A rauscher-virus-induced t-lymphocyte cell line. Induction of differentiation under influence of dimethylsulfoxide and phorbolesters (original) (raw)
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Leukemia Research, 1987
Rauscher murinc leukemia virus-induced erythroid, lymphoid and myeloid cell linc8 were characterized with respect to the expression of differentiation antigens using a panel of monoclonal antibodies. The expression of differentiation antigens was measured in a two-step micro ELISA procedure. The cell lines exprcss a number of early markers and lack a number of mature markers characteristic for the respective cell lineages. Moreover they express a number of surface markers which are not or only rarely found on their normal counterparts. Thc expression of differentiation antigens indicates that the cell lines investigated are arrested in an immature stage of differentiation. This observation implies that the Rauscher virus preferentially transforms early hemopoietic cells.
Developmental Biology of T Lymphocytes
Annals of the New York Academy of Sciences, 1988
Immunology has experienced long-lasting debates on issues that could not be addressed experimentally at the time. For instance the debate over the source of antibody diversity was only closed after several decades by the unforeseen discoveries of Tonegawa' and Weigert et al.2 A similarly long-lasting debate concerns the self-nonself discrimination of the immune system or immunologic tolerance. Hypotheses were put forward at the end of the first half of this century and new ones have been added ever since. Two main streams of thought can be distinguished. One is best described as the deletion model and is associated with the names of Burnet, Lederberg, Cohn, and others. One particular version states that lymphocytes during their differentiation from stem cells pass through a stage in which contact with antigen results in lymphocyte death rather than lymphocyte activation. Other views-perhaps more compatible with Jerne's thinking-propose that autoreactive lymphocytes are not deleted but exist in a suppressed state within the network of the immune system. In early 1985, we decided to test the clonal deletion hypothesis in T-cell receptor transgenic mice. The idea was to construct mice that, in a large fraction of T cells, expressed a receptor with specificity for self antigens and to chose a receptor that could easily be detected by a monoclonal antiidiotypic antibody. Ideally, this experimental design should allow study of the fate of T cells expressing this receptor in mice lacking or expressing the relevant antigen. We therefore chose to isolate the a,@ T-cell receptor genes from a cytotoxic T-cell clone specific for the male antigen in the context of H-Db and to analyze female and male offspring from a,@ T-cell receptor transgenic mice. EXPRESSION OF a AND B T-CELL RECEPTOR GENES IN TRANSGENIC MICE Genomic DNA containing the functionally rearranged a and p genes from the male-specific cytotoxic T-cell clone B6.2.16 were injected in fertilized eggs. Initially, bOn sabbatical leave at
Journal of Experimental Medicine, 1990
The study of prethymic stages of T cell development has been limited because specific markers for mouse pro-T lymphocytes were not available. We developed a panel of rat monoclonal antibodies (mAbs) that bind to our pro-T lymphocyte clones obtained from bone marrow of young adult mice and the thymus of 14-d-old embryos. The mAbs, called Joro 30-8, Joro 37-5, and Joro 75, were found to bind to all pro-T clones tested but not to cell lines representing later stages of T cell development, B lymphocyte, or myeloid lineages. We determined the frequency and tissue distribution in normal and immunodeficient mouse strains as well as the ontogeny in liver and thymus of cells positive for these mAbs. The results were consistent with the pattern of reactivity observed with cell lines. We isolated Joro 30-8+, Joro 37-5 +, and Joro 75+ bone marrow cells by cell sorter and found that: (a) phenotypically, they are Thy-1+, CD4-, CD8-, CD3-, B-220-, IgM-, F4/80-, and PgP-1+; (b) they grew in response to the combination of interleukin 3 (IIr3) + IIr4 or IIr3 + IIr4 + IIr6; and (c) Joro 37-5+ and Joro 75+ marrow cells gave rise to mature T lymphocytes but not to B lymphocytes, while Joro 30-8+ marrow cells generated both T and B lymphocytes after 8-12 wk of transfer into severe combined immunodeficient (Scid) mice. In normal mice subjected to 600 rad of irradiation to induce a wave of thymus recolonization, we found by flow fluorocytometry analysis that Joro+ cells entered the thymus 2 d after irradiation, expanded during the next 4 d, and underwent further differentiation, and from day 8 up to day 21, post-irradiation Joro+ cells were no longer detectable in the thymuses. Immunohistochemical analysis of normal thymus shows the presence of very few Joro 30-8+, Joro 37-5+, and Joro 75+ lymphoid cells in the subcapsular area and outer cortex but not in the medulla. The kinetic analysis of tissue sections from thymuses at various days post-irradiation suggests that Joro+ cells enter the thymus via blood vessels through the subcapsular and outer cortex areas; subsequently, these cells seem to migrate to the inner cortex without reaching the medulla, and give rise to Joro thymocytes. We conclude that in bone marrow of young mice, cells expressing Joro 30-8 are very early hematopoietic precursor cells that are either committed to the lymphocyte pathway or are even less differentiated pluripotent cells. Commitment to T cell differentiation appears to be associated with additional expression of Joro 37-5 and Joro 75 .
