Variations in the expression of theta-C3H alloantigen on thymic and peripheral lymphocytes of newborn and young adult mice

The relative frequency of lymphocytes of mice showing varying degrees of surface θ-positivity (circumferential fluorescence) was recorded. Thymocytes were nearly 100% θ-positive. The relative proportions of θ-positive cells in Peyer's patches and lymph nodes of newborn mice varied in an almost identical fashion as a function of age. At 4 weeks of age and beyond, the relative numbers of θ-positive cells in Peyer's patches were consistently lower than in lymph nodes. As opposed to the predominance of thymocytes with complete rings, peripheralized thymic (T) lymphocytes showed a broad, age- and organ-dependent range of surface θ-positivity. These results suggest that surface θ may be lost rather rapidly upon emigration of lymphocytes from the thymus and/or that many θ-positive T cells with complete rings disappear within a short time. Variations in the relative proportion of complete rings on mesenteric lymph node cells on Days 1 and 4, were tentatively related to antigen-induced changes in the magnitude of thymocyte emigration.The pattern of surface θ-antigen of a given thymocyte or T cell with its size and DNA synthetic activity was compared. The findings suggest that incomplete ring fluorescence may especially be observed on proliferating lymphoblasts in the outer thymic cortex on their way to acquire the full complement of θ-antigen, and on medullary thymic lymphocytes or T cells having reentered mitotic activity, in response to antigenic and/or other microenvironmental stimuli. Our study yielded data consistent with the hypothesis that the progressive loss of surface θ-antigen does not represent a fully autonomous time-dependent process. Moreover, it is not clear if continued loss of θ-antigen by T cells below a certain threshold would render these cells undetectable by anti-θ sera.     

Further studies on Th-B, a cell surface antigenic determinant present on mouse B cells, plasma cells and immature thymocytes.

A goat antiserum (Goat anti-M104E) has been produced which contains antibodies selectively cytotoxic for mouse B cells and a subpopulation of thymus cells. It reacts with the Th-B antigenic determinant which has been shown by us (1–3) to be present on B cells and on plasma cells and on some cells in the thymus. It also is very cytotoxic for mouse B cells while a previously developed rabbit antiserum was not. The antiserum was obtained by immunization with cells of the BALB/c mouse myeloma MOPC-104E. When the antiserum was purified by in vivo absorption in mice, antibodies remained which were cytotoxic for cells of all of several myelomas at a titer between 1:128 and 1:1024 as determined by an in vitro complement dependent cytotoxicity test. The in vivo purified antibodies were also cytotoxic for about 70% of thymus cells, for about 70% of spleen cells, for about 50% of lymph node cells and for about 20% of bone marrow cells. They were very cytotoxic for splenic or lymph node B cells separated from T cells by a nylon wool column and only slightly cytotoxic for splenic or lymph node T cells. The antibodies were only weakly cytotoxic for one out of five T cell tumors tested and not cytotoxic for the remaining four. Irrespective of target cells used, the cytotoxicity of purified Goat anti-M104E was easily removed by absorption with cell suspensions from tissues which contain B cells, plasma cells or thymus cells. In order to confirm that the same anti-Th-B antibodies recognize the determinant present on spleen cells and on some thymocytes, the purified Goat anti-M104E serum was absorbed with either spleen cells or thymus cells. The absorbed sera were tested for ability to label thymocytes or spleen cells using the fluorescence activated cell sorter (FACS). Either absorption removed essentially all the antibody capable of binding to either cell population. In addition it was shown, using the FACS, that only B cells and not T cells of the spleen contain the Th-B determinant. The anti-Th-B antibodies have now been used for the rapid elimination of B cells from a mixed population of lymphocytes without affecting the function of mature T cells. Thus in vitro treatment of spleen cells from SRBC-immunized donors with purified Goat anti-M104E plus complement results in the killing of a high proportion of the B memory cells as shown by the reduction of PFC produced when the treated cells are transferred to irradiated recipients. The T cell helper function of the transferred cells is not affected by Goat anti-M104E treatment as shown by appropriate cell transfer experiments in which effective B cells are provided by an AKR anti-Thy-1.2-treated spleen cell population and effective T cells are provided by the Goat anti-M104E-treated spleen cell population. Antibodies detecting Th-B may serve as an approach to understanding the ontogeny of lymphocytes. Our results suggest that Th-B is a cell surface marker appearing early in the development of lymphoid cells, on the common precursor of B and T cells and that it is lost from T cells as they mature in the thymus.     

