Double labelled T suppressor lymphocytes in the thymus of a patient with myasthenia gravis: morphological and immunological characteristics

Double labelled lymphocytes were prevalent in the thymus of a 32-year-old woman with myasthenia gravis. The simultaneous presence of both E and C3b surface receptors was demonstrated. Focal paranuclear acid phosphatase activity of the cells supported their T cell origin. Histological analysis revealed a hyperplastic thymus with germinal centres in one lobe and an unusual transition to "thymoma-like" diffuse sheets of thymocytes and large thymic epithelial cells in the other lobe. Immunological studies of isolated cells from the "thymoma-like" region revealed a high percentage of E rosettes (96%), EAC rosettes (87%) and IgG-EA rosettes (60%). The large number of "active" rosettes (94%) indicated high avidity of receptors for sheep red blood cells. The postoperative accumulation of acid phosphatase positive T cells with C3b surface receptors in the peripheral blood was striking. These characteristics suggest that this cell population represents an activated T subset (TG) with a certain degree of immaturity similar to a particular stage of foetal T cell development (prothymocytes).     

Human lymphocyte subpopulations. Effect of corticosteroids

Normal subjects given 60 mg of prednisone orally at 8:00 a.m. developed a transient lymphopenia at 2:00 p.m. To define the populations of lymphocytes affected the number and type of lymphocytes in the peripheral blood were assayed. "Late" and "early" spontaneous sheep red blood cell rosettes were used as markers for thymus-derived (T) lymphocytes and one of its subpopulations, respectively. Receptors for aggregated gammaglobulin and complement identified bursal-equivalent or bone marrow-derived (B) lymphocytes and one of its subpopulations, respectively. 6 h after administration of 60 mg of prednisone, the blood samples showed a decrease in proportion of T cells from 69.2 +/- 2.1% to 55.9 +/- 2.8% (average +/- SE) and an increase in B-cell proportion from 21.3 +/- 2.0% to 44.8 +/- 4.1%. The changes of "early" rosettes and complement receptor lymphocytes also paralleled these. In all cases the absolute numbers of T cells and of B cells were decreased by prednisone. The density gradient distribution of the lymphocytes did not change after prednisone. These data indicate that both T and B lymphocytes are affected by the prednisone but that the T cell lymphopenia was more pronounced. The lymphopenia might reflect either sequestration in the marrow and/or transient arrest of recirculation.     

Malignant lymphoma with dual B and T cell markers. Analysis of the neoplastic cells with monoclonal antibodies directed against T cell subsets

In the course of analyzing human lymphoma tissue with conventional surface marker techniques and with monoclonal antibodies directed against T cell subsets, five tumors were encountered with dual B and T cell determinants. All bore on their surface membrane IgM of kappa light chain type, complement receptors, and the Ia-like antigen. In each of the five cases, the neoplastic lymphocytes reacted with a monoclonal antibody that detects the sheep erythrocyte receptor (OKT11); all but one reacted with a monoclonal antibody for peripheral T cells (OKT3); and all but one reacted with a monoclonal antibody specific for either the inducer-helper (OKT4) or the cytotoxic- suppressor (OKT8) T cell subsets. In addition, lymphocytes from two of the five cases formed spontaneous rosettes with sheep erythrocytes (E- rosettes). These tumors with dual B and T surface characteristics were confined to human malignant lymphomas that originate from B lymphocytes of the follicle center.     

