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A cDNA sequence coding for mouse mast-cell growth-factor (MCGF) has been cloned from a cDNA library prepared from mRNA derived from a concanavalin A-activated mouse T-cell clone. Cloning was achieved using the pcD vector that permits expression of cDNA inserts in mammalian cells. The DNA sequence codes for a polypeptide of 166 amino acid residues including a putative signal peptide. The supernatant fluid obtained after transfection of COS-7 monkey cells with the pcD-MCGF plasmid had mouse MCGF activity. The MCGF amino acid sequence between amino acids 33 and 41, deduced from the nucleotide sequence of its cDNA, is identical to the NH2-terminal sequence of the mouse lymphokine, interleukin 3, suggesting that MCGF is related to interleukin 3.

We have prepared a cDNA probe that detects genes that are rapidly and abundantly expressed after exposure of inducer T-lymphocyte clones to antigen or mitogen. All inducer cells tested express a characteristic set of new mRNA, and these mRNAs are not expressed after activation of other lymphocytes. This initial burst of mRNA synthesis is paralleled by synthesis and secretion of a family of polypeptides that mediate inducer cell activity, including T- and B-cell growth factors, interferon, and molecules that bind to antigen. Expression of this initial genetic program precedes mitosis and is replaced within 74 hr by a different genetic program, which may control further cell division. The action of these sequential sets of genetic programs defines two stages of the cell's differentiation and accounts for altered expression of the cell's immunological functions.

When B mice are supplied with Ly1 or Ly23 cells they acquire, over the next 6 mo, only the immune functions associated with each of these T-cell subclasses, respectively. The T-cell population of these "B-Ly1" and "B-Ly23" mice mice also remains restricted to the Ly1 and Ly23 subclass phenotypes. Thus the Ly1 and Ly23 populations are derived from two separate lines of differentiation and are not sequential stages of a single differentiative pathway.

Relatively large numbers of nonimmune spleen cells do not protect against the local growth of two lymphomas. However, this heterogeneous population of splenic lymphocytes contains a subset of cells that efficiently protects against in vivo tumor growth. This cell population (cell-surface phenotype Thyl.2(-)Ig(-)Ly5.1(+)) represents less than 5 percent of the spleen cell population and is responsible for in vitro NK-mediated lysis. Although these studies clearly and directly demonstrate that Ly5(+) NK cells selected from a heterogeneous lymphoid population from nonimmune mice can protect syngeneic mice against local in vivo growth of two different types of tumor cells (in contrast to other lymphocyte sets within the spleen), they do not directly bear upon the role of NK cells in immunosurveillance. They do indicate that highly enriched Ig(-)Thyl(-)Ly5(+) cells, which account for virtually all in vitro NK activity, can retard tumor growth in vivo. It is difficult to ascribe all anti-tumor surveillance activity to NK cells, because they probably do not recirculate freely throughout the various organ systems of the body. Perhaps NK ceils may play a role in prevention of neoplastic growth within discrete anatomic compartments where there is rapid differentiation of stem cells to mature progeny (e.g., bone marrow, spleen, and portions of the gastrointestinal tract)and may normally act to regulate the growth and differentiation of non-neoplastic stem cells. Long-term observation of chimeric mice repopulated with bone marrow from congenic or mutant donors expressing very low or very high NK activity may help to answer these questions.

We describe a sequence of reciprocal interactions between cloned inducer T cells and antigen-presenting cells (APC) that results in selective depletion of the antigen-reactive inducer cells. We show that corecognition of antigen and I-A by hapten-reactive inducer T cell clones results in (a) release of macrophage-activating factor (MAF) and other lymphokines, (b) expression of lytic activity by a subset of MAF-sensitized APC after triggering, and (3) lysis (mediated by the activated and triggered macrophage) of the inducer T cell clone and other cells in the vicinity. We suggest that this sequence of steps may limit the extent of macrophage-mediated tissue destruction by depleting the specific inducer T cell clones that initiate the response.

We have analyzed a panel of T cell clones that corecognize defined epitopes of the insulin molecule in association with Ia for their patterns of recognition of alloantigens. A striking correlation is observed between recognition of the I-Ab gene product and cow insulin alpha loop and recognition of I-Eu of the PL/J haplotype. These results are consistent with the notion that reactions to foreign major histocompatibility complex (MHC) products reflect molecular mimicry by foreign class II antigens of 'physiologic' complexes formed by autologous class II MHC molecules and antigen.

We previously described a cloned cell line that combines information for a unique display of cell surface antigens and specialized function similar to activated natural killer (NK) cells. In addition to conventional cellular targets such as the YAC-1 and MBL-2 lymphomas, this cloned line also lysed lipopolysaccharide-activated B lymphocytes. To determine whether some NK cells can inhibit B cell function, we tested the ability of NK-like clones to suppress Ig secretion in vitro and in vivo. These cloned cells suppressed Ig secretion when they constituted as few as 0.2% of the total cell population and inhibition did not require identity at the H-2 locus. We suggest that some NK cells might recognize non-major histocompatibility complex gene products on activated B lymphocytes and lyse these cells, and this might represent a fundamental cell-cell interaction that regulates antibody secretion by activated B cells.

