Human and mouse lymphoid cells, stimulated by phytohemagglutinin (PHA) or lipopolysaccharide W (LPS), release supernatant factor(s) which are mitogenic for mouse thymocytes and which potentiate their responses to PHA or concanavalin A (Con A), The term LAF (lymphocyte-activating factor) is proposed for this activity. LAF not only enhances the mitotic responses of the less dense thymus subpopulations (A, B, and C) separable on discontinuous bovine serum albumin (BSA) gradients but also gives substantial responses in the otherwise inert cells of the denser fractions D and P. LAF does not exert a potentiating stimulatory effect on the responses of unfractionated mouse spleen cells, but does act synergistically with PHA on nonadherent spleen cells and on spleen cells of mice of several strains 5 days after irradiation and injection of thymocytes. Similarly LAF, which has no visible effect on unfractionated human peripheral blood cells, strongly potentiates the PHA response of column-purified lymphocytes, when these are cultured at low concentration. We conclude that LAF stimulates both central and peripheral T lymphocytes and enhances their responses to other stimulants.

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Immunization of mice with low doses of sheep red blood cells (SRBC) which are suboptimal for antibody responses, leads to sensitization for delayed foot pad reactions within 4 days (1-3) . These delayed-type hypersensitivity (DTH) reactions cannot be elicited in mice which have been immunized with larger doses of SRBC that are more optimal for antibody responses (3, 4) . It has been suggested that B-cell responses to high dose immunization cause suppression of T-cell-mediated delayed foot pad reactions (3) . In support of this notion, animals pretreated with B-cell depleting doses of cyclophosphamide (2-300 mg/kg) have markedly suppressed antibody responses and augmented delayed reactions (4-7) . We have studied delayed foot pad reactions of mice immunized with SRBC and found that by lowering the dose of cyclophosphamide pretreatment to 20 mg/kg, delayed-type hypersensitivity can be augmented without affecting antibody responses . Thus antibody is not the sole regulatory factor in mice immunized with high doses of SRBC .Groups of five to six male mice (5 wk old) were obtained from Jackson Laboratories (Bar Harbor, Maine), and were immunized intravenously with 0.2 ml of different concentrations of washed SRBC 1 wk after arrival . Cyclophosphamide (Cytoxan, Mead Johnson & Co ., Evansville, Ind.) was dissolved in sterile saline immediately before use and injected by the intraperitoneal route as a single dose of 20 mg/kg 1 day before immunization, or 200 mg/kg 2 days before immunization . 4 days after immunization, one foot pad per mouse was measured for thickness with a micrometer (Brown & Sharps Mfg. Co ., No . Kingston, R.I .) and was then tested by injection of 0.03 ml of 20% SRBC in sterile saline . The original foot pad thickness was compared to subsequent measurements to compute the percent increase in thickness. Animals were re-tested in the contralateral foot pad 10 days after immunization . Blood samples were obtained by capillary tube puncture of the retro-orbital plexus 5 and 10 days after initial immunization . The resulting serum from individual mice was twofold diluted in 0.025-ml volumes in microtray wells for measurement of hemagglutinating antibody (HA) titers . Student's t test was used to compute statistical significance (P < 0.05) of HA titers and DTH.

The T-lymphocyte population is divisible into several subclasses; each subclass possesses a distinctive genetic program which combines information for cell-surface phenotype and function (1). In the mouse, there is evidence that T cells which express the Thyl+Lyl+Ly23 -surface phenotype CLyl cells") are programmed for helper (TH) 1 function. In contrast, T cells that express the Thyl+Lyl-Ly23 ÷ surface phenotype CLy23 cells") are programmed for suppressor (Ts) function (1). Isolation of these two T-cell subclasses in mice depleted of T cells CB mice") has indicated that each belongs to an independent line or branch of thymus-dependent differentiation (2). A third major T-cell subclass, expressing the surface phenotype Lyl+2+3 +, can react to antigen and differentiate to Ly23 + cytotoxic effector cells (3), suggesting that this subclass probably contains precursor cells that have acquired receptors for antigen but have not yet become committed to either TH or Tc~s function (3).These findings, and others, are consistent with the view that functionally distinct T-cell sets carry cell surface components that are invariably associated with particular immunologic function. According to this idea, cells carrying the Lyl+Ly23 -surface phenotype are programmed for helper and not suppressive activity regardless of external conditions, such as the mode or type of antigen stimulation. To test this hypothesis we have stimulated purified populations of Lyl+2 -T cells with antigen in vitro, by using conditions devised to induce unselected T cells to express optimal levels of antigen-specific suppressive activity (4). We find that (a) stimulation of purified Lyl cells under these conditions results in the generation of TH but not Ts activity and (b) such hyperimmune Lyl cells also induce a subset of nonimmune T cells to exert potent suppressive effects upon the antibody response. The surface phenotype of the T-cell set responsible for "feedback" inhibition is described in this study.

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