SURFACE MARKERS ON HUMAN T AND B LYMPHOCYTES : I. A LARGE POPULATION OF LYMPHOCYTES FORMING NONIMMUNEROSETTES WITH SHEEP RED BLOOD CELLS

By using the two criteria (a) high density of immunoglobulin determinants on the cell surface and (b) presence of receptors for C'3 on the cell surface for defining bone marrow-derived lymphocytes, it is indirectly shown that all or at least a major population of human thymus-derived lymphocytes under certain conditions will form nonimmune rosettes with sheep red blood cells (SRBC). Almost all thymocytes tested from two different donors formed rosettes. The SRBC rosettes are not formed by virtue of immunoglobulin receptors and form only around living cells. Positive bivalent ions are required for rosette formation since EDTA will block rosette formation. Sodium iodoacetate will also block rosette formation demonstrating the dependence on an intact glycolytic pathway. Rosette formation is temperature dependent and will not appear at 37°C. Trypsin treatment of lymphocytes will abolish their SRBC-binding ability which cannot be restored by treating them with fresh donor serum or fetal calf serum, but which will reappear after culturing the lymphocytes. It is suggested that these rosettes are formed by a rapidly released or metabolized receptor substance on the living cell surface which behaves as a trypsin-sensitive structure produced by the cells themselves.     

A subpopulation of normal human peripheral B lymphcytes that bind IgE.

Isolated normal human peripheral lymphocytes were analyzed for their ability to bind IgE as shown by rosette formation with aldehydefixed ox red cells coated with an IgE myeloma protein (Eo'-IgE) as indicator cells. An average of 4.3% of the cells in the lymphocyte preparations of 12 donors formed Eo'-IgE rosettes. The lymphocyte preparations contained an average of 0.36% basophils which also formed Eo'-IgE' rosettes, suggestiing that about 4% of the lymphocytes bound IgE. The rosette formation was inhibited by IgE myeloma protiens or IgE Fc fragments but not by IgE Fab fragments or Ig of the other classes. On the average, the lymphocytes of the 12 donors contained 70.5% cells forming spontaneous rosettes with sheep erythrocytes (E), 10.6% cells having surface immunoglobulin (SIg), and 15.5% bunding IgG as shown by rosette formation with IgG-coated ox red cells (EoA). Fractionation of the lymphocytes into populations rich in spontaneously E-rosetting cells and cells with SIg indicated that the majority of the lymphocytes forming Eo'-IgE rosettes belonged to the SIg-positive lymphocytes. Analyses of lymphocyte populations lacking cells forming EoA rosettes and of rosetting with mixed indicator cells both demonstrated that over 90% of the lymphocytes forming Eo'-IgE rosettes did not form EoA rosettes and apparently had no Fc receptors for IgG. Pretreatment of the lymphocytes with trypsin in amounts that did not alter the number of EoA-rosetting cells abolished the Eo'-IgE rosette formation. These data indicate that a subpopulation of normal human peripheral lymphocytes, probably belonging to the B-cell type, binds IgE presumably via trypsin-sensitive receptors specific for the Fc fragment of IgE. The surface markers of these lymphocytes resemble those of cultured human lymphoblastoid cells that have recently been shown to bind IgE.     

Characteristics of human large granular lymphocytes and relationship to natural killer and K cells

Recent evidence, has demonstrated an association between a subpopulation of peripheral blood mononuclear cells, morphologically identified as large granular lymphocytes (LGL), and natural killer (NK) activity. We have now evaluated more directly the role of LGL in both NK activity and antibody- dependent cellular cytotoxicity (ADCC), by using highly enriched populations of LGL, obtained by centrifugation of peripheral blood mononuclear cells on Percoll discontinuous density gradients. Both spontaneous and interferon- augmented NK and ADCC activities were exclusively associated with the LGL- enriched, low density fractions. The majority of LGL formed conjugates with NK-susceptible and antibody-coated target cells. Approximately 20 percent of small conventional lymphocytes also formed conjugates with the target cells for NK, but this was not associated with cytotoxic activity. Virtually all LGL were found to have receptors for the Fc portion of IgG (FcγR). The frequency of LGL among blood leukocytes was 2-6 percent. LGL could be enriched to an average purity of 95 percent by combining discontinuous density gradient centrifugation with subsequent adsorptions of the low density fractions on monolayers of immobilized immune complexes. About 50 percent of LGL were found to be FcγR-bearing T cells (T(G)), forming low affinity rosettes with sheep erythrocytes at 4 degrees C. Only 10-20 percent of LGL formed high affinity rosettes with sheep erythrocytes at 29 degrees C. LGL could be enriched to a purity of more than 90 percent by depleting high affinity rosette-forming cells from low density Percoll fractions. LGL were only a subpopulation of T(G) cells, because some lymphocytes with conventional morphology also adhered to the immobilized immune complex monolayers and formed high affinity rosettes with sheep erythrocytes. Separation of these cells from LGL by discontinuous density gradient centrifugation indicated that they are not cytotoxic, suggesting a morphological and functional subdivision of T(G) cells. The verification in this study that virtually all human NK and K cells have a characteristic morphology adds a useful parameter to the monitoring of human lymphocytes, and the ability to purify these cells by simple physical procedures should be invaluable in their further characterization.     

