The Journal of Experimental Medicine

Kasai, M; Takashi, T; Takahashi, T; Tokunaga, T

doi: 10.1084/jem.159.4.971pmid: 6608576

A mouse monoclonal antibody (IgM) was obtained by cell hybridization between X63-Ag8.653 myeloma cells and spleen cells from a BALB mouse that was immunized with GRSL leukemia cells of the GR strain. This antibody identified a unique fetal antigen, which is expressed exclusively on embryonic thymocytes of all strains tested. Therefore, the antigen defined was named fetal thymus antigen-1, FT-1. The proportion of FT-1+ fetal thymocytes detected by immunofluorescence assay sharply decreases as gestation time increases, and finally they disappear from the thymus. On the other hand, Thy-1+ cells increase in inverse proportion. The immunofluorescence studies and absorption tests showed that FT-1 antigen is not detectable on brain, liver, kidney, or lymphoid tissue cells of adult mice. However, it is expressed on some leukemia cells of various mouse strains, which demonstrated that this is the first example of an oncofetal antigen of a mouse leukemia. The molecular weight of FT-1 antigen on leukemia cells was estimated to be 130,000 by means of biosynthetic labeling with 3Hgalactose and 35Smethionine. The two-dimensional gel electrophoresis pattern of FT-1 antigen shows a family of glycoproteins with extensive charge heterogeneity. It was also shown that the FT-1 antigen molecule carries the receptor for DBA lectin.

Takasaki, Y; Fishwild, D; Tan, E M

doi: 10.1084/jem.159.4.981pmid: 6142919

A nuclear antigen associated with cell proliferation (proliferating cell nuclear antigen PCNA) and blast transformation is recognized by autoantibodies in the sera of some patients with systemic lupus erythematosus. Using this autoantibody as a reagent, PCNA was purified 120-fold by ammonium sulfate fractionation, DEAE chromatography, and Sephadex G200 gel filtration. The antigenicity of PCNA was sensitive to trypsin but resistant to ribonuclease and deoxyribonuclease, suggesting that the antigenic determinant resided in protein and not nucleic acids. PCNA was inactivated at 56 degrees C for 30 min. Isoelectrophoretic focusing showed that the pI was 4.8. Analysis of immunoprecipitates on polyacrylamide gels showed the presence of IgG heavy and light chains and a single polypeptide band of 33,000 mol wt. This polypeptide band was the reactive antigen in immunoblotting (Western transfer) assays.

Phillips, J H; Le, A M; Lanier, L L

doi: 10.1084/jem.159.4.993pmid: 6231353

Lymphoid cells with natural killer (NK)-like function, morphology, and antigenic phenotype have been identified in a mixed lymphocyte culture generated by co-culture of human peripheral blood mononuclear cells with an allogeneic human B lymphoblastoid cell line CCRF-SB. The majority of these mixed lymphocyte (MLR)-response activated NK cells express the Leu-11 surface antigen, but do not express certain T cell-associated antigens (Leu-1, Leu-3, and Leu-4) or the mature monocyte specific antigen, Leu-M3. Unlike most freshly isolated Leu-11+ human NK cells, the MLR-activated Leu-11+ cells expressed class II major histocompatibility antigens, DR and DC. Concomitant with expression of class II gene products, the Leu-11+,DR+ NK cells demonstrate enhanced cytotoxicity against the NK-sensitive tumor cell line K562. The presence of mitotic cells in the Leu-11+,DR+ population and the acquisition of increased levels of transferrin receptor on the cell surface were further indicators of activation of these cells. The direct precursors of the MLR-activated Leu-11+,DR+ cell were Leu-11+ cells that lacked expression of another NK-associated antigen Leu-7, i.e., Leu-7-,11+. These studies provided a definitive identification of the "NK-like" cell in MLR cultures and thus allow quantitation and isolation of these cells for further study.

Neilson, E G; McCafferty, E; Phillips, S M; Clayman, M D; Kelly, C J

doi: 10.1084/jem.159.4.1009pmid: 6231352

Antiidiotypic immunity can successfully inhibit the development of antitubular basement membrane (alpha TBM) disease that produces interstitial nephritis. Rats normally immunized to produce disease, however, do not develop this regulatory and protective antiidiotypic effect. The failure to see such a regulatory response is functionally related to the influence of a nonspecific, RT7.1+, OX8-suppressor T cell that appears shortly after immunization. While this suppressor cell system can partially reduce the intensity of disease, it also limits the host's ability to specifically regulate the alpha TBM immune response and, hypothetically, leaves the disease process in an operationally active mode.

