The Journal of Experimental Medicine

Rous, P

doi: 10.1084/jem.150.4.729pmid: 229185

In this paper is reported the first avian tumor that has proved transplantable to other individuals. It is a spindle-celled sarcoma of the hen, which thus far has been propagated into its fourth tumor generation. This was accomplished by the use of fowls of pure blood from the small, intimately related stock in which the growth occurred. Market-bought fowls of similar variety have shown themselves insusceptible, as have fowls of mixed breed, pigeons and guinea-pigs. The percentage of successful transplantations has been small, but in the individuals developing a tumor its growth has been fairly rapid. Young chickens are more susceptible than adults. The reinoculation of negative fowls has never resulted in a growth. Throughout, the sarcoma has remained true to type. It is infiltrative and destructive. Metastasis has been observed once (to the heart). Experiments to determine whether the growth may be transmitted by cell-fragments have not yet been made. Repeated bacteriological examinations have yielded negative results. In its general behavior, so far as tested, this avian tumor closely resembles the typical mammalian neoplasms that are transplantable.

Yamazaki, K; Yamaguchi, M; Baranoski, L; Bard, J; Boyse, E A; Thomas, L

doi: 10.1084/jem.150.4.755pmid: 512584

Previous studies of mating preference signified that mice can sense one another's major histocompatibility complex (MHC) types, probably by olfaction. This conclusion has now been substantiated by the use of a Y-maze whose two arms were differentially scented with currents of air conducted through boxes occupied by B6 (H-2b) males and by B6-H-2k congenic males. Four B6 mice, two males and two females, were successfully trained, by water deprivation and reward, to enter the arm scented by B6 or B6-H-2k males. One of the males and one of the females were trained to select the B6-scented arm; the other male and female were trained to select the B6-H-2k-scented arm. Untrained mice showed no MHC discrimination in the maze. The performance of the trained mice in distinguishing between MHC congenic homozygous F2 segregants derived from a cross of B6-H-2k with B6 was as good as their performance in distinguishing the respective inbred strains, thus essentially eliminating alternative and significant additional explanations of MHC-associated sensory discrimination. The data further indicate that chemosensory discrimination of MHC types can be entirely dissociated from sex differences and from the circumstances of mating.

Weinberger, J Z; Germain, R N; Ju, S T; Greene, M I; Benacerraf, B; Dorf, M E

doi: 10.1084/jem.150.4.761pmid: 92517

The ability of NP-coupled syngeneic spleen cells to induce antigen-specific T-suppressor cells capable of binding to NP-BSA-coated Petri dishes and mediating transfer of specific suppressive activity to NP was demonstrated. Furthermore, in strains of mice bearing the Ig-1b allotype, including SJL, and in (non-Ig-1b x Ig-1b)F1 hybrids, the NP-specific suppressor cells also interferes with expression of immunity after priming with NIP-BGG. Anti-NPb anti-idiotype antiserum plus complement treatment effectively abrogated the ability to transfer suppression. Formal genetic mapping of the fine specificity of cross-reactivity with Ig-1 allotypic congenic mice implies that expression of this trait is linked to the Ig-1b heavy chain linkage group. The sensitivity of NP-suppressor cells of appropriate strains to anti-idiotype treatment was also consistent with the formal mapping data. These experiments suggest that there are shared V-region structures on antibody and T cells that are crucial in the suppression pathway for the same antigen.