Identification of human B and T lymphocytes by scanning electron microscopy

In this study a variety of human lymphocytes of known B or T cell type, obtained from multiple sources, were prepared for scanning electron microscopy (SEM) by the critical point drying method. Distinction between normal B and T lymphocytes was relatively easy in most instances, on the basis of their surface architecture. Using immunological methods, between 20 and 30% of normal peripheral blood lymphocytes (PBL) were identified as B cells and from 69 to 82% as T cells. SEM results showed that 20% of the PBL had a complex villous surface and approximately 80% of cells were smaller and had a relatively smooth surface. Comparison of the above data and enrichment of B cells from PBL, by centrifugation after T cell rosettes had formed, indicated that the "villous" cells were B lymphocytes and the "relatively smooth" cells were T lymphocytes. T cells obtained from two human thymuses were also of the generally smooth cell type. Further evidence for the distinction of B and T lymphocytes, on the basis of surface morphology, was obtained from the examination of cultured lymphoid cell lines of known B or T cell derivation. Cells from cases of chronic lymphocytic leukemia also provided support for the above interpretations. Five of six untreated cases were clearly of B cell type by immunologic and SEM criteria. One unusual case showed the presence of T and B lymphocytes in almost equal numbers by SEM and a mixture of B and T cells by immunologic markers. An additional case that had received chemotherapy showed numerous atypical cells that were difficult to classify by SEM. Detailed examination of the smoother T cells showed that at least half of them had a moderate number of surface digitations and a small proportion had an intermediate surface morphology with a relatively large number of surface digitations. The latter presented difficulties in classification and may correspond to different stages of differentiation and represent subpopulations of lymphocytes. The distinction between human B and T lymphocytes on the basis of their surface architecture can be made by SEM of critical point dried samples, with relative ease in most but not all instances. The effects of stimulation, cell cycle, differentiation, intercellular contact, and density of cell population, on the surface architecture of lymphoid cells, remain to be determined.     

Maturation of bone marrow lymphocytes. II. Development of Fc and complement receptors and surface immunoglobulin studied by rosetting and radioautography