Goldman, D W; Goetzl, E J

doi: 10.1084/jem.159.4.1027pmid: 6323613

Human polymorphonuclear (PMN) leukocytes bound 3Hleukotriene B4 (3H-LTB4) specifically, as assessed by the displacement of 88% or more of the bound radioactivity by a 15,000-fold higher concentration of nonradioactive LTB4 or by micromolar concentrations of structural isomers of LTB4. The specific binding of 3HLTB4 by PMN leukocytes was characterized by rapid association and dissociation, and was saturable at 800 nM LTB4. The results of computer analyses of the concentration dependence of binding of 3HLTB4 were consistent with the expression of two classes of receptors having respective mean affinities of 3.9 X 10(-10) M and 6.1 X 10(-8) M and mean densities of 4.4 X 10(3) and 2.7 X 10(5) per PMN leukocyte. Structural isomers of LTB4 inhibited the binding of 3HLTB4 to PMN leukocytes at concentrations similar to those required to elicit chemotaxis, while chemotactic peptides did not inhibit binding. PMN leukocytes that were deactivated by prior exposure to LTB4 lost high affinity binding sites selectively and concurrently with a reduction in the chemotactic response to LTB4. Chemotactic deactivation altered, but did not eliminate, the low affinity receptors for LTB4 and reduced only minimally the lysosomal degranulation elicited by LTB4. The high affinity receptors for LTB4 on normal human PMN leukocytes appear to transduce the chemotaxis evoked by LTB4 without substantially modifying lysosomal degranulation.

Tsao, B P; Fair, D S; Curtiss, L K; Edgington, T S

doi: 10.1084/jem.159.4.1042pmid: 6368733

In the present study we demonstrate that human monocytes can be induced by the model stimulus, lipopolysaccharide (LPS), to produce and assemble on their surface functional Factor VII/VIIa. This protease was not induced in relatively purified monocytes alone following exposure to LPS; but was induced in the presence of Leu-3a positive helper/inducer T cells. The Factor VII/VIIa protease activity represented 35-40% of the potential initiating activity for the extrinsic coagulation pathway and was demonstrated using functional coagulation assays, as well as in amidolytic assays for the activation of Factor X. This activity of cell-bound Factor VII/VIIa appeared to involve a tight adduct of calcium. The identity of the Factor X-activating protease as Factor VII/VIIa was confirmed by the capacity of antibody specific for Factor VII/VIIa to neutralize the cell-bound protease. Further propagation of the extrinsic pathway following generation of Factor Xa required addition of exogenous Factor Va. These results expand the repertoire of proteases that have been identified with appropriately triggered cells of the monocyte/macrophage series, and suggest that initiation and propagation of the extrinsic coagulation protease network on induced monocytes involves not only expression of the initiating cofactor molecule, tissue factor, but also production of Factor VII and its organization into the molecular assembly. Thus, in the absence of exogenous Factor VII/VIIa a directly proteolytic effector cell can be generated. Further molecular assembly of the extrinsic pathway on the monocyte surface sequentially expands the proteolytic capacity of this response. The synthesis and assembly of the extrinsic activation complex by the monocyte and its derived progeny, the macrophage, provides a mechanism by which coagulation is initiated under T cell instruction at sites of immunologic responses.

Hind, C R; Collins, P M; Renn, D; Cook, R B; Caspi, D; Baltz, M L; Pepys, M B

doi: 10.1084/jem.159.4.1058pmid: 6707579

Serum amyloid P component (SAP) is a normal plasma protein that is of interest because of its presence in amyloid deposits, its presence in normal human glomerular basement membrane, and its stable evolutionary conservation. It has calcium-dependent ligand-binding specificity for amyloid fibrils, fibronectin (Fn), C4-binding protein (C4bp), and agarose. Although the binding to agarose, a linear galactan hydrocolloid derived from some marine algae, is unlikely per se to be related to the physiological function of SAP, it does provide a model system in which to explore the precise ligand requirements of SAP. We report here that the amount of SAP from human, mouse, and plaice (Pleuronectes platessa L.) serum able to bind to agarose from different sources reflect precisely their pyruvate content. Methylation with diazomethane of the carboxyl groups in the pyruvate moiety of agarose completely abolishes SAP binding to agarose. The pyruvate in agarose exists as the 4,6-pyruvate acetal of beta-D-galactopyranose. We have therefore synthesized this galactoside, using a novel procedure, established its structure by analysis of its nuclear magnetic resonance spectra, and shown that it completely inhibits all known calcium-dependent binding reactions of SAP. The R isomer of the cyclic acetal, methyl 4,6-O-(1-carboxyethylidene)-beta-D-galactopyranoside (MO beta DG) was effective at millimolar concentration and was more potent than its noncyclic analogue, while pyruvate, D-galactose, and methyl beta-D-galactopyranoside were without effect. The autologous protein ligands of SAP presumably, therefore express a structural determinant(s) that stereochemically resembles MO beta DG. Availability of this specific, well-characterized, low molecular weight ligand for SAP should facilitate further investigation of the function of SAP and its role in physiological and pathophysiological processes.