Rothenberg, E; Boyse, E A

doi: 10.1084/jem.150.4.777pmid: 315985

Thymus-leukemia (TL) antigens are expressed in murine lymphocytes under strict developmental regulation. To elucidate the molecular basis of TL expression, we have identified the molecular species that react with TL antiserum. At least three species can be resolved by metabolic radiolabeling of thymocytes and ASL1 leukemia cells, lysis, immune precipitation, and sodium dodecyl sulfate-polyacrylamide. After a brief incubation with 35Smethionine, the only radioactive molecule recognized by TL antiserum is a homogeneous species with an apparent Mr of 45,000 daltons. This molecule, 45K TL, includes high-mannose-type carbohydrate attached to a 45,000 dalton glycosidase-resistant backbone. In this form, 45K, it is never exposed on the cell surface. If pulse-labeled cells are further incubated with nonradioactive methionine before lysis, however, radioactivity disappears from the 45K TL species and appears in the slower migrating species 46K and 48K TL. Thus, 46K and 48K appear to represent products generated from the 45K TL precursor by posttranslational modification. These TL forms are displayed on the cell surface; they lack high-mannose carbohydrate but evidently include acidic complex-type carbohydrate. Normal thymocytes from Qa:Tla-negative mice lack not only the surface forms of TL but also the intracellular 45K TL form. Peripheral lymphoid cells of Qa:Tla-positive mice synthesize none of these TL species. But the TL antiserum, which contains Qa antibody, recognizes a distinct gene product in spleen and thymus of Qa-Tla-positive mice. In its pulse-labeled form, this molecule, which may represent Qa-1, has an apparent Mr of 44,000 daltons, and consists of a glycosidase-resistant polypeptide core of only 35,000 daltons linked to more high mannose carbohydrate than 45K TL.

Kubagawa, H; Vogler, L B; Capra, J D; Conrad, M E; Lawton, A R; Cooper, M D

doi: 10.1084/jem.150.4.792pmid: 92518

IgA myeloma proteins of kappa- and lambda-types were isolated from two patients. These were used to produce and purify anti-idiotype antibodies of both broad (myeloma-related) and narrow (individual myeloma) specificities. The anti-idiotype antibodies were conjugated with fluorochromes and used as immunofluorescent probes to trace in the patients clonal expansion at different levels of B-cell differentiation. Our results (a) confirm that B lymphocyte precursors in IgA plasma-cell myelomas are involved in the malignant process, (b) show that B lymphocytes of the malignant clone include those expressing each of the major heavy-chain isotypes, mu, delta, gamma, and alpha, and (c) provide strong circumstantial evidence that pre-B-cell members of the malignant clone are also increased in frequency. T cells expressing idiotypic determinants were not detected. These findings argue that the initial oncogenic event may occur in a B-stem cell and is not influenced through stimulation by antigen. An interesting association was the increased frequency of related clones of B lymphocytes as detected by their reactivity with anti-idiotype antibodies of broad specificity. Neither plasma cell nor pre-B-cell members of these related clones were increased in frequency. Anti-idiotype antibodies or helper T cells reactive with myeloma-related idiotypes could be responsible for this phenomenon. We discuss other implications of these findings and speculate that all of the various phenotypes of B-lineage malignancies may result from oncogenic processes affecting stem cell targets.

Schrater, A F; Goidl, E A; Thorbecke, G J; Siskind, G W

doi: 10.1084/jem.150.4.808pmid: 315986

Although athymic mice make an excellent immune response to the thymus-independent antigen trinitrophenyl-lys-Ficoll (TNP-F), nude mice of AKR/J and BALB/c strains lack the anti-idiotypic response that occurs in euthymic mice of both of these strains within the first 1--2 wk after injection of more TNP-F. Anti-idiotypic antibody-blocked (hapten-augmentable) anti-TNP splenic plaque-forming cells (PFC) do not occur at any time and serum anti-idiotypic antibody is absent in both congenitally athymic mice, and thymectomized, irradiated, bone marrow-reconstituted mice. Nevertheless, nu/nu mice do have PFC which can be inhibited by exposure to anti-idiotypic antibody produced in +/+ mice. As a consequence of the failure to produce anti-idiotypic antibodies, the anti-TNP PFC response is athymic as compared to euthymic mice is of greater magnitude, declines less precipitously, and shows an increase rather than a decrease in affinity between days 4 and 7 after antigen injection. It is concluded that the anti-idiotypic antibody response is thymus dependent and that athymic mice lack a helper cell required for the induction of anti-idiotypic antibodies.