Ly phenotype and mechanism of action of mouse neonatal suppressor T cells
The Journal of …, 1977
Ly PHENOTYPE AND MECHANISM OF ACTION OF SUPPRESSOR T CELLS DBA/2N males were used to breed the (CBA/N x DBA/2)FI male (defective) and FI female (normal) mice used in some experiments. Culture Conditions. Single cell suspensions of mouse thymus or spleen were prepared and cultured by using a microculture adaptation of the Mishell and Dutton method as previously described (19). Briefly, a total of 1 x 108 cells were cultured in each well of a flat-bottomed microtiter plate (Falcon 3040, Falcon Plastics, Division of BioQuest, Oxnard, Calif.) for periods of 3-4 days in a humidified 5% CO2-95% air atmosphere. Antibody formation against the trinitrophenyl (TNP) determinant was stimulated by the addition of TNPss aminoethylcarbamylmethyl (aecm)s3 Ficoll, TNP-lipopolysaccharide (TNP-LPS), or TNP.Brucella abortus. These antigens appear to be thymic-independent in the newborn as well as the adult mouse (D. E. Mosier. Manuscript in preparation) and their preparation has been described previously (20, 21). Direct plaque-forming cells (PFC) were assayed against TNP-modified sheep erythrocytes prepared as described by Rittenberg and Pratt (22). All groups were assayed in triplicate and the data logarithmically transformed to obtain the geometric mean. Data are expressed as the antilog of the geometric mean ×/-the relative standard error (the antilog of the log standard error). Proliferation assays were performed as described previously. Virus Injection. Mouse thymic virus was obtained by serial passage from a stock maintained
Immunology, 1977
An investigation has been made of the development of various T cell functions in lethally irradiated mice reconstituted with anti-0 treated spleen or bone marrow cells. Evidence is presented to show that both organs contain a post-thymic precursor pool able to regenerate by 15 days limited T cell responses in thymectomized recipients. A prethymic pool also exists in each organ able to regenerate, at a later date, first a suppressor T cell population and probably later, mature functional T cells involved in helper functions and cell mediated lympholysis. The spleen is apparently a better source of precursors of the suppressor cells than bone marrow, while a poorer source of precursors of the other T cell functions. All T cell functions investigated apparently first appear in large cells which undergo a reversion to small cells without necessarily maturing to their full potential reactivity. By following the kinetics of appearance of T cell functions, and the physical parameters of th...
Comprehensive Phenotyping of Peripheral Blood T Lymphocytes in Healthy Mice
Cytometry Part A, 2020
T lymphocytes play a central role in antigen-specific immune responses. They modulate the function of different immune cells both through a direct contact (receptor binding) and through the secretion of cytokines. At the same time, they are deeply involved in the direct killing of aberrant target cells. T lymphocytes derive from a bone marrow precursor that migrates in the thymus where the main differentiation steps take place. Mature CD4 and CD8 single-positive cells, then, leave the thymus to reach the secondary lymphoid organs. T cell subsets and their maturation steps can be identified mainly based on the expression of extracellular markers, intracellular transcription factors and cytokine production profiles. In this review, we report, from a cytometric point of view, an overview of the most important T cell subpopulations and their differentiation state. T cell identity card: development and functions T lymphocytes belong to the adaptive immune system and play an integral role in executing and controlling humoral and cell-mediated immune responses. T lymphocytes are distinguished from the other lymphocytes (B and NK cells) by the expression of a T-cell receptor (TCR) on their cell surface (1). T lymphocytes derive from a bone marrow precursor (T cell precursor: TCP) which, in the thymus gland, starts its differentiation through four stages, all characterized by a CD4-CD8phenotype, the so called Double Negative (DN) cells. DNs differentiation stages (DN1-DN4) are distinguishable based on their differential expression of CD44, CD25 and CD117 (2). (Figure 1). Expression of the pre-TCR at DN3 leads to DN4 stage, followed first by expression of CD8 (ISP: Intermediate Single Positive) and, right after, of CD4, thus progressing to the CD4 + CD8 + Double Positive (DP) stage (3). A positive selection, then, leads to a commitment toward CD4 or CD8 T cell lineage. Mature CD4 and CD8 single-positive cells leave the thymus and, through the peripheral circulation, reach the secondary lymphoid organs as naïve T cells, each one with a unique antigen specificity (Figure 1). In the thymus, a similar differentiation pathway takes to a DN expressing TCR + which will give rise to mature T lymphocytes, as clearly described by Buus et al in OMIP-057 (4). Moreover, also NKT cells, which express -TCR coupled with a CD1d glycolipid and NKp46, derive from the same precursors (5) (Figure 1). Upon specific antigen encounter, both CD4 + and CD8 + T cells are activated and differentiate into a specific subtype, which expand, to fulfil its unique effector role, for example by migrating into various tissues and organs. The immunological memory will be guaranteed by some of the antigenactivated T cells which differentiate into various memory T cell subtypes.