Radioautographic DNA labeling and rosetting techniques were combined to study the development of surface IgM, Fc, and complement receptors (FcR, CR) on small lymphocyte populations in mouse bone marrow. [3H]thymidine was either infused continuously to label newly formed cells for periods up to 4 days, or injected daily, 21--35 days before use, to label a sample of long-lived cells. Bone marrow cells were incubated with sensitized sheep erythrocytes to detect surface IgM, FcR, and CR, respectively, and examined radioautographically after cytocentrifugation. During [3H]thymidine infusion, marrow small lymphocytes lacking surface markers were the first to show [3H]thymidine labeling. Most of these cells became labeled by 4 days (IgM--ve, 89%; FcR--ve, 92%; Cr--ve, 88%). Labeling of small lymphocytes bearing surface IgM, FcR, and Cr began after an initial lag and increased to high values by 4 days (IgM + ve, 73%; FcR + ve, 82%; CR + ve, 83%). Labeled IgM + ve small lymphocytes formed progressively larger rosettes as cell age increased. Some proliferating large lymphoid cells formed rosettes for IgM, FcR, and CR. Labeled long-lived small lymphocytes expressed surface IgM, FcR, and CR, the incidence of each receptor being uniformly high (38--43%) and the rosettes tending to be larger than those formed by newly formed lymphocytes. In double- surface marker studies, FcR and CR rosettes were formed by some IgM--ve small lymphocytes as well as IgM + ve cells in the marrow. After transfusion of marrow cells from donor mice infused with [3H]thymidine for 24 h, many labeled newly formed lymphocytes homed into the splenic red pulp of unlabeled syngeneic recipients. Subsequently, these cells showed a rapid increase in the incidence of rosettes for surface IgM, FcR, and CR, together with a progressive enlargement of each type of rosette. Although all the labeled small lymphocytes recovered from the spleen developed both surface IgM and FcR by 3 days, only approximately one-half developed CR. The results demonstrate that most of the small lymphocytes bearing FcR, CR, and surface IgM in mouse bone marrow are newly formed indigenous cells. Each receptor becomes detectable by rosetting soon after the small lymphocytes are first produced. The newly formed, marrow-derived small lymphocytes are able to continue their development of surface IgM, FcR, and CR after migrating into the spleen, consistent with a maturation of primary B lymphocytes. In addition, the data indicate the genesis in mouse marrow of a non-B lineage of lymphocytes (notably, IgM--ve FcR + ve cells.). A minority of small lymphocytes bearing IgM, FcR, and CR in mouse marrow are long- lived cells, presumptive recirculating immigrants, differing in receptor status from the newly formed cells. The results are discussed with regard to the heterogeneity of marrow lymphocytes and proposed models of primary B lymphocyte and null lymphocyte production.     

Ia-bearing T lymphocytes in man. Their identification and role in the generation of allogeneic helper activity

The presence of Ia-like antigens was demonstrated on a small population (2-6%) of highly purified human circulating T lymphocytes by immunofluorescence with a rabbit anti-Ia serum raised against the isolated bimolecular Ia structure. The Ia+ T lymphocytes have no surface or intracellular immunoglobulins. The expansion of this Ia+ T- cell population was encountered in certain patients. Ia antigens were also found on T blasts grown in long-term cultures with conditioned medium generated by phytohemagglutinin-stimulated lymphocytes. In addition, leukemia blasts which stained for Ia antigens and formed E rosettes were identified in the peripheral blood of two leukemic patients. This evidence further supports the existence of Ia-bearing T cells in man. The Ia+ T-lymphocyte population was shown to contain cells responsible for the generation of allogeneic helper activity. Elimination of Ia+ lymphocytes from a purified T-cell population by the anti-Ia antiserum and complement abolished its ability to help an allogeneic B-cell preparation to generate plaque-forming cells against sheep erythrocytes in vitro in the presence of the antigen.     

The interaction between human monocytes and red cells. Binding characteristics

Red cells coated with IgG globulin were bound firmly to human mononuclear cells and formed rosettes. Rosette formation occurred when red cells were coated with IgG attached either immunologically (anti-D, anti-penicillin, or Donath-Landsteiner antibodies) or nonimmunologically with chromic chloride; no attachment was observed with cells coated with albumin. Rosette formation was blocked by pretreatment of white cells with sulfhydryl-binding reagents. Metabolic inhibitors did not prevent red cell adherence. White cells of other primates demonstrated a high degree of species specificity. Ultrastructural studies showed that the predominant leukocytes involved in rosette formation were monocytes, but some cells with characteristics of lymphocytes also formed rosettes. Considerable interdigitation of cell surfaces occurred at attachment sites and bound red cells appeared deformed. Thus, these studies confirm the presence of specific surface receptors for IgG on human monocytes and suggest that such receptors may provide a mechanism by which large numbers of red cells are eventually destroyed.     