Yamamoto, H; Nonaka, M; Katz, D H

doi: 10.1084/jem.150.4.818pmid: 92519

Delayed-type hypersensitivity (DTH) responses specific for the phosphorylcholine (PC) hapten were induced in BALB/c mice by immunization with syngeneic peritoneal exudate cells (PEC) coupled with diazotized phenyl-phosphoryl-choline. PC-specific DTH responses were elicited in such immunized mice after footpad challenge with PC-derivatized syngeneic spleen cells. Moreover, PC-immune lymph node cells could passively transfer PC-specific DTH responses to naive BALB/c mice and it was possible to demonstrate that the cells responsible for such passively transferred responses were T lymphocytes. Because the T-15 idiotypic determinant displayed on the TEPC-15 PC-binding myeloma protein is known to be a dominant idiotype associated with anti-PC antibody responses in BALB/c mice, an analysis was made of the effects of anti-T-15 idiotypic antibodies on the induction and expression of murine PC-specific DTH responses. Repeated injections of anti-T-15 idiotypic antiserum, raised in A/J mice by immunization with TEPC-15 myeloma protein, into recipient BALB/c mice both immediately before and after sensitization with PC-PEC virtually abolished the development of PC-specific DTH responses. Although administration of anti-T-15 antiserum effectively inhibited the induction phase of PC-specific DTH responses, these anti-idiotypic antibodies had no suppressive activity at the effector phase of these responses. The inhibition observed with anti-T-15 antibodies was highly specific for the PC hapten, and for PC-specific DTH responses of BALB/c but not A/J mice. Studies were conducted to address the possibility that anti-Id treatment induced suppressor T lymphocytes capable of specifically inhibiting the activity of PC-specific T cells participating in DTH responses. The results demonstrate that idiotype-specific suppressor T cells are, indeed, induced by treatment with anti-Id; moreover, such suppressor T cells, once induced, are highly effective in abrogating both the induction and the effector phases of PC-specific T cell-mediated DTH responses in BALB/c mice.

Solinger, A M; Ultee, M E; Margoliash, E; Schwartz, R H

doi: 10.1084/jem.150.4.830pmid: 92520

The T-lymphocyte proliferative response to pigeon cytochrome c was studied in the mouse. H-2a and H-2k strains were responders to this antigen whereas H-2b, H-2d, H-2f, H-2ja, H-2p, H-2q, H-2r, H-2s, and H-2u strains were low or nonresponders. Genetic mapping demonstrated that two major histocompatibility complex (MHC)-linked Ir genes control the response, one in I-A, the other in I-E/I-C. The major antigenic determinant recognized in this response was localized by cross-stimulations with species variants and cyanogen bromide cleavage fragments of cytochrome c. It was found to be a topographic surface determinant composed of an isoleucine for valine substitution at residue 3, a glutamine for lysine substitution at residue 100 and a lysine for glutamic acid substitution at residue 104. Tobacco hornworm moth cytochrome c, which contains a glutamine at residue 100 but a terminal lysine at residue 103 (one amino acid closer to the glutamine), stimulated pigeon cytochrome c immune T cells better than the immunogen. This result demonstrates for the first time a functional T-cell heteroclitic proliferative response in a system under Ir gene control. Immunization with the cyanogen bromide cleavage fragments revealed that only pigeon cytochrome c fragment 81-104 was immunogenic. This fragment primed for a T-cell proliferative response whose specificity was nearly identical to that of the T-cell response primed for by the whole molecule, suggesting that the glutamine at 100 and the lysine at 104 form the immunodominant portion of the antigenic site. Furthermore, mixing experiments using the two cross-reacting antigens, hippopotamus cytochrome c and Pekin duck or chicken cytochrome c fragment (81-104), each of which contains only one of the two immunodominant substitutions, demonstrated that the T lymphocytes responding to the major antigenic determinant comprise a single family of clones that recognize both amino acids as part of the same determinant. Thus, two complementing MHC-linked Ir genes can control the immune response to a single antigenic determinant.