Fractionation of mouse T and B lymphocytes by preparative cell electrophoresis. Efficiency of the method

Mouse lymphocytes have been fractionated in preparative cell electrophoresis into two functionally viable populations, a high mobility cell (HMC) and a low mobility cell (LMC) population. The distribution of HMC in CBA spleen, blood, and lymph node corresponds to known proportions of θ-positive cells in these organs. The HMC carry the θ-isoantigen, respond to phytohemagglutinin in vitro, and induce a graft-versus-host reaction in newborn F₁ hybrid mice. Nearly all spleen LMC have complement receptors on their surface. About 70% of spleen LMC are sensitive to anti-MBLA serum and form “caps” when incubated with FITC-conjugated anti-Ig. Only LMC respond to E. coli lipopolysaccharide. Thus, T cells localize in the HMC population and B cells in the LMC population. There is no detectable contamination of T lymphocytes among the LMC, nor of B lymphocytes among the HMC.     

Thymus cell subpopulations separated on discontinuous BSA gradients: antigenic properties and circulation capacity

Subpopulations of thymus cells were separated on discontinuous bovine serum albumin (BSA) gradients. The cells in each fraction were tested for their sensitivity to the cytotoxic effect of normal guinea pig serum (GPS), and of antisera against the TL, θ, and H-2 alloantigens. The more buoyant fractions of strain A thymus cells were found to possess a high sensitivity to H-2 antibodies, but a low sensitivity to GPS, and to TL and θ-antisera. In contrast, high density A thymus cells were highly sensitive to the cytotoxic effect of GPS, TL-antibody and θ-antibody, and resistant to the cytotoxic effect of H-2 antibodies. Among subpopulations of C57BL thymus cells, GPS showed a marked different activity on buoyant and dense fractions. Subpopulations of thymus cells varied in their capacity to migrate to lymph nodes of isogeneic recipients. ⁵¹Cr-labeled cells of one of the more buoyant subpopulations showed the highest capacity of localizing in lymph nodes, whereas the densest cell subpopulation had the lowest capacity of migrating to lymph nodes.     

Oligocellular mechanism in the production of antibodies: in vitro response to a haptenic determinant

Spleen explants from mice tolerant to rabbit serum albumin (RSA) failed to react in vitro to dinitrophenyl (DNP)-RSA; antibodies to DNP were, however, produced by such spleens, when stimulated with α-DNP-poly(Lys). To study the function of T and B cells in recognition of carrier determinants, spleen explants from X-irradiated mice, which had been inoculated with combinations of thymus and bone marrow cells from normal and from RSA tolerant donors, were tested for their reactivity in vitro to the DNP-RSA conjugate. A significant response was obtained only by spleens of mice containing bone marrow and thymus from normal donors. Spleens of mice treated with thymus from tolerant and bone marrow from normal or with thymus from normal and bone marrow from tolerant donors did not respond to DNP-RSA. The absence of the response to DNP-RSA by tolerant B cells combined with normal T cells was unexpected. It could not be attributed to binding of the tolerogen to B cells which would have prevented the interaction with T-cells. Neither could the result be attributed to an inhibition of normal cells by RSA-tolerant B-cells. θ-positive cells in the bone marrow are not the cells controlling the recognition of carrier determinants in the B population, since elimination of θ-positive cells did not affect the reactivity of spleens repopulated with B and T cells. Nor are bone marrow macrophages responsible for the lack of reactivity in spleens containing tolerant B cells, since normal macrophages did not restore reactivity. Hence, the production of antibodies to DNP is based on the recognition of carrier determinants not only by T cells, as previously established, but also by B cells. Whether this indicates a B-B in addition to the T-B cell cooperation is an inviting possibility.     

IgM complex receptors on subpopulations of murine lymphocytes.

Murine lymphocytes from spleen, lymph node, and thymus were examined for IgM complex receptors. Lymphocytes from all three organs were found to bind SRBC sensitized with IgM from various sources including: primary anti-SRBC serum, murine and rabbit anti-Escherichia coli LPS sera, and a murine IgM myeloma (MOPC 104E). Rosette formation by lymphocytes with IgM-sensitized SRBC was inhibited by soluble antigen-IgM complexes but not by IgM or antigen alone. Rosette formation was also inhibited by human IgM (Fc)5mu but not by Fab mu. Antiserum and complement treatment of the cells and subsequent recovery of the viable cells by trypsinization, filtration, and washing revealed the IgM rosette-forming cell (RFC) in the thymus to be a T cell. Spleen on the other hand was found to contain both B and T cells capable of binding IgM sensitized SRBC. Removal of both B and T cells from spleen cell suspensions eliminated all IgM RFC. The IgM complex receptor was found to be trypsin insensitive. Anti-Ig column fractionation enriched IgM RFC in spleen and lymph node suspensions passed through the columns, whereas cells bearing surface Ig, IgG complex receptors, and C3 receptors were retained in the columns.     