Characterization of human thymic lymphocytes forming rosettes with stromal cells.

The interaction of thymic lymphocytes and stromal cells is believed to be important for T cell development in thymus. In this study, thymic rosettes (TR), which are cell-cell complexes of thymic lymphocytes and stromal cells, were isolated from human thymic tissue, and were characterized. Treating human thymus with collagenase in mild condition, human TR were successfully isolated. Subsequently, TR were purified by the 1G sedimentation method. Human TR consisted of a stromal cell in center surrounded by lymphocytes. The stromal cells were positive for CD14, CD11b, and HLA-DR but negative for thymic epithelial cell specific mAb, UH-1, suggesting that they are macrophage/dendritic cells. The lymphocytes which formed TR (TRL) were mainly double positive (CD4+CD8+) and CD1+ cells, and few of them expressed bright CD3, indicating that TRL are in the intermediate maturation stage. TRL expressed activation markers (Ta1 and HLA-DR) in a significantly higher percentage of cells than did unselected thymocytes. Blocking test revealed that CD11a and CD2 are involved in the binding of TRL and the stromal cells as adhesion molecules.     

SHEEP RED CELL BINDING TO HUMAN LYMPHOCYTES TREATED WITH NEURAMINIDASE; ENHANCEMENT OF T CELL BINDING AND IDENTIFICATION OF A SUBPOPULATION OF B CELLS

Neuraminidase treatment of normal human lymphocytes enhances their capacity to form SRBC rosettes; more red cells are bound and the rosettes are more stable. Under these conditions approximately 90% of peripheral blood lymphocytes form rosettes. In addition to the effects on T cells, another population of lymphocytes which possess surface immunoglobulin and have the Fc receptor acquire the rosette-forming property after neuraminidase. This subpopulation of ‘B’ cells represents approximately half of the lymphocytes with surface immunoglobulin but is not found among the leukemic lymphocytes of patients with chronic lymphocytic leukemia. Electron microscope observations indicate close approximation and intimate association of the red cell and lymphocyte membranes after neuraminidase.     

Immune function in patients with multiple myeloma

Summary Immunologic function was evaluated in 27 patients with multiple myeloma (MM). When tested for their ability to develop delayed hypersensitivity to a panel of skin test antigens, 2 out of 15 such patients were found to be anergic. The property of human peripheral blood T lymphocytes to form rosettes with sheep [total E and active or early or early rosette-forming cells (RFC)] and human erythrocytes (H-RFC) was also studied. In addition, rosette formation with mouse erythrocytes (M-RFC) was investigated as a B cell marker. Decreased proportions of total E-RFC were found in one third of the patients with MM when compared to normal volunteers. By contrast, both mean percentages and ranges of active E-RFC, H-RFC, and M-RFC in the MM patients overlapped those revealed in healthy controls. The rosette-forming ability of peripheral blood lymphocytes was tested before and after treatment with levamisole, thymosin, and transfer factor in healthy controls, as well as in some groups of patients with MM, active systemiclupus erythematosus and Hodgkin’s disease. A positive effect of all three immunomodulating agents could only be demonstrated on cells from T cell-deficient patients. It is suggested that the immunodeficiency syndrome associated with MM and related malignancies may reflect quantitative and possibly selective defects of lymphocyte subpopulations. Supported in part byConsiglio Nazionale delle Ricerche, Roma, Italy.     

Interactions of the third component of complement (C3) with cross-linked dextran. IV. Adherence of lymphocytes to C3 coated dextran gel particles.

