or in the absence of known viral infection. The macrophage appears to be the responsible producing cell. Cultures prepared from sterile peritoneal exudates, which contained about 90% macrophages, are at least as efficient as cultures of rabbit kidney (RK) cells in their capacity to synthesize NDV-induced interferon. Interferon can be detected in the medium by 2 hr after viral infection and the titers usually reach a peak of 10,000 PDD 50 /ml by 4–6 hr. Exposure to actinomycin prior to or shortly after viral induction effectively blocks interferon synthesis by cells of both types. However, macrophages become refractory to actinomycin by 30–60 min compared with 607–120 min for RK cells, a finding which suggests earlier and more rapid transcription of interferon-specific messenger RNA in macrophages. Macrophages harvested from the peritioneal cavity of rabbits injected intravenously with NDV 48 hr previously also exhibit slightly reduced capacity to synthesize interferon, but this tolerant state is less marked than is tolerance to production of circulating interferon in intact rabbits. Interferon is also synthesized by rabbit macrophages not infected with virus but simply incubated at 37°C in medium with or without added bacterial endotoxin. Uninfected polymorphonuclear leukocytes, rabbit kidney and spleen cells produced no detectable interferon under similar conditions of cultivation. No interferon was released by intact macrophages incubated at 4°C or by ultrasonically disrupted macrophages incubated at 37°C. Although interferon titers were found to be higher when uninfected cultures were exposed to 10–100 µg/ml of E. coli lipopolysaccharide, unavailability of suitable pyrogen-free maintenance media precluded answering the question whether macrophages can continually synthesize and release interferon spontaneously. Interferon yields from uninfected macrophages were only 1% or less of the yields from NDV-infected macrophages, but the rates of synthesis were similar under both conditions. Studies with actinomycin and puromycin revealed that sequential transcriptive and translational events are required for de novo interferon synthesis by uninfected cells in a manner similar to virus-induced interferon synthesis. The physical properties and molecular weights of these rabbit interferons are discussed in the following report (12). Footnotes Submitted: 28 October 1966

Smith, Thomas J.; Wagner, Robert R.

doi: 10.1084/jem.125.4.579pmid: 6066967

Antiviral factors present in cultures of rabbit peritoneal macrophages or rabbit kidney (RK) cells infected with Newcastle disease virus (NDV) and those in cultures of uninfected macrophages all fulfilled the biological and physicochemical criteria for classification as interferons. Virus-induced macrophage and RK interferons were slightly more stable to heat or acid than "spontaneously produced" or endotoxin-induced macrophage interferon. Interferon activity in serum of NDV-infected rabbits was decidedly more labile than NDV-induced macrophage interferon. However, these differences in lability were too slight to serve as a useful basis for distinguishing one rabbit interferon from another. Rabbit interferons from various sources could be differentiated by filtration through Sephadex G-100 and their molecular weights estimated by comparison with elution profiles of a series of marker proteins of known molecular weight. Each of four different preparations of rabbit interferons was found to contain more than one molecular component. Elution peaks for three NDV-induced interferons were equivalent to the following molecular weights: RK ≃44,000–45,000 and > 134,000 (variable and < 1% when present); macrophage ≃37,000, 44,000–45,000, and > 134,000 (variable and <1% when present); and serum ≃50,000–52,000 and > 134,000 (∼10% and heat labile). NDV-induced serum interferon may also contain another molecular component of mol wt ≃45,000 represented by a trailing shoulder from the major 51,000 mol wt peak. Endotoxin-induced macrophage interferon proved to be polydisperse. Sephadex filtration of this interferon did not reveal clear and consistent elution patterns, partially owing to its low initial titer and lability. However, variable peaks of biological activity could be detected in Sephadex fractions equivalent to approximate molecular weight values of > 134,000, 72,000–78,000, 33,000–38,000, 28,000–30,000, and possibly a component of 42,000–45,000. A major component of mol wt ≃37,000 was present in all samples of endotoxin-induced macrophage interferon. The other constituents may be biologically active subunits or polymers. These data indicate that rabbit macrophages produce two primary kinds of interferon: ( a ) an RK-like component of mol wt ≃45,000 that is synthesized in greatest amount after viral induction, and ( b ) a different species of mol wt ≃ 37,000 that can also be synthesized in the absence of viral induction. The presence of major interferon constituents of mol wt ≃51,000 and > 134,000 in rabbit serum after viral induction suggests that macrophages are not the principal interferon-producing cells that respond to intravenous injection of NDV. Footnotes Submitted: 28 October 1966

Rothbard, Sidney; Watson, Robert F.

doi: 10.1084/jem.125.4.595pmid: 4289739

Rabbit serum containing antibody to human collagen, perfused through human infant kidneys obtained at autopsy, gives an immunofluorescent reaction with an antigen in the basement membranes of the glomeruli and the tubules. This reaction was shown to be specific by the absence of reaction with normal rabbit serum, antibody to carp collagen, or anti-human collagen serum absorbed with human collagen. Slight cross-reactions were found in the human kidneys with antibody to chicken or rat collagen. There was no evidence that a collagen-like protein in human serum or an antigen common to human erythrocytes and renal glomeruli enters into this immunofluorescent reaction. Perfusion of the kidneys with purified collagenase before the introduction of the antibody to human collagen altered the antigen so that antibody could not be demonstrated in the basement membranes by immunofluorescence. Testicular hyaluronidase used in the same way did not affect the immunofluorescent reaction. This method of perfusion provides another means for studying antigens in human organs by immunofluorescence. These observations in human kidneys extend our earlier findings in laboratory animals and indicate that an antigen, collagen, is also present in human renal glomerular and tubular basement membranes. Footnotes Submitted: 30 November 1966