Watson, J; Gillis, S; Marbrook, J; Mochizuki, D; Smith, K A

doi: 10.1084/jem.150.4.849pmid: 315987

Murine spleen cells activated by concanavalin A (Con A) in culture produce a class of lymphokine molecules which possess biological activity in a number of lymphocyte response assays. Lymphokines with a mol wt of 30,000, as estimated from gel filtration studies, can be resolved into two components which differ by charge, with isoelectric point (pI) values of 4.3 and 4.9, respectively. Both components stimulate (a) the growth of established T-cell lines in culture, (b) the proliferation of thymocytes in the presence of Con A under culture conditions where Con A alone is nonmitogenic, (c) the induction of antibody responses to heterologous erythrocyte antigens in athymic (nude) spleen cultures, (d) the generation of cytotoxic T lymphocytes (CTL) in thymocyte cultures, and (e) the generation of CTL in nude spleen cultures. In each of these culture systems we suggest that the assays are detecting a single class of lymphokine which acts directly on activated T cells. Nonactivated T cells must be stimulated by either antigen or mitogen before becoming responsive to lymphokine, but do not require antigen or mitogen for continued growth with lymphokine. The two molecular species, separable by isoelectric focusing are referred to as the T-cell growth factor (TCGF). A lymphokine, similar in size (30,000 daltons) to TCGF but heterogeneous in charge (pI 3.0--4.0), stimulates immune responses to erythrocyte antigens in T-cell-depleted spleen cultures but has no stimulatory activity in the other lymphocyte assay systems described. The data have been interpreted as showing the two molecular forms of murine TCGF (pI 4.3 and 4.9) are responsible for many of the lymphokine activities described elsewhere as thymocyte mitogenic factor, nonspecific T-cell-replacing factor and killer helper factor or costimulator. The other lymphokine, separable from TCGF by charge, appears to have true T-cell-replacing activity.

Beachey, E H; Stollerman, G H; Johnson, R H; Ofek, I; Bisno, A L

doi: 10.1084/jem.150.4.862pmid: 390084

We tested the ability of pepsin-extracted, highly purified M protein to induce type-specific immunity in experimental animals and humans. M protein was prepared from limited peptic digests of whole group A type 24 streptococci and was purified to chemical homogeneity as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, quantitative amino acid analysis, and Edman degradation. For vaccination, the lyophilized M24 protein preparation (pep M24) was precipitated in aluminum hydroxide. When injected into laboratory animals, alum-precipitated pep M24 produced type-specific protective antibodies and was free of non-type-specific immunoreactivity. In man, skin tests with 1-microgram doses of pep M24 were negative in all 37 adults tested. 12 adult human volunteers received two-four subcutaneous injections of 100-200 micrograms of alum-precipitated pep M24 at intervals of at least 2 wk. The immune response to pep M24 was measured by a variety of assays designed to detect (a) type-specific humoral antibodies (opsonophagocytic, long chain, and mouse protection tests); (b) total humoral antibodies (complement fixation and enzyme-linked immunosorbent assay); (c) cellular immunity (skin tests); and (d) heart cross-reactive antibodies (immunofluorescence). Type-specific opsonic antibodies developed in 10 of the 12 vaccinees, and positive delayed-type skin tests developed in 11. Immune sera from two of the vaccinees were effective in mouse-protection tests against challenge with M24 but not M6 streptococci. None of the volunteers developed heart-reactive antibodies or antibodies to non-type-specific M protein antigens. Alum-precipitated pep M24 was well-tolerated in man, and no serious local or systemic reactions were observed. Thus, pep M24 induces type-specific, protective antibodies in doses that are well-tolerated in man.