In previous studies we could show that the third component of complement (C3) is bound from normal serum to cross-linked dextran (Sephadex) after activation via an alternate pathway. Data presented in this paper demonstrate that a subpopulation of human lymphocytes adheries to C3 coated Sephadex particles. The adhering subpopulation of lymphocytes is identical with or overlaps extensively with lymphocytes bearing easily detectable membrane Ig. This phenomenon can be used for the fractionation of lymphocyte subpopulations. After a single passage of tonsillar lymphocytes through C3 coated Sephadex bead columns, no membrane Ig positive lymphocytes were detectable in the effluent. 97-100% of effluent cells formed rosettes with untreated sheep red blood cells, thus resembling pure T cells. Apart from that, the use of C3 coated dextran gel particles allows also the recovery of adherent cells after dextranase digestion of the dextran gel particles. An investigation of retained cells showed enrichment of B cells, but also T cells were demonstrable. The retention of these T cells is probably due to nonspecific interactions.     

In vitro generation of NK-like activity in human PBL co-cultured with fresh T leukaemia cells

Generation of cytotoxic activity attributable to natural killer (NK)-like effector cells, against the NK-sensitive erythroleukaemia cell line K562 was estimated in peripheral blood lymphocytes (PBL) from normal donors, following stimulation with fresh leukaemic cells (FLC) of T-origin. One of 9 FLC induced anti-K562 cytotoxicity which was considerably higher than that of unstimulated control PBL. Cells derived from this patient possessed acid phosphatase (AP) and TdT activity. The reacted with the monoclonal antibodies RFA1 (directed against early and mature T cells) and A10 (specific for lymphohaemopoietic precursors). However, the cells did not form E rosettes, nor did they react with the monoclonal antibodies OKT3, OKT4, OKT8 and OKT11 to human T cell surface antigens. OKT6, which recognizes 70% of thymic lymphocytes was also negative. The high stimulatory activity of these cells may be related to the expression of surface determinants characteristic of early stages of the T-cell pathway, detected only by the monoclonal antibody RFA1.     

E-receptors mediate the attachment of activated T-lymphocytes to human red blood cells: evidence from studies with monoclonal antibodies and simple sugars

Lectin-stimulated human T-lymphocytes display the capacity of forming stable E-rosettes with sheep red blood cells (SRBC) and of forming rosettes with allogeneic and autologous erythrocytes (RBC). Fractionation experiments indicated that the same subpopulation of lymphocytes displays both of these characteristics. Exposure of T-lymphocytes stimulated with concanavalin A (Con A) or phytohemagglutinin (PHA) to T11 monoclonal antibody (directed against the E-receptor) completely blocked the formation of rosettes with human RBC. Monoclonal antibodies to other T-cell surface markers (Leu 4, T4, T8 and OKT9) had no such effect. The formation of human RBC rosettes and stable E rosettes by PBL stimulated for 2 to 8 days by Con A was inhibited to a similar extent by dilutions of T11 monoclonal antibody. Lectin-stimulated PBL were exposed to various sugars to determine their effect on human RBC rosette formation. Rosette formation was inhibited over 80% with 2-deoxy-D-glucose and by 50-70% with D-mannose, while the other sugars tested had either less effect or none. The present findings support the concept that E-receptors on the surface of human T lymphocytes play a role in the recognition of carbohydrate containing "self" structures.     

Lymphocytes in Patients with Variable Immunodeficiency and Panhypogammaglobulinemia: EVALUATION OF B AND T CELL SURFACE MARKERS AND A PROPOSED CLASSIFICATION

Peripheral blood lymphocytes from 15 patients with variable immunodeficiency and severe panhypogammaglobulinemia were evaluated for B and T cell surface markers. B cells were enumerated by immunofluorescent detection of both surface immunoglobulin (Ig) and the ability to bind aggregated Ig complexes. T cells were identified by their ability to form nonimmune rosettes with sheep red blood cells. Four distinct patterns were observed which were designated types I-IV. Type I: six patients had normal percentages (8.5-19.0%) of Ig-bearing B lymphocytes. Type II: four patients were observed to have B lymphocytes (4.5-15.0%) which lacked fluorescence-detectable surface Ig. Type III: the peripheral blood of these four patients contained a subpopulation (11.3-20.0%) of lymphocytes which apparently lacked both B and T cell markers ("null" cells). Type IV: one patient's blood was characterized by a subpopulation (18.0-22.0%) of lymphocytes which bore both B and T cell markers. Patients of each type had some clinical features in common. It is concluded that evaluation of lymphocyte surface markers provides a means of separating patients with variable immunodeficiency and panhypogammaglobulinemia into distinct groups which appear to differ in the nature of their fundamental defect.     