Prostaglandin E 1 as a regulator of lymphocyte function. Selective action on B lymphocytes and synergy with procarbazine in depression of immune responses

The possibility that prostaglandin (PG) E₁ can regulate lymphocyte function in vivo was studied. In conjunction with procarbazine hydrochloride, a powerful immunosuppressive agent whose main target is the thymus and thymus-derived (T) cells, PGE₁ had a synergistic effect in prolonging homograft survival, though alone it had no effect on this immune response. The main lymphocyte target of PGE₁ appeared to be bone marrow-derived (B) as determined from direct assessment of total, complement receptor and θ-positive lymphocytes in the spleen and by histological evaluation.     

Thymus dependency of cells involved in transfer of delayed hypersensitivity to listeria monocytogenes in mice

Delayed-type hypersensitivity (DH) to Listeria antigens was induced in inbred C3Hf/Umc mice by intravenous injection of a sublethal dose of viable Listeria monocytogenes. Bone marrow, spleen, and lymph node cells from the immune mice were capable of passive transfer of DH to syngeneic neonatally thymectomized or lethally (900 R) irradiated recipients. Immune thymus cells as well as immune serum were ineffective in transferring DH to irradiated animals. In vitro treatment with antitheta isoantibody (anti-θ) and complement abolished the capacity of spleen and bone marrow cells from immune donors to transfer DH to irradiated hosts, indicating the thymus dependency of this cell population. The results with bone marrow indicate the existence of a small, but biologically significant, thymus-dependent population in this tissue.     

Graft-versus-host induced immunosuppression: depressed T cell helper function in vitro.

The cause of graft-versus-host (GVH) induced suppression of the plaque forming cell (PFC) response to sheep erythrocytes (SRBC) was investigated by in vitro restoration experiments employing a double compartment culture vessel. The two culture compartments were separated by a cell impermeable membrane. Restoring cells were placed in one chamber and responding GVH spleen cells plus SRBC were placed in the other chamber. It was demonstrated that thymus, lymph node, and spleen cells restored the PFC response whereas bone marrow cells did not. Treatment of the restoring cells with anti-theta serum plus complement abrogated restoration. Supernatants obtained from antigen free cell cultures restored nearly as well as whole cell suspensions. The degree of restoration was not increased by allogeneic or xenogeneic antigenic stimulation of the restoring cells. Thymus and lymphoid cells obtained from animals experiencing a GVH reaction restored as well as normal cells, however spleen cells were unable to restore by day 5 post-GVH induction. The results suggest that GVH induced immunosuppression of the PFC response is due, at least in part, to a depressed T cell factor production by splenic T cells.     

Studies of the mouse Ly-6 alloantigen system

Three monoclonal antibodies were produced by fusing mouse myeloma cell line NS-1 with spleen cells from C3H/An mice hyperimmunized with B6-H-2^k spleen cells. These antibodies recognized an alloantigen displaying a similar strain distribution pattern to the Ly-6.2 and Ala-1.2 alloantigens. Analysis of C×B and B×H recombinant inbred mice revealed close linkage of genes controlling Ly-m6 and Ly-6. The monoclonal antibodies lysed 70 percent of cells in lymph nodes and 60 percent in spleen in direct cytotoxicity assays, but did not lyse significant numbers of cells of thymus and bone marrow. Separated T and B cells were reactive with the antibodies, but T cells were more sensitive to the antibody and complement than B cells. Virtually all cells in cultures of cells activated in the mixed lymphocyte reaction or by Concanavalin A were reactive with the monoclonal antibodies. Direct plaque-forming cells were completely eliminated by the monoclonal antibody and complement. By absorption tests, cells from all organs tested so far (thymus, lymph node, spleen, bone marrow, brain, kidney and liver) were shown to express the Ly-m6 determinant. Tumor cell lines with T, B or stem cell characteristics were reactive with the monoclonal antibody by direct cytotoxicity and absorption assays.     

Mitogen induction of murine C-type viruses. I. Analysis of lymphoid cell subpopulations.