Winterbauer, Richard H.; Gutman, Laura T.; Turck, Marvin; Wedgwood, Ralph J.; Petersdorf, Robert G.

doi: 10.1084/jem.125.4.607pmid: 5335676

1. After injection into the renal medulla of rats Escherichia coli 06 variants reverted rapidly in vivo in the absence of penicillin. These variants had previously been shown to be stable in vitro. 2. Variants failed to survive following intramedullary injection when animals were receiving penicillin. 3. Late reversion of variants also failed to occur in animals treated with penicillin for only 1 or 2 days. 4. Variants survived and reverted more readily when injected in the renal medulla, compared with liver and spleen. Classical bacteria injected into the kidney, liver, and spleen were recovered in approximately equal numbers. 5. The histologic response to nonreverting variants, medium not containing variants, and killed variants was similar and was characterized by a fibrotic reaction with moderate round cell infiltration. 6. In contrast, the histologic response to reverting variants and to classical E. coli was characterized by an intense, acute, polymorphonuclear leukocytosis typical of acute pyelonephritis. Footnotes Submitted: 11 December 1966

Morse, Stephen I.; Riester, Sallie K.

doi: 10.1084/jem.125.4.619pmid: 4289740

The 24 hr volume flow, cell concentration, and total cell output of thoracic duct fluid from mice with pertussis-induced hyperlymphocytosis were markedly reduced when compared with values obtained in normal animals. An increase in the number of circulating lymphocytes occurred in several of the pertussis-treated mice despite the presence of an indwelling thoracic duct cannula. The drainage from such animals also showed a reduced cell concentration and total cell output. It is suggested that lymphocyte recirculation may be minimal in pertussis-induced lymphocytosis, and the evidence obtained also suggests that lymphocytes may enter the blood stream by direct routes during the course of the reaction. Footnotes Submitted: 14 December 1966

Black, Paul H.; White, Beverly J.

doi: 10.1084/jem.125.4.629pmid: 4289741

Primary weanling hamster kidney cultures were transformed with the adeno 2-SV40, adeno 3-SV40, and adeno 12-SV40 hybrid viruses and with adenovirus type 12. The transformed cell lines which were established were characterized with respect to morphology, virus and antigen content, and chromosome aberrations. The adeno 2 and adeno 3-SV40 hybrid transformed cells had the morphology and T antigen content characteristic of SV40 transformations; cells transformed by the former hybrid had cytogenetic changes typical of SV40-transformed cells as well. The adeno 12-SV40 transformed cells were similar morphologically to adeno 12-transformed cells, contained both the SV40 and adeno T antigens and demonstrated the karyotypic instability of SV40-transformed cells, indicating that both viral genomes are operative in these cells. Although the results indicate that the SV40 genome in hybrids derived from the moderately or nononcogenic adenoviruses supplies the determinants for most of the characteristics investigated, and perhaps for oncogenesis, evidence was presented which suggests that a portion of a nononcogenic adenovirus genome may be integrated in adeno 2-SV40 transformed cells and directs the synthesis of adenovirus T antigens. Footnotes Submitted: 19 October 1966

Igel, H. J.; Black, P. H.

doi: 10.1084/jem.125.4.647pmid: 4289742

Tumors induced with hamster kidney cells transformed by the adeno 2-, adeno 3-, adeno 7-, and adeno 12-SV40 hybrid viruses, and by adenovirus type 12, were examined histologically. The tumors induced with adeno 2-, adeno 3-, and adeno 7-SV40-transformed cells were similar to tumors induced with SV40-transformed hamster kidney cells but contained cells intermediate in morphology between SV40 and adenovirus tumor cells and occasionally contained nests of adenovirus-like cells. Cells transformed by the adeno 12-SV40 hybrid and by adenovirus type 12 gave rise to morphologically similar tumors. The results suggest that both viral genomes are operative in hybrid-transformed cells but that one genome is apparently responsible for the predominant morphology of the tumor. Evidence that the morphology of a single transformed target cell is determined by the transforming genome was discussed. Footnotes Submitted: 19 October 1966

Churchill, Winthrop H.; Weintraub, Ronald M.; Borsos, Tibor; Rapp, Herbert J.

doi: 10.1084/jem.125.4.657pmid: 6020007

The titer of late-acting complement components in sera from male mice is 8–10 times higher than the titer of sera from female mice. Using assays developed to measure the serum content of two of the late-acting components, we have shown that this difference is due to the effect of androgen and estrogen on these two late-acting complement components. These two components have been tentatively identified as C'5 and C'6. Androgen and estrogen have greater effect on C'6 than on C'5. The possibility has not been excluded that still other of the late-acting complement components are affected by androgens and estrogens. The course of homograft rejection was unchanged in mice deficient in C'5 and C'6. Footnotes Submitted: 20 November 1966