Autologous rosette-forming T cells regulate responses of T cells. Phenotypic and functional analysis of suppressor cells generated from autologous rosette-forming T cells after autologous mixed lymphocyte reactions.

An average of 5--9% of human peripheral blood of T lymphocytes from rosettes with autologous erythrocytes (ARFT). This population responded only slightly against autologous and allogeneic non-T cells. In contrast, T cells that did not form rosettes with autologous erythrocytes (NRFT) proliferated to a greater degree in auto- and allogeneic mixed lymphocyte reactions (MLR) and also in reactions to trinitrophenyl (TNP) modified autologous non-T cells (TNP-auto-MLR) as compared with ARFT or unfractionated T cells. The ARFT populations could suppress the increased allogeneic (allo)MLR and TNP-auto-MLR of NRFT when the ARFT were added to the NRFT at the beginning of the cultures. Fluorescence-activated cell-sorter (FACS) analysis of these freshly obtained T cell fractions using monoclonal antibodies to subpopulations of T cells did not demonstrate any selective gain or less of T cell subsets in the ARFT and NRFT as compared with unfractionated T cells. But when each T cell fraction was cultured separately for a week in the presence of autologous non-T cells (auto-MLR) and the cells were again analyzed by fluorescence-activated cell sorter, there was an increase in OKT8-positive cells (suppressor/cytotoxic subset) only in the ARFT fraction. The above findings strongly suggest that suppressor T cells are generated from the ARFT fraction during an auto-MLR, these may then regulate the responses on NRFT.     

Immunological features in chronic lymphocytic leukaemia (CLL) of T cell origin.

Peripheral blood lymphocytes from a patient with chronic lymphocytic leukaemia of T cell origin were studied. The thymus derived nature of these lymphocytes was confirmed by surface markers, mitogen cultures, mixed lymphocyte reaction, cytotoxicity studies, and cytochemical stains. This case is notable for several clinical and laboratory findings. Among these, the benign clinical course, the reduced rate of serum immunoglobulins, the elevated number of active E rosettes, the increased PHA-induced response to low mitogen doses, the absence of PHA mediated cellular cytotoxicity, and the thy-like positivity to ANAE should be pointed out. Emphasis should be placed, however, on the loss of stimulatory ability in MLR. This last feature supports the hypothesis that these cells proliferate as a clone.     

Rosette formation between murine lymphocytes and erythrocytes. A new locus in the H-2 region

More than 5% of murine splenic lymphocytes form rosettes with syngeneic erythrocytes. This property was maximally expressed when the lymphocytes were cultured for 24 h before rosetting. About 70% of the rosetting lymphocytes were B cells and 30% were T cells on the basis of surface immunoglobulin and the Thy-1-antigen. Capping surface immunoglobulin had no effect on the capacity of lymphocytes to form rosettes, indicating that the receptor in question was not immunoglobulin. The capacity of lymphocytes to form rosettes with erythrocytes from other strains of mice was H-2 restricted. Extensive pairings of congenic and recombinant strains as donors of lymphocytes and erythrocytes showed that none of the known loci within the H-2 region-controlled rosetting. The involvement of regions on chromosome 17, telomeric or centromeric to H-2, was also excluded. The data were only compatible with the conclusion that this form of self-recognition is associated with a new locus (or loci) mapping between H-2G and H-2D.