We reported previously in vitro induction of endogenous C-type viruses from normal mouse spleen cells by lipopolysaccharide (LPS) as well as by combination treatment with concanabalin A and 5-bromo-2'-deoxyuridine (Con A/BrdU). To identify the cell types sensitive to virus induction and to study the relationship of mitogenicity to virus induction we have compared T cell populations (BALB/c thymus cells and cortisone-resistant thymus cells), B cell populations (nu/nu spleen cells and lymph node cells), adherent BALB/c peritoneal cells and mixed populations (BALB/c spleen cells, macrophage-depleted BALB/c spleen cells, and lymph node cells). LPS-induction occurred only in B cell-containing populations. In contrast, induction by Con A/BrdU depended on the presence of both T and B cells. In both instances, neither macrophages nor hemopoietic cells appeared to be a major source of virus. Treatment with anti-Ig serum and complement reduced virus induction by LPS/BrdU but not by Con A/BrdU suggesting that different cell populations produce virus after stimulation with these two different mitogens.     

Identification of human peripheral T cell antigens.

A new type of differentiation antigens on human T cells was demonstrated by using a heterologous anti-human T cell serum (ATS). This type of antigen, referred to as human peripheral T cell antigen (HPTA), was found on peripheral T cells and medullary thymocytes, but not on cortical thymocytes and B cells. The percentage of ATS-reactive lymphocytes in human peripheral lymphoid organs was correlated with that of cells rosetting with sheep erythrocytes, but contrasted with the number of B cells defined by the presence of a complement (C) receptor or by rabbit anti-human B cell serum (ABS). ATS also reacted with T cells purified by nylon fiber column filtration but ABS did not. Chronic lymphocytic leukemia cells rosetted with either sheep erythrocytes or erythrocyte-antibody-complement complexes were lysed by ATS and ABS, respectively. Mitogenic responses of blood lymphocytes to phytohemagglutinin (PHA) and concanavalin-A (Con A) were abrogated by treating them with ATS and C, whereas ABS suppressed only their response to Con A. Although numerous thymus cells rosetted with SRBC, only 14% were reactive with ATS. Quantitative absorption studies demonstrated that HPTA content of the thymus cells was much lower than that of lymph node cells. Anatomical localization of ATS-reactive lymphocytes in human lymphoid organs studied by immunofluorescence indicated that they were present in the thymus-dependent paracortical areas of lymph node and in the medullary region of thymus. ABS, on the other hand, did not stain thymocytes but reacted selectively with the cells located in the lymphoid follicles of lymph node. These data, together with that from cell suspension studies, confirmed that HPTA were shared between medullary thymocytes and peripheral T cells.     

Subpopulations of human T lymphocytes. III. Distribution and quantitation in peripheral blood, cord blood, tonsils, bone marrow, thymus, lymph nodes, and spleen.

Human T cell subpopulations (Tμ and Tγ) were examined for their distribution in the peripheral blood, cord blood, bone marrow, tonsils, thymus, lymph nodes and spleen. The proportions of Tμ and Tγ cells are comparable in the peripheral blood, tonsils and bone marrow. The proportions of Tγ cells in cord blood are significantly higher than those in the peripheral blood. Almost complete lack of Tγ cells was observed in lymph nodes. Spleen has very high proportions of Tγ cells. Thymuses have very low proportions of both Tμ and Tγ cells when compared with peripheral blood, cord blood, tonsils, and bone marrow. The receptors for IgM on Tμ cells appear to be masked by passively absorbed IgM and require prior in vitro incubation in medium containing fetal calf serum for the full expression of this marker.     

Ex vivo imaging of T cells in murine lymph node slices with widefield and confocal microscopes

Naïve T cells continuously traffic to secondary lymphoid organs, including peripheral lymph nodes, to detect rare expressed antigens. The migration of T cells into lymph nodes is a complex process which involves both cellular and chemical factors including chemokines. Recently, the use of two-photon microscopy has permitted to track T cells in intact lymph nodes and to derive some quantitative information on their behavior and their interactions with other cells. While there are obvious advantages to an in vivo system, this approach requires a complex and expensive instrumentation and provides limited access to the tissue. To analyze the behavior of T cells within murine lymph nodes, we have developed a slice assay ¹, originally set up by neurobiologists and transposed recently to murine thymus ². In this technique, fluorescently labeled T cells are plated on top of an acutely prepared lymph node slice. In this video-article, the localization and migration of T cells into the tissue are analyzed in real-time with a widefield and a confocal microscope. The technique which complements in vivo two-photon microscopy offers an effective approach to image T cells in their natural environment and to elucidate mechanisms underlying T cell migration.     