Drizlikh, G; Schmidt-Sole, J; Yankelevich, B

doi: 10.1084/jem.159.4.1070pmid: 6142918

Irradiated (H-2b X H-2k)F1 and (H-2b X H-2d)F1 recipients strongly resist the growth of H-2b parental bone marrow cells and do not resist marrow grafts from non-H-2b parents such as C3H and BALB/c. This phenomenon of hybrid resistance has been shown to be under genetic control of the H-2D-linked loci and was interpreted by Cudkowicz (9) as due to the existence of H-2D-linked recessive hemopoietic histocompatibility genes. To check whether the H-2D-linked loci are solely responsible for the fate of bone marrow allografts, we measured the strength of resistance of irradiated (B6 X C3H)F1 and (B6 X BALB/c)F1 recipients toward bone marrow grafts from a set of H-2 recombinant and F1 hybrid donors carrying either the H-2b, H-2d, and H-2k alleles. We found that growth of all H-2b grafts was resisted, although to different degrees. Resistance was minimal when donors shared with the input strain of a corresponding F1 hybrid the H-2K and H-2I regions, or when both F1 donors and F1 recipients formed identical unique hybrid Ia molecules. In addition, H-2b grafts were resisted by congenic, H-2D-identical, H-2K-and H-2I-incompatible recipients. The fate of grafts from H-2Dd donors seemed to depend on the incompatibility of the combinatorial determinant Ia.22. If both donor and recipient expressed such a determinant (either in the cis or in the transposition), or if neither could form such a determinant, grafts were not resisted. The H-2Dk allele is not the main or only factor that confers on the C3H parental bone marrow cells the ability to grow unresisted in (B6 X C3H)F1 recipients. Grafts from congenic C3H.OH donors, carrying the same H-2Dk alleles and differing in the left part of the H-2 complex, were resisted by the F1 recipients. We conclude that both class I (K and D) and class II (I-A and I-E) major histocompatibility complex genes, rather than hypothetical hemopoietic histocompatibility genes control hemopoietic resistance. To reconcile codominant inheritance of classic H-2 antigens with the apparent recessive inheritance of hybrid resistance, we assume that there exist parental determinants that are not formed in some F1 hybrids due to preferential association of either Ia alpha chains with allogeneic beta chains or of class I antigens with allogeneic or hybrid class II restriction elements.

Fischetti, V A; Jones, K F; Manjula, B N; Scott, J R

doi: 10.1084/jem.159.4.1083pmid: 6368734

Type 6 streptococcal M protein produced by E. coli bearing plasmid pJRS42.13 (ColiM6) accumulates in the periplasmic space of this new host. No immunoreactive M protein was found either on the surface of the organism or in the culture medium. The ColiM6 protein was purified from the periplasm and the final preparation consisted of three protein bands of apparent molecular weight 55,000, 57,000, and 59,000. These three bands were identical in migration in SDS PAGE to that of the M protein present in freshly prepared crude periplasm. The amino acid composition of the ColiM6 protein was nearly identical to that of M protein isolated from streptococci with phage lysin (LysM6). Furthermore, except for the amino terminal residue of the LysM6 molecule, the amino terminal sequence of the ColiM6 molecule was identical to those of both LysM6 and M protein released from the streptococcus by limited peptic digestion (PepM6). These results reveal that the molecule produced in the E. coli and transported into the periplasm may be the complete M protein as it exists on the streptococcus. The results also indicate that the systems that process M protein for transport through the cytoplasmic membrane are similar in the streptococcus and E. coli. The purified ColiM6 protein was able to remove opsonic antibodies from both human and rabbit serum, as well as to stimulate the production of opsonic antibodies in rabbits, indicating that the immunodeterminants on this molecule are the same as those found on streptococcal-derived M molecules.

Taniguchi, M; Tokuhisa, T; Itoh, T; Kanno, M

doi: 10.1084/jem.159.4.1096pmid: 6200561

The functional roles of the two polypeptide chains that compose the T cell suppressor factor (TsF) that mediates the antigen-specific and genetically restricted suppressor function were studied by using the heavy or light chains isolated from the conventional TsF or the 11S and 13S mRNA translation products of TsF. Either the heavy or the light chain of mRNA translation products reconstitutes the active TsF that suppresses the antibody response in an antigen-specific and genetically restricted manner when it is combined with the isolated heavy or light chain from the conventional TsF. As a consequence, the antigen-binding heavy chain mediates the antigen specificity of TsF. On the other hand, the I-J-positive light chain works as an element to determine the genetic restriction specificity. Thus, the identity of the histocompatibility between the I-J haplotypes on the light chain and the responding cell is essential for the functional expression of TsF. No genetic preference, however, was observed, in the association of the heavy and light chains of TsF.