Proliferation of thymic stem cells with and without receptors for interleukin 2. Implications for intrathymic antigen recognition.

We have tested the dividing cells in the mouse thymus for expression of interleukin 2 (IL-2) receptors (IL-2-R) using the rat monoclonal antibody 7D4. A discrete subpopulation of the lymphoblasts clearly expressed IL-2-R at levels comparable to those on mitogen-activated peripheral T cells. This subpopulation, however, represented a small minority of the proliferating cells. IL-2-R-bearing cells were depleted from the PNA+ (peanut agglutinin) lymphoblast population, which contains the direct precursors of most of the cells in the thymus. The majority of receptor-bearing cells were found in the PNA- lymphoblast population, where they constituted only approximately 12% of the cells. Thus, virtually all the PNA+ and most of the PNA- blast cells were in cycle without detectable IL-2-R expression. This indicates that they were not dividing in response to IL-2, and implies that they were not dividing in response to antigen, but rather to novel thymus-specific mitogenic stimuli. On the other hand, the proliferating cells that do express IL-2-R were enriched 4-5-fold in the rapidly growing neonatal thymus, suggesting that they may also play a key role in T cell development.

HIV-1 production from infected peripheral blood T cells after HTLV-I induced mitogenic stimulation.

The human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type I (HTLV-I) are two distinct human retroviruses that infect T cells. Recent epidemiologic studies have identified a cohort of individuals that are coinfected with both viruses. It is reported here that human peripheral blood leukocytes infected with HIV-1 in vitro can be induced to produce large quantities of HIV-1 after mitogenic stimulation by noninfectious HTLV-I virions. It is also shown that HTLV-I virions may exert this effect prior to, immediately following, or well after the cells are infected with HIV-1. These results provide further impetus for epidemiologic studies of dually infected individuals to determine whether HTLV-I may act as a cofactor for acquired immunodeficiency syndrome (AIDS).

Progression to development of lymphoma in the thymus of AKR mice treated neonatally with SL 3-3 virus.

All AKR mice develop thymic lymphoma between 60 and 90 days of age after neonatal treatment with the oncogenic retrovirus SL 3-3. At 40-50 days of age, in the normal-sized thymus of virus-treated mice, cells appear that produce lymphoma when inoculated intrathymically but not when inoculated s.c. These cells are designated as thymus-dependent (TD) lymphoma cells. TD cells progress to cells that form tumors after both intrathymic and s.c. inoculation; these are designated as thymus-independent (TI) lymphoma cells. In this report, we show that the TD and TI cells can be distinguished as two distinct cell populations. Experiments show that the TD cells reside within the immature CD4- CD8- thymocyte population of the virus-treated mice. In addition, we also show that CD4- CD8- thymocytes from SL 3-3 virus-treated mice do not mature in fetal thymic stromal rudiments. Using three-color flow cytometry to trace maturation of CD4- CD8- thymocytes after intrathymic inoculation into irradiated syngeneic hosts, disregulated thymocyte maturation of this population from virus-treated mice is demonstrated. Thus, altered maturation of and the appearance of TD lymphoma cells in, the most immature population of thymocytes appears to be a first step in a multistep process of thymic lymphomagenesis caused by SL 3-3 virus.

Early precursor thymocytes can produce interleukin 2 upon stimulation with calcium ionophore and phorbol ester.

T-cell precursors were stimulated with a conventional T-cell mitogen or with the calcium ionophore A23187 in order to determine whether pre-T cells acquire the ability to produce interleukin 2 (IL-2) before they acquire the ability to respond to antigen or mitogenic lectins. Immature T cells were obtained by eliminating mouse thymocytes that expressed the Lyt2 and L3T4 cell surface proteins. The remaining Lyt2-, L3T4- cells were stimulated for IL-2 production by using concanavalin A (Con A) or A23187, together with phorbol 12-myristate 13-acetate (PMA). We found that these "double-negative" thymocytes were unresponsive to Con A plus PMA but produced substantial amounts of IL-2 when stimulated with A23187 plus PMA. In contrast, both stimulation regimens induced more mature T-lymphocyte populations to produce IL-2. This implies that developing T cells acquire the ability to make IL-2 upon induction before they acquire the ability to be triggered by Con A. Day-15 fetal and cortical thymocytes were also tested for their ability to make IL-2. Both populations failed to synthesize this growth factor, even when stimulated with A23187 and PMA. For cortical thymocytes, this result, together with the finding that A23187 plus PMA fails to activate these cells, suggests that this population is immunologically inert rather than immature. On the other hand, the inability of day-15 fetal thymocytes to produce IL-2 indicates that these T-cell precursors are developmentally distinct from adult Lyt2-, L3T4- thymocytes, which they phenotypically resemble.

Isolation and partial characterization of concanavalin A receptors on cloned cytotoxic T lymphocytes.

A small set of concanavalin A (Con A)-binding glycoproteins was isolated from the surface membrane of cloned cytotoxic T lymphocytes (CTL) and partly identified using monoclonal antibodies. The binding of Con A by these glycoproteins on the CTL surface results in the secretion of gamma-interferon and in blocking the effector functions of the cells-namely, antigen-specific and lectin-dependent cytotoxicity. The Con A is evidently bound tightly to some surface structures ("Con A-receptors") that are required for the activation and cytotoxic activity of CTL. To isolate and identify these receptors, antibodies to Con A were used. After Con A was allowed to bind to radiolabeled cloned CTL (labeled with 125I or [35S]methionine or 3H-labeled amino acids), the cells were washed thoroughly, lysed in detergents and anti-Con A antibodies were added to bind to the Con A-receptor complexes. The resulting aggregates were adsorbed with protein A-bearing Staphylococci and the receptors were then specifically released from the pelleted bacteria by alpha-methyl-D-mannoside and analyzed by polyacrylamide gel electrophoresis under reducing conditions. Eight to nine labeled components were seen by autoradiography and with the aid of monoclonal antibodies to known T-cell surface molecules, four were identified as T200, lymphocyte function-associated antigen (LFA)-1, alpha- and beta-chains, and (on some clones) Lyt-2. Other components with Mr congruent to 160,000, 120,000, 46,000, 42,000, and 23,000 have not been identified. The procedures described here may have general application in the studies of the functional properties of other cell surface molecules.