Contact sensitivity to picryl chloride: the occurrence of B suppressor cells in the lymph nodes and spleen of immunized mice.

Mice were immunized for contact sensitivity and antibody production by painting the skin with picryl chloride. Lymph node and spleen cells taken 4 days later transferred contact sensitivity. However, cells taken at 7–8 days failed to transfer but were able to block the transfer by 4 day immune cells. These suppressor cells occurred in the regional lymph nodes, spleen and thymus. The suppressor activity of lymph node and spleen cells was due to B cells as shown by the effect of anti-θ serum and complement, nylon wool filtration and separation of EAC positive and negative cells by centrifugation on a discontinuous gradient. The transfer of fractions rich or poor in macrophages showed that the suppressor cell in the transferred population was not a macrophage. Separation using EAC rosettes suggested that B cells were responsible for the suppressor activity in the thymus.T cells isolated from the lymph nodes and spleen 7–8 days after immunization transferred contact sensitivity although the initial population was inactive. This indicates that passive transfer cells are present in the regional lymph nodes and spleen at later times after immunization but cannot be demonstrated because of the presence of suppressor B cells. However, no passive transfer cells were found in the thymus. The production of B suppressor cells required little or no T cell help and following immunization the spleens of reconstituted (B) mice were at least as active as control cells in causing suppression. There are several different suppressor cells which act in the picryl system and the B suppressor cells in immunized mice described here are distinct from the T suppressor cells in mice injected with picryl sulphonic acid.     

A novel cell-surface antigen expressed on most leukocytes and a minor cortisone-resistant population of thymocytes in rats: characterization by monoclonal antibodies

A cell-surface antigen on rat lympho-hemopoietic cells was determined by using a monoclonal antibody, R2-1B3 (1B3). The 1B3 antibody, when tested for its reactivity with different hemopoietic cells by cytofluorography with a FACS analyzer, labeled more than 80% of lymph node, spleen, and bone marrow cells and 10-20% of thymus cells. Cytofluorographic analysis performed on purified rat T cells, B cells, macrophages, and granulocytes demonstrated that the antigen defined by 1B3 was readily detectable on all of these cell types, with the greatest expression on B cells. A minor population of thymocytes that were labeled by 1B3 appeared to be cortisone-resistant and were located mainly in the thymic medulla. These 1B3 positive thymic cells seemed to be functionally more mature than 1B3-negative thymus cells as suggested by the fact that the cytotoxic treatment of thymus cells with 1B3 antibody and complement (C) resulted in significant reduction of their responsiveness to phytomitogens and lymphokines derived from concanavalin A (con A) activated rat spleen cell cultures. Immunochemical data showed that 1B3 antibody recognized the broad ill-defined band with a molecular weight of 32K to 47K daltons as estimated by SDS-polyacrylamide gel electrophoresis. These data indicate that the 1B3 defined antigen is distinct from other, previously reported, antigens on rat lymphoid cells including leukocyte-common (L-C) and MRC OX-22 antigens, and that this 1B3 antibody is a useful reagent for analyzing the intrathymic differentiation of T cells in rats.     

Synergy among rat T cells in the proliferative response to alloantigen.

A synergistic interaction in the proliferative response to alloantigen is described for mixtures of rat thymus and lymph node cells. The optimal conditions for synergy are quantitatively defined. Regression analysis of the slope of the dose-response curve has been utilized to estimate the degree of interaction in thymus-lymph node cell mixtures. The slope of the response of cell mixtures was noted to be significantly greater than the slope for the response of lymph node cells alone. Irradiation was shown to have a differential effect on the response of thymus and lymph node cells in mixtures. Irradiated thymus cells retained the capacity for synergy in mixtures, whereas irradiated lymph node cells did not. Additional studies have demonstrated that both de novo protein synthesis and specific antigen recognition by both responding cell populations in mixtures was required for maximal synergy. These studies demonstrate that synergy cannot be explained as an artifact of altered cell density in vitro. They establish that thymus cells and lymph node cells represent distinct subsets which manifest qualitatively different functions in the proliferative response to alloantigen. Thymus cells can respond directly to alloantigen by proliferation but also have the capacity to amplify the proliferative response of lymph node cells—a capacity which is resistant to X irradiation but requires recognition of alloantigen and de novo protein synthesis. Lymph node cells may similarly respond by proliferation to alloantigen but lack the amplifier activity of thymus cells. Synergy for rat lymphoid cells, like mouse lymphoid cells, has been shown to involve an interaction of thymus-derived lymphocytes.     