We describe clones of hapten-specific inducer T cells from (BALB/c X A/J)F1 mice that respond to the p-azobenzenearsonate hapten conjugated to carrier proteins or directly conjugated to antigen-presenting cells. Some of the clones are also activated by haptens structurally related to arsonate. All activating analogues are recognized by each clone in association with the same major histocompatibility complex (MHC) protein as is arsonate. Weakly activating and nonactivating analogues are immunogenic in D2.GD amd (BALB/c X A/J)F1 mice, since they can effectively activate primed lymph node cells or long-term hapten-reactive cell lines. Hence the specificities of these clones may reflect their intrinsic recognition of arsonate and its analogues, rather than more efficient presentation of certain analogues than of others by antigen-presenting cells, or differential recognition of associated MHC epitopes by the clones. We compare the activation specificities of the clones with the binding specificities of monoclonal antibodies to arsonate, and discuss structural features of the analogues that may be important for activation and binding. Our results suggest that a site (or subsite) on arsonate-reactive T cell clones may interact directly with hapten, and may be experimentally separable from the site (or subsite) for MHC determinants.

Feedback regulation of the primary humoral immune response to sheep erythrocytes (SRBC) was studied in vitro. Whole spleen cells or spleen cell subpopulations were incubated with antigen for 4 d under Mishell-Dutton conditions (education) and the surviving cells tested for regulatory activity in fresh anti-SRBC spleen cell cultures assayed by measuring plaque-forming cells on day 4. The data indicate that (a) whole spleen cells educated with SRBC exert potent antigen-specific suppression in the assay culture, (b) surface Ig- (sIg-) cells (T cells) prepared by either nylon-wool separation or fractionation on rabbit anti-mouse-Ig-coated polystyrene Petri dishes failed to generate suppressive activity when educated alone, in 2-mercaptoethanol, or in the presence of additional macrophages, (c) surface Ig (sIg+) (B) cells educated alone also failed to generate suppressor cells, and (d) mixing sIg- (T) and sIg+, Lyt 123- (B) cells reconstituted the ability to induce suppressor cells under these conditions. The antigen-primed cell actually required to transfer suppression was also characterized by separating cells using anti-Ig coated dishes, by fluorescence-activated cell sorting and by anti-Lyt treatment. All these methods clearly identified sIg+ (B) and not sIg+ (T) cells as the important educated cells. It is concluded that under our conditions, T cell-dependent B cells triggered by antigen during primary in vitro cultures cause potent specific feedback suppression of humoral responses. Possible mechanisms for this suppression, including antigen blockade or anti-idiotypic responses, are discussed.

We have shown that (a) purified T-helper cells induce cells of another T-cell set-, expressing the Ly123+Qa1+ surface phenotype, to exert potent suppressive activity, (b) this T-T interaction plays an important role in regulating in vivo immune responses, and (c) this interaction represents an important barrier to protocols intended to augment the immune status of individuals by adoptive (or active) immunotherapy. Our results also indicate that the Ly123+ T-cell set mediating feedback suppression in vivo is sensitive to both low doses of cyclophosphamide and removal of the thymus in adult life. The importance of this T-T interaction to normal, physiologic regulation of the immune system is emphasized by the finding that the major T-cell deficit of NZB mice (an inbred strain of mice that spontaneously develops an autoimmune disorder) is the absence or malfunction of an Ly123+ T-cell set responsible for feedback inhibition.

These experiments test the hypothesis that cells carrying the Ly1+23- surface phenotype are programmed exclusively for helper and not suppressive activity regardless of external conditions such as the mode or type of antigen stimulation. To this end, we have stimulated purified populations of Ly1 cells with antigen in vitro using conditions devised to induce unselected T cells to express optimal levels of antigen specific T-suppressor activity. We find that after such stimulation, Ly1 cells generate SRBC-specific T-helper activity but not T-suppressive activity. These findings establish that the Ly1.2+,2.2/3.2- surface phenotype is a stable, and probably invariant, marker of T cells that are programmed to express only helper activity and have lost the capacity to directly suppress the antibody response. These findings support the concept that the genetic program for a single differentiated set of cells combines information for cell surface phenotype and function. We also demonstrate that antigen-stimulated Ly1 cells, in addition to inducing B cells to secrete antibody, can induce or activate other sets of resting T cells to develop profound suppressive effects. The surface phenotype of this feedback suppressive T-cell set is shown to be: Ly1+2+3+Qa1+. These findings, taken together, indicate that activation of resting Ly123 cells by immune Ly1 TH cells may represent an important homeostatic immunoregulatory mechanism.