Neonatal infection with mouse thymic virus. Differential effects on T cells mediating the graft-versus-host reaction.

Neonatal infection with mouse thymic virus (TA), a murine herpes virus, produced extensive but temporary necrosis of the thymus which was maximal at 10 to 14 days of age. Studies of precursor and amplifier cells mediating graft-vs-host (GVH) reactivity of thymocytes, spleen cells (SC), and lymph node cells (LNC) of normal and TA-infected mice were made at 4 and 8 weeks of age. Infection with TA resulting in a profound reduction (70 to 80%) in the direct GVH reactivity of thymocytes at both ages; by comparison, the capacity of thymocytes to produce synergy when combined with normal LNC was normal at 8 weeks. Direct GVH reactivity of SC was depressed 90% 4 weeks after infection with TA but returned to near normal at 8 weeks. Direct GVH reactivity of LNC from TA-infected mice was normal at 4 and 8 weeks of age, but amplifier T cell activity in LNC was markedly depressed at 8 wekks. These results demonstrate that TA has highly selective effects upon subpopulations of T cells in thymus and lymph node.     

Pituitary Adenylate Cyclase Activating Peptide Deficient Mice Exhibit Impaired Thymic and Extrathymic Regulatory T Cell Proliferation during EAE

We have shown that mice deficient in pituitary adenylate cyclase-activating polypeptide (PACAP, gene name ADCYAP1) manifest enhanced sensitivity to experimental autoimmune encephalomyelitis (EAE), supporting the anti-inflammatory actions described for this neuropeptide. In addition to an increased proinflammatory cytokine response in these mice, a reduction in regulatory T cell (Treg) abundance in the lymph nodes (LN) was observed, suggesting altered Treg kinetics. In the present study, we compared in PACAP deficient (KO) vs. wild type mice the abundances and rates of proliferation FoxP3⁺ Tregs in three sites, the LN, central nervous system (CNS) and thymus and the relative proportions of Th1, Th2, and Th17 effector subsets in the LN and CNS. Flow cytometry analyses revealed a decrease in Treg proliferation and an increased T effector/Tregs ratio in the LN and CNS of PACAP KO mice. In the thymus, the primary site of do novo natural Treg production, the total numbers and proliferative rates of FoxP3⁺ Tregs were significantly reduced. Moreover, the expression of IL-7, a cytokine implicated in thymic Treg expansion during EAE, failed to increase at the peak of the disease in the thymus and LN of PACAP KO mice. In addition to these Treg alterations, a specific reduction of Th2 cells (about 4-fold) was observed in the lymph nodes in PACAP KO mice, with no effects on Th1 and Th17 subsets, whereas in the CNS, Th1 and Th17 cells were increased and Th2 decreased. Our results suggest that endogenous production of the neuropeptide PACAP protects against EAE by modulating Treg expansion and Th subsets at multiple sites.     

T cell lineages in the thymus of lpr/lpr mice. Evidence for parallel pathways of normal and abnormal T cell development

Autoimmune mice homozygous for the lpr/lpr (lpr) gene develop a profound lymphadenopathy resulting from the accumulation of immature Thy-1+ Lyt-2- L3T4- cells in peripheral lymphoid tissues. The source of these cells is not known although the presence of a thymus is necessary to manifest both the lymph node enlargement and the autoimmunity. For this reason and the fact that the abnormal lpr cell phenotypically resembles immature thymocytes, we studied the thymus in lpr mice. Adult lpr thymuses were found to contain an immature population phenotypically identical to the peripherally accumulating cells, including the expression of B220 and Pgp-1 antigens as well as the presence of surface T cell receptor molecules as defined by the antibody KJ16-133. Evidence is presented that some of these lpr precursor T cells are capable of intrathymic differentiation, whereas the vast majority are exported unchanged to the lymph nodes where a portion differentiate further into mature T cells. This lpr-specific lineage could be distinguished from a normal component of the lpr thymus by surface phenotype and immunohistology. The results suggest that the massive accumulation of cells in lpr lymph nodes is not so much the result of abnormal proliferation of T cells as abnormal intrathymic differentiation. In addition, a minor subpopulation of normal Lyt-2- L3T4- thymocytes was identified that resembles the phenotype of the lpr cell and similarly expresses surface T cell receptor molecules. The presence of two parallel lineages in the lpr thymus thus also provides insight into normal T cell development.